Official Journal of the Romanian Society of Physiological Sciences

CHIEF EDITOR
CO-CHIEF EDITORSASSOCIATE EDITORSEXECUTIVE EDITORSFRANCISC SCHNEIDERIOANA SISKACARMEN TATUMIHAI NECHIFORSORIN RIGA
FLORINA bOjIN
GAbRIELA TANASIEDACIANA NISTORCALIN MUNTEANEDITORIAL BOARD
ARDELEAN AURELBADIU GHEORGHEBĂDĂRĂU ANCA
BENEDEK GYÖRGYBENGA GHEORGHEBUNU CARMEN
COJOCARU MANOLECUPARENCU BARBUCONSTANTIN NICOLAE
HAULICĂ ION
IANCĂU MARIAMIHALAŞ GEORGETA
MUNTEAN DANINAMUREŞAN ADRIANANESTIANU VALERIUOPREA TUDOR(Arad)(Constanţa)(Bucureşti)(Szeged)(Cluj)(Timişoara)(Bucureşti)(Oradea)(Bucureşti)(Iaşi)(Craiova)(Timişoara)(Timişoara)(Cluj)(Craiova)(New Mexico)PĂUNESCU VIRGILPETROIU ANA
POPESCU LAURENŢIU
RÁCZ OLIVERRIGA DANSABĂU MARIUS
SIMIONESCU MAIA
SIMON ZENOSAULEA I. AURELSWYNGHEDAUW BERNARDTANGUAY M. ROBERTTATU FABIAN ROMULUS
VLAD AURELIANVOICU VICTORZĂGREAN LEON

(Timişoara)(Timişoara)(Bucureşti)(Košice)(Bucureşti)(Tg. Mureş)(Bucureşti)(Timişoara)(Chişinău)(Paris)(Canada)(Timişoara)(Timişoara)(Bucureşti)(Bucureşti)AccREDITED By cNcSIS - B+cATEgORy - cODE 240
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Fiziologia(80)‑ Physiology 2010supplement
2013.24.4
2014.24.2(82)
 Fiziologia- Physiology

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11

Official Journal of the Romanian Society of Physiological Sciences

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Fiziologia- Physiology 2014.24.2
2013.24.4(82)(80)
Fiziologia‑ Physiology 2010supplement

STNETNOC
4................................................................................... recnaC fo skramllaH eht gniylrednU erucsbO revewoH ,romuT fo stnatibahnI – stsalborbiF detaicossA-recnaC :weiveR .1
C ucsetianaP ,V ucsenuaP ,G eisanaT ,M acieD ,F nijoB ,L tanreC
31.................................... sitirhtrA decudnI-tnavujdA fo ledoM taR a ni ecnalaB tnadixoitnA-tnadixO eht no sdicA yttaF detarutasnuyloP 3-agemO dna nituR fo stceﬀE ehT .2
A naşeruM ,D dalV ,R navodloM ,D utsopA ,C uicacoS ,A notraM ,CI şihC ,D uratlaB

CONTENTS

81.................................................................. staR ni stceﬀE evitcetorpoidraC trexE dna aixopyH cirabopyH fo stceﬀE eht esreveR nasotihC dna murabraB muicyL ,nitecreuQ .3
P afloB ,G azroB ,D dalV ,R navodloM ,A notraM ,A naşeruM ,CI şihC ,A icivortimuD
32........................................................................................................................................ yhtapolahpecnE citapeH fo ygolohtapoisyhP eht ni lotisonI-oyM fo ecnatropmI ehT .4
IA uărădăB ,AD noI ,R aecocapaP ,C ienroiC ,M aenuB ,M răzaL ,C uaehcS

CONTENTS

62............................................................................................................................................ ksiR ralucsavoidraC fo noitacﬁitartS eht ni ecnacﬁingiS citsongorP :erusserP esluP .5
S șacnaM ,A rimogarD ,G ucsenoI ,D aţâbiâB
13....................................................................................................................... skramdnaL lacimehcotsihonummI dnA lacigolotsiH - amorbﬁoignA laegnyrahposaN elinevuJ .6
D tahroH ,S aeblutoC ,M atumoD ,MR narihG ,G ucsenavoI ,FD aediG

1. TheEffectsof Cornus Mas, Prunella
Vulgarisand Crataegus
Monogyna
Extracts
on Viabilityof Neonatal
Rat Cardiomyocytes......................................................4
Review:
Cancer-AssociatedFibroblasts– Inhabitantsof Tumor,
However Plant
ObscureUnderlying
the Hallmarksof Cancer...................................................................................4Bădălică-Petrescu
M, Tudor A, Stoichescu-Hogea
G, Therapy
Socaciu C,..............................................................................................................................................................4Drăgan S
1. Review: Metformin-From
Diabetes to Cancer
53........................................................................................................................................................................................................... sitilucsaV yranomluP eht otni thgisnI weN .7
MI uracojoC ,DC eibarV ,E usuR ,M uracojoC
83............................................................................................................................................................................. ?gnihT laeR a tI sI ,yhtaporueN citpO detaicossA enoradoimA .8
I goloZ ,M naetnuM ,N ucseluciN

Cernat L, Bojin F, Deica M, Tanasie G, Paunescu V, Panaitescu C
Deica
M, Sima R,Laparoscopic
Cernat L, Bojin
F, Tatu C,ofTanasie
G,Panaitescu
V Liver Metastases – Initial Experience.........................................................................10
2.
Simultaneous
Approach
Colorectal
Cancer andC,ofPaunescu
Synchronous
SNIRPUC

4................................................................................................................................................. recnac ni uirotinﬁed lor uc elaromut etnenopmoC – elaromutirep eletsalborbiF .1
C ucsetianaP ,V ucsenuaP ,G eisanaT ,M acieD ,F nijoB ,L tanreC

Brebu
S, LazarTarța C, DuțaC, Lazar F Fatty Acids on the Oxidant-Antioxidant Balance in a Rat Model of Adjuvant-Induced Arthritis ....................................13
2. The D,
EﬀPantea
ects of Rutin
andC,Omega-3
Polyunsaturated
31............................................... tnavujda ed ăsudni ătirtra uc inaloboş al iţnadixoitna-iţnadixo ieţnalab arpusa 3-agemO iţarutasenilop işarg rolizica la iş iulunituR lutcefE .2
A naşeruM ,D dalV ,R navodloM ,D utsopA ,C uicacoS ,A notraM ,CI şihC ,D uratlaB

2. Adipose-Tissue Derived Stem Cells: State of the Art Tissue Engineering .................................................................................................................................1181............................ naloboş ed ledom nu-rtnî rotcetorpoidrac snupsăr nu ăticrexe iş erabopih ieixopih eletcefe ăzaeracartnoc lunasotihC iş murabraB muicyL ,lunitecreuQ .3
P afloB ,G azroB ,D dalV ,R navodloM ,A notraM ,A naşeruM ,CI şihC ,A icivortimuD

Baltaru
D, Chiş IC, Marton
A, Socaciu
C, Apostu
D, Moldovan
R, Vlad D, Mureşan A
3.
Patient-Perceived
Qualityof Medicalin Timis
HurmuzM, Bojin F, Miu
C, Ungurean
C,Services
Tatu C, Ionac
M,County
Tatu F Public Health Hospitals.........................................................................................................................14
Bogdan C, Bucur A
32............................................................................................................................................................... ecitapeh ieitapolafecne aigolotapoizﬁ nî iululotizoni-oim aţnatropmI .4
IA uărădăB ,AD noI ,R aecocapaP ,C ienroiC ,M aenuB ,M răzaL ,C uaehcS
62................................................................................................................................... ralucsavoidrac iulucsir aeracﬁitarts urtnep acitsongorp aitacﬁinmes :iuluslup aenuiserP .5
S șacnaM ,A rimogarD ,G ucsenoI ,D aţâbiâB

3. Long-Term
Quercetin,
Lyciumand Chitosan
ReverseEﬀectsBehaviorsof Hypobaric
Hypoxia
and Diseases..........................................................................................................20
Exert
CardioprotectiveEﬀects in Rats ..................................................................18
4.
EffectsBarbarumof Simvastatin
TherapyEffects
on Chronic
Inflammatoryand3.
Human Recombinant
Inteferon-AlphaontheActivein Autoimmune
theRat Forced
Swimming
Test.....................................................................................17
Ciurariu
Dumitraşcu
V RI,A,Baca
Dumitrovici
A, Chiş IC,MureşanMartonA, Moldovan
Zahiu
D,E,Goganau
I, Braga
L, Zagrean
L R, Vlad D, Borza G, Bolfa P
13...................................................................................................................................... ecimihcotsihonumi iş ecigolotsih icitsirecarac – linevuj naignirafosan lumorbﬁoignA .6
D tahroH ,S aeblutoC ,M atumoD ,MR narihG ,G ucsenavoI ,FD aediG
53........................................................................................................................................................................................................................ ăranomlup atilucsav nî iţătilautcA .7
MI uracojoC ,DC eibarV ,E usuR ,M uracojoC

83................................................................................................................................................................ ?etatilaer o etse – anoradoima uc iulutnematart ătaicosa aitaporueN .8
I goloZ ,M naetnuM ,N ucseluciN

5. Prospective Study of Laparoscopic Great Curvature Plication – Effects on Weight Loss and Hunger Hormone Levels............................................................................23
Importance
Myo-Inositol
inD,thePhysiopathology
ofofHepatic
Encephalopathy........................................................................................................................................23
4. TheCorticalHeart
Rate
ChangesMarkers
Stimulation Response
during Chloral Hydrate Anesthesia ...................................................22
Dobrescu
A,Activity
VerdeșofG,andTarța
C, Brebu
StoicaL,asLazăr
C, Duță
CNoxious3

ygoloisyhP - aigoloiziF  )77( 1.32.3102

PavelCălinM,A,CiorneiRădulescu
Am, RoscaAe,
BragaRi, Zahiu
ScheauB,C,DăneasaLazăr M,A,Bunea
C, Papacocea
R, Ion DA,Bădărău
AI D, Zăgrean L

6. Telomere Lenght Changes in Alzheimer Disease....................................................................................................................................................................................27
Groza S, Anghel S, Cristea M, Tatu C, Tanasie G, Panaitescu C, Gavriliuc O, Paunescu V, Bojin F

5.
Arguments
forPrognostic
Using Thrombelastography
in thecation
StudyofofCardiovascularHemostasis Comparedto Other Laboratory Tests .........................................................................26
5. Pulse
Pressure:
Signiﬁcance in the Stratiﬁ
Risk ............................................................................................................................................26

7.
Gadolinium-BasedContrast Agents:
Safety,N,Efficacy
And
Potential AC,
AdverseEvents.
Oniceanu
I, Adumitresi
Stefanescu
Chirica
R ..............................................................................................................................31
Bâibâţa
D, FM,Ion
Ionescu
A,C,Radulescu
Mancaș
S Farcas C,
Minca
DM,
Enache G,
EL,Dragomir
Tatu F, Tatu
C

6. Regular
Patient Related Prognostic
and Therapeutic
Strategy inOutcomes
Larynx Squamous
Cell Carcinoma.........................................................................................31
8.
Activity Angioﬁ
AndFactors
Ablation
Lone Atrial And
Fibrillation
In Young
Patients.............................................................................................................38
6. Juvenile PhysicalNasopharyngeal
broma Of- Histological
Immunohistochemical
Landmarks.......................................................................................................................31
ProdeaAG,
M,Ivănică
HorhatG,D,Balica
N, Boruga
O, Poenaru
Negru
Andronache
M, Bucur
A, NegruMS, Ivănică A, Luca CT, Ionac A, Petrescu L, Drăgulescu SI

Gidea DF, Iovanescu G, Ghiran RM, Domuta M, Cotulbea S, Horhat D

9. BCL-2 Expression as a Negative Prognostic Factor in Hodgkin Lymphoma.............................................................................................................................................43
7. In VitroCharacterisation
Solid Tumors..............................................................................................................35
Potre-Oncu
O, Ionita I, Ionita of
M,Tumor-AssociatedFibroblastsfromDifferent
Calamar D, Sorica C, Ionita H

7. New L,Insightthe Pulmonary
Vasculitis...........................................................................................................................................................................................................35
CernatBojininto
F, Deica
M, Anastasiu
D, Tanasie
G, Ilina R, Paunescu V, Panaitescu V

10.
OtitisM,
Media
Effusion
In Children
Cojocaru
RusuWith
E, Vrabie
CD, Cojocaru
IM – A Clinical Trial.......................................................................................................................................................................48
Marin
K, Vintila R, andPopescu
Marin AH,
ProdeaM, Poenaru
M
8. Comorbidities
RiskR,Factors
in Heart
Failurewith Preserved
Ejection Fraction...................................................................................................................39

MavreaA, BucurA, Tomescu
MNeuropathy,
8. Amiodarone
Associated
It a Real Thing?
.............................................................................................................................................................................38
11.
MultiplemyelomaandOptic
stent
restenosis inIsobstructive
coronary
artery disease...................................................................................................................................52
Valcovici
NiculescuM,
N, Pascalau
Muntean L,M,Iancu
ZologOICUPRINSCUPRINSCUPRINS1.
extractelorde plante
din Cornus mas,
Prunellasi Crataegus
monogyna asupra viabilitatii cardiomiocitelor neonatale de sobolan .............................4
1.
Metformin:dela diabetla terapiaantitumorală
.......................................................................................................................................................................4
1. Efectele
Fibroblastele
peritumorale
Componente
tumoralecu rolvulgaris
nitoriuBădălică-Petrescu
M, Tudor A,–Stoichescu-Hogea
G, SocaciuC,deﬁDrăganS in cancer .................................................................................................................................................4

Deica
SimaF,R,Deica
CernatBojin G,F, Tatu
C, Tanasie
G,Panaitescu
C, Paunescu V
CernatM,
L, Bojin
M, L,Tanasie
Paunescu
V, Panaitescu
C

2. Rezectia laparoscopica simultana a cancerului colorectal si a metastazelor hepatice sincrone – studii initiale.....................................................................................10
Brebu
D, Pantea
Lazar C, Tarța
C, Duța C, Lazaractualităţiîningineriatisulară
F
2. Celulele
stemS,derivate
din ţesutuladipos:.................................................................................................................................11

2. Efectul Rutinului şi al acizilor graşi polinesaturaţi Omega-3 asupra balanţei oxidanţi-antioxidanţi la şobolani cu artrită indusă de adjuvant ...............................................13

HurmuzM, Bojin F, Miureferitoare
C, Ungurean C, Tatu C,serviciilor
Ionac M, Tatu F in spitalele judetului Timis.................................................................................................................143.
Perceptia
Baltaru
D, Chişpacientilor
IC, Marton A, SocaciulaC,calitatea
Apostu D, Moldovanmedicale
R, Vlad D, Mureşan A
Bogdan C, Bucur A

3. Efecteleinterferonului-alfa recombinant umanasupracomportamentelor active la testulînotuluiforţat, la şobolan ...............................................................17

4.
pe termenlungale
cucontracarează
in afectiunile
inflamatorii
autoimune.
.........................................................................................20
3. Efectele
Quercetinul,
LyciumBarbarumşi Chitosanul
hipoxiei
hipobare şicronice
exercităsiun
răspuns cardioprotector
într-un model de şobolan ............................18
Zahiu
D, Goganau
I, Braga
RI, tratamentului
Baca
L, Zagrean
LSimvastatin efectele
Ciurariu
E, Dumitraşcu
V
Dumitrovici
A, Chiş IC, Mureşan
A, Marton A, Moldovan R, Vlad D, Borza G, Bolfa P

4. Studiu
Modificărileactivităţii
corticaleşialeritmuluicardiac
ca markeri
la stimulareanociceptivăîntimpulanestezieicucloralhidrat
5.
prospectiv privind
plicaturarea
laparoscopica a marii
curburiai–răspunsuluiefectele asuprapierderii in greutate si a nivelului hormonilor de foame ...............................22...............................23
Dobrescu
A, Verdeș
TarțaA,C, Rădulescu
Brebu
D, StoicaLazărC, Duță
4. Importanţa
mio-inozitoluluiîn ﬁziopatologia
encefalopatiei
PavelB, Dăneasa
A,G,Călin
Am,L,RoscaAe,
BragaChepatice
Ri, Zahiu...............................................................................................................................................................23D, Zăgrean L

ScheauC, Lazărlungimii
M, Buneatelomerelor
M, Ciornei C,inPapacocea
R, Ion DA, Bădărău AI
6.
Modificarileboala Alzheimer...........................................................................................................................................................................27
5. Argumentepentruutilizareatrombelastografieiînstudiulhemostazeicomparativ
cuFalteteste de laborator .............................................................................26
Groza
S, Anghel S, Cristea M, Tatu C, Tanasie G, Panaitescu C, Gavriliuc O, Paunescu V, Bojin
Oniceanu FM,Ion I, Adumitresi C,Radulescu N, Farcas C, Stefanescu AC, Chirica R

5. Agenți
Presiunea
semniﬁprognostica
pentru stratiﬁ
carea riscului...................................................................................................................................26
7.
de pulsului:
contrast pebazăcatia
de gadoliniu:
siguranță,eficacitateși efectecardiovascularadverse potențiale.
................................................................................................................31
BâibâţaDM,D, Ionescu
A, MancașS
Minca
Enache G,
EL,Dragomir
Tatu F, TatuC

6. Factori de prognostic legaţi de pacientşistrategiiterapeutice în carcinomullaringian cu celulescuamoase .............................................................................31

8.
Activitatea
fizicaD,Balica
regulataN,si Boruga
rezultatele
ablatiei fibrilatiei
atriale la pacientii tineri...............................................................................................................................38
ProdeaM, Horhat
O, Poenaru
M
6. Angioﬁ– caraceristici
şi imunohistochimice......................................................................................................................................31
Negru
AG,bromulIvănicănasofaringianG, Andronachejuvenil
M, Bucur
A, Negru S,histologiceIvănică A, Luca
CT, Ionac A, Petrescu
L, Drăgulescu SI

Gidea DF, Iovanescu G, Ghiran RM, Domuta M, Cotulbea S, Horhat D

7. Expresia
Caracterizare
fibroblastelorperi-tumoraleizolatedin diferite tumori solide ............................................................................................................35
9.
BCL-2incavitro
factora deprognostic negativ
in limfomul
Hodgkin....................................................................................................................................................43
Cernat L, Bojin
F, Deica
M, Anastasiu
IlinaHR, Paunescu V, Panaitescu V
Potre-Oncu
O, Ionita
I, Ionita
M, CalamarD,D,Tanasie
Sorica C,G,Ionita

7. Actualităţi în vasculita pulmonară ........................................................................................................................................................................................................................35

10.
Otita medie
la copii–riscîninsuficienţacardiacă
studiuIMclinic........................................................................................................................................................................................................ 48
Cojocaru
M, Rususeroasa
E, Vrabie
CD, Cojocaru
8.
Comorbidităţişifactorii
MarinK, Vintila R, Popescu R,deMarin
AH, Prodea M, Poenaru M cu fracţie de ejecţieprezervată ..............................................................................................................39

Mavrea A, Bucur A, Tomescu M

11.
Mielomul multiplu
restenoza intrastentin boala –coronariana
obstructiva.....................................................................................................................................................
52
8. Neuropatia
asociată sitratamentului
cu amiodaronaeste o realitate?
................................................................................................................................................................38
Valcovici
M,
Pascalau
L,
Iancu
O
Niculescu N, Muntean M, Zolog I
2014.24.2(82) 
Fiziologia - Physiology
2013.24.4
2013.23.1 (80)(77) 
 Fiziologia - Physiology

333

THE EFFECTS OF CORNUS MAS, PRUNELLA VULGARIS AND
CRATAEGUS MONOGYNA PLANT EXTRACTS ON VIABILITY
OF NEONATAL RAT CARDIOMYOCYTESBĂDĂLICĂ-PETRESCU MARIUS1, TUDOR ANCA2*, STOICHESCU-HOGEA GHEORGHE3, SOCACIU CARMEN4, DRĂGAN SIMONA1
1
Department of Cardiology, University of Medicine and Pharmacy “Victor Babeș” Timișoara
2
Department of Medical Informatics, University of Medicine and Pharmacy “Victor Babeș” Timișoara 3City Hospital Timisoara
4
University of Agricultural Sciences and Veterinary Medicine, Faculty of Food Science and Technology, Cluj-Napoca

ABSTRACT

Aim: Our study aimed to evaluate the effects of three aqueous plant extracts rich in polyphenols (Crataegus monogyna, Cornus mas
and Prunella vulgaris) on cardiomyocytes taken from newborn rats, in normoxic conditions.
Materials and methods: Hearts from 270 newborn Wistar rats(1-2 days) were prelevated and introduced in ice cold phosphatebuffered saline solution (PBS). The ventricles were separated, minced, and introduced in trypsin solution(0,25%) for 25-30 minutes.
The resulted cell suspension was centrifuged for 15 minutes, at 2000 rpm and 4oC, then the infranatant was separated and mixed
with Dulbecco’s Modified Eagle Medium (DMEM). Several 96-well plates were prepared and introduced in the incubator for 72
hours(37oC, 95% O2, 5 %CO2). Using the Folin-Ciocâlteu method, we were able to determine the total polyphenol content of eachplant extract. Starting from the original stocks, we prepared six consecutive dillutions using bidistilled water for each plant extract.
The dillutions were used for treating the cells in normoxic conditions. The viability measurement was performed using FLUOstar
Optima plate reader, after calcein and propidium iodide-digitonine staining.
Results and conclusions: Only Cornus mas extract showed statistically significant improvement in cell viability versus the control
group. Crataegus monogyna and Prunella vulgaris extracts had no significant effect on cell viability.
Keywords: plant extracts, polyphenols, normoxia, viability

INTRODUCTION
For centuries, various plants have been used for preventing or treating diseases. The leaves and/or fruits of medicinal
plants proved to be rich in active constituents like polyphenols
or flavonoid glycosides, thus having a major impact on reducing
the incidence of chronic diseases(1).
As Burta et al.(2) showed, extracts of Crataegus species, also
known as “hawthorn”, due to their high content in flavonoids,
were used in traditional medicine for decades in preventing or
treating moderate heart diseases. Many studies were conductedby Ammon (3) to explain the relationship between the composition and the cardiotonic activity of Crataegus species extracts.
A number of 23 active constituents such as phenolic acids, anthocyanins and flavonoid glycosides were identified in Crataegus
monogyna fruits (4).Therefore, due to its high concentrations ofphenols, phenolic acids and proven in vitro antiradical activity,
the Crataegus monogyna extract was evaluated as possible
source of antioxidant phytochemicals (5).
Extracts obtained from Crataegus leaves and flowers using
ethyl acetate were found to have the highest phenolic content(343.54 mg of gallic acid equivalents/g) and the highest DDPH
radical scavenging activity. Worldwide known for its medicinal
uses, the phytopharmaceutical characteristics of Crataegus oxycantha leaves, flowers, berries and bark were recently rewied (6).
Cornus mas, also known as “Cornelian cherry”, is a medicinal

plant used for its cardiotonic and anticancer effects, as well as
for treating inflammation (7).
The chemical characterization of Cornus mas fruits showed
high amounts of phenolic and antioxidant compounds. Flavonoids, anthocyanins, vitamins E, C, thiamine (B1) and riboflavin(B2) were also found in its fruits (8). A recent study revealed the
cardioprotective effect of Cornus mas fruit extract against cardiotoxicity induced by carbon tetrachloride (CCl4) in a rat model (9).
The antioxidant activity of Cornus mas fruits was observed
during antioxidant assays in Turkey (10, 11), in Greece (12)and in other countries (13, 14). The DPPH scavenging activity
was also high and shown to be increased by salycilic acid treatment due to significant increase of the total phenols, flavonoids,
anthocyanins and ascorbic acid content (15).
The antioxidant capacity expressed as DPPH scavenging effect and the total phenolics expressed in gallic acid equivalents
in both Crataegus monogyna and Cornus mas extracts (mgGAE/100 ml) were demonstrated in another study (16).
Once used in traditional medicine from all around Europe andAsia for treating fever, wounds and throat infections (17), Prunella
vulgaris is now a rediscovered plant due to its potential uses inmedicine. Its compounds can be used as antibacterial, antibiotic
and antiseptic, as well as antipyretic, diuretic and hypotensive
agents (18, 19).

Received 17th of March 2014. Accepted 20th of May 2014. Address for correspondence: Tudor Anca, Department of Medical Informatics, “Victor Babes”
University of Medicine and Pharmacy Timisoara, Eftimie Murgu Square No. 2A, RO-300041, Timisoara, Romania, phone/fax: +40256490288; e-mail:atudor@umft.ro.

4

Fiziologia - Physiology  2014.24.2 (82)Using High Performance Liquid Chromatography (HPLC)
analysis, it was possible to identify the main compounds in
Prunella vulgaris i.e. phenols like quercetin, rutin, caffeic androsmarinic acid (20). The rosmarinic acid found in Prunella
vulgaris extract was correlated with the cardioprotective effectagainst oxidative stress induced in rats (21). Furthermore, theeffect on eNOS gene expression of the Prunella vulgaris extract
resulted in favourable influences on blood vessels. By acting
as an eNOS upregulating agent, the extract proved to have apromising cardioprotective potential (19).
Crataegus monogyna, Cornus mas and Prunella vulgaris are
three medicinal plants well known for their cardiotonic effect.
Rich in polyphenols, the antioxidant qualities of these plants isunquestionable.
Our study aimed to compare the effects of these plant extracts
on neonatal rat cardiomyocytes used in cell culture, while being
subjected to normoxic conditions.
MATERIALS AND METHODS
Aqueous plant extracts preparation
Only fresh leaves of Crataegus monogyna and Cornus mas
and inflorescences of Prunella vulgaris were harvested from
Cluj-Napoca Botanical Garden in June 2012. To prepare theextracts, 15 g of each plant were infused in 100 ml bi-distilledwater in a sonication bath for 30 minutes, then centrifuged at2000 rpm for 10 minutes. Before performing HPLC-DAD-ESI(+)-MS analysis, the supernatant was collected and filteredthrough a 0,45 µm filter.
Total polyphenols determination
Total phenolic content was determined using Folin-Ciocalteu
method, adapted by Singleton (1999). 1 ml filtered extract ofeach plant was mixed with 5 ml Folin reagent, homogenized,then 15 ml Natrium carbonate 7,5% were added to the mixture.
The absorbance of the solution was observed after 2 hours, at
λ= 750 nm. All results were expressed in milligrams of gallic acidequivalents (GAE)/100 ml extract.
The highest quantity of polyphenols was found in Cornus
mas extract (105,03 mg GAE/100 ml), followed by Prunella
vulgaris (39 mg GAE/100 ml) and Crataegus monogyna (31,5mg GAE/100 ml).
Cardiomyocyte isolation and culture
All the experiments were conducted in accordance with the
National Institutes of Health Guide for the Care and Use of Laboratory Animals (22) and were approved by the Ethics Committeeof University of Medicine Szeged, Hungary.
The isolation protocol was performed on 1-2 days newborn Wistar
rats, in accordance with a method described by Gorbe et al. (23).
Hearts from 270 newborn Wistar rats were excised and placedin ice-cold phosphate-buffered saline solution (PBS) pH 7.2. The
ventricles were separated from the atria, minced using forceps
and washed three times with sterile PBS to rinse away tissuedebris and erythrocytes.
For dissociation into single cells, the cut pieces of the ventricles were collected in 0.25% trypsin solution and introduced
in 37oC water bath for 30 minutes (24, 25). After the digestion
period, the cell suspension was centrifuged for 15 minutes, at
2000 rpm and 4oC.
2014.24.2 (82)  Fiziologia - Physiology

The supernatant was removed using 5 ml Pasteur pipettes.
The cell pellet was re-suspended in Dulbecco’s modified Eagle’s medium (DMEM I), which was supplemented with 1%
L-Glutamine, 1% Antibiotic and Antimycotic mixture and 10%
fetal bovine serum (FBS). The resulted cell suspension wasthen collected in Falcon tubes and diluted in 4-5 ml DMEM I.
We performed the pre-plating stage by adapting the method
described by Blondel et al. in 1971(24). Thereby, the nonmuscular cells were eliminated after the cell suspension wasplated onto 6-well plates at 37°C for 90 min.
After the pre-plating period finished, the suspension containing non-adherent myocytes was collected in Falcon tubes andagitated.
The cell counting was performed manually using a Burker
chamber and an inverted microscope. Only the round-shaped
medium-sized cells from 3 big squares were counted and theaverage was took into consideration for further calculations.
In order to obtain a 75-80% confluence, the cell suspension
was homogenized using a pipette and plated at a chosen density of 2x104/well on non-mobile 96-well plates. Each well was
supplemented with 100 μl DMEM I and the plates were gently
shaked horizontally and vertically for better distribution of thecells, before being placed into the incubator.
The plates were left for 24 hours in an incubator, being subjected to a humidified atmosphere at 37oC, 95% air and 5% CO2.
After the first incubation period finished, the DMEM I medium
was substituted with DMEM II (Dulbecco’s modified Eagle’s
medium with 1% fetal bovine serum (FBS), 1% L-glutamine and
1% Antibiotic-Antimycotic mixture).
Using the inverted microscope, the contractile activity of the
cardiomyocytes was checked at 24, 48 and 72 hours.
The normoxic experiments started 72 hours after plating.
Preparation of normoxic control solution
As described by X Li et al (26), to obtain the normoxic solution,
we used the following (Table I):
Table I. Normoxic solution. The MgCl2 saturated solution contains
0,3774 g/1 ml.

NORMOXIC SOLUTION
MW
mM
Miligrames/100
ml
NaCl
58.4
125
730
KCl
74.5
5.4
40.2
NaH2PO4.1 H2O 137.9
1.2
16.6
CaCl2. 2 H2O 174.02
1
17.4
MgCl2 x 6 H2O
95.2
0.5
12.6 UI(saturated solution)
Glucose
198.1
15
297.2
Taurine
125.1
5
62.6
Creatine mono- 149.1
2.5
37.2
hydrate
HEPES
238.3
20
476.6
BSA
0.1%
100

Grames/
1L
7.305
0.402
0.16550.174
126 UI
2.972
0.625
0.372
4.766

5

Before using the normoxic solution, the pH was set to 7.4.
During the experiments, a dilution of 1:1000 was obtained bymixing 1 µl bi-distilled water + 999 µl normoxic solution. This
served as a control solution, it’s effects on cardiomyocytes being
compared with the effects of the three plant extracts.
Preparation of the Cornus mas, Crataegus monogyna andPrunella vulgaris dillutions
As we have shown earlier in another study (27), after applying
the Folin-Ciocalteu method, the total polyphenols content foreach plant extract was determined (Table II):
Table II. Total polyphenols content for Cornus mas, Crataegus
monogyna and Prunella vulgaris

Extract sample
Cornus mas
Crataegus monogyna
Prunella vulgaris

Mg GAE/100 ml
105.0331.5
39Because of the different content in polyphenols, the initialextract stocks of all three plant extracts had to be standardized
to a desired concentration of 0,1 mg polyphenols/ml solution,using bi-distilled water.
Starting with the original stocks, we have obtained six consecutive dilutions for each extract, as follows: 0.01 mg polyphenols/
ml, 0.001 mg polyphenols/ml, 0.1 µg polyphenols/ml, 0.01 µg
polyphenols/ml, 0.001 µg polyphenols/ml and 0.0001 µg polyphenols/ml (Table III).
Table III. Cornus m., Crataegus m. and Prunella v. dilutions (mgpolyphenols/ml solution); pps= polyphenols
CO1
CO2
CO3
CO4
CO5
CO6
0.01
0.001 0.1 µg 0.01 µg 0.001 0.0001
mg
mg pps/ pps/ pps/ml µg pps/ µg pps/
pps/mlml
ml
ml
ml
CrataeCR1
CR2
CR3
CR4
CR5
CR6
gus mo- 0.01
0.001 0.1 µg 0.01 µg 0.001 0.0001
nogynamg
mg pps/ pps/ pps/ml µg pps/ µg pps/
dilupps/ml
ml
ml
ml
ml
tions
Prunella PR1
PR2
PR3
PR4
PR5
PR6
vulgaris 0.01
0.001 0.1 µg 0.01 µg 0.001 0.0001
dilumgmg pps/ pps/ pps/ml µg pps/ µg pps/
tions
pps/mlml
ml
ml
ml
Cornusmas dilutions

For preparing the dilutions, we used bi-distilled water obtainedin the laboratory using the Millipore Direct-Q3 water distiller.
Normoxic experiments
For the normoxic experiments, we used a protocol adapted
after Gorbe A. et al. (23) and X Li et al. (26) which involved treatment of the cells with the desired dilutions of the plant extracts
and incubation for 4 hours at 37oC, 5% CO2, 95% air. After the 4hours incubation, we changed the applied treatment with DMEM
I and we let the cells for another 2 hours, in the same conditions(37oC, 5% CO2, 95% air), in the incubator.
The protocol used for normoxic experiments is expressed in
6

Figure 1:

Fig.1. Working protocol for the normoxic experiments

The plating for each of the three plant extracts was performed
after a fixed scheme, as shown in Figure 2:
Row 1 Row 2 Row Row Row Row Row Row Row Row Row Row
3
4
5
6
7
8
9
10
11
12
DMEM Norm PE 1 PE PE PE 4 PE PE 6 Norm Norm DMEM No
I
ctrl+veh235
ctrl+ ctrl+I
cells
veh vehFig. 2. Plating example for 96-well plates. PE= plant extract dilution;
Norm ctrl+veh= normoxic solution + bi-distilled water (1:1000)

Treatment with Cornus mas extract. The six dilutions obtained(see Table No. 3) were used for treating six rows of cells oneach 96-well plate (from row 3 to row 8). We used 100 µl/well,at a dilution of 1:1000 (1 µl stock + 999 µl normoxic solution).
The same dilution was used for obtaining the Normoxic
control+vehicle (1 µl bi-distilled water + 999 µl normoxic solution), which was applied over rows 2, 9 and 10, whereas rows 1
and 11 were left untreated, and row 12 had no cells to begin with.
The Cornus mas dilutions and the Normoxic control+vehicle
solution were applied over the cells just before the 4 hours incubation period started. After the first incubation period finished,
we changed the solution of each row with DMEM I and the plates
were left for another 2 hours in the incubator.
Treatment with Crataegus monogyna and Prunella vulgaris
extracts. We followed the same steps as in Cornus mas extract
treatment.
Viability assay
For the viability assessment, we used the calcein/propidium
iodide (PI)-digitonine staining and the FLUOStar Optima plate
reader. Both calcein and PI-digitonine stocks were prepared in
our laboratory, half an hour before the viability assay started.
The viability assessment using calcein is based on theconversion of the cell permeant non-fluorescent calcein AM
dye to the fluorescent calcein dye due to intracellular esterase
activity in living cells (27). Propidium iodide (PI) is membrane
impermeant, therefore it is not usually found inside viable cellswith intact membranes. However, when it is applied over deadcells, it penetrates the cellular membrane and gains access tothe nucleic acids, and the fluorescent signal increases (28).
After the last incubation period which lasted for 2 hours, theplates were removed from the incubator and the cells were gently
washed three times using 200 µl Dulbecco’s Phosphate Buffered
Saline solution (DPBS)/well.
When the washing was finished, 3µM calcein AM was added
Fiziologia - Physiology  2014.24.2 (82)over the cells, 200 µl/well and the plates were left for 30 minutes
at 37oC in a dark room. The 3µM calcein AM was changed with
200 µl DPBS /well and the plates were introduced in the plate
reader to evaluate the live cells.
Having evaluated the percentage of living cells, the plates
were removed from the plate reader, the DPBS was removed
and 2.5-5 µM PI-digitonine was added over the cells, 200 µl/
well. The plates were left again for 30 minutes, at 37oC in a darkroom, then the Pi-digitonine dye was replaced with 200 µl DPBS
/well and viability measurement using the plate reader followed.
All data was collected and interpreted by the FLUOStar Optima
plate reader, then transposed on Excel working sheets.
Statistical analysis
The results have been statistically analyzed and expressedin the form of mean ± SD. Mann-Whitney nonparametric test
and SPSS15 software were used to examine the results andto compare the mean values for different experimental groups.
A value of P < 0.05 was considered as significant. The graphs
were realized using Excel software.
RESULTS
Cornus mas vs Normoxic control+vehicle
All six dilutions of Cornus mas extract proved to have asignificant effect in increasing the cell viability vs Normoxic
control+vehicle group (p<0.05).
The best increase in cellular viability was observed whentreating the cells with CO1 (0.01 mg pps/ml) (p= 0.012) and CO3(0.1 µg pps/ml) ( p= 0.001) dilutions.
Prunella vulgaris vs Normoxic control+vehicle
None of the six dilutions of Prunella vulgaris extract used fortreating the cells had a significant effect in increasing the viability
vs Normoxic control+vehicle treatment (p=0.05).
Moreover, the PR1 dilution proved to be toxic and killed all
the cells.
Crataegus monogyna vs Normoxic control+vehicle
Although the cardioprotective effect of Crataegus spp.extracts
used in hypoxic condition was demonstrated (29), none of thesix dilutions used for treating cells in normoxic conditions had
any effect in increasing cell viability.
Cornus mas vs Crataegus monogyna vs Prunella vulgaris
Using the Mann-Whitney non-parametrical test, the dilutionsof the three plant extracts were compared.
Cornus m. vs Prunella v. : after comparing each dilution (CO1vs PR1, CO2 vs PR2, etc.) the Cornus mas extract was found tobe significantly better than Prunella vulgaris extract in increasingcell viability when used in normoxic conditions. The p values are
expressed in Table IV.
Table IV. Cornus m. vs Prunella v.- Mann-Whitney nonparametric test
Dilutions

CO1 vs CO2 vs CO3 vs CO4 vs CO5 vs CO6 vs
PR1
PR2
PR3
PR4
PR5
PR6

p

<0.001

0.005

0.004

<0.001<0.001

0.004

α

0.001

0.01

0.01

0.001

0.001

0.01

2014.24.2 (82)  Fiziologia - Physiology

Cornus m. vs Crataegus m. : the Cornus mas dilutions proved
to have a significant effect in increasing cell viability vs Crataegus
m. dilutions, as expressed in Table V.
Table V. Cornus m. vs Crataegus m.- Mann-Whitney nonparametric
test
Dilutionspα significance level

CO1 vsCR1

CO2 vs CO3 vs CO4 vs CO5 vs CO6 vs
CR2
CR3
CR4
CR5
CR6

<0.001 <0.001<0.001<0.001<0.001<0.001

0.001

0.001

0.001

0.001

0.001

0.001

Crataegus m. vs Prunella v.: as shown previously, none of thetwo plant extracts had a significant effect in increasing cell viability vs Normoxic control+vehicle treatment. Still, after comparing
their dilutions, a significant difference was observed. Except forPR1 dilution, all Prunella v. dilutions were significantly superior
compared with Crataegus m. dilutions (Table VI).
Table VI. Prunella v. vs Crataegus m.- Mann-Whitney nonparametrictest
PR1 vsCR1

PR2 vsCR2

PR3 vsCR3

PR4 vsCR4

PR5 vs PR6 vs
CR5
CR6<0.001p
α significance
0.001
level

<0.001

0.002<0.001

0.019<0.001

0.001

0.01

0.001

0.05

0.001

Dilutions

A schematic representation of the effect of the three plant
extracts and Normoxic control+vehicle treatment on neonatal
rat cardiomyocytes can be seen in Figure 3.

Fig. 3. Comparison between mean values of each dilution of plant
extract and Normoxic Control+Vehicle solution

DISCUSSIONSThe effect of three plant extracts on neonatal rat cardiomyocytes used in cell cultures was studied. As proven, only the
Cornus mas dilutions had a significant effect in increasing cell
viability, while being applied in normoxic conditions vs Normoxic
control+vehicle treatment (p<0.05)
The fact that Cornus mas plant extract was found to have thehighest content in polyphenols (see Table II) is not correlatedwith the increase of cell viability because all three plant extracts
were standardized and a concentration of 0,1 mg polyphenols/
ml solution was used for the initial stocks. This allows the7speculation that it is not the quantity, but indeed the quality ofpolyphenols that might be correlated with the increase of cell
viability in normoxic conditions.
On the other hand, after performing HPLC-DAD-ESI (+) MS
analysis in a previous study, we have shown that these threeplant extracts contain a lot of other constituents like flavonoids
and flavonoid derivates that are well known for their antioxidant
capacity (16).
The most complex plant extract was represented by Cornusmas, followed by Prunella vulgaris and Crataegus monogyna
extracts.
Cornus mas extract contained a mixture of flavonoid glycosides
and phenolic derivatives, represented mainly by epi-catechin,quercetin, kaempherol and derivatives, caffeic acid and derivativesof coumaric acid. Prunella vulgaris was foud rich in rosmarinicacid, rutin, betulinic and oleanolic acids, ursolic acid and tannins(30), while Crataegus monogyna extract contained mainly vitexin
isomers like 2’ and 4’ –O-rhamnoside (16).
As for the antioxidant activity of each extract, the highest stability
and potential of flavonoids to express scavenging potential against
DPPH in hydrophilic environment was shown by the Cornus mas
extract (16).
Therefore, the antioxidant activity of Cornus m. extract canbe attributed to polyphenols like epi-catechin and flavonoids likequercetin, two proven antioxidant agents that were studied before,both in in-vitro and in in-vivo experiments (31, 32).
Rosmarinic acid seems to be involved in conferring antioxidant
capacity to the Prunella v. extract, as shown in a study conductedby R. Rasool (30).
After performing the HPLC-DAD-ESI (+) MS analysis that revealed the content of each plant extract and knowing the effectsof each compound, it is easy to understand why the Crataegus
monogyna extract was less effective in increasing cell viability.
Although the antioxidant effect of Crataegus monogyna andCrataegus oxycantha was demonstrated in numerous studies,
we could not reproduced it. This failure could be due to the factthat we have used only aqueous extracts, instead of the alcoholic
extracts. As proof stands the fact that there was a major difference
between the compounds found in the two types of extracts. Therefore, while the aqueous extract of Crataegus m. contained mainly
vitexin isomers like 2’ and 4’ –O-rhamnoside (16), the alcoholic
extract was rich in proanthocyanidin, vitexine-2-O-rhamnoside,
hyperoside, anthocyanin, chlorogenic acid and epicatechin (33).
Leaving aside the antioxidant capacity of each compound thatcan be isolated from the plant extracts that we have studied, an
important role in increasing the cell viability could be played by thesynergic effect of all compounds and their interactions.
Another important aspect is represented by the fact that obtaining an increase in cell viability is a matter of dose dependence.
Therefore, further studies are needed in order to elucidate the appropriate dose, as well as the exact compounds and mechanismsthat could be correlated with the viability increase.
CONCLUSIONSIn our experiments, all dilutions of Cornus mas extract had
8a significant effect in increasing the cell viability vs Normoxic
control+vehicle group (p<0.05). The best increase was observed
when the cells were treated with CO1 dilution (0.01 mg pps/ml)(p= 0.012), respectively CO3 dilution (0.1 µg pps/ml) (p= 0.001).
Neither Prunella vulgaris, nor Crataegus monogyna dilutions
used for treating the cells, had a significant effect in increasing
the viability vs Normoxic control+vehicle treatment (p≥0.05).
Acknowledgements
The research was supported by POSDRU 107/1.5/S/78702
and by HU-RO TRANSMED 0901/137/2.2.2. projects.
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method for increasing the proportion of myoblasts. Experientia 1971;
27: 356-358

25. Grynberg A, Athias P, Degois M. In Vitro Cell Dev Biol. 2004;
22(1):44-50.
26. X Li, F R Heinzel. Role of Cx 43 in ischemic preconditioning does
not involve intercellular communication through gap junctions. J. Molec.
Cell. Card., 2014.
27. Kwan J, Ansar H, Crumbie H, Vanezis A, Al-Azzawi F, Rodrigo
G. The impact of menopause and oestrogen replacement therapy on
cardioprotection in remote ischaemic preconditioning. Heart, 2014;
100(3):A109.
28. Simon J, Nguyen V, Coder D. Assessment of Cell Viability:
Current Protocols in Cytometry Unit Number: Unit 9.2, 2013; DOI:
10.1002/0471142956.cy0902s64Online Posting
29. Garjani A, Nazemiyeh H, Maleki N, Valizadeh H. Effects of extracts
from flowering tops of Crataegus meyeri on ischaemic arrhythmias inanaesthetized rats. Phytother Res 2000; 14: 428-431.
30. Rasool R, Ganai BA. Prunella vulgaris: A literature review on its
herapeutic potential. Pharmacologia 2013; 4(6): 441-448.
31. Othman A, Jalil AMM., Weng KK, Ismail A, Ghani N, Adenan I.
African Journal of Biotechnology, 2010; 9(7): 1052-1059.
32. Zhang M, Swarts SG, Yin L, Liu C, Tian Y, Cao Y, Swarts M, Yang
S, Zhang SB, Zhang K, Ju S, Olek DJ, Schwartz L, Keng PC, Howell
R, Zhang L, Okunieff P. Antioxidant properties of quercetin. Adv Exp
Med Biol, 2011; 701: 283-9.
33. Dinesh K, Vikrant A, Zulﬁ A, Nisar AK., Deo NP. Pharmacognosy
The genus Crataegus: chemical and pharmacological perspectives.
Brazilian Journal of Pharmacognosy, 2012.

EFECTELE EXTRACTELOR DE PLANTE DIN CORNUS MAS, PRUNELLAVULGARIS SI CRATAEGUS MONOGYNA ASUPRA VIABILITATII CARDIOMIOCITELOR NEONATALE DE SOBOLAN
REZUMAT

Scop: Studiul nostru isi propune evaluarea efectelor a trei extracte apoase bogate in polifenoli (Crataegus monogyna, Cornus mas
si Prunella vulgaris) asupra cardiomiocitelor recoltate de la sobolani nou-nascuti, in conditii de normoxie.
Materiale si metode: Au fost prelevate inimile a 270 de sobolani Wistar nou-nascuti (1-2 zile) si au fost introduse in solutie salina
rece (PBS). Au fost indepartati ventriculii si fragmentati, iar piesele maruntite au fost introduse in solutie de tripsina (0,25%) timp de
25-30 minute. Suspensia celulara rezultata a fost centrifugata 15 minute, la 2000 rpm si 4oC, sedimentul fiind amestecat cu mediul decultura Dulbecco’s Modified Eagle Medium (DMEM). Au fost pregatite placi de cultura cu 96 de godeuri si introduse in incubator timp
de 72 de ore (37oC, 95% O2, 5 %CO2). Folosind metoda Folin-Ciocâlteu am determinat continutul total de polifenol al fecarui extract
de plante. Pornind de la solutiile stoc originale, am preparat sase dilutii succesive, folosind apa bidistilata pentru fiecare extract deplante. Dilutiile au au fost folosite pentru tratarea celulelor in conditii de normoxie. Au fost efectuate masuratori ale viabilitatii celulare
folosind FLUOstar Optima plate reader, dupa marcarea cu iodura de propidiu-digitonina.
Rezultate si concluzii: Doar extractul de Cornus mas a indus o crestere a viabilitatii celulare semnificativa statistic comparativ cu
grupul de control. Extractele de Crataegus monogyna si Prunella vulgaris nu au indus efecte semnificative asupra viabilitatii celulare.
Cuvinte cheie: extracte de plante, polifenoli, normoxie, viabilitate2014.24.2 (82)  Fiziologia - Physiology

9

SIMULTANEOUS LAPAROSCOPIC APPROACH OF COLORECTALCANCER AND OF SYNCHRONOUS LIVER METASTASES –INITIAL EXPERIENCEBREBU DAN, PANTEA STELIAN, LAZAR CAIUS, TARŢA CRISTI, DUŢA CIPRIAN, LAZAR FULGERSurgery Department, University of Medicine and Pharmacy “Victor Babes” Timisoara, Romania
Second Surgery Clinic of Timisoara County Hospital

ABSTRACTThe simultaneous resection of colorectal cancer (CRC) and of synchronous liver metastases is a debate topic related to the postoperative morbidity as compared to the staged resection. The purpose of this study is to analyse the results found in a series ofeight patients with CRC and synchronous liver metastases who benefitted from combined colorectal and liver laparoscopic surgery.
A prospective assessment was performed in terms of patients’ characteristics, tumours’ characteristics, operative variables andpostoperative results. The primary tumour was localized at the level of the colon in six patients, and at the level of the rectum in twopatients, while the synchronous liver metastases were predominantly solitary. A laparoscopic approach was applied to all 8 patients,
who underwent colorectal resection (R0), as well as metastasectomies and hepatic wedge resections. There was no need to convert
to classic surgery. The sample was extracted by pararectal incision or by transverse suprapubic incision. The median operative time
was approximately 313 minutes (range 151-394 minutes), with an average blood loss of 600 ml (range 200-850 ml).The postoperative
length of stay varied between 6 and 14 days. On the basis of this initial experience of a single centre, the laparoscopic simultaneous
resection of colorectal cancer and the hepatic resection can be performed on the selected patients suffering from CRC and SLM,
with satisfactory short-term results.
Key words: colorectal cancer, liver metastases, laparoscopic surgical approach

INTRODUCTION
Liver is the most frequent site of hematogenous disseminationof colorectal cancer (CRC) (1, 2). Liver metastases are present
in 10-25% of the patients during the surgical intervention forcolorectal malignancy (3).The surgical resection is the most efficient and potentially curative treatment for the liver metastasesof CRC (4).The treatment strategies and the results for thesepatients developed and underwent several changes due to thetechnical innovations (5-6).Laparoscopic surgery improves theaccuracy of the surgical approach, the postoperative recovery, it
minimizes the postoperative pain, reduces the rate of abdominal
wall infection, decreases the hospital length of stay, allows a more
rapid return to normal activities, has better cosmetic results andit does not compromise the oncologic result (7,8).
There are several treatment options for the patients sufferingfrom CRC with synchronous liver metastases (SLM), dependingon the location of the primary tumour (rectum or colon) and on
the extent of liver disease. The controversies are related to theconcept that the staged resection of the primary tumour and of
the liver metastases is better tolerated by the patient. Recently,
a systemic analysis has shown that synchronous colorectal andhepatic resection led to a shorter hospital stay and to fewer complications, as compared to a staged resection (9).The specialised
literature shows that the staged laparoscopic resection, initiallyof the primary tumour and subsequently of the hepatic lesions
is more documented than the combined laparoscopic resectionof CRC and SLM, for which there are only a few references.
The purpose of this study is to assess the initial experienceof the simultaneous laparoscopic resection of CRC and SLM

and to monitor the evolution and the prognosis of these caseson the short term.
MATERIAL AND METHODS
Between January 2012 and March 2013 the Second Surgery
Clinic of Timisoara County Hospital had 48 laparoscopic surgery
cases for patients suffering from CRC, of which 15 also sufferedfrom SLM. Of the 15 patients, 8 patients with CRC and SLM wereselected to benefit from combined laparoscopic colorectal andliver surgery. The other 7 patients required major hepatectomy
and they were excluded from the study. The data were prospectively collected and analysed for this study. The assessedparameters were the following: the patients’ characteristics,
the tumours’ characteristics, the operative variables and the
postoperative results.
Multimodal treatment of CRC and SLM
For the patients suffering from CRC and SLM, the decision forthe best treatment was established depending on the location of
the primary tumour, the complexity of the primary tumour resection,
the location of the liver metastases, as well as their size and number, the feasibility of the laparoscopic approach and the generalcondition of the patient. For the series under study we performed
simultaneous resection in patients undergoing colorectal resection with anastomosis in elective settings (T1-T3) and a single ormultiple liver lesions (up to 3 lesions) with a peripheral location on
segments 2-6 which allowed a limited hepatic resection.
Neoadjuvant therapy was administered for the primary rectal
tumours (2 patients), consisting of short-course radiotherapy (5 x
5 Gy), followed by 3 to 5 systemic chemotherapy cycles (capecit-

Received 12th of March 2014. Accepted 15th of May 2014. Address for correspondence: Dan Brebu, MD, PhD Student Department of Surgery, University of Medicine
and Pharmacy “Victor Babeș” Timișoara, Iosif Bulbuca Street, No. 10, Timișoara, România; Phone: +40755012719, e-mail: dr.brebudan@gmail.com

10

Fiziologia - Physiology  2014.24.2 (82)abine). All the cases have been analysed by a multidisciplinary
team, the treatment being adapted to each particular case andwell argumented. The protocol of our clinic was respected interms of laparoscopic approach. The other patients underwent
classic surgery or a staged approach.
Surgical approach
The laparoscopic approach of colorectal tumours was a medial-to-lateral approach. A right pararectal incision was performed
for the right hemicolectomy in order to extract the resection
specimen, as well as to perform an extracorporeal anastomosis.
A Pfannenstiel incision was performed on the left colon in order
to extract the resection specimen, followed by an intracorporeal
colorectal anastomosis with circular stapler. The laparoscopic
liver resection was performed by the surgeon standing between
the legs of the patient. Hepatic wedge resections were performed
for the peripheral metastases and metastasectomies. For thelaparoscopic liver resection, 2-3 additional trocars were mainly
used, as compared to the ones necessary for the approach of
the colorectal primary tumour. The 10 mm trocar situated at
the level of the umbilicus to ensure the view, initially used forthe approach of the primary lesion, is used for the subsequent
approach of the liver lesions. 2 additional trocars of 5 mm areplaced in the upper abdomen, left and right. When needed, an
additional 5 mm trocar was used.
Enseal® advanced technology instruments (advanced bipolar
technology) or LigaSure™ instruments were used for the transection of the hepatic parenchyma, and Tachosil® absorbablefibrin
sealant patch was used for the additional hemostasis of thehepatic section surface. The hepatic resection specimen wasplaced in a plastic bag and extracted through the pararectal
incision and the Pfannenstiel incision, respectively.
RESULTS
6 men and 2 women diagnosed with CRC and SLM were
included in the study. The preoperative diagnosis of CRC andSLM on the basis of the investigations performed was assigned
to 6 patients. During the laparoscopic surgery, two patientswere diagnosed with SLM in segments 2 and 6 and they were
included in the protocol for simultaneous resection. The average age was 68 years (range 54-76 years). The characteristics
of each patient are shown in Table I. The average body massindex was 29.0 (range 24.9-31.2) kg/m2. A patient suffered an
emergency laparoscopic cholecystectomy 2 months before, fora phlegmonouslithiasic acute cholecystitis. During laparoscopy,
a hepatic lesion was identified and subsequently investigated.
The colonoscopy revealed a tumour in the sigmoid colon. The
preoperative treatment of the rectal cancer consisted of a shortcourse radiotherapy (5 x 5 Gy) and an associated systemic
chemotherapy (capecitabine). All the 8 laparoscopic resections
were successfully completed. There were no conversions toclassic surgery. The surgical results are shown in Table II.
The following procedures have been performed: laparoscopic
right hemicolectomy and wedge resection of segment 2/3 withextraction of the specimens through the right pararectal incision, laparoscopic sigmoidectomy with metastasectomies insegments 3, 4, and 6 with extraction of the specimens throughthe Pfanennstiel incision, rectal anterior resection and metasta2014.24.2 (82)  Fiziologia - Physiology

sectomies in segments 3 and 6 with extraction of the specimens
through the Pfanennstiel incision. The incision used to extract
the specimens varied between 5 and 10 cm.
Table I. Patient characteristics and preoperative data
Patient Sex Age
NumberMedicalhistory

Primary Hepatic No. of Neoadtumor metastasis me- juvant
location location tasta- treatsis
ment
Apendectomy Sigmoid Segment 6
1
No
Hypertension

1

M

63

2
3

M
M

54
72

None
Sigmoid Segment 3
Hypertension Cecum Segment
Anemia
2/3

1
3

No
No

4

F

68

Sigmoid Segment 2

1

No

5

M

77

AsSegment 3
cending
colon

1

No

6

F

70

7

M

67

Acute cholecistytis
Laparoscopiccholecystectomy
Hypercolesterolemia
Coronary
heart disease
Anginapectoris
Hypertension
Hypothyroidsm
Apendectomy

8

M

69

Hypercolesterolemia
Atrial fibrillation

AsSegment 2
cendingcolon

Rectum Segment 6

1

Rectum Segment 31
2

5x5 Gy5x5 GyNo

Table II. Surgical results
Patient
numberType of operation

1

Sigmoidectomy andmetastsectomy ofsegment 62

Sigmoidectomy andmetastasectomy ofsegment 3
Right hemicolectomy
and atipicrezection ofsegments 2/3
Sigmoidectomy andmetastasectomy ofsegment 2
Right hemicolectomy
and metastasectomyof segment 3
Rectal anterior resection and metastasectomy of segment 6
Rectal anterior resection and metastasectomy of segment 3
Right hemicolectomy
and metastasectomyof segment 2

3
4
5
6
7
8

Operation
time
(min)
275

Blood
loss
(ml)

181

200

6

30

230

500

9

48

260

450

8

36

190

550

14

72

354

600

9

48

315

750

8

48

210

350

10

72

650

Postop- Resumperativetion ofhospital
bowel
stay (days) (hours)
8
36

11

The median operative time was 280 minutes (range 181-354
minutes), with a total estimated blood loss of 600 ml (range 200750 ml).The intraoperative complications consisted of the bleedingat the level of the section surface, solved with suture with Prolene
4-0 and by the introduction of an additional trocar. A patient showeda Pfannenstiel wound infection, and 2 patients suffered fromdelayed gastric emptying that required a nasogastric tube. Other
postoperative complications: a patient developed a lower urinary
tract infection and hypertension episodes that were managed by
the administration of antibiotics and antihypertensives. The average hospital stay was 9 days, ranging between 6 and 14 days.
There were no records of postoperative mortality. The R0 resectionof the primary tumour and of the hepatic lesions was performed
to all patients. Details regarding the pathological examination are
shown in Table III.

AuthorYear

Nr. of Laparo- Timepascopic(min)tients liverresection(type)

Geiger et al. [10]Leung et al. [11]Vibert et al. [12]Law et al. [13]Bretagnol et al. [14]

2006
2006
2006
2008
2008

1
1
8
4
1

Pessaux andPanaro [15]Sasaki et al. [16]2009 32009 9Casaccia et al. [17] 2010 1
Lee et al. [18]2010 10*

Table III. Pathological examination
Patient
number

p/yTN
stage

Radicality

Diameter SLM(cm)

Resection
marginSLM (mm)

1
2
3
4
5
6
7
8

pT2N1
pT3N0
pT2N2
pT3N0
pT2N1
yT1N1
yT2N0
pT2N2

R0
R0
R0
R0
R0
R0
R0
R0

1.5
2.4
2/1.5/0.9
1.8
1.5
1.3
2
2.1/1.6

6
8
>10
8
7
5
6
5

DISCUSSION
The advantages of the laparoscopic approach in colorectal
surgery or the hepatic resections performed by experienced
surgeons (8) led to a significant increase in the number of suchinterventions during the last years. The separate laparoscopic
approach of colorectal cancer and of colorectal cancer metastases was proven and resulted in a more rapid recovery and a low
morbidity rate, with oncologic results similar to those obtained byopen surgery (7,8). This suggests that a combined laparoscopic
approach both for the primary tumour and for the liver metastases
may be beneficial for patients who candidate for a simultaneous
resection.
Moreover, the open surgery for the resection of the colorectal
primary tumour and of synchronous metastases may require an
extended incision, which may sometimes be lacerating, especiallywhen the tumours have opposite location (for instance the rectum
and the right liver lobe).Using the laparoscopic approach, theexposure can be improved even when from an anatomical point ofview there is a narrow pelvis and sites to which access is difficult,in the upper abdominal area. The feasibility of the simultaneous
laparoscopic approach was proven by our initial experience in
8 cases, and the results obtained confirm the data found in thespecialised literature for this topic, presenting series of a comparable size (Table IV).
Table IV. NR: not reported, LH: left hemihepatectomy, LLS: left lateral
sectionectomy, M: metastasectomy, S: segmentectomy, RH: righthemihepatectomy, RFA: radio frequency ablation, *13 resections in 10
patients.

12

Hayashi et al. [19] 2011 2
Tranchart et al. [20] 2011 2

LLS
LLS
NR
NR
1 LLS
2M
1M+
RFA
2 LLS
7M
1 LLS
6 LLS
5M
1S
1 RH
NR
1 LH
1 RH

330
350
NR
NR
NR

Blood loss Post(ml)operative
hospitalstay(days)
600
4
500
7
NR
NR
NR
NR
NR
NR

NR

NR

NR

418
(215–520)
455
401(230-620)
NR
NR(270-575)
310
345

219
(32-745)NR
500(60-1000)
NRNR(40-330)200
200

9(7-26)
1210(7-15)
NR
NR(7-14)
4
6

The patients with a single metastasis located in segments
2-6 are the ideal candidates for the simultaneous laparoscopic
resection. Besides the standard location of trocars in laparoscopic
colorectal surgery, 2 additional trocars are required, for a good access to perform the hepatic resection. Both resection specimens
can be extracted through a single incision. In the case of lesions
located in segments 7-8, or in the case a major hepatectomy was
required, the simultaneous resection combined approach wasaborted, and these patients were excluded from the study. However, certain authors with experience in laparoscopic liver surgery
have proven that the laparoscopic major hepatectomy performed
simultaneously with the laparoscopic colorectal resection is also
feasible (18,20). The major hepatectomy specimen is extracted
through a Pfannenstiel incision, which has been proven to havethe lowest rate for the occurrence of an incisional hernia (21).
The theoretical arguments against simultaneous resection would
be the combination between a clean operation and a contaminatedone, and the deficiency on protein synthesis following the hepatic
resection could lead to an increase of the infection risk and could
compromise the healing of the anastomosis. Another hypothesis
is related to the Pringle manoeuvre, when the resulted venous
congestion could lead to intestinal edema. Nevertheless, thespecialised literature based on 14 comparative studies reveals thatthe combined resection shows a lower morbidity rate (9). This led
to the conclusion that simultaneous resection can be performed topatients selected by surgical teams specialised both in colorectal
surgery, and in hepatobiliary surgery. The correct assessment of
the patients, as well as their selection are essential for this typeof complex surgery, and the multidisciplinary team must decideupon the optimal moment when the multimodal treatment must
be applied.
As a result of the improvement of the surgical techniques inliver surgery, of the medical technical innovations (LigaSure™,
Harmonic®, Enseal®) and of the systemic chemotherapy, the life
expectancy of the patients with colorectal metastases increased.
An initial laparoscopic approach results in a reduced formation of
Fiziologia - Physiology  2014.24.2 (82)

peritoneal adhesions and it facilitates a repeated hepatic resection,
which was proven in the case of patients with HCC that subsequently needed a liver transplant (22). The benefits of laparoscopy
in this case can be seen in the improved oncologic results, in the
improvement of the quality of life, the integrity of the abdominal
wall and the cosmetic advantages.
CONCLUSION
The simultaneous resection of CRC and SLM continues to bea controversial issue. There are no randomized clinical studies
comparing the simultaneous resection to the staged one. The
recently communicated data of the prospective series are difficult
to interpret because of the selective character of the patients.
Our initial experience, correlated with the specialised literature,indicates that simultaneous resection of CRC and SLM is feasible
and can be chosen for the patients selected, provided that it isperformed by a team with adequate experience in this field.
REFERENCES
1. Cummings LC, Payes JD, Cooper GS. Survival after hepatic resection in metastatic colorectal cancer: a population-based study. Cancer.
2007; 109(4):718-726.
2. Grundmann RT, Hermanek P, Merkel S, et al. Diagnosis and treatment of colorectal liver metastases-workflow. Zentralblatt fur Chirurgie.
2008;133(3):267-284.
3. Popescu I. Rezecţia hepatică. In: Popescu I, editor. Chirurgia ficatului.
Bucureşti: Editura Universitară „Carol Davila”, 2004: 537-662.
4. Popescu I, Ionescu M, Alexandrescu S, Ciurea S, Hrehoreţ D,
Sârbu-Boeţi P, Boroş M, Croitoru A, Anghel R.Surgical treatment of liver
metastases from colorectal cancer. Chirurgia 2006; 101(1):13-24.
5. González HD, Figueras J. Practical questions in liver metastases ofcolorectal cancer: general principles of treatment. HPB. 2007; 9(4):251258.
6. Yang AD, Brouquet A, Vauthey JN. Extending limits of resection formetastatic colorectal cancer: risk benefit ratio. Journal of Surgical Oncology. 2010; 102(8):996-1001.
7. Kuhry E, Schwenk WF, Gaupset R, Romild U, Bonjer HJ. Long-term
results of laparoscopic colorectal cancer resection.Cochrane Databaseof Systematic Reviews.2008; (2, article CD003432)
8. Reddy SK, Tsung A, Geller DA. Laparoscopic liver resection. World
Journal of Surgery. 2011; 35:1478-1486.
9. Hillingsø JG, Wille-jørgensen P. Staged or simultaneous resection ofsynchronous liver metastases from colorectal cancer-a systematic review.
Colorectal Disease. 2009;11(1):3-10.
10. Geiger TM, Tebb ZD, Sato E, Miedema BW, Awad ZT. Laparoscopic
resection of colon cancer and synchronous liver metastasis.Journal of Laparoendoscopic and Advanced Surgical Techniques A. 2006; 16(1):51-53.
11. Leung KL, Lee JFY, Yiu RYC, Ng SSM, Li JCM. Simultaneous laparoscopic resection of rectal cancer and liver metastasis.Journal of Laparoendoscopic and Advanced Surgical Techniques A. 2006; 16(5):486-488.
12. Vibert E, Perniceni T, Levard H, Denet C, Shahri NK, Gayet B. Laparoscopic liver resection. British Journal of Surgery. 2006; 93(1):67-72.
13. Law WL, Fan JKM, Poon JTC, Choi HK, Lo OSH. Laparoscopic bowel
resection in the setting of metastatic colorectal cancer.Annals of Surgical
Oncology. 2008; 15(5):1424-1428.
14. Bretagnol F, Hatwell C, Farges O, Alves A, Belghiti J, Panis Y. Benefitof laparoscopy for rectal resection in patients operated simultaneously
for synchronous liver metastases: preliminary experience. Surgery. 2008;
144(3):436-441.
15. Pessaux P, Panaro F. Advantages of the first-step totally laparoscopic
approach in 2-staged hepatectomy for colorectal synchronous liver metastasis. Surgery. 2009; 145(4): 453.
16. Sasaki A, Nitta H, Otsuka K, Takahara T, Nishizuka S, Wakabayashi
G. Ten-year experience of totally laparoscopic liwer resection in a single
institution. British Journal of Surgery. 2009; 96(3):274-279.
17. Casaccia M, Famiglietti F, Andorno E, di Domenico S, Ferrari C,
Valente U. Simultaneous laparoscopic anterior resection and left hepatic
lobectomy for stage IV rectal cancer. Journal of the Society of Laparoendoscopic Surgeons. 2010; 14(3):414-417.
18. Lee JS, Hong HT, Kim JH, et al. Simultaneous laparoscopic resectionof primary colorectal cancer and metastatic liver tumor: initial experienceof single institute. Journal of Laparoendoscopic and Advanced SurgicalTechniques. 2010; 20(8):683-687.
19. Hayashi M, Komeda K, Inoue Y, et al. Simultaneous laparoscopic
resection of colorectal cancer and synchronous metastatic liver tumor.
International Surgery. 2011; 96(1):74-81.
20. Tranchart H, Diop PS, Lainas P, et al. Laparoscopic major hepatectomy can be safely performed with colorectal surgery for synchronouscolorectal liver metastasis. HPB. 2011; 13(1):46-50.
21. DeSouza A, Domajnko B, Park J, Marecik S, Prasad L, Abcarian H.
Incisional hernia, midline versus low transverse incision: what is the ideal
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Laparoscopic liver resection facilitates salvage liver transplantation forhepatocellular carcinoma. Journal of Hepato-Biliary-Pancreatic Surgery.
2009; 16(3):310-314.

REZECTIA LAPAROSCOPICA SIMULTANA A CANCERULUI COLORECTAL SI AMETASTAZELOR HEPATICE SINCRONE – STUDII INITIALEREZUMAT

Rezecția simultană a cancerului colorectal (CCR) și a metastazelor hepatice sincrone (MHS) este un subiect de dezbatere legat demorbiditatea postoperatorie în comparație cu rezecția în etape. Scopul acestui studiu a fost analiza rezultatelor unei serii selectate deopt pacienți cu CRC și metastaze hepatice sincrone care au beneficiat de chirurgie laparoscopică combinată – colorectală și hepatică.
Au fost evaluate prospectiv caracteristicile pacienților, caracteristicile tumorilor, variabilele operatorii cât și rezultatele postoperatorii.
Tumora primară a fost localizată la nivelul colonului la șase pacienți și la nivel rectal la doi pacienți, iar metastazele hepatice sincrone
au fost majoritar solitare. Prin abord laparoscopic la toți cei 8 pacienți,a fost realizatărezecția colorectala (R0) cât șimetastazectomii
și rezecții hepatice atipice limitate. Nu a fost necesară conversia la chirurgia clasică. Extragerea piesei de rezecțies-a făcut printr-o incizie pararectală sau suprapubiană transversală. Timpul operator mediu a fost de aproximativ 313 minute (interval 151-394
minute), cu o pierdere medie de sânge de 600 ml (interval 200-850 ml). Durata spitalizarii postoperatorii a variat între 6 și 14 dezile. Din această experiență inițială a unui singur centru, rezecția laparoscopică simultană colorectală și hepatică poate fi efectuată
la pacienții selectați cu CCR și MHS, cu rezultate satisfacatoare pe termen scurt.
Cuvinte cheie: cancer colorectal, metastaze hepatice, chirurgie laparoscopică

2014.24.2 (82)  Fiziologia - Physiology

13

PATIENT-PERCEIVED QUALITY OF MEDICAL SERVICES IN
TIMIS COUNTY PUBLIC HEALTH HOSPITALSBOGDAN CIPRIAN, BUCUR ADINA“Victor Babes” University of Medicine and Pharmacy Timisoara

ABSTRACTIntroduction: The primary objective of hospital reform proposed by the Ministry of Health is to improve the quality of health services
provided to insured persons and to increase access of all categories of population, including the disadvantaged. Currently, theconcept of medical services clients’s/users’s response reaction is particularly emphasised in the public health system along with theidea of providing what he/she desires.
The aim of this paper is to assess patients’ perceptions on the medical services and medical personnel professionalism in Timis
County public health facilities.
Methodology: Three hundred seventy-three questionnaires were applied during 2013 year to randomly selected in-patients in Timis
County hospitals. A set of questions were asked, out of which we selected for the purpose of this paper those highlighting the behaviour
and professionalism of medical personnel, as well as aspects regarding medical care provided during continuous hospitalisation.
Responses were collected in a database and processed by the means of SPSS 17.0 programme.
Results: Most patients were women and the best represented group was 41-50 years. Most patients showed a higher degree of
dissatisfaction at questions about various aspects of medical care delivered. A higher proportion of patients declared themselves
satisfied with the behaviour and professionalism of medical personnel. Regarding the general quality of medical services, 94.40%
of patients are satisfied.
Conclusions: Although 90% of the interviewed patients address the same hospital and are satisfied with the quality of medical
services in 94% of cases, they consider some aspects related to medical care and personnel should be improved.

INTRODUCTION
The primary objective of hospital reform proposed by the Ministry of Health is to improve the quality of health services provided
to insured persons and to increase access of all categories of
population, including the disadvantaged (1).
In the context of current reform, health facilities managers are
more and more concerned with the improvement of the qualityof services provided by the facilities they are managing (2,3).
Continuous quality improvement primarily aims to increasesatisfaction of clients (both external and internal), as well as todecrease the cost of each delivered service (4).
Patient satisfaction mirrors the quality of health services.
Currently, the concept of medical services clients’s/users’s
response reaction is particularly emphasised in the public healthsystem along with the idea of providing what he/she desires (5,6).
Patient satisfaction is expressed as cognitive evaluation andemotional reaction to the elements of structure (human, material,
financial resources and hospital environment), process (technicaland interpersonal aspects), as well as to the results of medical
services delivered to them (7).
Research in the field shows satisfaction is related to the perceived technical abilities, intelligence and qualification of medical
personnel. However, patients mostly appreciate interpersonalcommunication skills of medical personnel medical (8).
First of all, patients’ needs must be identified in order to enable

medical personnel to address them using appropriate medical
techniques and procedures (2,9).
Patients should be asked about their expectations from thephysician. One frequently studied aspect of medical care is theextension and accuracy of physician-patient communication(2,4,10). Patient satisfaction is closely associated with accuracyof information provided and this could be an important indicationof physician-patient communication quality (11).
The aim of this paper is to assess patients’ perceptions on
the medical services and medical personnel professionalism
in Timis County public health facilities.
METHOD
In order to apply the questionnaires for patients, we haveselected public hospitals in Timis County, except for children’sand psychiatric ones. We have applied a non-probability sampling
method, namely the fixed quota method, the monthly number ofhospital admissions in each hospital being known. An averageof 11.959 patients per month resulted from the total numberof admissions in all the studied hospitals, from which we havecalculated, by the means of SPSS 17.0 programme, a sample
of 373 patients, with a +/- 3% error. The questionnaires were
distributed among hospitals based on their proportion from thetotal number of admissions in all the studied hospitals. The
questionnaire comprised preformed response questions, while

Received 24th of April 2014. Accepted 13th of May 2014. Address for correspondence: Adina Bucur, MD, PhD, Department of Functional Sciences, “Victor Babes” University of Medicine and Pharmacy Timisoara, Eftimie Murgu Sqare No. 2A, RO-300041, Timisoara, Romania, phone/fax: +40256490507, e-mail: adina.bucur@gmail.com

14

Fiziologia - Physiology  2014.24.2 (82)

three were open-ended questions. Questionnaires were applied
during 2013 year.
Table I. Distribution of patients in the sample that received questionnaires
Hospital

Patient
percentage

County Clinical Emergency Hospital Timisoara

29.59%

City Clinical Emergency Hospital Timisoara

22.69%

Obstetrics-Gynaecology Clinical Hospital Timisoara

5.41%

Pneumo-phtisiology and Infectious Diseases Clinical 4.66%
Hospital Timisoara
Cardiovascular Diseases Institute Timisoara

4.69%

Railway Hospital Timisoara

4.65%

Military Hospital Timisoara

5.55%

City Hospital Faget

3.81%

City Hospital Sannicolau Mare

5.41%

City Hospital Jimbolia

3.68%

City Hospital Deta

2.76%

City Hospital Lugoj

7.79%

RESULTSThe questionnaires were randomly distributed to in-patientsin the Timis County public hospitals.
We have analysed the questions on patients’ satisfaction
regarding the quality of medical services provided to them during hospitalisation. The medical services quality is perceived
by the patient as improvement of his/her health, which is conditional upon the correct assessment of patient’s needs, so as the
medical personnel to be able to address them using appropriate
medical techniques and procedures. All these are based on theeffective physician (medical personnel)-patient communication.
In the studied group, women represented the greatest proportion of admitted patients (Figure 1), with people between
41-50 years being the best represented age group, while two
aggregated age groups (51-60 years and 61-70 years) wereapproximately equal with the 31-40 years age group.

Fig. 2. Distribution by age groups

At the question regarding prior hospitalisation in the same
health facility, 91% of patients chose the same hospital, which
is most probably due to hospital proximity and not to medical
services. Furthermore, when only one department for a certain
medical specialty is available in the county, the patient will be
re-admitted in the same department for the same pathology.

Fig. 3. History of admissions in the same health facility

In defence of the previously mentioned reasons for choosing
the same hospital, only 42% would choose the same healthfacility if free option, with no financial or service uniquenessconstraints would be available.

Fig.4. Distribution of patients according to the choice for the same
health facility

Fig. 1. Distribution of in-patients by gender
2014.24.2 (82)  Fiziologia - Physiology

Regarding the admission method, only 12% have been admitted through the emergency department.
15

pitalisation, the proportions were almost equal for accompanied
versus not accompanied patients. Among patients, 23.60% wereaccompanied by relatives.

Fig. 5. Distribution of patients according to admission method

Most of the patients (77.20%) were waiting between one andtwo hours to be examined for hospitalisation.

Fig. 8. Distribution of answers regarding the method of accompaniment to various medical procedures

Treatment is another important component of health services. On the question whether patients have purchased or not
medicines for their treatment in the hospital, only 4.4% said theirtreatment was exclusively received from the hospital, 28.80%
have both received and bought medicines, but the vast majority(66.80%) have bought their medicines.

Fig. 6. Distribution of answers regarding waiting time for examination

Out of the total number of patients, 89.20% assert medical
tests were performed for them and only 10.80% came with medical tests already done in the outpatient setting; patients proved
to be satisfied in this respect.

Fig. 9. Distribution of answers regarding treatment source

In respect of the information received on various requests,
patients are fully dissatisfied (91.20%).

Fig. 7. Distribution of answers according to medical tests performed

On question whether they were accompanied by medical
personnel to the various procedures recommended during hos16

Fig. 10. Distribution of answers regarding information received on
various requests

Fiziologia - Physiology  2014.24.2 (82)Interaction with medical personnel and their professionalism
is also a component of the medical services quality. The attending physician’s attitude during his/her interaction with thepatient was considered good and very good (94.40%), eventhough approximately 50% were satisfied with the time devoted
to consultation (Figure 12).

Fig. 14. Distribution of answers regarding disease-related information

Behaviour of nurses was judged as well, approximately 50% of
patients being satisfied by their behaviour, 30% being dissatisfied
with their professionalism and 70% judging it as appropriate.
Fig. 11. Distribution of answers regarding physician’s attitude­­Fig. 12. Distribution of answers regarding the time devoted toconsultation

Fig. 15. Distribution of answers regarding nurses’ behaviour

Physician’s professionalism is difficult to judge. However, 90%consider the attending physician was well qualified, although only
64% consider they have received enough information regarding
their disease.

Fig. 16. Distribution of answers regarding nurses’ professionalism

Fig. 13. Distribution of answers regarding physician’s qualification
2014.24.2 (82)  Fiziologia - Physiology

Regarding the general quality of medical services, 94.40% of
patients are satisfied.
17

the physician’s attitude (94%), although 50% are satisfied withthe time devoted to consultation and 90% with the physician’squalification. A great part of patients are dissatisfied with nurses’
behaviour (43.6%), even though 70% are satisfied with the medical care they provide.
The quality of medical services provided by Romanian hospitals is judged good and very good by approximately one third of
respondents (12). Similarly, one third of respondents judge thequality of medical services as average and one quarter consider
they are poor or very poor (12). In our study, 50.8% of patients
consider the quality of medical services is average and only
43.6% judge it as good and very good.
Fig. 17. Distribution of answers regarding quality of medical services

DISCUSSIONSThe public opinion barometer conducted in 2006 among Romanian citizens shows that people with a less or more severe
health problem choose primarily the family/specialist physician’s
office (60.47%) and only 17.6% address emergency services(emergency room - UPU) (12). This is in accordance with ourstudy, where admissions through emergency services account for
12% of total admissions within the study group. In our study, 90%of people would choose the same hospital, which is in agreementwith data in the literature showing three quarters of the individuals
in the general population hospitalised during 2006 year would
choose the same hospital in the event of a re-admission.
During hospitalisation, 90% of the questioned patients in thegeneral population received explanations about their diagnosis,
85% about the procedures and interventions they underwent
or about disease prognosis and evolution (12). Only half of thehospitalised individuals received information regarding theirrights to refuse a medical intervention, to receive an alternate
medical opinion or about alternatives to the proposed procedures(12). In our study, 86% of the interviewed people are dissatisfied
with the information received to various requests, while 86.8%
are satisfied with information they received about their disease
compared to general population.
Half of the in-patients assert there were certain negative
aspects during hospitalisation related to services delivered, but
only one in ten individuals has complained about this to someother person (family member, nurse or physician) (12).
One of five persons considers medical personnel attention
and behaviour is the most important aspect that should bechanged in hospitals (12). Fifteen percent of the respondents
consider endowment with medical equipments and unofficial
payments are the most important aspects that need to bechanged (12).
Medical personnel professionalism is an important aspectasserted by 8.5% of patients (12). These data are in supportof our study, in which the interviewed patients are satisfied with

18

CONCLUSIONS
Although 90% of the interviewed patients address the same
hospital services and 94% of them are satisfied with the quality of
medical services, there are also aspects that should be improved
regarding the medical services and the medical personnel:
better communication with the patient of both physicians andnurses, enough time devoted to patient and better qualificationof personnel.
Regarding medical services, patients’ discontent is mirrored by
the fact that they must purchase their medicines for in-hospitaltreatment, which should be incumbent to the health facility.
REFERENCES
1. Legea 95/2006
2. Vlădescu C. Sănătate Publică şi management Sanitar, Ed. Cartea
Universitară, Bucureşti, 2004.
3. Vlădescu C. Managementul serviciilor de sănătate, Ed.
Expert,Bucuresti, 2000.
4. Dayton NA. The demise of total quality management. The TQM
Magazine, 2003; 15(6): 391-396.
5. Berwick DM. A Primer on Leading the Improvement of Systems.
British Medical Journal, 1996; 312: 618-22.
6. Arndt M, Bigelow B. Reengineering: Dej. vu All Over Again! HealthCare Management Review, 1998; 23: 58-66.
7. Chassin MR. Is Health Care Ready for Six Sigma? Milbank Quarterly, 1998; 76: 565-91.
8. Hughes JM. Total Quality Management in a 300-Bed Community
Hospital: The Quality Improvement Process Translated into Patient
Care! Quality Review Bulletin, 1998; 18: 293-300.
9. Ancona DG. Outward Bound: Strategies for Team Survival in an
Organization. Academy of Management Journal, 1990; 33: 334-65.
10. Gann MJ, Restucci DJ. Total Quality Management in Health Care:
A View of Current and Potential Research. Medical Care Review, 1994;
51 (4): 465-500.
11. Bradley EH, Holmboe ES, Mattera JA, Roumanis SA, et al. The
roles of senior management in quality improvement efforts: What are
the key components? / Practitioner application. Journal of Healthcare
Management, 2003; 48(1): 15-28.
Barometrul de opinie privind Serviciile de sănătate realizat în rândul
populaţiei din România, Centrul pentru Politici şi Servicii de Sănătate
Bucureşti, decembrie 2006

Fiziologia - Physiology  2014.24.2 (82)PERCEPTIA PACIENTILOR REFERITOARE LA CALITATEA SERVICIILOR MEDICALE IN SPITALELE JUDETULUI TIMISREZUMAT

Introducere: Obiectivul principal al reformei spitalelor propusa de Ministerul Sanatatii este acela de a imbunatati calitatea serviciilor
medicale oferite persoanelor asigurate si cresterea accesului pentru toate categoriile populationale, inclusiv persoanele dezavantajate.
In prezent, este promovat mai ales conceptul de servicii medicale bazat pe reactia de raspuns a clientului/utilizatorului in sistemul
de sanatate publica, impreuna cu idea de a asigura ceea ce doresc acesti utilizatori
Scopul acestui studiu este de a testa perceptia pacientilor asupra serviciilor medicale si a profesionalismului personalului medical
din institutiile de sanatate publica ale judetului Timis.
Metodologie: Au fost aplicate 373 de chestionare in decursul anului 2013 pacientilor spitalizati in spitalele judetului Timis. Au fostformulate o serie de intrebari, din care am ales pentru scopul acestui studiu pe acelea care subliniau comportamentul si profesionalismul personalului medical, precum si aspectele legate de asistenta medicala oferita in timpul spitalizarii. Raspunsurile au fost
introduse intr-o baza de date si procesate cu ajutorul programului SPSS 17.0.
Rezultate: Majoritatea pacientilor au fost de sex feminin si cel mai bine reprezentat grup a fost cel cu varste cuprinse intre 41-50 ani.
Majoritatea pacientilor au prezentat un grad crescut de insatisfactie la intrebarile legate de diferite aspecte ale serviciilor medicale
oferite. O proportie crescuta de pacienti s-au declarat satisfacuti de comportamentul si profesionalismul personalului medical. Referitor
la calitatea generala a serviciilor medicale, 94.40% dintre pacienti au fost satisfacuti.
Concluzii: Cu toate ca 90% dintre pacientii intervievati s-au adresat aceluiasi spital si au fost satisfacuti de calitatea serviciilor medicale
in 94% din cazuri, acestia considera ca anumite aspecte legate de asistenta medicala si personalul medical ar trebui imbunatatite.

2014.24.2 (82)  Fiziologia - Physiology

19

LONG-TERM EFFECTS OF SIMVASTATIN THERAPY ON
CHRONIC INFLAMMATORY AND AUTOIMMUNE DISEASESELENA CIURARIU1, VICTOR DUMITRAŞCU2,3
1
Department of Functional Sciences,” Victor Babes” University of Medicine and Pharmacy, Timisoara, Romania
2
Pharmacology Department,” Victor Babes” University of Medicine and Pharmacy, Timisoara, Romania
3
Timisoara County Emergency Clinical Hospital, Romania

ABSTRACT

Objective. To examine the immunosuppressive effects mediated by long-term therapy with simvastatin in a trial of patients withchronic inflammatory and autoimmune diseases..
Patients and Methods. Fifty-five eligible patients participated in this randomized, open, retrospective and comparative survey(study group). The patients, ranging in age from 16 to 76 years, redeemed a prescription for simvastatin for impaired lipidic profile.
Inclusion criteria: patients were classified in three subgroups, accordingly with their certified chronic inflammatory and autoimmune
disease: rheumatoid arthritis (RA) (n = 23), lupus erythematosus (LES) (n = 22), and respectively sclerosis multiple (SM) (n = 10).
Simvastatin was administered during a 6-month period in a daily evening dose of 40 mg, as an adjunct to existing disease-modifying
antirheumatic drug therapy.
Patients were followed up over 6 months. Clinical disease activity variables and acute-phase reagents were analyzed comparatively.
Results. Three major diseases have been revealed in the study group, respectively: 67 % cases of rheumatoid arthritis (RA), 22%cases of lupus erythematosus (LES) and 11% of sclerosis multiple (SM). After simvastatin treatment, pain relief mobilization was
significantly improved in 53% of cases of RA, 27% cases with LES and 20 % cases of SM. 63 % of the patients with RA improved
their physical activity. C-Reactive Protein (CRP) and erythrocyte sedimentation rate declined by 67% and 35%, respectively after
6-month simvastatin therapy and interleukin -6 (IL-6) decreased from 107.22 pg/mL to 18.16 pg/mL.
Conclusion. In addition to the beneficial effects of cholesterol reduction, immune modulation in chronic inflammatory and autoimmune diseases may contribute to the cardio protective effect of simvastatin.
Key words: simvastatin, immunomodulatory role, chronic inflammatory and autoimmune diseases

INTRODUCTION
Simvastatin belongs to the most widely class of drugs used
as lipid-lowering agents. Based on their pleiotropic spectrumof activity, several studies revealed complex immunologicalpathways modulated by statin therapy (1,7).
Simvastatin acts as a 3-hydroxy-3methylglutamyl-coenzyme
A (HMG-CoA) which catalyses the conversion of HMG-CoA tomavelonic acid (Figure 1):
HO

O

H
O

O
O H

Simvastatin

Fig. 1. Chemical structure of simvastatin

Published studies revealed immunological effects that indicate
the impact of statin therapy on immune inflammatory responses(3). Simvastatin decreases chemotaxis and lipopolysaccharide
mediated release of tumour necrosis factor-α (TNT- α).
Previous researchers have pointed out the important role ofsimvastatin on bone remodeling by the finding that nitrogencontaining bisphosphonates exert their cytotoxic effects on
osteoclasts by interfering with the mevalonate pathway (5,9).
Long-term simvastatin treatment exerts a dual beneficial pharmacological and clinical effect on the lipidic profile and bonemineral density, demonstrating positive consequences in clinicalpractice, as we revealed in a 2-years research (2).
Systematically review of the literature data point out the nonlipidic mechanisms – called “pleiotropic”, which have as a result
the lowering of inflammation by simvastatin therapy. Most of thesestudies focus on the rheumathoid arthritis (RA), because this disease is associated with a higher risk of cardiovascular disease (12).
The aim of the present study was to retrospectively evaluate
the immunosuppressive effects mediated by long-term therapy
with simvastatin in a trial of patients with chronic inflammatory
and autoimmune diseases.
MATERIALS AND METHODS
Fifty-five eligible patients participated in this randomized,
open, retrospective and comparative survey (study group).
The patients, ranging in age from 16 to 76 years, redeemed aprescription for simvastatin for impaired lipidic profile.
Inclusion criteria: patients were classified in three subgroups,
accordingly with their certified chronic inflammatory and autoimmune disease: rheumatoid arthritis (RA) (n = 23), lupus
erythematosus (LES) (n = 22), and respectively sclerosis multiple

Received 25th of April 2014. Accepted 15th of May 2014. Address for correspondence: Elena Ciurariu, MD, PhD student, Department of Functional Sciences, ”Victor
Babes” University of Medicine and Pharmacy, 2 Eftimie Murgu Square, 300041 Timisoara, Romania, phone: +40 723081655; e-mail: ciurariu.elena@gmail.com

20

Fiziologia - Physiology  2014.24.2 (82)(SM) (n = 10).
Exclusion criteria: patients with multiple co-morbidities, drug
abuse, intolerance to simvastatin or previous statin therapy within
the last 6 month.
This study was performed between February 2013- July 2013, in
accordance with the ethical standards of the responsible institutional
committee on human experimentation and with the declaration of
Helsinki, as revised in 2000. All the participants gave informed consent and were recruited from the patient lists of family healthcare
providers from Timisoara, Romania.
Simvastatin was administered during a 6-month period in a daily
evening dose of 40 mg, as an adjunct to existing disease-modifying
antirheumatic drug therapy.
Patients were followed up over 6 months. Clinical disease activity
variables and acute-phase reagents were analyzed comparatively.
Sera from all the 55 patients from the study group were analyzed
for interleukin-6 (IL-6) by enzyme-linked immunosorbent assay(ELISA) using commercially available kits; baseline plasma levels
were compared with those obtained after 6 month of simvastatin
therapy.
All statistical analyses were performed using the SPSS software
package (version 6.0 for Windows, SPSS Inc, Chicago, IL.).
RESULTS AND DISCUSSIONSOf the patients from the study group, 5 (9.09 %) were aged between 16 and 30 years, 9 (16.36 %) between 31 and 40 years, 12 (21.82 %) between 41 and 50 years, 22 (40 %) between 51 and 60
years , 7 (12.73%), between 61 and 74 years. The group consistedof 55 patients, 33 females (60 %) and 23 male patients (40 %).
The study revealed maximum incidence of chronic and autoimmune disorders which corresponds to the period 51-60 years, female
sex predominance.
From the viewpoint of clinical diagnosis, three major disease
have been revealed in the study group, respectively: 67 % casesof rheumatoid arthritis (RA) , including bilateral form prevailed in
53% of cases compared with the unilateral - 18 % of cases, 22%cases of lupus erythematosus (LES) and 11% of sclerosis multiple(SM) (Figure 2).

Legend: RA = Rheumatoid Arthritis; LES = Lupus Erythematosus;
SM = Sclerosis Multiple
Fig.2. Patient distribution in the study group according to the type of
autoimmune disease

In terms of clinical simvastatin treatment administered forsix month, pain caused by joint mobilization was observed in
all cases studied initially (100 % ); after simvastatin treatment,
pain relief mobilization was significantly improved in 53% ofcases of rheumatoid arthritis (RA), 27% cases with lupus
erythematosus (LES) and 20 % cases of sclerosis multiple(SM) (Figure 3).
2014.24.2 (82)  Fiziologia - Physiology

Legend: RA = Rheumatoid Arthritis; LES = Lupus Erythematosus;
SM = Sclerosis Multiple
Fig. 3. Pain relief mobilization according to the type of autoimmune
diseaseFunctional impotence associated with the degree of limitationof physical activity of the patient under investigation was significantly improved after 6 month treatment with simvastatin, andthe best results were the following (figure 4):− 63 % of the patients with rheumatoid arthritis (RA) improved their physical activity;− 42 % of the patients with lupus erythematosus (LES) and− 22 % of the patients with sclerosis multiple (SM).
These results highlight the immunomodulatory role of simvastatin in rheumatoid arthritis (RA), which acts by decreasing the
inflammatory synovitis, articular destruction, and accelerated
atherogenesis, characteristic for all the chronic inflammatory
and autoimmune diseases (11).

Legend: RA = Rheumatoid Arthritis; LES = Lupus Erythematosus;
SM = Sclerosis Multiple
Fig. 4. Improving physical activity according to the type of autoimmune disease

All these effects contribute to improving patient’s lives, translated by reducing pain at rest and on mobilization, increased
general joint mobility, improved walking tolerance and decrease
the patient’s debilitating physical dependence.
Compared to baseline, C-Reactive Protein (CRP) and erythrocyte sedimentation rate declined by 67% and 35%, respectively
after 6-month simvastatin therapy. In the study group, CRP decreased from 21 to 8.40 mg/dl;a significant drop in plasma CRP
level was achieved after the six month of simvastatin treatment.
A special marker of inflammation is Interleukin-6 (IL-6), which
had as baseline a mean serum concentration of 107.22 pg/mL(range 68.52 - 146.40 pg/mL). From these, 22 cases (40%)
recorded levels higher than100 pg/mL, ten times the upper limit
of normal. Cases with lupus erythematosus (LES) recorded thesehigh values of IL-6 at baseline, as described in the literature (10).
After 6 month of simvastatin therapy, the IL-6 levels decreased21significantly, achieving a mean concentration of 18.16 pg/mL(range 8.78 - 84.12 pg/mL).
Interleukin-6 (IL-6) variations are revealed in Figure 5:

Legend: IL-6 = Interleukin-6
Fig.4. Comparison of Interleukin-6 levels at baseline and after 6
month of simvastatin therapy in patients with autoimmune disease

Extensive experimental studies and more recently many clinical
trials have strongly suggested statins to possess an important rolein chronic inflammatory and autoimmune diseases, mainly mediated by their anti-inflammatory and immunumodulatory properties(4,8).
Our results are in accordance with those of previous researches,
who revealed that simvastatin reduces proinflammatory cytokine
production in vitro by T cell contact-activated macrophages inperipheral blood and synovial fluid (6).
The greatest improvements have been revealed in patients withmore active autoimmune diseases.
The presented study has some limitations: One limitation is thesmall sample size and the broad.confidence interval. The precise
mechanisms of simvastatin action on autoimmune diseases will
be clarified by further studies.

CONCLUSIONSIn addition to the beneficial effects of cholesterol reduction,
immune modulation in chronic inflammatory and autoimmune diseases may contribute to the cardio protective effect of simvastatin.
REFERENCES
1. Buemi M, Allegra A, Corica F, et al. Effect of fluvastatin on proteinuria
in patients with immunoglobulin A nephropathy. Clin Pharmacol Ther
2000; 67: 427-431.  
2. Ciurariu Elena, Dumitraşcu V, Matusz Anca-Alexandra. Beneficial
effects of 2-year treatment with simvastatin on bone mineral density ina cohort of postmenopausal women with impaired plasma lipidic profile.
Archives of the Balkan Medical Union 2013; 48(3): 271-274.
3. Holstein SA, Wohlford-Lenane CL, Hohl RJ. �������������������������
Isoprenoids influence expression of Ras and Ras-related proteins. Biochemistry 2002; 41: 13698-704.
4. Jury EC, Ehrenstein MR. Statins:mmunomodulators for autoimmune
rheumatic disease? Lupus 2005; 14:192-196.
5. Laaksonen R, Jokelainen K, Laakso J, et al. The effect of simvastatin treatment on natural antioxidants in low-density lipoproteins andhigh-energy phosphates and ubiquinone in skeletal muscle. Am J Cardiol
1996; 77: 851–854.
6. Leung BP, Sattar N, Crilly A, et al. A novel anti-inflammatory role forsimvastatin in inflammatory arthritis. J Immunol. 2013; 170:1524-1530.
7. Maron DJ, Fazio S, Linton MF. Current perspectives on statins.
Circulation 2000; 101: 207-223.
8. McCarey DW, Sattar N, McInnes IB. Do the pleiotropic effects ofstatins in the vasculature predict a role in inflammatory diseases? Arthritis
Res Ther 2005; 7:55-61.
Montagnani A, Gonnelli S, Cepollaro C, et al. Effect of simvastatin
treatment on bone mineral density and bone turnover in hypercholesterolemic postmenopausal women: a 1-year longitudinal study. Bone,
2003; 32(4): 427-433.
Noel B. Statins and lupus erythematosus. Rheumatology 2004; 43(3):397-398.
Shepherd J, Hunnighake DB, Barter P, Mckenney, Hutchinson HG.
Guidelines for lowering lipids to reduce coronary artery disease risk: A
comparison of rosuvastatin with atorvastatin, pravastatin and simvastatin for achieving lipid lowering goals. Am J Cardiol 2003; 91: 11C-19C.
Van Doomum S, Mc Coll G, Wicks IP. Accelerated atherosclerosis. Are
extra-articular feature of rheumatoid arthritis? Arthritis Rheum 2002;
46: 862-873.

EFECTELE PE TERMEN LUNG ALE TRATAMENTULUI CU SIMVASTATIN IN
AFECTIUNILE INFLAMATORII CRONICE SI AUTOIMUNEREZUMAT

Obiective. Investigarea efectelor imunosupresoare mediate de terapia pe termen lung cu simvastatina, pe un lot de pacienti cu
afectiuni inflamatorii cornice si autoimmune.
Pacienti si Metode. Au participat 55 de pacienti eligibili la acest studiu randomizat, deschis, retrospectiv si comparativ (grup destudiu). Pacientii, cu varste cuprinse intre 16 si 76 de ani au primit prescriptii medicale pentru simvastatin in cazuri de alterare a
profilului lipidic. Criterii de includere: pacientii au fost clasificati in trei subgrupuri, in functie de diagnosticul pozitiv pentru afectiuni
inflamatorii cornice si autoimune: artrita reumatoida (RA) (n = 23), lupus eritematos (LES) (n = 22), si respectiv scleroza multipla(SM) (n = 10). Simvastatina a fost administrata pe o perioada de 6 luni in doza zilnica de 40 mg, seara, ca tratament asociat terapiei
anti-reumatice existente. Pacientii au fost urmariti pe o perioada de 6 luni. Au fost analizate comparativ variabilele de activitate alebolii si parametrii de faza acuta.
Rezultate. Au fost evidentiate trei afectiuni majore in grupul de studiu: 67 % cazuri de artrita reumatoida (RA), 22% de cazuri delupus eritematos (LES) si 11% de scleroza multipla (SM). Dupa tratamentul cu simvastatin, mobilizarea ca urmare a reducerii dureriia fost imbunatatita semnificativ in 53% din cazurile de RA, 27% din cazurile de LES si 20 % din cazurile de SM. 63% dintre pacientii
cu RA au prezentat o imbunatatire a activitatii fizice. Preoteina C reactiva (CRP) si viteza de sedimentare a eritrocitelor au prezentat
o scadere cu 67% si respectiv cu 35%, dupa 6 luni de terapie cu simvastatina, iar interleukina 6 (IL-6) a prezentat o scadere de la107.22 pg/mL la 18.16 pg/mL.
Concluzie. Aditional efectelor benefice asupra reducerii valorii colesterolului, modularea imuna in afectiunile cornice inflamatorii si
autoimun ar putea avea o contributie la efectele cardioprotectoare ale simvastatinei.
Cuvinte cheie: simvastatina, rol imunomodelator, boli cornice inflamatorii şi autoimune.

22

Fiziologia - Physiology  2014.24.2 (82)PROSPECTIVE STUDY OF LAPAROSCOPIC GREAT CURVATURE
PLICATION – EFFECTS ON WEIGHT LOSS AND HUNGERHORMONE LEVELSDOBRESCU AMADEUS, VERDEŞ GABRIEL, TARŢA CRISTI, BREBU DAN, STOICA LAURIAN,LAZĂR CAIUS, DUŢĂ CIPRIAN
Second Surgery Clinic of Timisoara County Hospital,”Victor Babeș” University of Medicine and Pharmacy Timișoara

ABSTRACTIntroduction: Laparoscopic greater curvature plication (LGCP) introduces no implantable device, requires no gastric or intestinal
resection or intestinal bypass, and one of the most important advantages is that this technique is potentially reversible; also it can befollowed by more extensive procedures. Aim of our study is to evaluate the excess weight loss (EWL) and the ghrelin levels changes
in a series of obese patients that underwent LGCP.
Methods: This is a prospective non-randomized study and includes all the patients operated with LGCP during 2012 in Second
Surgical Unit of the Timisoara County Hospital.Data were collected from the patients’ files, questionnaires and from specified followup visits at 1, 3, 6 and 12 months. All the patients fulfilled the follow up at 12 months at the moment of this writing.Main intervention
was considered the LGCP. Main outcome was excess body weight loss (EWL), secondary outcomes were ghrelin levels, modified
hunger and satiety sensations, weight regain, surgical complications, readmissions, intra-operative data – operating time, blood loss.
Results:14 patients were included in the study – they underwent LGCP, 12 females, 2 males. All the procedures were performed
entirely laparoscopic. EWL was at 1 month - 25.7%, 3 months - 34.9%, 6 months - 44.9%, 12 months - 57.6%. Before surgery the
median level of ghrelin was 283.05 pg/ml, after surgery the level dropped at 160.60 pg/ml at 1 month and to 65.40 pg/ml at 3 months.
Comorbidities resolve in 4 patients out of 8 patients. There were two minor complications – nausea and vomiting that prolonged thehospital stay and needed conservative treatment. Two patients had less weight loss due to diet and lack of motivation.
Conclusion: LGCP can be a successful bariatric procedure performed by a trained bariatric surgeon on a population of selectedpatients with strong motivation. EWL is satisfactory; complications are rare and mild and usually need only conservative treatment.
Key words: ghrelin, laparoscopic placation, weight loss

INTRODUCTION
Of all treatments for obesity, metabolic/bariatric surgery has
shown the greatest sustained effectiveness in achieving weightloss and comorbidity improvement, with social reinsertion of thesubjects. The evolution of our society with the food not being
a concern for ordinary people results in an increased trend inprevalence of morbid obesity around the world (1,2). Same thing
happened in Romania since the introduction of the fast-foodculture. Influence of diet and exercise in morbid obese patients
is about 10% in long term period; thus, in case of life-stylemodification failure, bariatric surgery could be considered (3).
Traditionally, the primary mechanisms through which bariatricsurgery achieves its outcomes are believed to be the mechanicalrestriction of food intake, reduction in the absorption of ingested
foods, or a combination of both (4). More or less all the bariatricprocedure lead to a partial resection of the stomach, opening of
the stomach or of the small bowel, either leave implants consisting in different devices. The very few prior preclinical and clinicalstudies of laparoscopic greater curvature plication (LGCP) — thenewest operation in metabolic surgery — suggest the potential
for LGCP to provide safe, significant weight loss and improvement of metabolic parameters similar to that shown by currently

accepted restrictive metabolic procedures. LGCP introduces no
implantable device, requires no gastric or intestinal resection or
intestinal bypass, and one of the most important advantages isthat this technique is potentially reversible; also it can be followedby more extensive procedures (5).
Nevertheless the technique is the less expansive bariatricprocedure, a major advantage in a developing country with more
and more obese and no proportional health financing.
Since introduction of the technique by Talebpour in 2007 (6),there are few studies evaluating the technique and the outcomes,
the largest one published by the same author in 2012 consistingin about 800 patients (7).
Our study is a prospective series including all the patients
operated in our hospital in 2012 with a LGCP and followed fora year, we also determined the hunger hormone ghrelin at 0, 1
and 3 months post-operatively and applied questionnaires about
hunger and satiety feelings before and after the procedure.For
our knowledge this is the first series published from our country.
METHODSThis is a prospective non-randomized study and includes all
the patients operated with LGCP during 2012 in Second Surgical

Received 15th of June 2014. Accepted 20th of July 2014. Address for correspondence: Amadeus Dobrescu, MD, PhD Student, Department of Surgery, University of Medicine and Pharmacy “Victor Babeș” Timișoara, Iosif Bulbuca Street, No. 10, Timișoara, România; phone: +40740015355, e-mail address: amadeusdobrescu@gmail.com

2014.24.2 (82)  Fiziologia - Physiology

23Unit of the Timisoara County Hospital.Data were collected fromthe patients’ files, questionnaires and from specified follow-up
visits at 1, 3, 6 and 12 months. All the patients fulfilled the follow
up at 12 months at the moment of this writing.
Main intervention was considered the LGCP. Main outcome
was excess body weight loss (EWL), secondary outcomes wereghrelin levels, modified hunger and satiety sensations, weightregain, surgical complications, readmissions, intra-operative
data – operating time, blood loss.
Surgical technique
The patient was placed in 30° reverse Trendelenburg position, with the operating surgeon standing between patient legs.
Pneumoperitoneum was created inserting the Veress’ needle atthe Palmer’s point, left mid clavicular line below the last rib either
on the linea alba at two thirds below xyphoid or using the open
Hasson technique for the insertion of the first trocar if previous
major abdominal surgery was encountered.
The patient was placed in 30° reverse Trendelenburg position, with the operating surgeon standing between patient legs.
Dissection started at the greater curvature of the stomach fromthe middle of the antrum and continued to the left diaphragmaticcrus, and downwards to the level of the pyllor. Communicating
vessels were ligated by LigaSure (LigaSureTMCovidien, USA),
Ultracision (Harmonic® Ultrasonic, Ethicon, USA) or clips.
Continuous suturing from 2 cm below the cardia through theantrum, 4 cm distance above the pyllorus, was performed inthis stage, making one and then two layers of plicated stomach
from the anterior wall of the stomach to the posterior wall. For
suturing 2-0 prolene (2-0 PROLENE™ Polypropylene Suture,
Ethicon, USA) was used, and the bulk of each stitch was 1 cm
with a 1-cm interval. For gastric tube calibration we used a 36
Fr tube. Sutures were extramucosal, preventing absorption by
gastric acid (8).
Drainage was used for 24 hours in order to see if there wasany bleeding.
Post-operative we administrated to all the patients proton
pump inhibitors 2*40 mg/day, anti-inflammatory and anti-nausea/
vomiting drugs. Low molecular heparins were administrated for
up to three weeks to prevent deep vein thrombosis and superior
mesenteric vein thrombosis.
Postoperative diet was resumed 24 hours after surgery andconsisted only in fluids up to the 9th POD. Next three weeks all
the food will be semi-liquid.
Measurements were performed for all patients at the five study
time points (baseline, 1, 3, 6 and 12 months post-op), as per
protocol. Body weight was measured to the nearest 0.5 kg andheight to the nearest 1 cm. BMI was calculated as body weight inkilograms divided by the square of the height in meters. Percent
excess weight loss (%EWL) was calculated according to thefollowing equations:%EWL = [(preoperative weight − follow-up weight)/preoperative weight-ideal body weight] x100
24

Questionnaires were applied at 0, 1, 3, 6 and 12 months regarding hunger and satiety sensations. Patients were required toevaluate on a scale from 1 to 10 these sensations. Weak hunger
sensation was noted as 1, strong sensation as 10. Rapid satiety
was noted as 1, whereas 10 counts for slow satiety and the needfor large meals to reach satiety.
RESULTS14 patients were included in the study – they underwent
LGCP. All the procedures were performed entirely laparoscopic.
In the Table I we have the demographics and the peri-operative
data of the patients. 8 patients had comorbidities: hypertension,
diabetes mellitus type II, dyslipidemia, osteo-articular disease
and obstructive sleep apnea.
Table I. Demographics and the peri-operative data
Patients

n=14

Sex ratio F:M

12:2

Age (years) *43

Other comorbidities

57.14%

OR time(minutes)*

51 (40 - 102)

Hospital stay(days)*
3 (2 - 10)*median value

There was no intra-operative blood loss. There was only one
readmission on the 3rd post-operative day (POD) due to incontrollable vomiting and nausea, followed by dehydration. The
treatment was conservative with intra-venous anti-inflammatory
drugs; proton pump inhibitors; parenteral fluids and nutrients.
The patient was discharge 8 days later with resumed oral foodintake and with no nausea or vomiting. Proton pump inhibitors,
anti-inflammatory and anti-nauseadrugs were prescribed per
mouth. Another patient had vomiting and nausea during hospital
stay for 10 days – same treatment was prescribed.
The results, in terms of BMI and EWL, of the procedure aresummarized in Table II. There were 2 patients lost to follow-up,
one at 6 month and another one at 12 months. Two patients had
less EWL, a male and a female, after re-screening they prove
to be sweet eaters.
Table II. BMI and EWL results0

1 month

3 months

BMI (kg/m2) 38.6

33.9

31.8

30.2

28.6

25.7

34.9

44.9

57.6

EWL %

-

6 months 12 months

BMI – body mass index; EWL – excess weight loss; average value
was used

There were no long-term complications. All the patients answered the questionnaires about hunger and satiety sensationand the results are shownin Figure 1.
Fiziologia - Physiology  2014.24.2 (82)Fig. 1. Hunger and satiety sensation score

Ten patients had ghrelin blood levels determined at 0, 1 and 3
months. Before surgery the median level of ghrelin was 283.05
pg/ml, after surgery the level dropped at 160.60 pg/ml at 1 monthand to 65.40 pg/ml at 3 months.
Comorbidities diminished at the end of the follow-up, there
were present in 4 out of 12 patients. Number of deaths wererecorded.
DISCUSSIONS
There few advantages of the LGCP – potentially reversal
procedure, lower cost, no resection, no foreign body inside theabdomen and a low rate of complications. One of the disadvantages is the lack of a randomized trial.
No need for a reversal of LGCP in our study occurred, Talebpour et al reported that reversal is possible in the first six weeks
after surgery, later on dense fibrotic adhesions are formed
between folds (7). Other authors also reveal that LGCP could
be easily reversed in an early postoperative period, before formation of dense adhesions between the layers of the plication (9).
There were no major complications recorded in our 14 patients’series. The rate of complication was 14.28%, consisting only inminor complications – nausea and vomiting, percentage that is
mentioned in the literature, our study scarce number of patients
could be a reason for the relative high percentage of minor
complications, Talebpour et al reported 16 % complications.
First the LGCP was performed by a single row of sutures, theremaining volume of the gastric tube was difficult to assessed,then two rows were used and the gastric volume got smaller.
In a detailed description of the technique Talebpour describedthe difference between the two methods the average anatomic
volume of stomach in the operating room was 100 and 50 cc inone- and tow-row plication respectively; the functional volumeof stomach in one- and two-row LGP respectively was about 25
and 15cc at first, 50 and 25cc after 2 weeks, 75 and 45cc after6
months, 100 and 60cc after 1 year and 250 and 150cc after 4
years (7). Looking at the relative small volume of the remaining
gastric tube the post-operative gastric wall edema can easily
temporary induce nausea and vomiting. Same author describedthe vomiting as common encounter two days after surgery (7).
EWL was recorded at 0, 1, 3, 6 and 12 months. Largest amountof EWL was at the patients with BMI lower than 40 kg/m2, most
likely due to more rapid social reinsertion, possibility to practice2014.24.2 (82)  Fiziologia - Physiology

physical exercices. Talebpour recommended special filters forthe candidates willing to have performed LGCP, strong motivation and discipline can play a mjor role in weight loss and is the
key factor in maintaining the EWL. The longer the follow-up theworse are the EWL (10). In our sudy the results are improved
comparing to other series on short term – at one month 25.7%
versus 20% in Talebpour series, latter on the results are surpassed by other as most likely the lack of obese support group
determined patients to fail the program they were enrolled. In
two patients the EWL was low, after reviewing the interviews wefound out that they were sweet eater, a relative contraindicationof the restrictive method.
The 57.6% EWL a year after surgery qualified the LGCP asa good choice of bariatric surgery, main drawback being theselection of the patients; method has its limit when we crossthe border of 40 kg/m2.
Comorbidities rates dropped after EWL consecutive to LGCP.
In our study there is a direct relation between EWL and comorbidities resolution. At 12 months follow-up EWL was 57.6%, andthe comorbidities rate was lower with 40%.
Hunger sensation of the obese persons who underwent LCGP
dropped dramatically in the first month after surgery, most likely
due to reduced ghrelin secretion from the cells localized at thegastric fundus. Ghrelin is supposed to be the principal hunger
hormone. The diminished ghrelin secretion resulted probably as
a consequence of the blood flow reduction on the gastric wall,
secondary to high pressure induced by the edema of the folds.
But in our study ghrelin maintain a descending trend even after
edema resolution, the median levels at three months are fourth
time the base value.
Actual knowledge place the ghrelin’s producing cells inside
the oxyntic glands, situated near the basal membrane, adjacent
to arterial blood flow, most of these cells don’t have contact withthe gastric lumen (11). Once the gastric wall edema disappeared
the hunger sensation increased his level during the follow-up,
reaching the highest level at 12 months, but not reaching theinitial level before the surgery.
Satiety sensation was rapidly installed due to reduced gastricvolume following the plication and the edema of the folds, especially during the first month when the edema was at his peak.
Same way as the hunger sensation, once the gastric reservoir
increased his volume due to edema resolution and due to pressure resulted from the continuously increased amount of the foodintake, satiety sensation was installed slower.
Restrictive bariatric procedure induced a EWL mainly to
reduced gastric volume and reduced food intake, but once thegastric volume is increased as an adaptation of larger and larger
meals the slop of EWL decrease. Regain can be a possibility. The
motivation and the cooperation of the obese patients become
essential in maintaining the EWL. Others keys elements in the
EWL after LGCP are the hormonal changes promoted by the
reduced gastric volume, the reduced appetite induced by thelower ghrelin levels and the increase in the metabolism, thesechanges maintain the EWL and resolve some comorbidities thatare associated with the obesity.
25

CONCLUSION
LGCP is a new bariatric procedure that it seems to be safeand effective in the short-term results. It could be more attractive, particularly in developing countries, because of a lower
morbidity, lower cost and equal efﬁcacy compared with otherrestrictive procedures. We believe that LGCP could be an acceptable option especially for patients with BMI under 40kg/m2,
who refuse an irreversible procedure such as sleeve gastrectomy.
Postprandial plasma levels of ghrelin are decreased at 1 and 3
months after the LGCP, a finding that has also been noted after
other bariatricoperations such as sleeve gastrectomy, verticalbanded gastroplasty and biliopancreatic diversion with duodenalswitch (12,13).
Aknowledgements: The research of Dr. Dobrescu Amadeus
was supported by PhD fellowship POSDRU 107/1.5/S/78702.
REFERENCES
1. Finucane MM, Stevens GA, Cowan MJ et al. National, regional,
and global trends in body-mass index since 1980: systematic analysisof health examination surveys and epidemiological studies with 960
country-years and 9 - 1 million participants. Lancet 2011; 377(9765):557-67.
2. Ruesten A, Steffen A, Floegel A et al. Trend in obesity prevalencein European adult cohort populations during follow-up since 1996 andtheir predictions to 2015. PLoS One 2011, 6: e27455.
3. Himpens J, Cadiere GB, Bazi M et al. Long-term���������������������������
Outcomes of Laparoscopic Adjustable Gastric Banding. Arch Surg 2011; 146(7): 802-807.

4. DeMaria EJ. Bariatric surgery for morbid obesity. N Engl J Med.
2007; 356(21): 2176-83.
5. FriedM.;DolezalovaK.; Buchwald J.N. et al; Laparoscopic Greater
Curvature Plication (LGCP) for Treatment of Morbid Obesity in a Seriesof 244 Patients; Obes Surg 2012; 22: 1298-1307.
6. Talebpour M, Amoli BS. Laparoscopic total gastric vertical plicationin morbid obesity. J Laparoendosc Adv Surg Tech A 2007; 17: 793-8.
7. Talebpour M, Motamedi SMK, Talebpour A et al. �������������������
Twelve year experience of laparoscopic gastric plication in morbid obesity: developmentof the technique and patient outcomes. Annals of Surgical Innovation
and Research 2012; 6: 7.
8. Copaescu C. Plicaturarea marii curburi gastrice pe cale laparoscopicã (pentru tratamentul obezitãtii morbide). Chirurgia, 2011; 106: 91-97.
9. Atlas H, Yazbek T, Garneau PY, Safa N, Denis R. Is there a future
forlaparoscopic gastric greater curvature plication (LGGCP)? A Reviewof 44 Patients. Obes Surg 2013; 23:1397-403.
10. Ji Y, Wang Y, Zhu J et al. A systematic review of gastric plication
for the treatment of obesity. Surgery for Obesity and Related Diseases
2014.
11. Ichiro S, Mami Y, Takafumi S, et al. Ghrelin-producing cells exist
as two types of cells, closed-and opened-type cells, in the rat gastrointestinal tract. Peptides 2002; 23:531-6.
12. Kotidis EV, Koliakos GG, Baltzopoulos VG, et al. Serum ghrelin,
leptin and adiponectin levels before and after weight loss: comparisonof three methods of treatment-a prospective study. Obes Surg. 2006;
16(11):1425-32.
13. Scott WR, Batterham RL. Roux-en-Y gastric bypass and laparoscopic sleeve gastrectomy: understanding weight loss and improvements in type 2 diabetes after bariatric surgery. Am J Physiol Regul
Integr Comp Physiol. 2011; 301(1): R15-27.

STUDIU PROSPECTIV PRIVIND PLICATURAREA LAPAROSCOPICA A MARIICURBURI – EFECTELE ASUPRA PIERDERII IN GREUTATE SI A NIVELULUIHORMONILOR DE FOAMEREZUMAT

Introducere: Plicaturarea laparoscopica a marii curburi (LGCP) nu necesita introducerea dispozitivelor implantabile, nici rezectia
gastric sau intestinala sau bypass intestinal, iar unul dintre cele mai importante avantaje ale acestei tehnici este potential reversibilitate;
de asemenea, poate fi urmata de procedure extensive. Scopul studiului nostru a fost de a evalua pierderea in greutate (excessive
weight loss-EWL) si modificarile nivelului ghrelinei pe un lot de pacienti obezi care au fost supusi LGCP.
Metode: Acesta este un studiu prospectiv, ne-randomizat si a inclus toti pacientii operati prin LGCP in cursul anului 2012 in Sectia II
Chirurgie a Spitalului Judetean de Urgenta Timisoara. Datele au fost colectate din fisele pacientilor, chestionare si din consulturile deurmarire de la 1, 3, 6 si 12 luni. Toti pacientii au fost prezenti la consultul de control de la 12 luni. Interventia chirurgicala majora a fostconsiderata LGCP. Principalul rezultat a fost pierderea in greutate (EWL), iar efectele secundare au fost: nivelul ghrelinei, modificareasenzatiei de satietate si foame, cresterea in greutate, complicatiile chirurgicale, spitalizari ulterioare, datele intra-operatorii – durata
interventiei chirurgicale, pierderea sanguina.
Rezultate: 14 pacienti au fost inclusi in studiu – supusi interventiei LGCP, 12 pacienti de sex feminin si 2 de sex masculin. Toate
procedurile au fost efectuate exclusiv laparoscopic. La interval de 1 luna, EWL a fost – 25,7%, la 3 luni – 34,9%, la 6 luni – 44,9%,
iar la 12 luni – 57,6%. Inaintea interventiei chirurgicale, nivelul mediu al ghrelinei a fost 283,05 pg/ml, iar dupa interventia chirurgicala
a scazut la 160,60 pg/ml la interval de 1 luna si la 65,40 pg/ml la 3 luni. Comorbiditatile au fost solutionate la 4 din 8 pacienti. Au
existat si doua complicatii minore – greturi si varsaturi, care au prelungit spitalizarea si au necesitat tratament conservator. Doi dintre
pacienti au prezentat o scadere redusa in greutate datorat lipsei motivatiei.
Concluzie: LGCP poate fi o procedura bariatrica de succes, efectuata de chirurgi bariatrici instruiti, pe un lot de pacienti selectionati,
cu motivatie puternica. EWL este satisfacator; complicatiile sunt rare si usoare, necesitand de obicei doar tratament conservator.

26

Fiziologia - Physiology  2014.24.2 (82)TELOMERE LENGHT CHANGES IN ALZHEIMER DISEASEGROZA SABINE1, ANGHEL SIMONA1,2, MIRABELA CRISTEA2, CARMEN TATU1,2, GABRIELATANASIE1,2, CARMEN PANAITESCU1,2, OANA GAVRILIUC1, VIRGIL PAUNESCU1,2, FLORINA
BOJIN1
Department of Functional Sciences, “Victor Babes” University of Medicine and Pharmacy Timisoara
Immunophysiology and Biotechnologies Center, Clinical Emergency County Hospital Timisoara

ABSTRACT

Telomeres are regions of repetitive DNA at the end of eukaryotic chromosomes, which prevent chromosomal instability. Telomere
shortening in peripheral blood mononuclear cells (PBMCs) has been associated with biological age and several chronic degenerative
diseases. Telomere shortening is linked to age- related disease including Alzheimer’s disease (AD) and has been reported to be
reduced in leukocytes of AD patients. The aim of the present study was to measure telomere length in lymphocytes of patients with
AD compared to healthy subjects. We used a new flowcytometric method of hybridization, which provided data referring to relative
telomere length compared to control cells. Our data show significant shorter telomere length in AD patients (22.95%) compared tocontrols (55.49%). In conclusion, the telomere length is age-dependent in lymphocytes and decreased in AD patients, which could
mean that the AD pathology may contribute to telomere length shortening. The high variability of telomere lengths in individuals
suggests that it will not be useful as a general biomarker for AD. However, it could become a biomarker in personalized long-termmonitoring of an individuals’ health.
Key words: Alzheimer disease (AD), telomere length, flowcytometric hybridization, lymphocytes

INTRODUCTION
Prior to mitotic cell division, all cellular DNA is duplicated by
the action of a DNA polymerase. However, this enzyme does not
replicate the very ends of the chromosomes, the so-called telomere
region. As a consequence, the telomeres get shorter after eachcell division. In normal somatic cells, this shortening contributes
to cellular senescence and the telomeres act as a “mitotic clock”.
Telomeres in all vertebrates are composed of a sequence of six
nucleotides (TTAGGG) repeated from a few hundred to several
thousand times. The length of telomeres is species-specific andranges from 5-20 kilobases in man to 20-150 kilobases in mice.
Telomeres are considered to have a protective function, keeping
the chromosome ends intact, and thereby protecting the underlying
genes and avoiding fusion of chromosome ends. Some specialized cells maintain their capacity to divide. Examples are germ
cells, fetal cells, haematopoietic stem cells, and basal cells of theepidermis. These cells overcome the ageing problem by the actionof the enzyme, telomerase (1). Telomerase maintains telomere
length by adding hexameric repeats to the telomeric ends of thechromosomes, thus compensating for the continued shortening oftelomeres that would otherwise occur. The length of the telomeres
depends both on the age of the cell donor and on the number oftimes the cell has divided. However, all normal diploid cells are“mortal” and have a limited capacity to proliferate in culture. In
contrast, established tumor cell lines can divide forever and are“immortal” mainly due to reactivation of telomerase. Almost 90% of
human primary tumors express telomerase, while the cells of mostnormal tissues lack this enzyme activity. Recently, the hypothesisthat the activation of telomerase is necessary for extension of thelife-span of human cells by avoiding telomere shortening has been
proven (1). Telomerase activity and the preservation of telomere
length are, therefore, important for the cancerous process and forthe sustained growth of most solid tumors both in vivo and in vitro.
The telomere length can be measured by quantitative FISH
in both human (2) and other vertebrate cells (3). Chromosomespecific factors regulate the length of the individual telomere. Shorttelomeres on the human chromosome 17p might be responsible
for the frequent loss of 17p alleles in human cancer, e.g. thetumor suppressor gene for the p53 protein (4). The geneticallydetermined variation in telomerase activity between individuals(5) makes telomere length measurements relevant for the studyof age-related diseases. The average telomere length of chromosomes in a cell population can be estimated by quantitative FISH
in flowcytometry (6, 7).
Alzheimer’s disease is the most common cause of dementia
in the elderly, and is characterized by the extracellular depositsof amyloid plaques and intracellular neurofibrillary tangles (8,9).
Advancing age is the major risk factor for the development of
Alzheimer’s disease (10) and overall prevalence of Alzheimer’s
disease is 14 times higher after 85 years of age compared to 65–69
years of age. Despite the close intimacy between ageing and
Alzheimer’s disease, very limited knowledge is currently available
on this aspect. At the molecular level, ageing is associated withthe accumulation of nuclear DNA damage (11,12) and a variety
of studies have provided evidence for accumulation of nuclear
DNA damage and activation of DNA damage checkpoints (suchas apoptosis and senescence) in ageing tissues (13).
Telomere shortening represents a cell intrinsic mechanism foraccumulation of DNA damage that may increase the sensitivityof neurons towards oxidative stress and proteotoxicity. Further it
has been reported that oxidative stress can increase telomere
shortening (14). A number of studies have documented telomere
shortening from different organs during ageing (15,16). However,in agreement with very limited cell division, telomere shortening

Received 10th of March 2014.Accepted 25th of May 2014. Address for correspondence: Sabine Groza, MD, PhD student, Department of Functional Sciences, Immunology, “Victor Babes” University of Medicine and Pharmacy Timisoara, EftimieMurgu Square No. 2A, RO-300041, Timisoara, Romania, Phone/fax: + 40 256490507;
email: sabine_groza@yahoo.com

2014.24.2 (82)  Fiziologia - Physiology

27was not observed in whole human brain samples (17). Similar studies on patients with Alzheimer’s disease revealed divergent results,
while telomere shortening was accelerated in peripheral blood cells(18), opposite results were obtained from neurons (19). Telomeresshorten as a consequence of cell division and limit the replicative
potential of cells (17). In the adult mammalian brain, glia but notneurons can divide, although active neurogenesis takes place in thesubventricular zone of lateral ventricles and the subgranular zone ofthe dentate gyrus (20). According to this paradigm, age-dependent
telomere shortening and decrease in adult neurogenesis could
represent a predisposing factor for Alzheimer’s disease progression.
Apart from adult neurogenesis, telomere shortening could also
alter the proliferative capacity of microglia and inflammatory signaling
in the ageing brain. Studies on fibroblasts have shown that senescence induces an aberrant secretion of pro-inflammatory cytokines(21) and similar observation was also made from ageing telomere
dysfunctional mice (22).
Despite the intimate association of ageing with telomere shortening and Alzheimer’s disease development, its functional relevanceon Alzheimer’s disease progression has not been fully investigated.
The aim of the present study was to measure telomere length inlymphocytes of patients with AD compared to healthy subjects, andto conclude whether immune cells dysfunction can induce changes
in inflammatory mediators release, thus contributing to disease
maintenance and progression.
MATERIALS AND METHODS
We used peripheral blood mononuclear cells (PBMCs) harvested
from Alzheimer disease (AD) patients (n=30) and control patients(n=30). The age of patients with Alzheimer disease ranged from57 to 84-year old, while the control samples were harvested fromelder subjects with similar age range, with no prior diagnostic of
Alzheimer disease or other forms of dementia. After signing theInformed Consent form (elaborated based on a well-establishedprotocol, approved by the Ethics Committee of “Victor Babes”
University of Medicine and Pharmacy Timisoara), we proceddeed
to further analysis.
10 ml of total peripheral blood was used for separation of
PBMCs from both AD and control subjects. Peripheral blood was
harvested on anticoagulant (Heparin), further diluted with Dulbecco’s
Phosphate Buffered Saline (PBS; Sigma-Aldrich Company, St.
Louis, USA) with a dilution ratio of 1:1, and then we used gradientcentrifugation (Ficoll-Hipaque, GE Healthcare Bio-Sciences AB)
for 25 minutes at 2500 rpm for peripheral blood mononuclear cells(PBMCs) separation. All PBMCs samples were cryopreserved until
further use.
For determination of telomere length of both study and control
patients groups, we used Telomere PNA Kit/FITC for Flow Cytometry(Dako), which provides a convenient method for measuring telomericsequences in vertebrate interphase hematopoietic cells, according
to the manufacturer instructions detailed below. Instead of 1301 cellline (tetraploid), we used K562 as control cell line (triploid) and we
adjusted the DNA index accordingly (DNA index = 1.5).
In addition to the fluorescein-conjugated peptide nucleic acid
(PNA) probe in hybridization solution, the kit contains hybridization
solution without probe for correction of cell autofluorescence, wash
solution for post-hybridization washes and DNA-staining solution for
28

identification of G0/1-cells. In a mixture of sample cells and control
cells, the sample DNA is denatured at 82 °C for 10 minutes in an
Eppendorf tube in the presence of hybridization solution with orwithout fluorescein-conjugated PNA telomere probe. Then, hybridization takes place in the dark at room temperature overnight. The
hybridization is followed by 2 washes in wash solution at 40 °C for
10 minutes each. Finally the cells are resuspended in DNA-staining
solution and stored in the dark at 2-8 °C for 2-3 hours before analysis
by flow cytometry. The specific fluorescence from telomere staining
will be observed in FL1, and fluorescence from DNA staining will beobserved in FL3. Compared with the traditional telomere restriction
fragment (TRF) method, a major advantage of the Dako Telomere
PNA Kit/FITC assay is that it does not suffer from the interaction ofsubtelomere sequences.
The probe of the kit recognizes telomeric repeats, but also potential intrachromosomal TTAGGG repeats will be included in the
fluorescence signal. In the literature the prevalence of non-telomeric
TTAGGG sequences vary (10, 11), but were found to be few andshort using the sensitive Q-FISH method with a PNA probe (4).
These data and the good correlation with Southern blotting support the idea that intrachromosomal telomere sequences are ofless significance for the total hybridization signal detected by flow
cytometry (6).
The samples are run in duplicate and that the mean values ofthe duplicate determinations are used for data analysis. PNA is asynthetic DNA/RNA analog capable of binding to DNA/RNA in asequence-specific manner obeying the Watson-Crick base pairingrules. In PNA the sugar phosphate backbone has been replaced
by a neutral peptide/polyamide backbone keeping the distances
between the bases exactly the same as in DNA (8). The neutral
peptide backbone gives PNA excellent properties for hybridizing to
DNA/RNA. In addition, PNAs are highly resistant to degradation by
DNases, RNases, proteinases, and peptidases. The method is optimal for estimation of telomere length, as the fluorescence intensityof the cells is directly correlated to the length of the telomeres (6, 7).
After flow cytometric analysis, the data obtained can be used for
determination of a relative telomere length (RTL). The RTL value iscalculated as the ratio between the telomere signal of each sample
and the control cell (K562 cell line) with correction for the DNA index
of G0/1 cells. Sample cells and control cells should are analyzed
separately for DNA ploidy in order to be able to make an accuratecompensation for the cellular DNA content, according to Vindeløv
et al. (12). This correction is performed in order to standardize thenumber of telomere ends per cell and thereby telomere length per
chromosome.
RESULTS
Samples hybridized with the Telomere PNA Probe/FITC should
exhibit a fluorescence signal in FL1, which is higher than the
background/autofluorescence signal obtained from the sample ofthe same cells hybridized with the hybridization solution without
probe. When analyzing flow cytometry data for cells hybridized
with or without the Telomere PNA Probe/FITC it is important only
to look at cells in the G0/1-phase of the cell cycle where the cellhas one copy of its genome. This is achieved by setting the correct
gates. If cells in S or G2/M-phase are not removed by gating, theestimated RTL will not be per genome equivalent.
Fiziologia - Physiology  2014.24.2 (82)We analyzed human the PBMCs isolated from blood by gradient centrifugation, and extracted the lymphocytes fraction fortelomere length determination. The cells were mixed 1:1 with
control cells (K562 cell line). The gated cells were then displayedin the FL3-height versus FL1-height dot plot. We identified cells in
the G0/1-phase and set gates around these populations. Statistical data on these cells, together with the DNA index of the cells
determined by traditional DNA measurement, were then used for
calculation of RTL of the sample cells compared to the control
cells in the following way:
Fig. 5. Statistic analysis of telomere length in control and AD subjects

Fig. 1. Dotplot analysis of control subjects’ PBMCs without hybridization probe

Fig. 2. Dotplot analysis of control subjects’ PBMCs with hybridization
probe

Fig. 3. Dotplot analysis of AD patients’ PBMCs without hybridization
probe

Fig. 4. Dotplot analysis of AD patients’ PBMCs with hybridizationprobe
2014.24.2 (82)  Fiziologia - Physiology

CONCLUSION AND DISCUSSION
The integrity of the telomere structure and its DNA hexamer(TTAGGG)n repeat sequence is critical for the protection of theends of chromosomes from degradation and in maintaining overall
genomic stability (23,24). The number of DNA hexamer (TTAGGG)
n repeats is reduced during each cell division in differentiated cells,
and as a consequence telomere length (TL) often decreases inmost differentiated cells throughout the lifespan of the organism(23). Shortening of telomeres can result in telomere end fusions
and an increased level of chromosome instability (CIN), which isin turn a key initiating event in numerous cancers (including lung,breast, colon, and prostate cancers, as well as certain leukaemia’s) (25,26). It has been shown that telomere shortening canbe accelerated by environmental factors such as psychologicaland physiological stress, cigarette smoking, obesity and highhomocysteine. Efficiency of TL maintenance is also affected bygender. TL has been shown to be associated prospectively withincreased risk of myocardial infarction, coronary artery disease,breast cancer free survival, clear cell renal cell carcinoma survival,
post-stroke mortality, dementia and cognitive decline, as wellas total survival independent of genetic influences (27-30). The
strong association between Alzheimer’s disease and ageing suggests that molecular events associated with ageing may promote
development of the disease. Given the close intimacy between
ageing and telomere shortening, it was important to determine
whether telomere shortening would have any direct influence on
Alzheimer’s disease development and/or progression.
In conclusion, the telomere length is age-dependent inlymphocytes and decreased in AD patients, which could mean that
the AD pathology may contribute to telomere length shortening.
The high variability of telomere lengths in individuals suggeststhat it will not be useful as a general biomarker for AD. However,it could become a biomarker in personalized long-term monitoringof an individuals’ health.
Acknowledgements
This work was supported by UEFISCDI, PNII-Idei, ProjectNo. 259/2011.
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11. d’Adda di Fagagna F, Reaper PM, Clay-Farrace L, Fiegler H, Carr P,
Von Zglinicki T, et al. A DNA damage checkpoint response in telomereinitiated senescence. Nature 2003; 426: 194-8.
12. Sedelnikova OA, Horikawa I, Zimonjic DB, Popescu NC, Bonner
WM, Barrett JC. Senescing human cells and ageing mice accumulate
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13. Nalapareddy K, Jiang H, Guachalla Gutierrez LM, Rudolph KL.
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stem and progenitor cell aging: what markers can we use? Exp Gerontol
2008; 43: 998-1004.
14. von Zglinicki T, Saretzki G, Docke W, Lotze C. Mild hyperoxia shortens
telomeres and inhibits proliferation of fibroblasts: a model for senescence?
Exp Cell Res 1995; 220: 186-93.
15. Aikata H, Takaishi H, Kawakami Y, Takahashi S, Kitamoto M, Nakanishi T, et al. Telomere reduction in human liver tissues with age andchronic inflammation. Exp Cell Res 2000; 256: 578-82.
16. Yang L, Suwa T, Wright WE, Shay JW, Hornsby PJ. Telomere
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in human adrenocortical cells. Mech Ageing Dev 2001; 122: 1685-94.

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Shay JW, et al. Telomere shortening is associated with cell division in vitro
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21. Rodier F, Coppe´ JP, Patil CK, Hoeijmakers WA, Mun˜ oz DP, RazaSR, et al. Persistent DNA damage signalling triggers senescenceassociated inflammatory cytokine secretion. Nat Cell Biol 2009; 11: 973-9.
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Johansson B, Pedersen NL. Telomere length predicts survival independent of genetic influences. Aging cell, 2007; 6(6):769-774.
28. Farzaneh-Far R, Cawthon RM, Na B, Browner WS, Schiller NB,
Whooley MA. Prognostic value of leukocyte telomere length in patientswith stable coronary artery disease: data from the Heart and Soul Study.
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29. Martin-Ruiz C, Dickinson HO, Keys B, Rowan E, Kenny RA, Von
Zglinicki T. Telomere length predicts poststroke mortality, dementia, andcognitive decline. Annals of neurology, 2006; 60(2):174-180.
30. Risques RA, Vaughan TL, Li X, Odze RD, Blount PL, Ayub K, Gallaher JL, Reid BJ, Rabinovitch PS. Leukocyte telomere length predicts
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2007; 16(12):2649-2655.

MODIFICARILE LUNGIMII TELOMERELOR IN BOALA ALZHEIMERREZUMAT

Telomerele sunt regiuni de ADN repetitiv situate la capetele cromozomilor eucariotelor, care previn instabilitatea cromozomiala.
Scurtarea telomerelor la celulele mononucleare din sangele periferic (PBMC) a fost asociata cu varsta biologica si cu o multitudinede afectiuni cronice degenerative. Scurtarea telomerelor este intalnita in afectiuni precum boala Alzheimer (AD), in care este indusa
la nivelul leucocitelor. Scopul prezentului studiu este de a determina lungimea telomerelor la limfocitele pacientilor cu AD comparativ
cu subiecti sanatosi de aceeasi varsta. Am utilizat o metoda noua de hibridizare flowcitometrica, care ofera date referitoare lalungimea relativa a telomerelor comparativ cu celulele de control. Datele noastre arata o scurtare semnificativa a lungimii telomerelorla pacientii cu AD (22,95%) comparativ cu subiectii control (55,49%). In concluzie, lungimea telomerelor la celulele limfocitare estedependenta de varsta si este scazuta la pacientii cu AD, ceea ce ar putea insemna ca patologia AD are o contributie la scurtarea
lungimii telomerelor. Narea variabilitate inter-individuala a lungimii telomerelor sugereaza ca acesta nu ar putea fi un biomarker
general pentru diagnosticul AD. Totusi, lungimea telomerelor ar putea deveni un biomarker personalizat pentru monitorizarea delunga durata a starii de sanatate in populatie.
Cuvinte cheie: boala Alzheimer (AD), lungimea telomerelor, hibridizare flowcitometrica, limfocite

30

Fiziologia - Physiology  2014.24.2 (82)GADOLINIUM-BASED CONTRAST AGENTS: SAFETY, EFFICACYAND POTENTIAL ADVERSE EVENTSDELIA MARIA MINCA1, ELENA LUMINITA ENACHE2, FABIAN TATU3, CARMEN TATU3
1Anatest Laboratory, Timisoara
2
University of Medicine and Pharmacy of Tirgu Mures
3
Victor Babes University of Medicine and Pharmacy Timisoara

ABSTRACT

Magnetic resonance imaging is a widely used diagnostic method, allowing for the simultaneous examination of the anatomy andphysiological parameters of tissues and organs by measuring the radio-frequency signals emitted by water protons in living tissues.
Signal resolution and sensitivity can be further enhanced by adding contrast agents. Nine gadolinium-based contrast agents (GBCAs)
are currently approved. The extracellular agents are most widely used in clinical practice and they can be further subclassified into:linear (ionic or non-ionic) and macrocyclic (ionic or non-ionic). GBCAs have been proved to be highly effective and showed favorable
clinical profiles. Several minor, transient adverse events were reported, such as headache and nausea, as well as asymptomatic risein serum iron and bilirubin levels. However, toxic gadolinium ion may be released in vivo by transmetallation, a process by which thegadolinium ion is displaced from its ligand by competing endogenous metals: zinc, cop­per, calcium, and iron. Linear GBCAs wereshown to be less stable than the macrocyclic ones under both normal and altered physiological conditions. The most severe complication associated with the use of GBCAs is nephrogenic systemic fibrosis (NSF), which is thought to be caused by the deposition offree gadolinium in tissues of patients with severe renal dysfunction and the subsequent inability of kidneys to eliminate it because ofpoor water solubility of the gadolinium. Even though nonionic linear GBCAs were proved to trigger this disease in special categories
of patients, the use of GBCAs remains safe provided measures are taken to prevent such an event.

INTRODUCTION
Magnetic resonance imaging (MRI) has become a widelyused diagnostic method over the past decades due to its ability
to produce three-dimensional representations of human tissues
with high spatial resolution and contrast, without using harmfulradiation (1). MRI allows for the simultaneous examination of the
anatomy and physiological parameters of tissues and organs bymeasuring the radio-frequency signals emitted by water protons
in living tissues (2). Signal resolution and sensitivity can be further
enhanced by adding contrast agents (CAs), with approximately
35% of MRI scans being performed with the use of CAs (3, 4).
Two major categories of CAs are available, paramagnetic andsuperparamagnetic, each of them being further classified into
extracellular, organ-specific and blood pool contrast agents(3). Most widely used paramagnetic CAs contain gadolinium,
a rare-earth metal of the lanthanide series; due to its toxicity,
the gadolinium (III) ion is administered in the form of chelatecomplexes, thus preventing the release of metal ion (5).
In MRI, relaxation is the process by which water proton spins
are returning back to the equilibrium state once the radio frequency pulse (that disturbed the equilibrium) is turned off (6), theability of contrast media to increase the relaxation being known
as relaxivity (2, 5) . Spin-lattice or longitudinal relaxation (T1)and spin-spin or transverse relaxation (T2) are the two main
relaxation processes. T1 and T2 relaxation times of water protons
are changed by contrast agents that generate a local fluctuating

magnetic field, thus producing changes in tissue contrast (2).
Even though all contrast agents shorten both T1 and T2, thedifference between them consists in the contrast image they areable to produce, that is positive (T1 contrast agents) or negative(T2 contrast agents) (8).
The Gd(III) ion, with its 7 unpaired electrons, produces a large
local magnetic field and it therefore has the strongest effect on the
T1 relaxation times (7, 8). However, the free gadolinium ion has
cardiovascular and neurologic toxicity, being deposited in liver,bone and lymph nodes (9). For this reason, it is administered in
the form of stable chelates, which eliminate its insolubility andtoxicity by using a multidentate organic ligand (3, 7, 10). The
resulting complex must be highly stable and resistant to therelease of free Gd(III) ion through transmetallation (3), a process
by which the gadolinium ion is displaced from its ligand by competing endogenous metals: zinc, cop­per, calcium, and iron (11).
Gadolinium-based contrast agents
Based on their distribution, Gd-based contrast agents (GBCA)
are divided into extracellular with renal elimination, extracellularwith mixed renal and hepatobiliary elimination and intravascularwith renal elimination or blood-pool agents (12), the first group
being the most widely used in clinical practice (10). All the nine
approved GBCA contain an octadentate co-ligand binding togadolinium and a single coordinated water ligand (7, 12).
The extracellular agents were the first approved GBCAs

Received 12th of May 2014. Accepted 11th of June 2014. Address for correspondence: Delia Minca, MD, Anatest Laboratory, Inului No.4 Street, Timisoara, Romania,
phone/fax: +40256 293192, e-mail: delia.minca@yahoo.com

2014.24.2 (82)  Fiziologia - Physiology

31

(12) and they can be further subclassified into: linear (ionic or
non-ionic), and macrocyclic (ionic or non-ionic) (10, 13). After
intravenous injection, they rapidly distribute in the blood and theextracellular space (12, 13) and are normally eliminated unmetabolized via kidneys through glomerular filtration (13, 14), withhalf-lives of approximately 1.5 hours in people with normal renal
function (5, 12, 13). However, the plasma half-life differs among
the available Gd chelates and is dependent upon the volume of
distribution and the glomerular filtration rate of the agent, while inpatients with impaired renal function is higher (13), thus caution
is recommended in these patients.
The extracellular GBCAs do not bind to proteins (10), but threeof them (gadoxetic acid disodium, gadobenate dimeglumine andgadofosveset trisodium) were designed as targeted agents (5)and have partial hepatobiliary excretion (10, 13), making them
useful for liver imaging (gadoxetic acid and gadobenate) or angiography (gadofosveset) (5, 12,13). The latter binds to humanserum albumin and its increased intravascular retention provides
greater contrast enhancement (5), although albumin binding isachieved also by the first two of them at a much lesser extent (15).
The first GBCA approved for clinical use in 1986 in Europe
and 1988 in the USA and Japan was Gd-DTPA (Magnevist) (7,16). The nine currently available GBCAs are DTPA (diethylenetriaminepentaacetate) or DOTA (1,4,7,10-tetraazacyclododecane1,4,7,10-tetraacetic acid) derivatives (7). They can be classified
according to various criteria, including their structure, relaxivity
and distribution (10, 11, 13, 14, 15) (Table I).
Table I. Classification of gadolinium based contrast agents and theirclinical uses
CliniDistribucaltionuse
Extracel- CNS/
lular non- whole
specificbody
Extracel- CNS/
lular non- whole
specificbody
Extracel- CNS/
lular non- whole
specificbody
Extracel- CNS/
lular non- whole
specificbody
Extracel- CNS/
lular non- whole
specific
body

Contrast agent

Trade
name

Structure Relaxivity

Gadopentetate
dimeglumine(Gd-DTPA)

Magnevist

Linear,ionicStandard

Gadoteridol(Gd-HP-DO3A)

ProHance

Macrocyclic, ionicStandard

Gadodiamide(Gd-DTPA-BMA)

OmniscanLinear,nonionic

Standard

Gadoteratemeglumine(Gd-DOTA)

Dotarem,
Magnescope

Macrocyclic, ionicStandard

Gadobutrol(Gd-BT-DO3A)

Gadovist,
Gadavist

Macrocyclic, ionicStandard

OptiMARK

Linear,nonionic

Standard

Extracel- CNS/
lular non- whole
specific
body

MultiHance

Linear,
ionic

High

Mixed

CNS/
whole
body

Primovist, Linear,
Eovist
ionic

High

Mixed

Liver

Vasovist,
AblavarHigh

Intravascular

Blood
Pool

Gadoversetamide(Gd-DTPABMEA)
Gadobenate
dimeglumine(Gd-BOPTA)
Gadoxetic aciddisodium(Gd-EOB-DTPA)
Gadofosveset
trisodium(Gd-DTPA)

Linear,ionic

CNS – central nervous system

32

According to the approved GBCAs producers, several dosages are recommended, the most common being 0.2 mL/kg(0.1 mmol/kg) body weight (bw); other dosages are to be used
for certain products, as follows: 0.12 mL/kg (0.03 mmol/kg) bw
for Ablavar/Vasovist, 0.1 mL/kg (0.1 mmol/kg) bw for Gadavist/
Gadovist, 0.1 mL/kg (0.025 mmol/kg) bw and 0.1 mL/kg (0.05mmol/kg bw when Omniscan is used for the purpose of kidney
imaging. Regardless of dosage, all the approved GBCAs areadministered by (rapid) intravenous bolus injection at a flowrate of 1 – 2 mL/second (intravenous infusion is also used incase of ProHance), followed by a normal saline flush (5-30 mL,
depending on product). The dose is delivered either by manual or
power injection and the imaging procedure should be completed
within 1 hour after the delivery of contrast agent.
Efficacy and safety of the nine approved GBCAs
Early studies (17, 18, 19) evaluating the safety and efficacy
of gadopentetate dimeglumine (Gd-DTPA), the first available
GBCA, have concluded it has higher diagnostic value comparedto the non contrast-enhanced MRI and exhibits high levels ofsafety and tolerance. In the multicenter double-blind randomized
clinical trial conducted by Russell et al. (17), for instance, GdDTPA improved diagnostic ability in 65% of patients and led tochanges in presumptive diagnosis, along with visualization of
more lesions in some of those patients. Adverse reactions insome patients were reported in these early studies, such as
asymptomatic rise in serum iron and bilirubin levels, hyper- or
hypotension, weakness, conjunctivitis, local burning at injection
site, headache, nausea and vomiting, all of which were minor
and transient. These adverse reactions were comparable tothose related to administration of iodinated non-ionic roentgen
contrast media (20).
In a phase III clinical trial (21), gadoteridol yielded morediagnostic information than unenhanced images in more than
70% of patients with suspected intracranial or spinal disease,
while the most common adverse reactions were dysgeusia andmild nausea in a minority of cases. Similarly, other studies (22,23) found gadoteridol is safe and well tolerated by patients andbesides the adverse reactions mentioned above, also reported
taste perversion and headache (≤1% of patients).
Gadodiamide was shown to be safe and well tolerated in a
European multicentre trial (24) in which a range of adverse reactions were recorded in a small number of patients: sensation ofheat or coldness, pain or pressure at the injection site, headache,nausea etc. This was in accordance with previous studies (25,26) reporting mild adverse reactions and minor transient elevations in serum iron. All studies reported gadodiamide injection
enhanced visualization of lesions provided more diagnostic
information. However, gadodiamide was the first GBCA linked
with the development of nephrogenic systemic fibrosis (27), arare disease that affects patients with renal failure, which will
be further discussed.
Postmarketing surveillance studies in Germany (28) and Japan(28) indicate gadoterate meglumine has excellent diagnostic efficacy and favorable clinical safety profile. Mild adverse reactions
Fiziologia - Physiology  2014.24.2 (82)

were reported in approximately 0.4% of cases and most of themwere gastrointestinal in nature.
An integrated analysis of gadobutrol safety conducted by Vothand colleagues (30) based on data from 34 prospective clinicalstudies concluded in favor of its safety that was well tolerated by
all categories of patients, including those with impaired liver orkidney function or cardiovascular disease, with a 4% incidenceof minor adverse reactions.
Similarly, gadoversetamide proves to be safe and effective,
while the occurrence of adverse reactions following its administration is comparable to other GBCAs (31, 32). In people withrenal impairment, however, elimination of gadoversetamide wasprolonged, but didn’t affect its overall efficacy (32).
A safety assessment report (33) on gadobenate dimeglumine
synthesizing data from 65 clinical trials conducted in Europe and
USA found generally mild transient adverse reactions occurred,with only 0.2% serious adverse reactions possibly related to thecontrast agent.
Gadoxetic acid disodium, as a targeted GBCA, is more effective for radiological diagnosis of liver lesions compared to otherimagistic methods, allowing for better detection and localizationof such lesions (34, 35). The adverse reactions are the same
associated with the use of other GBCAs.
Finally, the only GBCA in the series specifically designed foruse in MR angiography (although gadobenate dimeglumine isalso successfully used for that purpose), gadofosveset trisodium
shows great efficacy and a safety profile similar to all the other
GBCAs (36, 37).
Comparisons across the nine approved GBCAs
Many studies have been performed to compare the efficacy
and safety of different GBCAs, most of them involving gadopentetate dimeglumine, although intercomparisons of similar
contrast agents as regards their structure or clinical use wereconducted, as well with iodinated, ferromagnetic or superparamagnetic media. Basically, all the other GBCAs were comparedagainst the first one in the series at some point in phase II or III
clinical trials. The results regarding both the efficacy and safety
were similar to that obtained with Gd-DTPA for gadoteridol (38,39) when given in equal doses, although in the earlier, dosecomparison study (38) gadoteridol proved to more effective at
higher experimental doses. The same efficacy was observed withgadodiamide and Gd-DTPA (40, 41), with significant changes
in serum iron 24 h after injection for both GBCAs (40). Similarsafety profiles were shown by these two agents and gadoteratemeglumine in two multi-centric studies with some mild adverse
reactions occurring in relation to each of the contrast agents(42). On the other hand, when compared to the performance ofgadobenate dimeglumine (Gd-BOPTA) at an equivalent dose,
gadodiamide proved to be inferior in terms of efficacy (43).
Gd-BOPTA and Gd-DTPA have been found to have a similarsafety profiles in 10 clinical trials conducted in Europe, UnitedStates and Asia over a period of 8 years (44). However, given its
higher relaxivity, Gd-BOPTA yields improved diagnostic information for a range of indications, including liver (45), brain (46) or
2014.24.2 (82)  Fiziologia - Physiology

MR angiography (47, 48). A retrospective study (49) comparing
reduced dose of gadobenate dimeglumine with standard single
dose of gadoterate meglumine showed Gd-BOPTA produced
equal or better enhancement in cranial MRI. Again, the highrelaxivity of Gd-BOPTA played a critical role in this respect andallowed for the dose reduction.
Nevertheless, when comparing Gd-BOPTA and gadoxetic
acid trisodium for liver MRI, the latter showed superior liver
enhancement in the overall population, as well as in the cirrhotic
subgroup in a phase III clinical trial (50)., which is not surprisingsince Gd-EOB-DTPA is the only GBCA specifically targeted forliver MRI. Likewise, Gd-EOB-DTPA had significantly higher sensitivity compared to gadopentetate dimeglumine in the detectionof small hepatocellular carcinoma (51).
Gadoversetamide was also compared with Gd-DTPA in phase
III studies in patients with liver (52) and central nervous system
pathologies (53) in which safety, tolerability and efficacy of thetwo GBCAs were assessed, with no statistical differences observed between the two gadolinium chelates with respect to confidence in diagnosis, level of conspicuity, and border delineation.
Gadobutrol provided higher conspicuity and enhanced detection of cerebral lesions than gadopentetate, both GBCAs being
administered either at equal (54) or double dose (55). When
compared to gadoterate meglumine in patients with suspectedchronic myocardial infarction, gadobutrol showed superior contrast (56).
Comparison of gadofosveset trisodium with gadopentetate
dimeglumine (57) or gadobenate dimeglumine (58) in angiography resulted in either same image quality for gadofosveset
at standard (0.03 mmol/kgbw)/high doses (0.05 mmol/kgbw)
and gadopentetate dimeglumine at increased dosage (≈0.25
mmol/kg), or slightly better performance of gadofosveset overgadobenate dimeglumine.
Nephrotoxicity of GBCAs
Gadolinium chelates are routinely used nowadays in magnetic resonance imaging, mostly for CNS pathology, followed bycontrast-enhanced MR angiography, liver imaging, and breast
evaluation for malignancy (59). Unfortunately, toxic Gd(III) ions
may be released in vivo by transmetallation with other metal
ions, thus the kinetic stability of the gadolinium chelates is very
important (3). Macrocyclic chelates have been shown to be morestable than the linear ones (60, 61). A study (62) on the stability
and dissociation rate of the approved GBCAs in human serum
under normal physiological conditions (pH 7.4 and 370C), as
well as in case of elevated serum phosphate levels (often seenin patients with end-stage renal disease) concluded nonioniclinear chelates are less stable than the ionic linear ones undernormal and particularly under altered physiological conditions,while the microcyclic agents remained stable.
The interference of gadodiamide and gadoversetamide
with colorimetric determinations of serum calcium has been
well documented (63, 64, 65, 66), the release of gadolinium
ion resulting in spurious hypocalcemia (especially in patientswith renal insufficiency), which may lead to unnecessary and33

potentially inappropriate treatment. Furthermore, transmetallation may occur with other endogenous metal ions, causing both
positive and negative interference with other serum assays: zinc,iron, magnesium, copper, angiotensin-converting enzyme andtotal iron binding capacity (13, 61, 67, 68, 69, 70). Gadolinium
deposits were detected in several organs, particularly in theskin, kidney, liver, heart and bones (71, 72, 73). Several GBCAs(namely Omniscan, MultiHance, ProHance and Magnevist) wereshown to stimulate dermal fibroblasts proliferation, along with
up-regulation of matrix metalloproteinase-1 and tissue inhibitorof metalloproteinases-1 (74, 75). Contrast-induced nephropathy,
defined as increase in creatinine concentration (76) or decreasein glomerular filtration rate (77) has been associated with GBCAs,
particularly at high doses and in patients with renal impairment.
However, in patients with normal renal function, their use remains
safe, due to the rapid excretion of gadolinium complex.
But the most severe and possibly life-threatening complication associated with the use of GBCAs is nephrogenic systemic
fibrosis (NSF), a fibroproliferative disease affecting people withsevere renal impairment which was first observed in 1997 (78).
NSF is thought to be caused by the deposition of free gadolinium
in tissues of patients with severe renal dysfunction and the subsequent inability of kidneys to eliminate it because of poor watersolubility of the gadolinium (79). A recent review by Zou et al.
(80) indicates dialysis, edema, hyperphosphatemia, epoetin use
and proinflammatory conditions as additional factors involved in
the onset of NSF.
This causative link between GBCAs and NSF has prompted
the USA Food and Drug Administration to request a warningbox to be added by manufacturers to the labels of all GBCAs in
2007 (81) stating the increased risk of NSF in patients with acuteor chronic severe renal insufficiency (glomerular filtration rate
<30mL/min/1.73m2), or acute renal insufficiency of any severity
due to the hepato-renal syndrome or in the perioperative livertransplantation period. Subsequently, the European Medicines
Agency has released in 2009 a set of recommendations (82)based on the various degrees of risk demonstrated for the approved GBCAs. Hence, for high-risk GBCAs, in addition to thecategories of patients in the FDA warning, contraindications
were also recommended for newborn babies up to four weeks ofage. Additionally, patients with unknown kidney problems should
always be screened for renal function and that women should
discontinue breastfeeding for at least 24 hours after a scan.
For medium- and low-risk GBCAs warnings in the prescribing
information concerning their use in patients with severe kidney
problems and patients receiving a liver transplant are also
recommended, and screening for kidney problems is advised,
while decision on breastfeeding discontinuation is to be takenby the doctor and mother.
So what led to this position? As highlighted above, GBCAs
were increasingly linked with NSF, while gadodiamide wasmost frequently incriminated. In 2006 Grobner (83) was the firstto suggest the development of NSF after exposure to gadodiamide. Shortly after, Marckmann et al. (27) reported 13 casesof NSF after exposure to the same contrast agent in patients34

with end-stage renal disease. After that, more and more reports
were published adding more evidence in support of this link (8,9, 13, 72, 73, 76, 77). Based on data from literature, Tweedle
and colleagues (11) have classified GBCAs into three groups
according to their association with NSF, gadodiamide, gadopentetate dimeglumine and gadoversetamide being linked withthe greatest number of cases. The macrocyclic GBCAs are theleast involved, while the protein-binding ones have low incidenceof NSF cases. In a retrospective analysis at two large medical
centers over a period of 10 years, Prince et al. (84) calculated
the incidence and risk factors of NSF for patients receiving
standard or high doses of GBCA, as well as in patients with renal
impairment. At standard dose used for patients not screened forrenal function, no case of NSF was found and an incidence of
0.17% for high-dose usage was recorded. In patients with renal
dysfunction, after exposure to high dose GBCAs, the incidence
was 0.4% (hemodialysis patients), 8.8% (eGFR<15mL/min, noton hemodialysis) and 19% (acute renal failure and increased
creatinine level). This led to the conclusion that, while in thefirst category of patients hemodialysis may prevent NSF, in theremaining two groups, the risk of NSF after administration ofa high dose GBCA is associated with delayed hemodialysis,
proinflammatory events, and hyperphosphatemia.
In conclusion, GBCA stability and dosage are important considerations in the selection of the particular agent to be used for
MRI scans, especially in renally impaired patients, because of
the possibility of the release of free gadolinium ion in the bodyunder certain physiological conditions, with the subsequentdeposition that may result in a series of adverse reactions andcomplications, out of which the most severe is the nephrogenic
systemic fibrosis. Even though nonionic linear GBCAs wereproved to trigger this disease in special categories of patients,
the use of GBCAs remains safe provided measures are taken
to prevent such an event.
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AGENŢI DE CONTRAST PE BAZĂ DE GADOLINIU: SIGURANŢĂ, EFICACITATE ŞI EFECTE ADVERSE POTENŢIALEREZUMAT

Imagistica prin rezonanță magnetic este o metodă de diagnostic larg utilizată, care permite examinarea simultană a anatomiei șia parametrilor fiziologici ai țesuturilor și organelor prin măsurarea semnalelor de radiofrecvență emise de protonii din țesuturile vii.
Rezoluția și sensibilitatea semnalelor poate fi sporită prin adăugarea de agenți de contrast. În prezent există nouă agenți de contrast
pe bază de gadoliniu aprobați (GBCA). Agenții extracelulari sunt cel mai folosiți în practică și pot fi clasificați în: lineari (ionici sau
non-ionici) și macrociclici (ionici sau non-ionici). S-a dovedit că GBCA sunt foarte eficienți și au profiluri clinice favorabile. Au fostraportate unele reacții adverse minore tranzitorii, cum ar fi dureri de cap și greață, precum și creșteri asimptomatice ale fierului seric
și bilirubinei. Cu toate acestea, ionul toxic de gadoliniu poate fi eliberat in vivo prin transmetalare, un proces prin care ionul de gadoliniu este desprins de ligandul său și substituit de către metale endogene competitoare: zinc, cupru, calciu și fier. S-a demonstrat că
GBCA lineari sunt mai puțin stabili decât cei macrociclici atât în condiții fiziologice normale, cât și alterate. Cea mai gravă complicație
ca urmare a utilizării GBCA este fibroza sistemică nefrogenă (NSF). Se crede că NSF este cauzată de depunerea gadoliniului înțesuturile pacienților cu disfuncție renală severă și de incapacitatea rezultată a rinichilor de a-l elimina, datorită solubilității scăzute
a acestuia. Deși s-a demonstrat că GBCA lineari nonionici declanșează această boală la anumite categorii de pacienți, utilizarea
GBCA rămâne sigură în condițiile în care se iau măsuri pentru prevenirea acesteia.

2014.24.2 (82)  Fiziologia - Physiology

37

REGULAR PHYSICAL ACTIVITY AND ABLATION OF LONEATRIAL FIBRILLATION OUTCOMES IN YOUNG PATIENTSALINA G. NEGRU1, 2, GABRIEL IVĂNICĂ¹, MARIUS ANDRONACHE3, ADINA BUCUR4, ŞERBANNEGRU2, ADRIAN IVĂNICĂ5, C.T. LUCA1,2, ADINA IONAC1,2 , LUCIAN PETRESCU1,2, ŞTEFAN
I. DRĂGULESCU1,2¹Timişoara Institute of Cardiovascular Medicine²Department of Cardiology, University of Medicine and Pharmacy “Victor Babeş” Timişoara
3
CHU Nancy Rythmologie Cardiaque4
Departament of Public Health and Health Management, University of Medicine and Pharmacy “Victor Babeş”, Timişoara
5
Robert-Bosch-Krankenhaus Kardiologie

ABSTRACTIntroduction: Over the years it has been shown that sports reduce the risk of heart disease, stroke, metabolic syndrome, diabetes
mellitus, hypertension, cancer, depression and other pathological conditions. At the same time there is growing evidence that long term
sustained athletic activity is associated with atrial fibrillation (AFIB). Until present there is no evaluation of the AFIB ablation outcomes inleisure time sport practitioners compared to sedentary young patients. Aim of the study: The present study aims to evaluate mainly the
association between regular physical activity and AFIB ablation outcomes in non-professionals (excluding endurance athletes or otherprofessional sports practitioners) compared to sedentary patients in terms of peri-procedural and long term success ablation success.
Methods: The study population consisted of 173 young patients (p) diagnosed with lone AFIB, admitted for the AFIB ablation and divided
in two groups- 88 p performing moderate to vigorous physical activity and 87p not implicated in any sports activity. Mean age was 41±4.
AFIB ablation was performed in all patients included after the completion of a personalized questionnaire, physical examination, ECG,
echocardiography (for LA volume, EF), blood testing of each individual. The patients were included during 36 months; the follow-up period
for each patient was 12 months while the subjects were evaluated for early and late recurrences, need for antiarrhythmic treatment in orderto maintain sinus rhythm. Other correlations between LA volume, total procedure time, age and relapse were performed. Results: The
percentage of men practicing sports and diagnosed with lone AFIB (76.1%) is significantly higher than the number of women included in
the same group (23.9%). On the other hand, in the sedentary patients, the percentage of men is even greater (10-fold) than the numberof women compared to the sports group. However, the gender does not play a role in the rate of recurrences. There was a percentage
of 50% from the athletic group and 67% from the non-athletic group that necessitated antiarrhythmic treatment for a maximum of 3
months after ablation in order to maintain sinus rhythm. There was no significant difference in the scores for antiarrhythmic medicationgroup and no medication group in terms of recidive t (84) =-1.3, p=0.2. Atrial fibrillation recidive for both groups was evaluated at 9%(11.5% from the physical activity group and 7.5% from the sedentary group). As for the scores for LA volumes (LA volume reference=90ml), there was noted no difference either, t (84) =1, p=0.3. Significantly data were obtained by the evaluation of the total procedure timein patients that received peri-procedural antiarrhythmic medication; the procedure was clearly shorter in patients under anti-arrhythmic
medication. A very important outcome was that compared with non-exercisers, the patients with regular physical activity had 35% lower
risk of AFIB recidive after the first procedure (P=0.02) (early and late recidive).
Conclusions: Our study data supports a statistically significant association between regular physical activity and lower incidenceof recurrences post AFIB ablation in non-professional physical activity performers. However the question how much is too much isstill to be answer.
Key words: atrial fibrillation ablation, non-professional athletes, physical activity, exercise, ablation outcomes.

INTRODUCTION
Atrial fibrillation (AFIB) is considered the most common arrhythmia
and its prevalence tends to increase along with aging of the population from 0.7% at younger ages to 17.8% around 80 years (1). The
same study (the Rotterdam study), revealed the prevalence andincidence as greater in men than in woman. An older study datingfrom 1994 revealed a prevalence of 0.5% below 40 years and 8% in
the patients with ages over 80 years (2). All this data was obtained
from the general population. Several studies have reported higher
rates of AF among professional athletes. Current practice of sportwith a lifetime greater than 1500h was associated with lone atrial

fibrillation, the proportion of patients found in this study was greaterthan double (31%) compared to the proportion observed in controls(14%) (3). Although the presence of AF in athletes has been discussed and described previously (4,5), actually there is poor data
availability and inconsistent information regarding the association ofregular physical exercise with atrial fibrillation (excluded the professional athletic activity and endurance sports) (6).
AIM OF THE STUDYThis is the first study that aims to evaluate the evidence of theassociation between regular physical activity and the outcomes ofReceived 12th of May 2014. Accepted 15th of June 2014. Address for correspondence: Alina G. Negru, MD, “Victor Babeş” University of Medicine and PharmacyTimişoara, No. 2 Eftimie Murgu Square, Timişoara, phone: +40-746-239777, e-mail: eivanica@yahoo.com

38

Fiziologia - Physiology  2014.24.2 (82)

atrial fibrillation ablation especially the efficiency of atrial fibrillation
ablation in non-professional athletes or exercisers compared to
non-exercise population in terms of early and late relapse.
METHODS
It is a prospective, case control study in patients undergoing
AFIB ablation. Professional athletes were excluded. The study
duration was 3 years (36 months) and the follow-up period after
ablation was 12 months for each patient.
Lone atrial fibrillation is a term that was first introduced in
1954 by Evans and Swan (8). Currently, lone atrial fibrillation isconsidered when clinical and echo evidence of cardiovascularor pulmonary disease has been excluded. Pathologic conditions
such as hypertension, diabetes, hyperthyroidism, acute infections,
recent cardiothoracic or abdominal surgery, and systemic inflammatory diseases must be also excluded in order to diagnose thelone AFIB (9).
Study population.
We included in the study a number of 173 from 210 initial
candidates for inclusion, with lone atrial fibrillation, selected for
atrial fibrillation ablation and filtered by the baseline information.
Participants were only included after signing the informed consent form and after the completion of a questionnaire containing
information about: the type of sport, number of hours spent training, medical history, lifestyle, cardiovascular risk factors. Otherbaseline evaluation consisted of standardized physical evaluation,
electrocardiogram (ECG), echocardiography with evaluation of theleft atrial volume and size and of the ejection fraction (EF), blood
tests including thyroid hormones FT3, FT4 and TSH.
We divided the patients in two equal groups:
The first group included 87 patients with the following characteristics: age below 45 years; sedentary (patients which are not
performing any type of controlled regular physical activity); diagnosed with lone AFIB (at least two separate documented episodesof AFIB); EF >55%; without associated pathologic conditions; withor without anti-arrhythmic medication; admitted in the hospital forthe catheter ablation of the AFIB.
The 88 patients included in the second group matched the following criteria: age below 45 years; moderate to vigorous regularphysical exercise performed between 3 and 7 times per week;
presence of lone AFIB (minimum two separate documented
episodes of AFIB); EF>55%; without any associated pathologic
conditions; with or without anti-arrhythmic medication; option forradiofrequency catheter ablation (RFA).
Physical activity assessment
The exercise was assessed by level of intensity. We includedin the exercise group all the patients that fulfilled the above
mentioned criteria and practiced moderate or vigorous physicalactivity of minimum 3 times per week, for 1 hour. The physicalactivities were categorized as moderate or vigorous on the basisof current knowledge of the overall level of intensity required for the
average person engaged in it, taking into account brief episodes
when the level of intensity required for the activity might increaseor decrease considerably10.
We defined as moderate activity in accordance with CDC andACSM Guidelines all the activities between 3.0 and 6 METs (3.5 to
7 kcal/min) 11. The patients classified in the moderate activity group2014.24.2 (82)  Fiziologia - Physiology

were performing the following activities based on the completed
questionnaire:− race walking less than 8 km/h
− bicycling 8 to 14 km/h, level terrain or with a few hills− stationary bicycling using moderate effort− recreational swimming
We defined as vigorous activity 11all the activities greater than
6.0 METs (more than 7 kcal/min). The patients classified in thevigorous activity subgroup were performing the following activities:− race walking and aerobic walking 8 km/h or faster− jogging or running− bicycling more than 16 km/h or bicycling on steep uphillterrain− stationary bicycling using vigorous effort− swimming steady paced laps
Study design
The patients were included in the study for 36 months and thefollow-up period was 12 months for each patient, divided in 4
visits every 3 months.

Fig.1. Study design. AFIB-atrial fibrillation ablation, ECHO- cardiac
echocardiography, ECG- electrocardiogram, LA- left atrium, TPT-total
procedure time.

Catheter ablation procedure
Pulmonary vein isolation was performed using a CARTO 3 imaging system after the integration of an atrial computer tomography.
The 3D reconstruction and mapping of the pulmonary veins wereobtained using a ten pole Lasso type catheter and an irrigated
NaviStar ThermoCool catheter (Biosense Webster). The Lasso
catheters were installed via an 8 F sheath (Mullins fixed curve,
St. Jude) and the NaviStar catheter was passed in the left atrium
using fluoroscopic guidance through the same puncture point as
the Lasso. During the entire procedure heparin was injected inorder to maintain an activated clotting time between 300 and 400s.
Antral pulmonary vein ablation was performed for all veins in each39

patient. The bipolar electrograms were filtered at 50-1000Hz anddisplayed at 100 mm/s sweep speed (Cardiolab System, Prucka
Engineering). Radiofrequency energy was applied using a Stockert
generator (Biosense Webster). The irrigation was between 17 and
30 ml/min with saline solution 0.9%. The temperature and power
limits were 45ºC and 35W for the pulmonary veins and left atrium
and 20ml/min and 20-25W for the interior of the coronary sinus.
Post-ablation management
After the procedure the heparin was stopped and the lowmolecular-weight heparin was administered concomitant with
dabigatran for 24h or with acenocumarol until the INR reached thetherapeutic range between 2 and 3. The anticoagulation therapy
was stopped after a minimum of 3 months in patients free of AFIB
recurrences. The antiarrhythmic therapy was used in most cases
during the first 3 months after ablation in order to stabilize to sinus
rhythm and totally interrupted after the mentioned blanking period.
The patients were evaluated at 1 month, 3 months, 6 months and
12 months after ablation by physical examination, ECG, ECG
Holter/24 h or 7 days depending of the described symptoms,
cardiac echocardiography.
RESULTS
Statistical analysis was performed using the SPSS program.
There was observed a male predominance between both groups(physical activity and sedentary groups). From the 88p included in
the athletic activity group there were a number of 67 men (76.1%)
and 21 women (23.9%), as for the 87 patients included in thesedentary lifestyle group there were a number of 79 men (91%)
and 8 women (9%) This finding may suggest on one hand thatthe young population diagnosed with lone AFIB consists mainly ofmen which can be ten times more numerous than women in thesedentary group of patients. However, the sex does not play a rolein the rate of recurrences which seems similar in women and men.
The peri-procedural and up to three months antiarrhythmic
treatment was necessary in 102p from 172 to stabilize to sinus
rhythm. After the 3 months treatment, the antiarrhythmic waswithdrawn. There were a number of 44 p (50%) from the athletic
group and a number of 58p (67%) from the non athletic groupthat has necessitated antiarrhythmic treatment, showing the clear
numeric superiority of the sedentary individuals that necessitatedmedication after the AFIB ablation.
The total number of relapse to AFIB for both exercisers andnon-exercisers was 33 (9%).
From a total number of 86p performing physical activity, there
were 13p (7.5%) with AFIB relapse at minimum 6 months. The
relapse among the 87 sedentary patients was 20p (11.5%). This
may suggest that sport can play a role in the outcome of the ablation procedure in the sense of increasing the chances of successin patients with lone AFIB which perform physical activity. This isa very important end-point of our study.
No. relapses
%P
Total
33
19%
Exercise individuals
13
7.5%
Non-exercise individuals
20
11.5%
Fig. 2. Relapse table showing the numbers of recurrences in theexercise or non-exercise groups and the percentage (%P) from total
number of patients included in the study, corresponding to eachnumber of recidives.

40

The Levene’s test for the equality of variances showed that thevariability in all couples of groups that we compared was about
the same. We compared several groups of patients in terms ofprocedural success translated by recidive evaluated at 6 months:
patients with and without antiarrhythmic medication for the first 3
months after the ablation, patients with left atrial volume below andabove 90 ml, respectively patients with left atrial volume below
and above 120ml.
For the first two groups with/without antiarrhythmic medication
we obtained the following data. As mentioned above, there werea total number of 86 patients undergoing moderate or vigorous
exercise. The late recidive among exercisers was independentof the presence or absence of the antiarrhythmic treatment that
was administered in some cases for a maximum of 3 months
after ablation.
An independent samples t-test was conducted to compare
recidive in the group of young patients sport practitioners andsedentary which were divided in medicated and non-medicatedafter the ablation procedure. There was not a significant difference in the scores for antiarrhythmic medication group (M=0.3,
SD=0.45) and no medication group (M=0.17, SD=0.4), in termsof recidive; t(84)=-1.3, p=0.2.
For the second couple of groups, with left atrial volume smalleror greater than 90 ml, we obtained the following data: from a total
number of 86 patients recidivated, 61 had a left atrial volume
greater than 90 ml and 25 patients had a value of the left atrium
volume smaller or equal to 90ml. However, there was no significant
difference in the scores for left atrial volume under 90 ml (M=0.16,
SD=0.4) and above 90 ml (M=0.26, SD=0.44); t (84) =1, p =0.3.
These results suggest that the left atrial size does not affect theoutcome of the AFIB ablation regarding the recidive at 6 months.
The third couple of groups compared for recidive were patients(mixed population of exercisers and non-exerciser) with left atrial
volume under 120 ml (36p) and above 120 ml (50p). The conducted t-test, again, showed no significant difference between thetwo groups- the group with left atrial size above 120 ml (M=0.25,
SD=0.43) and the group with left atrial size under 120 ml (M=0.22,
SD=0.41); t (84) =0.32, p=0.7.
A statistically significant data that obtained was from the last couple of groups. This consisted of evaluation of the total AFIB ablation
procedure time in patients that had received or not peri-procedural
medication. The procedure was clearly shorter in patients underantiarrhythmic medication (M=280, SD=84) compared with noantiarrhythmic medication (M=202, SD=51); t (84) =-5, p=0.001.
One of the most important outcomes of our study already mentioned but need to be reminded, was that compared with no regularexercise, individuals with moderate-intensity or vigorous exercise
had 35% lower risk of AFIB relapse after the first ablation (p=0.02).
DISCUSSIONS
The mechanisms by which sports could generate lone AFIB
were described before. They include the alteration of the balance
between sympathetic and parasympathetic nervous system having
as consequence increases in the vagal tone12 (with bradycardia
and shortening of the atrial refractory period) and the left atrial
Fiziologia - Physiology  2014.24.2 (82)fibrosis associated with left atrial enlargement as a substrate formicro-reentry circuits (13). There was also incriminated the lossof electrolytes due to sweating, a changing that may modify theelectrophysiological properties of the atrial tissue. Coumel (14)studied the influence of autonomic system in the appearance of
AFIB and atrial flutter with the specification that AFIB occurs more
frequently in men than in women. Looking at our study findings, thenumber of women with lone AFIB belonging to the athletic groupis 3 times greater than the number of women belonging to thesedentary population group. That can suggest that the exercise
may be responsible for the vagal mechanism of producing AFIBin both sexes. Regarding this aspect it also should be noted thatthe inclusion rate of men population is superior of that of women.
The limitation of our study regarding this aspect consists in thebasic number of males implicated in athletic activity which is known
to be greater in general than that of women. Once installed AFIB
can be treated conservatory or by catheter ablation targeting
the isolation of the pulmonary veins. After performing the AFIB
ablation for all the patients included in our study (sedentary andphysical activity performers), two interesting findings came out:
among the sedentary patients there was a significantly greaternumber of patients which required antiarrhythmic treatment for
the first three months in order to obtain the stabilization to sinus
rhythm than in the athletic group; on the other hand, the recidive
at 6 months was significantly greater in the athletic group with thespecification that after ablation, at 12 weeks, the athletes resumed
their activity but at a lower intensity.
For our two groups of patients, the correlation between the recidive and the left atrial volume remained without any signification
for the two values that we used as reference (90 ml and 120 ml).
However, GIRAFA study data (15) showed that patients with loneatrial fibrillation had a larger atrial size compared with controls.
The same study revealed that patients with a first episode of AFIB
had the same atrial size compared with those suffering recurrences. The cumulated data from the GIRAFA study and our own
study permitted the following conclusion: if the left atrial volume
is a predictor for the AFIB incidence, after the ablation procedurethe same parameter cannot be used to predict recurrences. The
echographic data from another study conducted by Pelliccia et alsuggest that structural remodeling due to exercise is often present
in the atrium of elite athletes (16). Their data showed that theindividuals implicated in regular endurance practice have a larger
atrium compared with sedentary controls. Our study which included
leisure time physical activity individuals performing moderate orvigorous activity did indeed showed an enlargement in the left
atrial volume in this population, significantly higher compared to
non-exercisers (61% in athletes compared to 29% in sedentaryindividuals). However, this outcome does not affect the number ofrecurrences in our study, suggesting that even there is a differencein the AFIB mechanisms of the two groups, once the ablation is
performed there is no influence of the left atrial volume on theperi-procedural or long term success.
Systematizing the data obtained in our study we may conclude
that leisure time sport even at moderate or vigorous intensity has
not the same effects on the heart as endurance training in terms
2014.24.2 (82)  Fiziologia - Physiologyof AFIB. Even we did not study the incidence but the ablation
outcomes we may conclude that long-term endurance training
profoundly affects heart and body’s physiology. Among other things
it is shown to reduce the heart rate and testosterone levels (17,18).
Greek researchers evaluated participants in a distance run overa period of 36 hours (19). They observed a 152-fold increase in
C-reactive protein levels and an 8000-fold increase in the level ofinterleukin-6, both important markers of inflammation. The conclusion is that increases at such a scale of the inflammatory markers
may induce a potent systemic inflammatory response which at its
turn may produce the enlargement of the left atrium with arrhythmic
complications. This is another theory of arrhythmogenesis in sport
practitioners, the less explored until now.
Regarding the detraining as a mean to prevent AFIB recurrences,
there is some evidence proving that the patients who have been
ablated for right atrial flutter are at risk to develop atrial fibrillation
post-ablation if they have a history(81% increased) and continuationof endurance sports (68% increased risk) (20). Our study revealed
very important data regarding this aspect, namely, the outcomethat compared with no regular exercise, individuals from the physical activity group had 35% lower risk of AFIB recurrence after the
first ablation procedure (early and late recurrence). Those data arecontradictory with the actual situation described in the literature, withthe specification that our physical activity patients are not endurance
professional athletes as the individuals included in almost all published studies. The difference in the intensity and duration of sports
activity, lighter in our study population might be the changing point,
raising again the question: how much is too much?
CONCLUSIONSIs exercise good or bad? This is the questions that rise after
each study evaluation. Given the fact that through our study we
did not evaluate the incidence of the AFIB in the included individuals, but the outcomes of the AFIB ablation, the answer at thisquestion might not be a direct one. All the patients included in ourstudy had been already diagnosed with AFIB. However, the mostimportant outcome was that compared with non-exercisers, theindividuals included in the physical activity group had 31% lower
risk of AFIB relapse after the first ablation procedure (early andlate recurrences included). This suggests that leisure time physicalactivity performed in moderate or vigorous way has a great impact
on the ablation success even in the conditions of continuation ofsports after the procedure (at lower intensity than usual). Anotherimportant outcome is that the antiarrhythmic medication given
peri-procedural shortens the total procedure time and stabilizes
to sinus rhythm and does not influence the rate of recurrences.
On the other hand, there is no influence of the left atrial volume,
gender, age on the ablation success.
Overall, our data support a statistically significant association
between regular physical activity and lower incidence of recurrences after the AFIB ablation procedure in leisure time physicalactivity performers.
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41

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ACTIVITATEA FIZICA REGULATA SI REZULTATELE ABLATIEI FIBRILATIEI ATRIALE LA PACIENTII TINERIREZUMAT

Introducere: De-a lungul anilor a fost dovedit ca exercitiul fizic reduce riscul aparitiei bolilor cardiovasculare, a accidentelor cerebrale,sindromului metabolic, diabetului zaharat, hipertensiunii, cancerului, depresiei si a altor patologii. In acelasi timp, exista dovezi ca
activitatea fizica pe termen lung este asociata cu fibrilatia atriala (AFIB). Pana in prezent nu exista o evaluare a rezultatelor ablatiei
AFIB la cei care efectueaza efort fizic moderat, comparativ cu persoanele tinere sedentare. Scopul studiului: Studiu prezent isi
propune in principal evaluarea asocierii dintre activitatea fizica regulata si rezultatele ablatiei AFIB la sportivii neprofesionisti (excluzand
sportivii de anduranta sau alte tipuri de sportivi de performanta) comparativ cu pacientii sedentari in ceea ce priveste etapele peri-procedurale si succesul pe termen lung al ablatiei. Metode: Populatia de studiu a fost formata din 173 de pacienti tineri (p) diagnosticati
cu AFIB, spitalizati pentru ablatia AFIB si impartiti in doua grupuri – 88 p care efectuau activitate fizica moderata sau intensa si 87
p care nu erau implicati in activitati sportive. Media varstei a fost 41±4. Ablatia AFIB a fost efectuata la toti pacientii inclusi in studiu
dupa completarea unui chestionar personalizat, examen clinic, ECG, ecocardiografie (pentru volumul AS, FE) si teste hematologiceale fiecarui individ. Pacientii au fost inclusi in studiu pe o perioada de 36 de luni; perioada de urmarire a fiecarui pacient a fost de
12 luni, timp in care pacientii au fost evaluati pentru aparitia recurentelor precoce si tardive, necesitatea tratamentului antiaritimic
pentru mentinerea ritmului sinusal. Au fost efectuate si corelatii intre columul AS, timpul total necesar procedurii, varsta si recaderi.
Rezultate: Procentul persoanelor de sex masculin care practica sport si au fost diagnosticati cu AFIB (76,1%) a fost semnificativ
mai crescut comparativ cu cel al persoanelor de sex feminin incluse in acelasi grup (23,9%). Pe de alta parte, la pacientii sedentari,procentul persoanelor de sex masculin este chiar mai mare (de10 ori) decat cel al femeilor comparativ cu grupul care practica sport.
Totusi, sexul pacientilor nu joaca un rol in aparitia recurentelor. A existat un procent de 50% din grupul atletic si 67% din grupul nonatletic care a necesitat tratament anti-aritmic pentru o durata de maxim 3 luni dupa ablatie pentru mentinerea ritmului sinusal. Nu a
existat o diferenta semnificativa intre grupul cu medicatie anti-aritmica si cel fara medicatie anti-aritmica in ceea ce priveste recidiva,t (84) =-1.3, p=0.2. Recidiva fibrilatiei atriale pentru ambele grupuri a fost evaluata la 9% (11,5% pentru grupul cu activitate fizica si
7.5% pentru grupul sedentar). In ceea ce priveste volumul AS (volum de referinta AS =90 ml), nu a fost determinata nici o diferenta,t (84) =1, p=0.3. Date semnificative au fost obtinute prin evaluarea timpului total necesar procedurii la pacientii care au fost tratati
cu medicatie anti-aritmica peri-procedural; procedura a fost de durata mai scurta la pacientii cu medicatie anti-aritmica. Un rezultat
foarte important a fost ca pacientii cu activitate fizica regulata au avut un risc de recidiva a AFIB dupa prima procedura cu 35% mai
scazut decat pacientii sedentari (P=0.02) (recidiva pe termen scurt si lunge).
Concluzii: Datele studiului nostru sustin asocierea semnificativa statistic intre activitatea fizica regulta si scaderea incidentei recurentelor post-ablatia AFIB la sportivii ne-profesionisti. Totusi, ramane de stabilit cat de mult este prea mult.
Cuvinte cheie: ablatiafibrilatiei atriale, sportivi neprofesionisti, efort fizic, efectele ablatiei

42

Fiziologia - Physiology  2014.24.2 (82)BCL-2 EXPRESSION AS A NEGATIVE PROGNOSTIC FACTORIN HODGKIN LYMPHOMA
POTRE-ONCU OVIDIU, IONITA IOANA, IONITA MIHAI, CALAMAR DESPINA, SORICA CRISTINA,IONITA HORTENSIA“Victor Babes” University of Medicine and Pharmacy Timisoara, Romania

ABSTRACTIntroduction: Although important progresses have been made in Hodgkin’s disease (HD) treatment, its prognosis is unfavourable inmany cases. This is why many analyses of factors able to positively influence disease evolution and prognosis have been performed
and it is believed their earliest detection after the actual onset of the disease allows for the appropriate therapy to be administered.
Molecular immunology studies have shown that in classic Hodgkin lymphoma HRS (Hodgkin Reed Sternberg) cells derive from thegerminal centre in the B cells with immunoglobulin genes rearrangement, but without B cell receptor surface expression. BCL-2 hasbeen identified as the first gene involved in HRS cells apoptosis. BCL-2 expression by HRS cells may prevent apoptosis inducedby the absence of B cells functional receptors, which explains tumorigenesis. BCL-2 expression may also explain the resistance totreatment-induced HRS cells apoptosis.
Aim: This study aims to determine BCL-2 expression in Hodgkin’s lymphoma patients included in the study, correlation of its
expression with biochemical and immunological changes, as well as evaluation of survival in these patients.
Methodology: A retrospective analysis has been conducted from May 2008 to April 2013 for 151 patients diagnosed with Hodgkin
lymphoma in the Hematology Department, Timisoara. The main diagnostic method was biopsy, followed by histopathological andimmunohistochemical examinations of harvested tissue.
Results: Mean age of patients included in the study is 49.69±17.46, with a minimum of 18 and a maximum of 89 years, out of
which 37.7% were women and 62.3% men. The follow up period since diagnosis was 13.92±6.24 months up to complete remission(33.7% of patients); partial remission was seen in 45.7% of patients, 17.2% had progressive disease, 0.6% relapsed and 2.5% died.
BCL-2 was detected in 82 patients (54.3%), the intensity varying according to the histological degree of the disease, as follows:
BCL-2 expression is seen in 19.8% of MC patients, 39.1% of NS patients and 5.3% of patients with lymphocyte depletion.
Survival is lower in patients with BCL-2 expression than in those not expressing the gene, and gene expression shows positive, statisticallysignificant correlation with increased levels of other immunological (CD15, CD20, CD30) and biochemical (fibrinogen) markers.
Conclusions: A reduced survival rate of patients with BCL-2 expression associated with intense expression of CD15, CD20 and CD30
markers, as well as with increased fibrinogen levels were seen in this study. Biochemical markers ESR, LDH and ceruloplasmin are
inversely correlated with presence of BCL-2, and survival rates are not significantly changing at increased levels compared to normal.

INTRODUCTION
Although important progresses have been made in Hodgkin’s
disease (HD) treatment, its prognosis is unfavourable inmany cases. This is why many analyses of factors able topositively influence disease evolution and prognosis have been
performed and it is believed their earliest detection after theactual onset of the disease allows for the appropriate therapy
to be administered (1).
Changes in disease activity tests (ESR, fibrinogenemia, LDH
and serum alkaline phosphatase) positively correlate with a moreaggressive form of disease and therefore with a more reduced
rate of complete remission and shorter survival (5-year survivalrate); ESR levels over 80 mm/h and serum alkaline phosphataseover 200 IU/ml are highly significant prognostic risk factors (2,3).
Taken separately, ESR and serum alkaline phosphatase havevery close discriminative values; their combined assessment,
however, proved to be one of the most important prognostic
factors, always considered in multifactorial studies and influencing
HD patients survival (4-7). It is worth mentioning that increases
in ESR and serum alkaline phosphatase levels should be judged

in close connection with other disease activity factors. Similarly,
their increased levels mirror biologically aggressive forms orpresence of HD not detectable by current diagnostic procedures,as the period of complete remission is longer than 36 months
in less than 30-40% of cases with these modified activity tests(8-10); thus, they might be a clue to applying since the very
beginning more aggressive therapeutic protocols, such as bonemarrow transplantation (11). Recently, high levels of serum TNF
and its soluble receptors (p and p75) were shown to be negative
prognostic factors, indicating an unfavourable evolution of HD
patients (12-14).
Molecular immunology studies have shown that in classic
Hodgkin lymphoma HRS (Hodgkin Reed Sternberg) cells derivefrom the germinal centre in the B cells with immunoglobulin
genes rearrangement, but without B cell receptor surface
expression (15). These changes have been attributed to
defects in gene transcription with the immunoglobulin genes
rearrangement in HRS cells. Some authors consider the lackof B cell receptors expression leads to apoptosis in the normal
cells of the germinal centre. BCL-2 has been identified as

Received 16th of March 2014. Accepted 20th of May 2014. Address for correspondence: Potre Ovidiu, MD, PhD student, “Victor Babes” University of Medicine andPharmacy Timisoara, Eftimie Murgu Square No. 2A, RO-300041, Timisoara, Romania, phone/fax: +40256220479; e-mail: scr-stin@umft.ro.

2014.24.2 (82)  Fiziologia - Physiology

43the first gene involved in HRS cells apoptosis. Mechanism of
BCL2 involvement is partly understood. BCL-2 expression by
HRS cells may prevent apoptosis induced by the absence of B
cells functional receptors, which explains tumorigenesis. BCL2 expression may also explain the resistance to treatmentinduced HRS cells apoptosis (15).
AIMThis study aims to determine BCL-2 expression in Hodgkin’s
lymphoma patients included in the study, correlation of its expression with biochemical and immunological changes, as well
as evaluation of survival in these patients.
METHODOLOGY
A retrospective analysis has been conducted from May 2008
to April 2013 for 151 patients diagnosed with Hodgkin lymphoma
in the Hematology Department, Timisoara. The main diagnostic
method was biopsy, followed by histopathological and immunohistochemical examinations of harvested tissue. The stage of
disease was established by the means of computed tomography(CT). Polychemotherapy and number of cycles were decided
based on the disease histological stage and grading. Patients’
data regarding medical history and laboratory tests performed
have been extracted from each patient’s medical record.
Data were collected in a database and processed by the means
of SPSS 17 programme. Survival rates were calculated using
Kaplan-Meyer curves and correlations were established between BCL-2 presence and changes in the other biochemicalparameters.
RESULTS
Mean age of patients included in the study is 49.69 ± 17.46, witha minimum of 18 and a maximum of 89 years, out of which 37.7%
were women and 62.3% men. The follow up period since diagnosis
was 13.92 ± 6.24 months up to complete remission (33.7% of
patients); partial remission was seen in 45.7% of patients, 17.2%
had progressive disease, 0.6% relapsed and 2.5% died.
BCL-2 was detected in 82 patients (54.3%), the intensity varying according to the histological degree of the disease, as follows:
BCL-2 expression is seen in 19.8% of MC patients, 39.1% of
NS patients and 5.3% of patients with lymphocyte depletion.
Table I. BCL-2 expression according to clinical and biologicalparameters
Parameter
Present BCL-2
Absent BCL-2
expression
expression
Age (mean)
54.3%
45.7%
Male gender
33.2%
29.1%
ESR (>10)
24.5%
39.7%
Ceruloplasmin (in0.7%
28.5%
creased level)
LDH (increased level)
11.9%
27.2%
Fibrinogen (increased
30.4%
4%level)
CD15
54.3%
45.7%
CD20
54.3%
45.7%
CD30
54.3%
45.7%

44

There is an inverse, but statistically significant correlation(p<0.01) between BCL-2 expression and ESR, ceruloplasminand LDH. Fibrinogen, CD15, CD20 and CD30 exhibit positive,
statistically significant correlation with BCL-2 (p<0.01).
Table II. Correlations between BCL-2 expression and the otherevaluated biochemical and immunological parameters
Parameter
ESR

Correlation-0.277p<0.01

Ceruloplasmin
LDH
Fibrinogen
CD15
CD20
CD30

-0.473-0.447
0.323
0.843
0.944
0.866

<0.01
<0.01
<0.01
<0.01
<0.01
<0.01

Survival rates in patients with present BCL-2 expression islower in those with more intense gene expression (p<0.01),
which is also true for CD15, CD20, CD30 markers.

Fig. 1. Survival according to BCL-2 expression

Fig. 2. Survival according to CD15 levels

Fiziologia - Physiology  2014.24.2 (82)Fig. 3. Survival according to CD20 levels

Fig. 4. Survival according to CD30 levels

Fig. 6. Survival according to ceruloplasmin levels

Fig. 7. Survival according to LDH levels

Increased ESR levels are associated with survival periods
equal to that of patients with normal ESR levels, being inversely
correlated with BCL-2 expression, which is also seen in case ofceruloplasmin and LDH.

Conversely, increased fibrinogen levels are associated withdecreased survival rates and show positive, statistically significant correlation (p<0.01) with BCL-2 expression.

Fig. 5. Survival according to ESR levels

Fig. 8. Survival according to fibrinogen levels

2014.24.2 (82)  Fiziologia - Physiology

45

DISCUSSION
It is shown in the literature that changes in diseaseactivity tests (ESR, fibrinogenemia, LDH and serum alkaline
phosphatase) are positively correlated with a more aggressive
form of disease and therefore with a more reduced rate ofcomplete remission and shorter survival (5-year survival rate);
ESR levels over 80 mm/h and serum alkaline phosphataseover 200 IU/ml are highly significant prognostic risk factors. Our
study shows reduced survival in patients with high fibrinogenlevels, the survival rate being less influenced by increased
ERS, ceruloplasmin and LDH levels.
Regarding the expression of immunological markers,
our study shows a significant reduction in survival rates ofpatients with BCL-2 expression, which is closely correlatedwith intense expressions of the other immunological markers,
namely CD15, CD20 and CD30, some authors consideringthat BCL-2 expression by HRS cells may prevent apoptosis
induced by the absence of B cells functional receptors, which
explains tumorigenesis. BCL-2 expression may also explain
the resistance to treatment-induced HRS cells apoptosis.
CONCLUSION
A reduced survival rate of patients with BCL-2 expression
associated with intense expression of CD15, CD20 and CD30
markers, as well as with increased fibrinogen levels were seenin this study. Biochemical markers ESR, LDH and ceruloplasmin
are inversely correlated with presence of BCL-2, and survival
rates are not significantly changing at increased levels compared
to normal.
REFERENCES
1. Ishida T, Ishii T, Inagaki A, et al. Specific recruitment of CC
chemokine receptor 4-positive regulatory T cells in Hodgkin lymphoma
fosters immune privilege. Cancer Res. 2006;66(11): 5716-5722.
2. Marshall NA, Christie LE, Munro LR, et al. Immunosuppressive
regulatory T cells are abundant in the reactive lymphocytes of Hodgkin
lymphoma. Blood. 2004;103(5):1755-1762.
3. Ma Y, Visser L, Blokzijl T, et al. The CD4_CD26- T-cell populationin classical Hodgkin’s lymphoma displays a distinctive regulatory T-cell
profile. Lab Invest. 2008;88(5):482-490.
4. Steidl C, Shah SP, Woolcock BW, et al. MHC class II transactivator

46

CIITA is a recurrent gene fusion partner in lymphoid cancers. Nature.
2011;471(7338):377-381.
5. Diepstra A, van Imhoff GW, Karim-Kos HE, et al. HLA class
II expression by Hodgkin Reed-Sternberg cells is an independent
prognostic factor in classical Hodgkin’s lymphoma. J Clin Oncol.
2007;25(21):3101-3108.
6. van den Berg A, Visser L, Poppema S. High expression of the CC
chemokine TARC in Reed-Sternberg cells. A possible explanation forthe characteristic T-cell infiltration Hodgkin’s lymphoma. Am J Pathol.
1999;154(6):1685-91.
7. Biggar RJ, Jaffe ES, Goedert JJ, Chaturvedi A, Pfeiffer R, Engels
EA. Hodgkin lymphoma and immunodeficiency in persons with HIV/
AIDS. Blood. 2006;108(12):3786-91.
8. Atayar C, Poppema S, Visser L, van den Berg A. Cytokine gene
expression profile distinguishes CD4_/CD57_ T cells of the nodular
lymphocyte predominance type of Hodgkin’s lymphoma from their
tonsillar counterparts. J Pathol. 2006;208(3): 423-30.
9. Tanijiri T, Shimizu T, Uehira K, et al. Hodgkin’s Reed- Sternberg cellline (KM-H2) promotes a bidirectional differentiation of CD4CD25Foxp3
T cells and CD4 cytotoxic T lymphocytes from CD4 naive T cells. J
Leukoc Biol. 2007;82(3):576-84.
10. Chemnitz JM, Eggle D, Driesen J, et al. RNA fingerprints provide
direct evidence for the inhibitory role of TGFbeta and PD-1 on CD4_ T
cells in Hodgkin lymphoma. Blood. 2007; 110(9):3226-33.
11. Chemnitz JM, Driesen J, Classen S, et al. Prostaglandin E2 impairs
CD4_ T cell activation by inhibition of lck: implications in Hodgkin’s
lymphoma. Cancer Res. 2006;66(2):1114-22.
12. Juszczynski P, Ouyang J, Monti S, et al. The AP1-dependent
secretion of galectin-1 by Reed Sternberg cells fosters immune
privilege in classical Hodgkin lymphoma. Proc Natl Acad Sci U S A.
2007;104(32):13134-39.
13. Oelmann E, Herbst H, Zuhlsdorf M, et al. Tissue inhibitor of
metalloproteinases 1 is an autocrine and paracrine survival factor,
with additional immune-regulatory functions, expressed by Hodgkin/Reed-Sternberg cells. Blood. 2002;99(1): 258-267.
14. Yamamoto R, Nishikori M, Kitawaki T, et al. PD-1-PD-1 ligand
interaction contributes to immunosuppressive microenvironment of
Hodgkin lymphoma. Blood. 2008;111(6):3220-24.
15. Rassidakis GZ, Medeiros LJ, Vassilakopoulos TP, Viviani S, Bonfante V, Nadali G, Herling M, Angelopoulou MK, Giardini R, Chilosi M,
Kittas C, McDonnell TJ, Bonadonna G, Gianni AM, Pizzolo G, Pangalis
GA, Cabanillas F, Sarris AH. BCL-2 expression in Hodgkin and ReedSternberg cells of classical Hodgkin disease predicts a poorer prognosis
in patients treated withABVD or equivalent regimens. Blood. 2002;
100: 3935-41.

Fiziologia - Physiology  2014.24.2 (82)

EXPRESIA BCL-2 CA FACTOR DE PROGNOSTIC NEGATIV IN
LIMFOMUL HODGKINREZUMAT

Introducere: Cu toate ca s-au facut progrese importante in ceea ce priveste tratamentul bolii Hodgkin (HD), prognosticul acesteia
este nefavorabil in multe dintre cazuri. De aceea, au fost effectuate o multitudine de analize ale factorilor care ar putea influenta
pozitiv evolutia si prognosticul bolii si se crede ca detectarea precoce dupa debutul bolii permite administrarea unei terapii adecvate.
Studii de imunologie moleculara au aratat au aratat ca in limfomul Hodgkin clasic HRS (Hodgkin Reed Sternberg) celulele deriva
din centrul germinativ al limfocitelor B cu rearanjamente genice ale imunoglobulinelor, dar fara expresia receptorilor de suprafata ai
limfocitelor B. BCL-2 a fsot identificata ca prima gena implicata in apoptoza celulara a HRS. Expresia BCL-2 de catre celulele HRS
poate preveni apoptoza indusa de absenta receptorilor functionali ai limfocitelor B, ceea ce explica tumorigeneza. Expresia BCL-2
poate explica de asemenea rezistenta la apoptoza celulelor HRS indusa de tratament.
Scop: Acest studiu isi propune sa determine expresia BCL-2 la pacientii cu limfom Hodgkin inclusi instudiu, corelarea expresiei
acesteia cu modificarile biochimice si imunologice, precum si evaluarea supravietuirii la acesti pacienti.
Metodologie: A fost efectuata o analiza retrospectiva pentru perioada mai 2008-aprilie 2013 pe unlot de 151 de pacienti
diagnosticati cu limfom Hodgkin in Departamentul de Hematologie Timisoara. Principala metoda de diagnostic a fost biopsia,
urmata de examinarea histopatologica si imunohistochimica a tesutului prelevat.
Rezultate: Varsta medie a pacientilor inclusi in studiu a fost 49,69±17,46, cu varsta minima de 18 ani iar varsta maxima de 89 deani, din care 37,7% au fost de sex feminin si 62,3% de sex masculin. Perioada de urmarire din momentul diagnosticului a fost de
13,92±6,24 luni pana la remisiunea completa (33,7% dintre pacienti); remisiune partiala a fost intalnta la 45,7% dintre pacienti,
17,2% au prezentat boala progresiva, 0,6% au avut recaderi si 2,5% au murit. BCL-2 a fost detectata la 82 de pacienti (54,3%),intensitatea expresiei fiind variabila si concordanta cu gradul histologic al bolii, dupa cum urmeaza: expresia BCL-2 a fost de 19,8%
dintre pacientii cu MC, 39,1% la cei cei cu NS si 5,3% la pacientii cu depletie limfocitara. Supravietuirea a fost mai scazuta lapacientii cu expresie a BCL-2 comparativ cu cei care nu exprimau aceasta gena, iar expresia genica a aratat o corelatie pozitiva,
semnificativa statistic cu niveluri crescute ale altor markeri imuni (CD15, CD20, CD30) si biochimici (fibrinogen).
Concluzii: In acest studiu am relevat o scadere a ratei de supravietuire la pacientii cu expresie a BCL-2, asociata cu cresterea
expresiei markerilor CD15, CD20 si CD30, precum si cu cresterea nivelului fibrinogenului. Markerii biochimici VSH, LDH si ceruloplamina sunt corelati invers proportional cu prezenta BCL-2, iar rata de supravietuire nu este modificata semnificativ de niveluri
crescute comparativ cu normalul.

2014.24.2 (82)  Fiziologia - Physiology

47

OTITIS MEDIA WITH EFFUSION IN CHILDREN – A CLINICALTRIAL
KARINA MARIN, ROXANA VINTILA, ROXANA POPESCU, ALIN HORIA MARIN, PRODEAMIHAELA, MARIOARA POENARU“Victor Babes” University of medicine and Pharmacy Timisoara

ABSTRACT

Otitis media with effusion (OME), or ‘glue ear’, is very common in children, especially between the ages of one and three years
following Eustachia tube impairment. OME is defined as middle ear effusion without signs or symptoms of an acute infection. OME
may occur as a primary disorder or as a sequel to acute otitismedia. The functional effect of OME is a conductive hearing loss ofabout 25 to 30 dB associated with fluid in the middle ear.
Materials and methods: In total 250 patients with otitis media with effusion were enrolled in this study. They were diagnosed with
OME between 2011- 2014 in Bega ENT Clinic Timisoara. Diagnosis was established by otomicroscopy. In children with allergic
symptoms prick tests were performed. All children underwent pure tone audiometry, tympanometry, acoustic reflex. All children
underwent nasal bacterial exam, nasal endoscopy. Treatment was conservative or surgical.
Results: 214 (85.6%) patients presented transmission hearing loss, with type B tympanogramm. Transmission hearing loss ranged
between 25- 40 dB. 24 (9.6%) patients had type C tympanogramm. All patients had absence of ipsilateral acoustic reflex. Nasal
bacterial exam showed following results: S pneumoniae was found in 35% of cases, H influenzae is found in 20%, M catarrhalis
was present in 1% of the cases. Tympanocentesis was performed in 156 cases. Typmanotomy and adenoidectomy was performed
in 176 children. Tympanotomy, adenoidectomy and ventilation tube insertion was performed in 20 cases. Complications were otorrhea, tympanosclerosis and perforation.
Key words: otitis media with effusion, transmission hearing loss, ventilation tube, adenoidectomy

INTRODUCTION
Otitis media with effusion (OME) is an inflammatory process
within the middle ear space consisting of fluid accumulation.
Otitis media with effusion (OME) nearly always follows acuteotitis media (AOM) as it resolves. It is due to a failure of theEustachian tube to ventilate, clear and drain secretions. Ligationof the eustachian tube in animals invariably leads to the formationof a persistent middle ear effusion. Middle ear effusions (MEE)
result from inflammation within the middle ear cleft (otitis media).
Non-inflammatory middle ear fluid can result from barotrauma,
head trauma or a cerebrospinal fluid leak. Symptoms of acuteotitis media AOM are congestion of the tympanic membrane,
bulging of the tympanic membrane, fever, pain. It can lead topurulent secretions.
The newer models describe the primary event as inflammation of the middle ear mucosa caused by a reaction to bacteria
already present in the middle ear. Indeed, Bluestone andothers have shown (using radiographic evidence) that reflux
up the eustachian tube is demonstrable in children prone tootitis media (1). Furthermore, Crapko et al demonstrated thepresence of pepsin in the middle ear space of 60% of children
with otitis media with effusion (2). Many factors have been
implicated in the failure of the clearance mechanism, including
ciliary dysfunction; mucosal edema; hyperviscosity of the effusion; and, possibly, an unfavorable pressure gradient. Otitis

media with effusion is ubiquitous in children who have a cleft
palate. The cause is simply the lack of proper insertion of thetensor velipalatini muscle in the soft palate. The muscle is,therefore, unable to open the eustachian tube on swallowing
or wide mouth opening.
There is also a possibility that allergy can contribute to otitis
media with effusion. Major classes of immunoglobulins, complement, and immune complexes of antigen and antibody are foundin MEE with their concentrations being higher in mucoid effusion
as compared to serous effusions.
These mediators are hypothesized to play role in the pathology of otitis media.
Mucocilliary dysfunction caused by allergy and impaired Eustachian tube function can lead to negative pressure in the middle
ear resulting in transudation of liquid from the mucosa. Persistent
middle ear fluid from OME leads to mobility impairment of the
tympanic membrane and serves as a barrier to sound conduction.
When conservative treatment has failed, ventilation tubes
are generally used for treatment and prevention of recurrence.
OME appears usually in children occurring up to 90% by theage of 2. Refractory OME can lead to hearing loss, late language
acquisition, behavioral and cognitive difficulties. The mostcommon bacteria in acute otitis media, in order of frequency,
are Streptococcus pneumoniae, Haemophilus influenzae, andMoraxella catarrhalis

Received 12th of March 2014. Accepted 15th of May 2014. Address for correspondence: Karina Marin, MD, PhD Student Department of ENT, University of Medicine
and Pharmacy “Victor Babeș” Timișoara, Eftimie Murgu Square No. 2A, Timisoara, RO-300041, Phone: +40723316869, e-mail: marinkrn@yahoo.com

48

Fiziologia - Physiology  2014.24.2 (82)MATERIAL AND METHODS
In total 250 patients with otitis media with effusion wereenrolled in this study. They were diagnosed with OME between
2011- 2014 in Bega ENT Clinic Timisoara.
In order to establish the diagnosis and therapy all children
were examined by otomicroscopy, and a complete ENT consult.
Children who presented a positive family history or symptoms forallergy were submitted to allergy tests and measurement of total
and specific IgE level. Skin tests were performed for perennial
and seasonal allergens. Children underwent skin prick test for
respiratory allergens like dust mite, pollen, mixture of herbs,
molds, cat and dog epithelium. Results were evaluated after 10
minutes. Wheals greater than 3 mm at the side of the negative
control were considered positive.
All children underwent pure tone audiometry, tympanometry
and ipsilateral acoustic reflex.
Tympanograms were divided into the following types: type A(+ 99 to – 99 mmH2O), type C1 (-100 to -199 mmH2O, type C2(more than 200 mmH2O).
All children underwent nasal bacterial exam.
All children underwent nasal endoscopy. Children were treatedconservatory with nasal corticosteroids and surgical (tympanotomy, tympantotomy and adenoidectomy, tympanotomy,
adenoidectomy and ventilation tube insertion). Follow up wasafter 2 weeks and after 3 months after.
RESULTS146 (58.4%) children presented with positive allergic tests.
The presenting complaints were hearing loss in 196 (78.4%)
patients, lack of attention in 50 patients (20%), otalgia in 187(74.8%) patients, fullness in 200 (80%) patients, and tinnitus in
28 (11.2%) patients.
Children presented retracted tympanic membrane, fluid level,
bubbles, hypervascularity.
The age of the children ranged from 4 years and 10 years
with a mean age of 7.
154 (61.6%) were boys and 96 (38.4%) were girls.
146 (58.4%) children presented with positive allergic tests.
The presenting complaints were hearing loss in 196 (78.4%)
patients, lack of attention in 50 patients (20%), otalgia in 187(74.8%) patients, fullness in 200 (80%) patients, and tinnitus in
28 (11.2%) patients.

214 (85.6%) patients presented transmission hearing loss,
with type B tympanogramm. Transmission hearing loss ranged
between 25- 40 dB.
24 (9.6%) patients had type C tympanogramm. All patients
had absence of ipsilateral acoustic reflex.
Nasal bacterial exam showed following results: S pneumoniae
was found in 35% of cases, H influenzae is found in 20%, M
catarrhalis was present in 1% of the cases.

Fig. 2. Results of nasal bacterial exam

Of children younger than 2 years, 17% had recurrent disease.
This disease is sometimes associated with delayed language
development in children younger than 10 years, and the loss is
usually conductive, with an average air conduction threshold of
27.5 decibels (dB), but otitis media with effusion has also been
associated with sensorineural hearing loss. Delayed language
development was discovered in 15 cases.
Additional findings were turbinate enlargement especially incases of allergy (52%), postnasal drip, rhinorrhea, environmental
allergies.
48 patients presented allergies for dust mites, 20 patientswere positive for pollen, 15 patients were positive for mixtureof herbs, 15 patients were positive for molds,10 patients werepositive for cats and dog epithelia.

Fig. 3. Allergies presented by the patients

Fig. 1. Complaints presented by the patients2014.24.2 (82)  Fiziologia - Physiology

In general, the prognosis for otitis media with effusion
was good. Most episodes spontaneously resolved without
intervention, and many resolved undiagnosed.
49

Following spontaneous tube extrusion, 20-50% of patients
will have a recurrence of otitis media with effusion, potentiallyrequiring the replacement of pressure equalization tubes (PETs)
and, in most cases, simultaneous adenoidectomy.
TREATMENT
Tympanocentesis was performed in 156 children serving
both as therapeutic procedure and a diagnostic procedure.
Typmanotomy and adenoidectomy was performed in 176
children, and was performed in cases of chronic nasal obstruction
complicated with mucoid otitis media persisting beyond 3 months.
Tympanotomy, adenoidectomy and ventilation tube insertion was
performed in 20 cases asscoiated with persistent mucous otitis
media or repeated otitis media with effusion.

Fig. 4. Patients’treatment

In 10 children the ventilation tube was eliminated after 2 or 3
months. Most cases of effusion resolved spontaneously. Ongoing
monitoring is mandatory, because the frequency of structural
damage increases with effusion duration.
The overall complication rate after placement of pressure
equalization tubes appeared in 7 patients. Persistent otorrhea isthe most common complication, occurring in 2 patients. Second
was tympanosclerosis occurring in 6 patients, which was notclinically significant unless it was extensive.

Fig. 5. Complications appearing in patients

Persistent perforation appeared in 1 patient.
Complication after adenoidectomy was bleeding which
occurred in 2 patients, temporary velopharyngeal insufficiency in
4 patients. Follow up was performed after, at 2 weeks and then 3months after. Grommets were removed after 1 year or 2 years.
50

DISCUSSIONSOME is defined as a fluid in the middle ear. It is very
common in children leading to hearing loss or surgicalintervention. OME rate has to peaks: one around 2 years ofage, and the otherat 5 years (Zielhui et al.). As for gender,some studies revealed that OME appeared more frequentin boy than girls. The different prevalence rate can depend
also on the high atopic history. Approximately 20- 30% of thegeneral population is atopic and 15 % of children. Caffarelli etal. compared 192 children with OME with a control group of
200 children in order to determine the prevalence of allergy.
There were no significant differences between the 2 groups
regarding the family history of allergy and skin tests. Althoughchildren with OME symptoms la rhinitis, asthma, conjunctivitis,
were more frequent.
van den Aardweg et al. (5) saw a significant benefit of adenoidectomy as far as the resolution of middle ear effusion inchildren with OME is concerned. However, the benefit to hearing
is small and the effects on changes in the tympanic membrane
are unknown.
Lous et al. (4) indicates that the benefits of grommets in children appear small. The effect of grommets on hearing diminishedduring the first year. Potentially adverse effects on the tympanic
membrane are common after grommet insertion. Therefore an
initial period of watchful waiting seems to be an appropriate
management strategy for most children with OME.
Boonacker et al. (3) found that adenoidectomy is mostbeneficial in children with persistent OME aged ≥ 4 years. A
smaller beneficial effect was found in children with recurrent
AOM aged < 2 years. Browning (2) explained that in children
with OME the effect of grommets on hearing, as measured bystandard tests, appears small and diminishes after six to ninemonths by which time natural resolution also leads to improved
hearing in the non-surgically treated children.
Numerous systematic reviews have confirmed the efficacy
of pneumococcal vaccine in preventing pneumococcal AOM.
Following introduction of the 7-valent pneumococcal conjugate
vaccine (PCV7), childhood incidence of AOM declined in the USA
and Canada. In children aged under 2 years, a 43% reduction
in AOM.
REFERENCES
1. Qureishi A, Lee Y, Belfield K, Birchall JP, Daniel M. Update on otitis
media - prevention and treatment. Infect Drug Resist. 2014;7:15-24.
2. Browning GG, Rovers MM, Williamson I, Lous J, Burton MJ.
Grommets (ventilation tubes) for hearing loss associated with otitis
media with effusion in children. Cochrane Database Syst Rev. 2010;(10): CD001801.
3. Boonacker CW, Rovers MM, Browning GG, Hoes AW, Schilder AG,
Burton MJ. Adenoidectomy with or without grommets for children withotitis media: an individual patient data meta-analysis. Health Technol
Assess. 2014;18(5):1-118.
4. Lous J, Burton MJ, Felding JU, Ovesen T, Rovers MM, Williamson
I. Grommets (ventilation tubes) for hearing loss associated with otitis
media with effusion in children. Cochrane Database Syst Rev. 2005;(1): CD001801.

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5. van den Aardweg MT, Schilder AG, Herkert E, Boonacker CW,
Rovers MM. Adenoidectomy for otitis media in children. Cochrane
Database Syst Rev. 2010; (1): CD007810.

6. Perera R, Haynes J, Glasziou P, Heneghan CJ. Autoinflation
for hearing loss associated with otitis media with effusion. Cochrane
Database Syst Rev. 2006; (4): CD006285.

OTITA MEDIE SEROASA LA COPII – STUDIU CLINICREZUMAT

Otita medie seroasa apare in special la copii cu varste cuprinse intre 1-3 ani ca urmare a unei disfunctii a tubei lui Eustachio.
Transudatul poate apare de prima intentie sau urmand unei infectii acute. Este insotita de hipoacuzie de transmisie de 25- 30 dB.
Material si metoda: In studiu au fost inclusi 250 de pacienti. Acestia au fost diagnosticati cu otita seroasa pe perioada 2011- 2014
in Clinica ORL Bega Timisoara. Diagnosticul a fost stabilit ptin otomicroscopie. In cazul copiilor cu simptome alergice s-au efectuat
prick teste. Toti copiii au fost supusi audiometriei tonale, timpanometriei, reflexului stapedian, endoscopiei nazale. Tratamentul afost conservator sau chirurgical.
Rezultate: 214 (85,6%) din pacienti au prezentat hipoacuzie de transmisie intre 25- 40 dB, cu timpanograma de tip B, 24 pacienti(9,6%) au prezentat timpanograma tip C. In cazul tuturor pacientilor reflexul stapedian a fost abolit. �����������������������������
Exudatul nazal a relevant urmatoarele: S pneumonia a fost pozitiv in 35% din cazuri, H influenza in 20% din cazuri, M catarrhalis in 1% din cazuri. In 156 decazuri s-a efectuat timpanotomie. Timpanotomie plus adenoidectomie s-a efectuat la 176 de cazuri. Adenoidectomie si insertie detub de dren s-a efectuat in 20 de cazuri. Complicatiile aparute au fost otoreea, timpanoscleroza si perforatia.
Cuvinte cheie: otita media seroasa, hipoacuzie de transmisie, tub de ventilatie, adenoidectomie

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MULTIPLE MYELOMA AND STENT RESTENOSIS IN
OBSTRUCTIVE CORONARY ARTERY DISEASEVALCOVICI M, PASCALAU L, IANCU O“Victor Babes” University of medicine and Pharmacy Timisoara

ABSTRACTThe cardiotoxicity of chemotherapeutics is a reality. The association between cardiovascular risk factors and coronary events risk, during
the chemotherapy may precipitate an acute coronary syndrome that requires a fast approach to myocardial ischemia. We present thecase of a multiple myeloma patient who 2 months after the initiation of chemotherapy treatment was associated with an acute coronary
syndrome which required interventional revascularization. The appearance of restenosis at the resumption of chemotherapeutic cureraises the problem of pathophysiological interpretation of the proinflammatory phenomenon.
Keywords: multiple myeloma, anthracyclines, acute coronary syndrom, restenosis.

INTRODUCTION
Cardiotoxicity is defined by the disfunction of parameterswhich alters the global systolic function, as well as theelectrophysical parameters of the heart during the treatment
with chemotherapeutic agents dose-dependent.
Cardiotoxicity induced by chemotherapeutic agents, forexample anthracyclines, is considered to be resistant toconventional treatments. For many of these patients, the heartdisease mortality risk will become equal or even higher than themalignancy recurrence risk. Certain forms of chemotherapy havebeen directly associated with cardiotoxicity.
Current guidelines recommend that during the treatment
with chemotherapeutic agents to evaluate the systolic function
through different imagistic techniques. The cardiotoxicity
consequences may interest the coronary vasomotricity inpresence of traditional risk factors. In some cases, interventionalor surgical revascularization of these patients is the standard andobligatory procedure. For those interventional revascularizedthrough endovascular prosthesis placement, the restenosis
remains the major complication of these procedures.
We present the case of a patient with multiple myeloma
history who develops after about 2 months after the initiationof the chemotherapeutic agents treatment (see 2 applications:vincristine 0.4 mg continuous perfusion, anthracycline 9
mg / m² continuous perfusion, dexamethasone 40 mg, on
day 1-4) in the first month, 1 month break then initiates theadministration of velcade 1.3 mg / m ² (days 1-4, and 8-11),develops an episode of acute coronary typical pain.
CLINICAL CASE
The patient, S.A., aged 67 years, with cardiovascular risk
factors presents on one side hemodynamic: average aortic
stenosis and arterial essential hypertension stage II, metabolic:
LDL c (167 mg / dl), ischemic through STEMI and throughstenosis on LAD, multiplied risk through chronic renal failure
stage III, with multiple myeloma IgG kappa III B, shows up
with costrictive retrosternal intense pain which succumbs
to nitroglycerin, being clinically, electrically and biologicallydiagnosed with previous acute myocardial infarction with ST
segment elevation.
The ECG can detect 1-2 mm ST segment elevation in V1-V2
and depression 2 mm in V4-V6 (Figure 1). The transthoracic
ecocardiography reveals normal size VS, with concentric left
ventricular hypertrophy, global systolic function slightly altered(ejection fraction of 48%), global hypokinesis, degenerativechanges of mitral aortic apparatus, mitral regurgitation level
III, average aortic stenosis, dilatation of the left atrium. It is
decided the performing of the coronary angiography that can
detect: left dominant coronary system, aortic and coronary
calcifications visible through fluoroscopy. ACS, ACX, ACD
trunk without significant damage. Stenosis 80% - 85%, shortwith inhomogeneous appearance average LAD, distal vessel
without angiographic significant lesions (Figure 2). From thestenosis zone emerges Diagonal I, which presents an ostial
stenosis of 30%, distal vessel without significant lesions. It
was conducted the interventional permeabilization to averagelevel LAD (according to secondary prevention guidelines of the
obstructive cardiovascular disease), practicing the direct stentingof the lesion from average level LAD with BMS Integryti 3.0 /18 mm stentys, with good final result, without residual stenosis,
without proximal or distal dissection (Figure 3).

Fig. 1. 1-2 mm elevation of ST segment in V1-V2 and 2 mm depression in V4-V6

Received 13th of March 2014. Accepted 14th of May 2014. Address for correspondence: Valcovici M, MD, PhD Student Department of Cardiology, University of
Medicine and Pharmacy “Victor Babeș” Timișoara, Eftimie Murgu Square No. 2A, Timisoara, RO-300041, Phone: +40740250950, e-mail: mihaeladanielacardio@
gmail.com

52

Fiziologia - Physiology  2014.24.2 (82)Fig. 2. Coronary vessel re-permeabilization

Fig. 4. Interventional procedure at the level of LAD

Fig. 3. Intrastent stenosis

Fig. 5. 80% lesion at the level of LAD/calcified LAD

The patient is discharged stable, hemodynamically,
asymptomatic, with the following recommendations: regularcardiological evaluation, including ultrasound monitoring, thecontinuing of the basic medical treatment associated with dual
anti-aggregation platelet. In about 4 months from the coronary
injury, the patient shows up in a specialized hospital unit forrecurrence of angina type pain accompanied by dyspnea oforthopnea nature which hardly succumbs to nitroglycerin. The
EKG can detect ST-segment depression of 2-3 mm in V4-V6(Figure 1). The performance of a new angiographic exploration
highlights intrasent restenosis, with 90% lumen reduction to
the average LAD (Figure 4). Initially, it is performed balloon
angioplasty with good angiographic final result. The postprocedural 24 hours, the patient accuses retrosternal pain of
constrictive nature, with changes the dynamics of the electricalrest circuit path, ischemic-lesional type changes in the previous
territory. It is resumed the angiography procedure and it isperformed direct stenting with pharmacological stent active
Xience 2.75 / 38 mm, at average level LAD with good final result,
without proximal or distal dissection (Figure 5).

Fig. 6. ST segment depression in V4-V6, 1-2 mm depression of ST
segment in L1-L2

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DISCUSSION
The presented case illustrates on one side the additional risk ofchemotherapeutic cardiotoxicity with anthracycline in presence ofa high cardiovascular risk, multiplied by the presence of chronic
kidney disease stage III. (RFG 38.94). Therefore, ischemic risk
through the angiocoronarography evaluation in presence of themetabolic risk through LDLs increased.
The conducted studies to this date, demostrate the direct
association relationship between cardiovascular risk factors andanthracycline cardiotoxicity (guideline ACC / AHA published in
JACC in 2014), reveals the possibility of early cardiovascularcomplications and insists on the prediction strategies ofsubsequent cardiotoxicity at the initiation of the treatment withanthracyclines.
The peculiarity of the case focuses on the intrastent restenosis
phenomenon, which is well-known complication after therevascularization procedure through coronary endovascular
prosthesis. It is clear that the inflammation plays an importantrole in the pathology of Pathway - proteasome restenosis,
remains crucial in the controlling of the biological process that

includes inflammation, cell proliferation and apoptosis, events
that contributes to the neo-intima formation.
REFERENCES
1. Thavendiranathan P, Poulin F,Lim K-D, Plana JC, Woo A, Marwick
TH. Use of Myocardial Strain Imaging by Echocardiography for the
Early Detection of Cardiotoxicity in Patients During and After Cancer
Chemotherapy- A Systematic Review. Journal of the American College
of Cardiology , 2014, doi :10.1016/j.jacc.2014.01.073.
2. Bonura F, Di Lisi D, Novo S, D´Alessandro N. Timely Recognition of
Cardiovascular Toxicity by Anticancer Agents: A Common Objective of
the Pharmacologist, Oncologist and Cardiologist. Cardiovasc Toxicol,
2014, doi 10.1007/s12012-011-9141-z.
3. Calvo-Romero JM, Fernandez-Soria-Pantoja R, Gil-Cubero M.
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MIELOMUL MULTIPLU SI RESTENOZA INTRASTENT IN BOALA
CORONARIANA OBSTRUCTIVAREZUMAT

Cardiotoxicitatea chimioterapicului este o realitate. Asocierea intre factorii de risc cardiovasculari si riscul evenimentelor coronariene,
in cursul chimioterapiei poate precipita un sindrom coronarian acut ce necesita o abordare rapida pentru miocardul ischemic. Prezentam cazul unui bolnav cu mielom multiplu care la 2 luni dupa initierea tratamentului chimioterapeutic, s-a asociat cu un sindrom
coronarian acut ce a necesitat revascularizare interventionala. Aparitia resetenozei la reluarea curei chimioterapeutice ridica problemade interpretarea fiziopatologica a fenomenului proinflamatoar.
Cuvinte cheie: mielom multiplu, antracicline, sindrom coronarian acut, restenoza.

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