FIZIOLOGIA
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ARDELEAN AURELBĂDĂRĂU ANCA
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Fiziologia - Physiology • 2022.1(103)CUPRINSREZISTINA ÎN BOALA ALZHEIMER – DE LA MECANISME FIZIOLOGICE LA MECANISME PATOLOGICE ȘI ÎNAPOI…….4
Oliver Daniel Schreiner, Thomas Gabriel Schreiner
FIZIOLOGIA MODIFICĂRILOR CARDIOVASCULARE ASOCIATE CU INFECȚIA CU COVID-19: O REVIZUIRE.................10
Olusoji Adeola Adalumo, Sekunola Elizabeth Akinoye
EVENIMENTELE TROMBOTICE ȘI ȚINTELE DE ANTICOAGULARE ÎN INFECȚIA COVID-19………………………………18
Iacob Daniel Goje, Greta Ionela Barbulescu, Valentina Gabriela Ciobotaru, Oana Elena Ancusa, Ivanka Marian Todor, Daniel
Florin Lighezan
IMPACTUL TRATAMENTULUI CU HGMM ASUPRA INCIDENTEI RCIU IN SARCINILE CU TROMBOFILIE EREDITARA…23
Diroescu Ciprian , Anghel-Savciu Nicoleta-Loredana, Pana Razvan Cosmin, Cernea Nicolae
MODIFICARI VASCULARE IN SARCINILE CU TROMBOFILIE. ANTICOAGULAREA FACE DIFERENTA?………………..27
Anghel-Savciu Nicoleta-Loredana, Pana Razvan Cosmin, Cernea Nicolae, Diroescu Ciprian
PROVOCĂRI ÎN MANAGEMENTUL CARCINOMULUI BAZOCELULAR INVAZIV AL DORSUM NASI..........………………..31
Ivan Vatamanesku, Viorel Parasca, Maria-Magdalena Smeu, Dragos Popa, Dan Calota, Tiberiu Tebeica, Marius Eugen Ciurea
METODĂ DE IDENTIFICARE A CONFIGURAŢIILOR CIS ŞI TRANS ÎN LIPIDELE BIOACTIVE UTILIZÂND PROFILURI
MASĂ-ENERGIE..........…………………………………………………………………………………………………………………..40
Cojocariu Carolina, Pascariu Mihai Cosmin, Dinca Nicolae, Onulov Robert,Sisu Eugen, Puscasiu Daniela, Serb Alina,
Georgescu Marius

CONTENTSRESISTIN IN ALZHEIMER’S DISEASE – FROM PHYSIOLOGICAL TO PATHOLOGICAL MECHANISMS AND BACK..........4
Oliver Daniel Schreiner, Thomas Gabriel Schreiner
THE PHYSIOLOGY OF CARDIOVASCULAR CHANGES ASSOCIATED WITH COVID-19 INFECTION: A REVIEW………10
Olusoji Adeola Adalumo, Sekunola Elizabeth Akinoye
THROMBOTIC EVENTS AND ANTICOAGULATION TARGETS IN COVID-19 INFECTION…………………………………….18
Iacob Daniel Goje, Greta Ionela Barbulescu, Valentina Gabriela Ciobotaru, Oana Elena Ancusa, Ivanka Marian Todor, Daniel
Florin Lighezan
THE IMPACT OF LMWH TREATMENT ON IUGR INCIDENCE IN PREGNANCIES WITH INHERITED THROMBOPHILIA…23
Diroescu Ciprian , Anghel-Savciu Nicoleta-Loredana, Pana Razvan Cosmin, Cernea Nicolae
VASCULAR CHANGES IN THROMBOPHILIA PREGNANCIES. DOES ANTICOAGULATION MAKE A DIFFERENCE?......27
Anghel-Savciu Nicoleta-Loredana, Pana Razvan Cosmin, Cernea Nicolae, Diroescu Ciprian
CHALLENGES IN THE MANAGEMENT OF INVASIVE BASAL CELL CARCINOMA OF DORSUM NASI………………......31
Ivan Vatamanesku, Viorel Parasca, Maria-Magdalena Smeu, Dragos Popa, Dan Calota, Tiberiu Tebeica, Marius Eugen Ciurea
IDENTIFICATION OF CIS AND TRANS CONFIGURATIONS IN BIOACTIVE LIPIDS USING MASS-ENERGY PROFILES…40
Cojocariu Carolina, Pascariu Mihai Cosmin, Dinca Nicolae, Onulov Robert,Sisu Eugen, Puscasiu Daniela, Serb Alina,
Georgescu Marius

Fiziologia - Physiology • 2022.1(103)3

RESISTIN IN ALZHEIMER’S DISEASE – FROMPHYSIOLOGICAL TO PATHOLOGICAL MECHANISMS
AND BACK
OLIVER DANIEL SCHREINER1, THOMAS GABRIEL SCHREINER1,2,3,*
1University of Medicine and Pharmacy „Gr. T. Popa”, Iași, Romania
2Neurology Department, Clinical Rehabilitation Hospital, Iași, Romania
3University of Medicine and Pharmacy „Carol Davila”, Bucharest, Romania

Abstract: Resistin is an adipokine of great interest for the medical world, being increasingly associated with multiple inflammatory and noninflammatory pathologies, including Alzheimer's disease. Originally discovered in mice, the presence of resistin was later demonstrated also
in humans, but with some structural and functional differences. In the context of the effervescence of research in recent years, this article
aims to explore in detail the association between resistin and neurodegenerative pathology. While in the first part the mechanisms via which
this adipokine exerts its role in various cells and tissues are presented, in the second part the correlations between resistin and Alzheimer's
disease are discussed. The neuroinflammation hypothesis is the central element that brings the two entities together, but other theories could
also explain the roles of resistin in neurodegeneration. Based on pathophysiological data and the preliminary results from human and animal
clinical trials, it remains to be seen whether resistin will become a reliable biomarker for early detection of Alzheimer’s disease in the near
future.
Key words: Alzheimer’s disease, resistin, adipokine, biomarker, neuroinflammation

INTRODUCTION

Resistin, a small molecule belonging to the adipokine group[1], is currently receiving increasing attention fromresearchers, as more studies demonstrated its correlation
with numerous inflammatory conditions [2]. First describedby Steppan et al., 2001 [3] in mice as adipose-tissue-specificsecretory factor (ADSF), further research has shown that
resistin is also found in humans, being easily determined in
the peripheral circulation [4].
There are, however, significant differences between theresistin found in humans and its counterpart in animals(mainly mice and rats). First, while mouse resistin weighs 11
kDa and consists of 94 amino acids, the human resistin is a
12.5 kDa polypeptide containing 108 amino acids andshares only 59% of the murine counterpart sequence
identity [5]. The secretory cell of this adipokine also differs
when comparing humans to murine species. Murine resistin
is produced by adipocytes, its secretion being upregulated
by glucocorticoids, growth hormones, testosterone, andprolactin, while insulin, epinephrine, and somatotropin haveinhibitory influence [6]. In humans, there are a multitude ofcells capable of producing resistin, from mononuclear cells
and macrophages responsible for the production of the mostimportant percentage of circulating resistin, to a myriad ofother cells for different organs such as the pituitary gland,
hypothalamus, the colonic epithelium, pancreas, spleen,and skeletal muscle [7]. In human adipose tissue, researchshows that resident non-adipocyte inflammatory cells are
the main secretion site for resistin, contrary to the situationin mice [8]. Finally, it is worth mentioning the different
structural conformations of resistin in humans. Two isoforms
are more frequently encountered, an oligomer weighting 660
kDa and a trimer with a molecular weight of 45 kDa [9].
These two different assemblies were reported to be more
active in humans than in mice [10], and seem to play animportant role as pro-inflammatory factors [11], however
their biological importance is still to be fully determined.
The growing interest for the better understanding ofresistin’s physiological roles and its behavior in pathologicalconditions derived from recent results showing a positive
correlation between various inflammatory, immune andautoimmune conditions and resistin levels [12]. Resistin is
involved in a myriad of pathological conditions where achronic inflammatory status was assessed. Among the moststudied disorders associated with increased resistin levels
we mention obesity [13], metabolic syndrome [14], diabetes
mellitus type 2 [15], and atherosclerosis [16]. In recent years,
research has expanded, covering additional pathologies
such as non-alcoholic fatty liver disease [17], Crohn's
disease [18], Alzheimer's disease (AD) [19], and also noninflammatory pathological processes such as as endothelial
dysfunction, thrombosis, and dysfunction of smooth muscle
cells [20].
In this context, this review aims to present in the first part

Received 31st of January, 2022. Accepted 14th of April 2022. Address for correspondence: Thomas Gabriel Schreiner, University of Medicine
and Pharmacy „Gr. T. Popa”, str. Universității, no. 16, 700115, Iași, Romania; Phone: +40744860474, e-mail:schreiner.thomasgabriel@yahoo.com

4

Fiziologia - Physiology • 2022.1(103)the biological mechanisms by which resistin exerts its rolesin physiological conditions and in various pathologies.
Subsequently, starting from the neuroinflammatory theory ofAD, the authors highlight the potential role of resistin within
the pathogenesis of this neurodegenerative disease.
Summing up currently available theoretical knowledge andpreliminary results from clinical trials, it remains to be
determined if resistin can become a valuable biomarker for
AD early detection in the near future.
THE MOLECULAR PATHWAYS AND BIOLOGICALROLES OF RESISTIN
Being a relatively recently discovered molecule, only a
few of the molecular mechanisms by which resistin exerts
its effects in physiological and pathological conditions havebeen highlighted so far.
Resistin acts on a multitude of cell and tissue types [21],through various, still incompletely elucidated mechanisms.
For example, in human macrophages, resistin promotes the
production of pro-inflammatory cytokines such as TNF-α, IL6, IL-12 and alters the expression of cell adhesion molecules[22]. By promoting the expression of vascular cell adhesion
molecule-1 (VCAM-1), intercellular adhesion molecule-1(ICAM-1), and monocyte chemotactic protein-1 (MCP-1),
resistin contributes to chemotaxis and recruitment ofleukocytes to inflammation sites [23]. Similar effects are also
observed in peripheral blood mononuclear cells and hepatic
stellar cells [24]. Another target for this adipokyne isrepresented by both human and mouse blood vessel wallcells. At this level, under resistin modulation, smooth muscle
cell proliferation and endothelial cell dysfunction can beobserved [25]. These are also preliminary important steps in
inflammatory conditions that sustain immune cells adhesion
and infiltration via endothelium.
One debatable fact remains the target receptor forresistin. As detailed below (see Table I), there arecomplementary, sometimes contradictory result in theliterature regarding the cellular receptors via which resistin
exerts its functions.
Some research shows that resistin binds to the toll-likereceptor 4 (TLR4), subsequently exerting its proinflammatory functions [26]. In competition withlipopolysaccharide, via binding to TLR4, resistin finally leads
to the production of pro-inflammatory cytokines andinterferons, thus launching inflammatory and / or immune
responses [27]. There are already studies that havedemonstrated the effect of resistin via TLR4 mediatedpathway in the induction of hypertension and insulin
resistance in mice [28], or in facilitating the metastasis oflong adenocarcinoma in humans [29].
Daquinag et al., 2011 suggested that decorin could also
act as a resistin receptor in adipose progenitor cells in mice[30]. Further research highlighted the possibility for decorin
to be a resistin receptor also in humans, as a study
conducted on a Japanese population that demonstrated thatspecific single nucleotide polymorphisms in the vicinity of
the human decorin gene are associated with plasma resistin
Fiziologia - Physiology • 2022.1(103)[31]. In a similar fashion, research on a Finnish cohortsuggested that decorin polymorphisms might havebiological relevance in human vascular pathophysiology,
resistin becoming a potential therapeutic target [32].
Other possible receptors include the adenylyl cyclaseassociated protein 1 (CAP1), the insulin growth factor-1
receptor (IGF-1R), and the tyrosine kinase-like orphan
receptor-1 (ROR-1). Via CAP1, resistin can promoteincreasing of intracellular cyclic AMP levels, subsequently
enhancing protein kinase A and NF-κB activity, this pathway
explaining the production of inflammatory cytokines and therelated inflammatory response [33]. Several otherintracellular signaling cascades may also be relevant forresistin activity in different cells, among the most studied
ones being the pathway related to the L-type voltage
sensitive calcium channel [34]. Both the external calcium
influx and the activation of phospholipase C lead to theenhancement of intracellular calcium, which might
subsequently conduct to oxidative stress and immune cells
recruiting, sustaining the chronic inflammation [35][36].
Clinical trials suggested a possible relation between
resistin and IGF-1R, making IGF-1R a potential receptor forthis adipokine. In a study conducted on humans sufferingfrom rheumatoid arthritis, Boström et al., 2011 revealed thatabrogation of resistin synthesis is associated with areduction in IGF-1R expression at the level of the synovium,
suggesting that resistin utilizes the IGF-1R pathway [37].
Another research conducted by Bjersing et al., 2013 showedthat physical exercise modulates both resistin and IGF-1R
levels in fibromyalgia patients, suggesting that the twomolecules might share a similar pathway [38].
Finally, one study conducted on mice demonstrated therole of ROR-1 in the mediation of some of the multiple
functions such as adipogenesis and glucose uptake that
resistin can exert in 3T3-L1 cells [39]. As recent datasuggest that ROR-1 is involved in the proliferation, survival,
and metastasis of several cancer cells [40], including
malignant B cells in chronic lymphocytic leukemia [41],better understanding of this signaling cascade is of great
importance for future drug development.
RESISTIN IN ALZHEIMER’S DISEASE – A
POTENTIAL
BIOMARKER
LINKEDTO
THE
NEUROINFLAMMATORY THEORY
Alzheimer's disease (AD), the most common cause of
adult-onset dementia [42], is still lacking an effective therapy,
despite numerous clinical trials conducted so far. In thiscontext, researchers are increasingly focusing on theprevention of AD, the detection of reliable biomarkers for the
early stages of the disease being essential. Resistin, alongother adipokines, seems to be an important molecule for AD
pathogenesis, as recent studies suggest [19]. For example,
the study conducted by Leung et al., 2015 showed that
cerebrospinal fluid resistin was significantly associated with
Aβ1-42 levels and to cognitive impairment changes insymptomatic AD patients [43]. Similar results were foundalso in peripheral blood samples, as AD patients associated5higher serum resistin levels compared to the controls [44].
There is, however, a great variability between different
cohorts that must be taken into account. One recent study
demonstrated the ethnicity-related differences regarding
neurodegenerative diseases. Molecular biomarkers such as
resistin were elevated in African Americans compared to
Caucasians, these results suggesting the influence ofgenetic and ethnic background on AD [45]. Moreover, as
antidementia drugs such as acetylcholinesterase inhibitors
seem to lower serum resistin levels [46], this adipokine
remains a therapeutic target worth to be studied in future
trials.
Several pathophysiological mechanisms may explain
the association between resistin and dementia.
Inflammation remains a constant event in AD onset anddevelopment [47], among other potential mechanisms suchas excessive accumulation of amyloid beta in the brain [48],
the impact of oxidative stress [49], or the role of the vascularcomponentinneurodegeneration
[50].
The
neuroinflammatory status found in the earliest stages of AD
is characterized by increased levels of cytokines andchemokines, resistin being demonstrated to facilitate thesecretion of these proinflammatory factors. Althoughneuroinflammation has both neurodestructive andneuroprotective effects, in case of AD, activation of microglia
and the inflammatory cascade has mainly destructive andtoxic effects. Moreover, the association between AD andother chronic inflammatory conditions such as obesity andtype 2 diabetes mellitus [51] suggests the involvement ofshared cellular and molecular factors, resistin being one
possible link. Lastly, the multifaceted connection between
diabetes and AD has led many researchers to consider AD
as ”diabetes of the brain” or “type 3 diabetes” [52]. Amyloid
beta toxicity is linked to impaired insulin signaling, theincreased insulin resistance forming a vicious cycle that
subsequently sustains amyloid plaques pathologicalaggregation. Clinical data has already showed that highplasma resistin levels portend the insulin resistance-related
susceptibility to early cognitive decline [53], the role ofresistin and other common factors between the twopathological entities still to be completely explained.
CONCLUSION
Associated in the first place with inflammatory conditions
and insulin resistance, it has been demonstrated during
recent years that resistin plays also relevant roles in otherpathological conditions, including AD. Although thepathophysiological mechanisms are still incompletely
understood, according to current knowledge and preliminary
results from clinical trials, it seems that resistin can become
a valuable biomarker for AD early detection in the near
future. Future studies are needed in order to fully determine
resistin’s pathway and its relevance and AD etiology.

6

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Table I: Potential receptors for resistin
Molecular pathway

Potential receptor

Relevant clinical impact•

Toll-like receptor 4 (TLR4)

•
•
•
•
•

Decorin

Adenylyl cyclase-associated protein
1 (CAP1)
Insulin growth factor-1 receptor (IGF-

•
•
•
•

1R)
Tyrosine kinase-like orphan receptor1 (ROR-1)

8••

Stimulationofpro-inflammatory
cytokines and interferon production
Hypothalamic Inflammation
Insulin ResistanceHypertension
Metastasis
facilitation(lungadenocarcinoma)
Receptor in adipose progenitor cells inmice
Decorin polymorphism relevant forhuman vascular pathologies
Cyclic AMP increase - protein kinase A/ NF-κB pathway
L-type voltage sensitive calcium
channel mediated pathway
Clinical impact in rheumatoid arthritis
and fibromyalgia diagnosis andtreatment monitoring
Mediation of adipogenesis and glucose
via 3T3-L1 mice cells
Clinical impact in chronic lymphocytic
leukemia

Reference(s)Kuzmich et al., 2017 [27]Jiang et al., 2016 [28]Gong et al., 2018 [29]Daquinag et al., 2011 [30]
Onuma et al., 2013 [31]Kunnas et al., 2016 [32]Avtanski et al., 2019 [33]Singh et al., 2018 [34]
Boström et al., 2011 [37]
Bjersing et al., 2013 [38]
Sánchez-Solana et al., 2012 [39]Aghebati-Maleki et al., 2017 [41]Fiziologia - Physiology • 2022.1(103)REZISTINA ÎN BOALA ALZHEIMERMECANISME PATOLOGICE ȘI ÎNAPOI– DE LA MECANISME FIZIOLOGICELAREZUMAT
Rezistina este o adipokină de mare interes pentru lumea medicală, fiind asociată tot mai mult cu multiple patologii inflamatorii și noninflamatorii, inclusive cu boala Alzheimer. Descoperită inițial la șoareci, prezența rezistinei a fost demonstrată ulterior și la oameni, dar cu
unele diferențe structurale și funcționale. În contextul efervescenței cercetărilor din ultimii ani, acest articol își propune să exploreze în detaliu
asocierea dintre rezistină și patologia neurodegenerativă. În timp ce în prima parte sunt prezentate mecanismele prin care această adipokină
își exercită rolul în diferite celule și țesuturi, în a doua parte sunt discutate corelațiile dintre rezistină și boala Alzheimer. Ipoteza
neuroinflamației este elementul central care reunește cele două entități, dar și alte teorii ar putea explica rolurile rezistinei în neurodegenerare.
Pe baza datelor fiziopatologice și a rezultatelor preliminare din studiile clinice pe oameni și animale, rămâne de văzut dacă rezistina vadeveni un biomarker de încredere pentru detectarea precoce a bolii Alzheimer în viitorul apropiat.
Cuvinte cheie: boala Alzheimer, rezistina, adipokina, biomarker, neuroinflamație

Fiziologia - Physiology • 2022.1(103)9THE PHYSIOLOGY OF CARDIOVASCULAR CHANGESASSOCIATED WITH COVID-19 INFECTION: A REVIEW
OLUSOJI ADEOLA ADALUMO1, SEKUNOLA ELIZABETH AKINOYE1
1Department of Physiology, School of Basic Medical Sciences (SBMS), College of Health Sciences (COHS)

Federal University of Technology, Akure (FUTA), PMB 704, Ondo-State, Nigeria.

ABSTRACT
Although the clinical manifestations of zoonotic human COVID-19 are well understood to be respiratory, notable accounts of significant
cardiac complications ranging from myocyte biomarker elevation, arrhythmias, and myocardial dysfunction to electrocardiographic
abnormalities manifestations are being widely published. A few pathology studies conducted so far on patients that died of COVID-19
revealed that up to 35% of them harboured the novel SARS-CoV2 virus in the middle layer of their hearts, the myocardium. As of now, aprecise understanding of the effects of COVID-19 on the heart as well as the cardiovascular system as a whole is yet to be well-established,but existing evidence from preceding human zoonotic viral epidemics such as SARS-CoV and MERS-CoV suggests that viral infections intheir category could act as triggers for cardiac abnormalities such as acute coronary syndromes, arrhythmias, and heart failure via direct orindirect mechanisms. This review aims to summarise findings on the influence of the zoonotic COVID-19 on human cardiovascular functions
so far.
Keywords: Keywords: Coronavirus, Cardiovascular, ACE2 receptors, Cytokine storm, Spike protein, COVID-19

INTRODUCTION

A Brief Review of the Human Cardiovascular System
The cardiovascular system or the circulatory system are twointertwining terms used to describe the intriguing circulating
conduit in man, the system primarily consists of a cardiac
pump, which is famously referred to as the heart, a serving
circulatory medium which is the blood, then, the intricately
designed vasculature network which functions as the
conduit through which the blood flows (1). The heart issituated in the middle mediastinum, posterior and slightly tothe left of the sternum (2). The myocardium, a middle layerof the heart is structured with a bulk of cardiac tissues which
endows it with the ability to pump blood easily (3). Actions of
the conducting medium, known as blood, include
transportation of nutrients, oxygen, and electrolytes to cells,
removal of harmful waste of metabolic processes, andmaintenance of the internal milieu (homeostasis) (4). The
intricate vasculature system (an extensive network thatcomprises a large number of vessels such as arteries,
capillaries, and veins) contributes to the circulation of blood
throughout the cardiovascular system (5). The distribution ofoxygen, carbon dioxide, nutrients, and waste products within
the body are the central functions of the cardiovascularsystem in summary. However, the body never ceases toundergo major and minor changes in the course of its life
cycle, an integral role and requirement remain the capabilityof the cardiovascular system to accommodate and execute

vastly increasing body demands (3).
HISTORY AND EVOLUTION OF CORONAVIRUSES
Coronaviruses (CoVs) are viral pathogens that possess
single-stranded positive-sense RNA and can both mutate
and recombine effectively (6). Reportedly, four groups ofcoronaviruses (α, β, γ, and δ) have been identified, with only
two out of the four groups understood as zoonotic: αcoronaviruses (HCoV-229E and NL63) and β-coronaviruses(MERS-CoV, SARS-CoV, HCoV-OC43 and HCoV-HKU1)(7). These zoonotic coronaviruses often are the causes ofmoderate, self-limiting respiratory infections, accounting for
up to15% to 30% of all common colds (8). Other influencesof these viruses apart from respiratory infections also
include, intestinal disorders, neurological diseases andhepatic disorders (9). Patients with persistent cough, highfever, and shortness of breath followed by acute respiratory
distress syndrome (ARDS) caused by an unexplained viral
infection were reported in Wuhan (China) towards the endof December 2019. The analysis of viral genomes of fivepneumonia patients admitted to hospitals between
December 18 and December 29, 2019 revealed theexistence of the previously unknown β-coronavirus strain in
all of them (10).
The newly discovered βcoronavirus presented an 88
percent homology with the sequences of two bat
coronaviruses, bat-SL-CoVZC45 and bat-SL-CoVZXC21,

Received 22nd of February, 2022. Accepted 14th of May 2022. Address for correspondence: Olusoji Adeola Adalumo, 1Department of
Physiology, School of Basic Medical Sciences (SBMS), College of Health Sciences (COHS), Federal University of Technology, Akure (FUTA), PMB
704, Ondo-State, Nigeria; Phone: +2347031083977, e-mail: oaadalumo@futa.edu.ng

10

Fiziologia - Physiology • 2022.1(103)along with a homology of 79.5% and 50% with SARS-CoVand MERS-CoV respectively (10). The SARS-CoV outbreakstarted on November 2002 in Guangdong Province,
southern China, and was most likely initiated by a zoonotic
occurrence in China's wild-animal markets. Further researchon wild animals revealed compelling evidence that SARSCoV could have originated from bats (11). While in June
2012, the MERS-CoV crisis broke out in Saudi Arabia (12).
Through close contact, the virus spread from infecteddromedary camels, the intermediate host, to humans.
MERS-CoV is also thought to have originated in bats andspread to dromedary camels in the distant past (12-13). The
World Health Organisation (WHO) as of February 11, 2020,
officially named the newly emerging zoonotic virus; COVID19, an acronym for “Coronavirus Disease-2019”. This viral
pandemic unlike MERS-CoV and SARS-CoV, is capable ofcausing lethal pulmonary infections in humans (14). The
COVID-19 virion, like other coronaviruses, has a
nucleocapsid that houses the viral RNA and phosphorylated
N protein (15). The nucleocapsid is encased in phospholipid
bilayers and protected by a variety of proteins, including the
spike glycoprotein trimer (S) (spike protein, S protein), which
is found in all forms of CoV, haemagglutinin esterase (HE),the membrane and envelope proteins (M and E), which arefound between the spike (S) proteins in the viral envelope(16).
BASIC MOLECULAR PATHOBIOLOGY OF SARS-COV2
VIRUS
The viral SARS-CoV2 presents from the sub-division,
Coronavirinae in the family of Coronaviridae. The virus isrelatively microscopic (sizes about 60-140nm in diameter)
and the genome is an enveloped single stranded positivesense ribonucleic acid (+ssRNA), which normally ranges
from 26 to 32kbs in length, and considerably larger than any
other RNA virus. The newly discovered β-coronavirus takes
on a round or elliptic form with pleomorphic abilities, andalso unique with a halo or crown like appearance due to thepresence of protruding spikes from its surface. A certain
study in molecular genetics have convincingly demonstrated
that the virus consists of six major open-reading frames(ORFs); a reading frame that consists of a continuous
stretch of codons (begins at a start codon and ends at a stopcodon) known to other coronaviruses, and a number of otheraccessory genes (17-18). Around two-thirds of viral RNA istranslated into two significant polyproteins by the first ORF(ORF1a/b) (19), while the remaining one-third of the viralgenome is associated with the other open reading frames of
SARS-CoV2, and encodes for four major structural proteins:a spike glycoprotein (S), an envelope protein (E), a
nucleocapsid protein (N) and a membrane protein (M) alongwith various alternative proteins with unknown roles and are
not involved in the replication of the virus (20).
The spike protein mediates virus attachment to the hostcell and can uniquely recognize the human angiotensinconverting enzyme 2 (ACE2) receptor on the host cellsurface (21- 23). In coronaviruses, the envelope protein is a
Fiziologia - Physiology • 2022.1(103)small integral membrane protein which can oligomerize andform an ion channel (24). The E protein has been shown toplay a number of roles in the viral replication cycle, including
viral assembly (25), virion release (26), and viralpathogenesis (27). The coronavirus nucleocapsid (N)protein is a structural protein that binds directly to viral RNA
and provides stability (28). The coronavirus membrane (M)protein is an integral membrane protein that plays asignificant role in viral assembly and has been shown tohave the ability to cause apotosis. In order to encapsidate
the RNA genome, the protein also interacts with thenucleocapsid (N) protein. (29-31).
While SARS-CoV2 contains four structural proteins (S,E, M, and N), it also consists of sixteen nonstructuralproteins (nsp1−16). Nsp1 is a leader protein that plays a role
in RNA processing and replication. Nsp2 influences the hostcell's survival signaling pathway by binding to two hostproteins (PHB1) prohibitin-1 and (PHB2) prohibitin-2. Nsp3,
a papain-like proteinase protein is the largest encoded bycoronaviruses and thought to be involved in the separationof the translated protein. Nsp4 actively modifies ER
membranes. Interaction between Nsp3 and Nsp4 isimportant for viral replication and both consists oftransmembrane domains. During replication, Nsp5
participates in the polyprotein process. Nsp6 is a likely
transmembrane domain. The combination of Nsp12 andtemplate-primer RNA was significantly increased when
Nsp7 and Nsp8 were present. Nsp8 is a peptide cofactor
that forms a heterodimer with NSP7 (the other peptide
cofactor) and complexes with Nsp12. An Nsp8 monomer
unit also complexes with Nsp12 to form the RNA polymerase
complex, in addition to the Nsp7-Nsp8 heterodimer. Nsp9 is
a protein that binds to ssRNA. Nsp10 is needed for viralmRNA cap methylation. The function of Nsp11is currently
unknown. Nsp11 is made of thirteen amino acids and thefirst nine amino acids are identical to the first nine in Nsp12.
The RNA dependent RNA polymerase (RdRp), which is a
critical component of coronavirus replication andtranscription, is found in Nsp12. Nsp13 binds to ATP andparticipates in the replication and transcription processes via
its zinc-binding domain. The Nsp14 has 3′-5′
exoribonuclease activity and N7-methyltransferase activity
which is common to coronaviruses. Mn (2+)-dependentendoribonuclease activity is found in Nsp15. Nsp16 is a 2'Oribose methyltransferase (32-36).
NSP16 binds to the mRNA recognition domains of the
U1 and U2 snRNAs to inhibit mRNA splicing after
SARSCoV-2 infection. Nsp1 binds to 18S ribosomal RNA in
the ribosome's mRNA entry channel, interfering with mRNAtranslation. Nsp8 and Nsp9 bind to the 7SL RNA, which isfound at the Signal Recognition Particle, causing protein
trafficking to the cell membrane to be disrupted (37).
THE SPIKE GLYCOPROTEIN, ACE2 RECEPTORS AND
TMRSS2 CONTRIBUTIONS IN SARSCOV
The virus binds to a host cell via its target receptor, thatis the spike protein (S), which binds to the host cell surface

11receptor, Angiotensin converting enzyme 2 (ACE2) as thefirst step in the SARS-CoV2 entry cascade (38-39).
Although ACE2 is widely known to be expressed in theupper airway (goblet and ciliated epithelial cells), alveolar(Type II) epithelial cells of the lungs and pulmonary
vasculature. It is also actively present in the heart(cardiofibroblasts, cardiomyocytes, endothelial cells,
pericytes, and epicardial adipose cells) and the vascularsystem (endothelial cells, migratory angiogenic cells, andvascular smooth muscle cells) (40). The enzyme presents
as a transmembrane protein (specifically Type 1) with an
enzymatic domain located on the extracellular surface ofcells where it performs the roles of modifying angiotensin I
to angiotensin 1-9 and angiotensin II to angiotensin 1-7
respectively. The resulting angiotensin 1-7 then functions asa counter-regulator of the renin-angiotensin system (RAS)(41-42).
The RAS system regulates both blood pressure andelectrolyte balance, and thus plays an important role incardiovascular physiology (43). ACE2, as a functional
receptor for SARS-CoV2 couples with the humanTransmembrane Protease Serine 2 (TMPRSS-2), whichprimes the spike S protein of the virus to facilitate viral entry,
hereby significantly increasing chances of viral potency (4445). Since proteolytic cleavage of the viral S protein isessential for virus binding to ACE2, infection by SARS-CoV2
necessitates coexpression of ACE2 and TMPRSS2 in thesame cell type (46). The S protein has been shown to be
essential for proper protein folding, modulation of host cell
protease substrate accessibility, and antibody binding due toits extensive glycosylation (47-48). Zhou et al concluded
from infectivity experiments using Hela cells that expressed
ACE2 receptors that ACE2 meditated viral entry into hostcells, and that ACE2 could not promote viral entry into cells
that couldn’t express its corresponding receptor. This
discovery without an lingering doubts proved ACE2 as thereceptor through which SARS-CoV2 gains entry in host cells(18).
The transmembrane spike glycoprotein (encoded by the
structural S gene) forms homotrimers that protrude from theviral surface and has two subunits (S1 and S2) that bind to
the receptor. Binding is initiated by subunit S1, and fusion to
the infected cell is controlled by a trimeric S2 stalk. The S1subunit is composed of the N-terminal domain (S1-NTD)
and C-terminal domain (S1- CTD) which consists ofconserved amino acid residues referred to as RBD (receptor
binding domain), while features situated within the S2
subunit are: fusion peptide (FP), heptad repeat 1 (HR1),
central helix (CH), connector domain (CD), heptad repeat 2(HR2), transmembrane domain (TM), and cytoplasmic tail(CT) (23, 49, 18). The S1/S2 protease cleavage site is
located on the boundary between the S1 and S2 subunits.
The first cleavage (S1/S2 site), known as “priming cleavage,”
produces S1 (surface) and S2 (transmembrane) functional
subunits that remain non-covalently bound in a “prefusion”
conformation (50). The human serine protease TMPRSS-2(Transmembrane Protease Serine 2), a trypsin-like protease

12that has been shown to cleave monobasic sites and isrequired for SARS-CoV2 S1/S2 “priming” cleavage between
the RBD and fusion peptide (51).
Also, host proteases cleave the spike glycoprotein at theS2 cleavage site (activation cleavage) for all coronaviruses,
activating the proteins that are needed to fuse viralmembranes and host cells via permanent conformationalchanges (23, 49). The S1 subunit's receptor binding domain(RBD) directly recognizes the ACE2 receptor (52). The RBM(carboxy-terminal half of the RBD) is the region of the RBD
that contains the residues that interact with the host ACE2
receptor (53). The majority of SARS-CoV2 and ACE2
binding sites are found in RBM. The binding site is createdby two lobes of RBM and ACE2 and is contained in a
peptidase domain at the N-terminus of ACE2. RBM attaches
to the ACE2's small lobe on the bottom side. RBM's surface
is slightly concave inward to accommodate ACE2 (54).
In an analytical study of a resolution structure of spike
protein trimer with a single receptor binding domain in the“up” conformation (receptor accessible), due to this process,the S1 subunit dissociates and the S2 subunit refolds into astable post fusion conformation, as a result of receptor
binding destabilizing the pre-fusion structure. When RBD
undergoes
conformationaltransformations,thedeterminants of the spike protein are hidden or exposed,
allowing it to engage a host cell receptor. Two states will
hereby result from this process: "down" conformation and"up" conformation. However, SARS-CoV2 couldn’t identify
the ACE2 on the host cells in the "down" conformation (49).
Understanding the SARS-CoV2's receptor recognition
mechanism is critical since it dictates the virus's infectivity,
host range, and pathogenesis (53, 55).
Cardiovascular Complications associated with SARSCoV2 Numerous investigations have acknowledged that theviral receptor, ACE2, is also found in a variety of cellularcomponents of the cardiovascular system (i.e.,
cardiomyocytes, cardiac fibroblasts, pericytes, vascularendothelium, and vascular smooth cells) (56-57). The
relationship of the rapidly emerging SARS-CoV2 virus withthe cardiovascular system is under research, and it is still
being investigated whether the virus will directly proliferate
in the heart (58-59), in contrast, the virus is connected to anumber of proinflammatory mediators that may have a rolein the pathogenesis of cardiac and arrhythmic problems.
Myocardial involvement could be caused by direct viral
infection, hypoxia-induced apoptosis, or cytokine stormrelated cell damage in the body (46). With the knowledge
that is currently available, it is also possible to assume that
persons with underlying cardiovascular comorbidities are
more sensitive to adverse cardiac events after COVID-19
infection (58). According to a study based on single cell RNA
sequencing, more than 7.5 percent of cardiomyocytes
express ACE2, which could enhance SARS-CoV2 invasion
and cause direct cardiotoxicity (60).
ACE2 is a critical regulator of cardiac function in vivo,
according to genetic evidence (42), moreover, in humans,
circulating ACE2, which is released by endothelial cells, is a
Fiziologia - Physiology • 2022.1(103)biomarker of hypertension, heart failure, and diabetes,
showing enhanced ACE2 activity (59). The activation of theRenin Angiotensin System and the inhibition of ACE2
expression are significant in a number of cardiovasculardiseases (56). Severity of COVID-19's health decline has
been linked to underlying cardiovascular risk factors anddisease, which are thought to be highly correlated with age.
COVID-19 is thought to have more catastrophic
consequences in older individuals than in younger
individuals, with hypertension and diabetes presenting as
the two most significant co-morbidities (38). Acute Coronary
Syndrome Patients with a suspicion or evidence of acute
myocardial ischemia or infarction are referred to as having
acute coronary syndrome (ACS). The three conventionalkinds of ACS are non-ST-elevation myocardial infarction(NSTEMI), ST-elevation myocardial infarction (STEMI), andunstable angina. In reference to the Fourth Universal
Definition of AMI, it can be described as the presence ofacute myocardial injury discovered with aberrant cardiac
biomarkers in clinical or laboratory evidence of myocardialischemia in the context of AMI, whether STEMI or NSTEMI(61). Serious viral infections can trigger a systemic
inflammatory response syndrome, which raises the risk ofplaque rupture and thrombus development, leading to an
ST-elevation MI or MI with no ST elevation (62).
In a restricted study of 75 patients hospitalized with
SARS, a predecessor of the viral SARS-CoV2, acute
myocardial infarction (AMI) caused deaths in 2 of 5 fatal
cases (63). Although there is still minute scientific evidence
linking COVID-19 to the development of ACS, multiple
investigations have found a temporary but significant link
between lower respiratory tract infections and acutecoronary syndromes, implying serious clinical implications
again for SARS-CoV2 virus (46, 64). High expression of
ACE2 in pericytes might lead to an onset of microvascular
dysfunction, (65) explaining greater propensity for acutecoronary syndromes (46). There have been reports ofcoronary artery involvement and ischemia, including STelevation and non-ST-elevation myocardial infarction(STEMI & NSTEMI), with numerous theories explaining
coronary artery involvement. Direct viral infection can cause
plaque instability and type 1 MI because ACE2 is expressedin vascular endothelial cells. The significant systemic
inflammatory response in the third phase of the disease canalso cause plaque instability and rupture. COVID-19 canalso cause type II MI by causing demand ischemia as aperson becomes ill. Hypoxemia coupled with increased
cardiac demand owing to systemic infection causes Type-II
AMI, which is explained by a myocardial oxygen demandthat is disproportionate to the supply (61).
CARDIAC ARRHYTHMIC RISK
Cardiac arrhythmias identified as irregular rhythms ofheart such as tachyarrhythmia and bradyarrhythmia areother cardiovascular complications that occur frequently inpatients with COVID-19 (66). Arrhythmias in COVID-19
patients can take several forms, including atrial-ventricular
Fiziologia - Physiology • 2022.1(103)

block, atrial fibrillation, polymorphic ventricular tachycardia,
and complete heart block presenting no escape rhythm (67).
Early indications of Arrhythmia might be the firstpresentation of COVID-19, and new-onset and/or
progressive arrhythmia could indicate cardiac involvement.
According to a study presenting 137 infected subjects from
Wuhan, 7.3% of the subjects had exhibited palpitations as
one of their early symptoms for COVID-19 (68). Arrhythmias,such as atrial fibrillation, are also more common in COVID19 cardiomyopathy, as inflammation is a substrate for atrial
arrhythmias (69). Cardiac arrhythmias may be attributed todirect myocardial injury or secondary to systemic alterations
due to acute viremia (e.g., metabolic derangement, hypoxia,
neurohormonal changes, and inflammatory stress).
Hypercytokinemia and arrhythmogenic potential are seen in
COVID-19 subjects who are in hyperinflammatory condition.
Cytokine storm and rising levels of Interleukin-1, Interleukin2, Interleukin-6, monocyte chemoattractant protein, andtumor necrosis factor-alpha (TNF-) are also linked to deadly
arrhythmias. Atrioventricular block and ST segment
elevation have been described as electrocardiogram
findings of cardiac arrhythmias. Malignant arrhythmias, suchas multifocal ventricular tachycardia/ventricular fibrillation,
were observed to develop during COVID-19 and were also
linked to increased Troponin T levels (70).
MYOCARDITIS
Although case reports of myocarditis, explained as
inflammation of the myocardium exist in COVID-19 infectedindividuals, it is unknown whether myocarditis is caused
directly by the virus or as a result of an inflammatory reaction.
Several examples of severe myocarditis with impaired
systolic function have been documented following SARSCoV2 infection (71). Xu et al. discovered sparse levels ofmononuclear inflammatory infiltrates in the interstitial spaceof a COVID-19 patient's myocardium, implying that the newcoronavirus may promote myocarditis (72). SARS-CoV2
may share the same pathophysiological mechanism, withdirect cardiac muscle injury contributing to myocarditis in
COVID-19 infected people. Increased cardiac biomarkers ina significant proportion of individuals have revealed cardiac
muscle involvement. An increase in cardiac markers may
not be entirely attributed to myocarditis, elevated levels of
troponin and brain natriuretic peptide however, arecorrelated with worse outcomes (73). Acute cardiac injury
caused by SARS-CoV2 infection can sometimes lead tofulminant myocarditis, a remarkable clinical crisis withhemodynamic compromise and significant fatality rates
ranging from 40 - 70%. Fulminant myocarditis isdistinguished by rapid and diffuse cardiac inflammation,
necrosis, and eventual ventricular dysfunction, which resultsin cardiogenic shock, pathological arrhythmias, multi - organ
failure, and death (74).
THROMBOEMBOLIC EVENTSHypercoagulability has previously been linked to anincreased risk of blood clot formation in individuals infected13with SARS and MERS virus respectively (75). COVID19 has
been linked to a hypercoagulable state as well. Endothelial
cell dysfunction caused by infection results in excessive
thrombin production and fibrinolysis shutdown. The hypoxia
associated with significant COVID-19 can potentiallypromote thrombosis by increasing blood viscosity. Increased
blood coagulability is also aided by a hypoxiainducible
transcription factor-dependent signaling cascade (76).
Individuals who did not survive COVID-19 had considerably
greater levels of D-dimer and other fibrin degradation
products (FDP), as well as longer prothrombin and activatedpartial thromboplastin durations, according to primarystudies on COVID-19 (77). Individuals with COVID19 havebeen reported to suffer coagulopathy, microvascular, andmacrovascular thrombosis (78). Numerous case reports ofthrombosis involving a diverse array of vascular beds havenow been published. Lower extremity arterial and venous
vessels, abdominal veins, bypass grafts, coronary arteries,
cerebral veins, pulmonary arteries, large vessel cerebral
and carotid arteries, and small distal arteries with vasoocclusive inflammatory clots have all been explained ashaving thrombus formation causing a chilblains-likepresentation (79).
HEART FAILURE
Heart failure is a leading cause of death in people
plagued by COVID-19. It is caused by a variety of
myocardial aggression mechanisms, including direct viral
myocardial injury, indirect and direct inflammatory damage,O2 supply–demand imbalance, and an increase inatherothrombotic events due to inflammatory destabilizationof atheromatous plaques, all of which gives an end result asacute myocardial dysfunction (70-71). Heart failure inindividuals with COVID-19 can range from mild heart failure
with conserved ejection fraction in the earliest stages of theinfection to severe end-stage heart failure and cardiogenic
shock with substantial death rates (80).
CONCLUSION
Although the clinical manifestations of zoonotic human
COVID-19 are well understood to be basically respiratory,
notable accounts of significant cardiac complications
ranging from myocyte biomarker elevation, arrhythmias, andmyocardial
dysfunction
toelectrocardiographic
abnormalities manifestations are being widely published.
Precise understanding of the effects of COVID-19 on theheart as well as the cardiovascular system in whole is yet tobe well-established, but existing evidences from preceding
human zoonotic viral epidemics such as SARS-CoV and
MERS-CoV suggest that viral infections in their category
could act as a trigger for cardiac abnormalities such as acutecoronary syndromes, arrhythmias and heart failure via direct
or indirect mechanisms. Further studies on thesehypothesized mechanisms are strongly required. Further
clinical researches are then required to learn more about the
impact of COVID-19 on the cardiovascular system.
Conflicts Of Interest

14

The authors declare that they have no conflicts of interest in
regards to this publication.

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Fiziologia - Physiology • 2022.1(103)FIZIOLOGIA MODIFICĂRILOR CARDIOVASCULARE ASOCIATE CU INFECȚIA CU COVID-19: O
REVIZUIREREZUMATDeși manifestările clinice ale COVID-19 uman zoonotic sunt bine înțelese ca fiind respiratorii, sunt publicate pe scară largă relatări notabile
despre complicații cardiace semnificative, de la creșterea biomarkerului miocitelor, aritmii și disfuncție miocardică până la manifestări aleanomaliilor electrocardiografice. Câteva studii de patologie efectuate până acum pe pacienții care au murit din cauza COVID-19 au arătat
că până la 35% dintre aceștia găzduiau noul virus SARS-CoV2 în stratul mijlociu al inimii, miocardul. În prezent, o înțelegere precisă aefectelor COVID-19 asupra inimii, precum și asupra sistemului cardiovascular în ansamblu nu a fost încă bine stabilită, dar există dovezi din
epidemiile virale zoonotice umane precedente, cum ar fi SARS-CoV și MERS. -CoV sugerează că infecțiile virale din categoria lor ar putea
declanșa anomalii cardiace, cum ar fi sindroame coronariene acute, aritmii și insuficiență cardiacă prin mecanisme directe sau indirecte.
Această revizuire își propune să rezumă constatările privind influența COVID-19 zoonotic asupra funcțiilor cardiovasculare umane până înprezent.
Cuvinte cheie: Coronavirus, cardiovasculare, receptori ACE2, furtuna de citokine, proteină Spike, COVID-19

Fiziologia - Physiology • 2022.1(103)17THROMBOTIC EVENTS AND ANTICOAGULATIONTARGETS IN COVID-19 INFECTION
IACOB DANIEL GOJE1,2, GRETA IONELA BARBULESCU3,4,*, VALENTINA GABRIELACIOBOTARU1,2,5, OANA ELENA ANCUSA1,2,5, IVANKA MARIAN TODOR5, DANIELFLORIN LIGHEZAN1,2,5
1Department of Medical Semiology I, “Victor Babes” University of Medicine and Pharmacy, No. 2 Eftimie Murgu Square, 300041
Timisoara, Romania
2Advanced Cardiology and Hemostaseology Research Center, “Victor Babes” University of Medicine and Pharmacy, No. 2
Eftimie Murgu Square, 300041 Timisoara, Romania
3Department of Clinical Practical Skills, “Victor Babes” University of Medicine and Pharmacy, No. 2 Eftimie Murgu Square,
300041 Timisoara, Romania
4Immuno-Physiology and Biotechnologies Center (CIFBIOTEH), Department of Functional Sciences, “Victor Babes” Universityof Medicine and Pharmacy, No. 2 Eftimie Murgu Square, 300041 Timisoara, Romania
5Department of Internal Medicine, Emergency Clinical Municipal Hospital, No. 5 Gheorghe Dima Street, Timisoara, Romania

ABSTRACT
Coagulopathy, vascular damage, and thromboembolic events contribute to the severity of COVID-19 disease. The lack of randomized
anticoagulant trials in patients with COVID-19 has made the initial approach rather empirical based on background from other similar diseases.
Although some observational studies show that prophylactic and therapeutic anticoagulation can decrease mortality and the number ofcritical cases, others fail to support this finding, suggesting an increased risk of bleeding. Studies show that the vascular endothelium plays
a vital role in the procoagulant status in patients with thrombosis infected with the SARS-CoV-2 virus. COVID-19 associated coagulopathy
is a unique condition slightly different from other coagulopathies found in various diseases because of its particularities. There is still
significant interest in the mechanism of thrombosis and anticoagulation therapy in patients with COVID-19. Clinicians from experienced
medical centers test escalating doses of anticoagulants to optimize current standards of care and understand the risk-benefit of bleeding.
Key words: coagulopathy, deep vein thrombosis, pulmonary embolism, anticoagulant therapy, endothelial dysfunction

INTRODUCTION

Coagulopathy, thromboembolic events, andendothelial damage are key elements observed in patientswith COVID-19 and contribute decisively to the severity
and progression of the disease. Venous thrombotic events,
such as deep vein thrombosis or pulmonary embolism,
and arterial thromboses, such as myocardial infarction or
stroke, are more common in patients hospitalized with
COVID-19, with some meta-analyses showing an
incidence of up to 23% in the case of venous
thromboembolism in patients admitted to intensive careunits (ICU) [1]. These elements are added observations
detected in microscopic studies that have shown thepresence of microvascular thrombosis in the lungs andother organs, which could explain the diffuse evolution oflung lesions and disease aggression [2].
COVID-19 associated coagulopathy is anprognostic and therapeutic implications. A particular
aspect of this pathology is represented by different
dynamics of the value of D-dimers compared to othersimilar pathologies. Some authors suggest that theirsystematic evaluation has prognostic and therapeutic
implications [3]. All these observations concluded that
antithrombotic therapy is essential in managing patientswith COVID-19. However, the lack of randomized studies
made the initial approach rather empirical at the beginningof the pandemic, based on experience from other similar
pathologies.
The antithrombotic and the non-anticoagulant
properties of unfractionated heparin / low molecular weight
heparins (LMWH) led to the hypothesis that they could
influence the evolution of the disease. Non-anticoagulant
mechanisms such as heparanase inhibition, competitive

Received 13th of March, 2022. Accepted 14th of June 2022. Address for correspondence: Greta Ionela Barbulescu, Department of Clinical
Practical Skills, “Victor Babes” University of Medicine and Pharmacy, No. 2 Eftimie Murgu Square, 300041 Timisoara, Romania; phone:+40733177583, e-mail: barbulescu.greta@umft.roessential element that characterizes this entity and has
18binding and inhibition of pro-inflammatory factor synthesis,
Fiziologia - Physiology • 2022.1(103)inhibition of histones and neutrophil traps [4], and last but
not least competitive inhibition of SARS-CoV-2 virus-cell
invasion [5], together with the established anticoagulant
mechanism, gives heparins an essential role in the
management of patients with moderate or severe disease.
Recommendations for thromboprophylaxis andthe use of anticoagulant therapy in hospitalized patientswith COVID-19 are mainly based on data from similar
pathologies. Some observational studies show that boththerapeutic and prophylactic anticoagulation decreases
mortality and the number of cases requiring intubation (6).
In contrast, others fail to support the same evidence,suggesting an increased risk of bleeding and thattherapeutic anticoagulation does not affect survival. [7].
This uncertainty has created confusion regarding the
optimal anticoagulant treatment, with significant variability
in the intensity of anticoagulation observed in most clinicaldecisions,
hospital
protocols,
andexpertrecommendations [8,9].
THE ROLE OF THE VASCULAR ENDOTHELIUM
The level of understanding regarding themechanisms involved in COVID-19 coagulopathy isconstantly evolving; current evidence demonstrates acascading relationship between endothelial dysfunction,
inflammatory / immune activation, and coagulation [10].
The SARS-CoV-2 virus enters cells by interacting with the
spike protein and the angiotensin two conversion enzyme
receptor (ACE-2), a receptor identified in the lung, heart,brain, and endothelial tissue [11]. The result of thisinteraction will be an increase in the level of angiotensin 2,
leading to a procoagulant status explained by theaccumulation of bradykinin and many other mechanismsof the renin-angiotensin-aldosterone system (RAAS)
involved in thrombosis [12].
Endothelial injury triggered by viral invasion
explains diffuse endotheliitis observed in the lungs, heart,kidneys, and intestines, a phenomenon that will underlie
immune activation and cytokine storm, thus promoting
hypercoagulability and thrombosis [13]. Physiologically,
the vascular endothelium possesses a whole series offunctions, such as controlling hemostasis, fibrinolysis,
inflammation, vascular permeability and oxidative stress,
mechanisms severely disturbed in patients with SARSCoV-2 infection.
The cytokine storm observed in patients withsevere disease is thus explained by the transformation ofphysiological mechanisms into mechanisms generating
endothelial injury and thrombosis. Excessive production ofcytokines such as IL-1, IL-6, and TNF-α, together with thealteration of counter-regulation mechanisms, underlie thisphenomenon. This process will not be limited only to the
local endothelial level. The expression will be systemic,
generating an imbalance between thrombotic andFiziologia - Physiology • 2022.1(103)antithrombotic mechanisms that favor the accumulation ofthrombotic material [14].
Another important mechanism is the action of
IL-6 on the hepatocyte, which will promote theexaggerated synthesis of fibrinogen, PAI-1, a fibrinolysis
inhibitor, and C-reactive protein, thus resulting in alteration
fibrinolysis that can be directly correlated withthromboembolic events [15]. Activation of the coagulationcascade by tissue factor, excessive release of vonWillebrand factor (vWf) that will promote platelet
aggregation, and altered endothelial fibrinolytic
mechanisms observed in patients with a severe diseasealso explain the formation of thrombotic material.
Therefore, we observe that the vascular endothelium has
an essential role in explaining the pathophysiologicalmechanisms involved in COVID-19. Moreover, theelements of endothelial dysfunction begin to be the target
of innovative therapeutic approaches [14]. Also, the link
between coagulation and inflammation, translated byparaclinical elements such as D-dimers, fibrinogen, proinflammatory cytokines, will be directly correlated with thedisease, managing to bring a much clearer picture in terms
of prognosis, evolution, and outcome of therapeuticintervention.
COVID-19 ASSOCIATED COAGULOPATHY
Initial reports from patients with COVID-19 andrespiratory failure revealed the presence of disseminatedintravascular coagulation (DIC) criteria (according to ISTH
criteria). The incidence is much higher in those who diedcompared to survivors (71% vs. 0.6%) [16]. Other studies
have found that DIC incidence is lower in the early stagesof the disease, although there are stigmas of coagulation
damage, which has led to the conclusion that a distinct
form of coagulopathy turns into DIC once the severity ofthe disease and septic complications occur [17]. COVID19 associated coagulopathy (CAC) is a distinct, intensely
studied entity, different from coagulopathy in otherrespiratory viral illnesses, sepsis-induced coagulopathy(SIC), or observed in DIC. In CAC, we observe higher
values of D-dimers and fibrinogen, but with minimalchanges in prothrombin time and platelet count [18]. Also,
consumptive coagulopathy, the primary element present in
CID, is rarely observed in the early stages of COVID-19.
There is no official definition of CAC, but it would
be imperatively needed for research and therapy guidance.
Some authors propose a definition that includes
confirmation of SARS-CoV-2 virus infection and two of thefollowing four elements:• A platelet count below 150 × 109 / L• A marked increase in the value of D-dimers• Prolongation of prothrombin time or INR• Documentation of micro or macro thrombosis

19

If patients meet only one of the four criteria mentioned
above and will have one or more of the following: (i)increased fibrinogen, (ii) increased vWf, (iii) the presenceof lupus anticoagulant or antiphospholipid antibodies, they
will be at high risk of CAC. The definition presented is not
an official or unanimously accepted one, but it manages toshed some light on the understanding of this complex
entity. Using this definition, only 10% of patients met the
CAC criteria at hospitalization, a percentage that
increases sharply at the time of transfer to intensive careto over 60% [19].
CAC presents similarities and clinical-biologicalcharacteristics common with CID, SIC, hemophagocytic
syndrome, antiphospholipid syndrome, or thrombotic
microangiopathy, but with specific unique characteristics[17], incompletely defined and elucidated. These elements
could underlie a better understanding of the disease andguide the therapeutic approach.
COVID-19 ANTICOAGULATION
Thromboprophylaxis of patients with COVID-19
has been marked by enthusiasm since the onset of thepandemic, which has been based on clinical observations,pathophysiologicalinvestigations,andinitialepidemiological data [8,9]. The primary concern was thatthromboembolic events had an increased incidence
despite applying the prophylaxis standards valid until then[20]. These elements have led some experts torecommend the empirical use of high-intensity
thromboprophylaxis [21]. The use of intensive
anticoagulation involves an increased risk of bleeding,requiring a closer assessment of the alleged benefit, withsome guidelines not recommending the empirical use of
escalating doses of anticoagulants [8,9].
The significant interest in this topic has led to theglobal launch of a series of multicenter randomized trials
to answer how combining antithrombotic agents or using
different doses could optimize current standards of careand understand the risk-benefit elements in terms of
bleeding [22]. Most of these studies included
anticoagulants with intermediate or therapeutic doses
compared with the usual prophylactic doses, antiplatelet,
fibrinolytic agents, and multiple other innovative
approaches [23].
The RECOVERY study shows that the additionof aspirin has no benefit in reducing mortality. This
statement started to clear up aspirin use in patients with
COVID-19 [24]. When it comes to anticoagulant therapy,
things are not as clear. The optimal doses of anticoagulant
agents in thromboprophylaxis of patients with moderateand severe forms of the disease were investigated inseveral randomized trials, their results being able to bringa certain degree of clarity.
Three randomized trials (REMAP-CAP, ACTIV20

4a, ATTACC) comparing the prophylactic dose of
anticoagulant (predominantly low molecular weight
heparin) versus the therapeutic dose in hospitalized
patients showed that the results differed according to theseverity of the disease [25,26]. Thus, in non-criticalpatients, defined as patients who do not require ventilatoryor cardiovascular support, the therapeutic dose increases
the probability of survival and reduces the use of cardiorespiratory organ support compared to usual
thromboprophylaxis [25]. However, a similar benefit was
not observed in critical patients admitted to ICU; the initialstrategy of using curative doses of anticoagulant did notincrease the probability of survival or the number of days
without organ support compared to usual pharmacologicalthromboprophylaxis [26].
Similar data were observed in the HEP-COVID
study, finding that the primary efficacy endpoints,
represented by venous/arterial thromboembolic eventsand deaths of any cause, were significantly reduced using
therapeutic doses in non-critical patients. However, thesame benefit was not observed in ICU patients [27].
Data from the INSPIRATION trial, which
compared intermediate-doses (enoxaparin, 1 mg/kg/day)versus the standard prophylactic dose (enoxaparin, 40
mg/day) in ICU patients, failed to show a clear benefit ofintermediate-dose use; there is even a trend of increasing
the risk of bleeding, which has not reached statisticalsignificance [28].
Direct oral anticoagulants (DOAC) have also
been studied in patients hospitalized for COVID-19. The
ACTION study compared the use of therapeutic doses ofrivaroxaban with standard thromboprophylaxis and failed
to demonstrate a clinical benefit. It pointed out that using
therapeutic doses of rivaroxaban significantly increased
bleeding events, with the authors concluding that DOAC
should be avoided in this category of patients unless an
explicit indication is present [29].
The results of these trials are promising, but
greater public availability and more rigorous validation
would be needed. The logical question is whether
guidelines and protocols should be changed to include
therapeutic anticoagulation in selected categories ofpatients [30].
At the time of writing, major guidelines forthromboprophylaxis recommend the use of prophylactic
doses of anticoagulant in patients hospitalized with
COVID-19 who do not have confirmed thromboembolic
events. The use of high-intensity anticoagulation is notsupported by currently available randomized studies, atleast for patients with severe forms of illness admitted to
ICU departments [9,31,32].
The use of therapeutic anticoagulation inpatients with moderate forms of the disease remains
controversial, with some guidelines recommending thisFiziologia - Physiology • 2022.1(103)approach based on the results of recent studies [33], whileothers await more rigorous validation by the international
scientific community [31].
CONCLUSION
The complexity of the balance between anticoagulation,
thrombosis, and hemorrhagic complications in patientswith COVID-19 makes the therapeutic decision very
difficult. The unique international collaboration in large
platform trials has managed to bring the first quality datato help clinicians optimize anticoagulant treatment forpatients with COVID-19. A whole series of studies will be
published in the coming period, which will most likely
respond to current uncertainties and help clinicians be
more efficient in managing patients with COVID-19.

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EVENIMENTELE TROMBOTICE ȘI ȚINTELE DE ANTICOAGULARE ÎN INFECȚIA COVID-19 (NTM)
REZUMAT
Coagulopatia, afectarea vasculară endotelială și evenimentele tromboembolice contribuie la severitatea bolii COVID-19. Lipsa studiilor
randomizate cu anticoagulante la pacienții cu COVID-19 a făcut ca abordarea inițială să fie destul de empirică, bazată pe studiul altor boli
similare. Deși unele studii observaționale arată că anticoagularea profilactică și terapeutică poate scădea mortalitatea și numărul de cazuri
critice, altele nu susțin această idee, sugerând un risc crescut de sângerare. Studiile arată că endoteliul vascular joacă un rol vital în statusul
procoagulant al pacienților cu tromboză, infectați cu virusul SARS-CoV-2. Coagulopatia asociată COVID-19 (CAC) este o afecțiune unică,ușor diferită de alte coagulopatii întâlnite în diferite boli, datorită particularităților sale. Există încă un interes semnificativ în studiul
mecanismului trombozei și terapiei anticoagulante la pacienții cu COVID-19. Clinicieni din centre medicale cu experiență testează diferite
doze de anticoagulante pentru a optimiza standardele actuale de îngrijire și pentru a înțelege relația risc-beneficiu în ceea ce privește
sângerarea.
Cuvinte cheie: coagulopatie, tromboză venoasă profundă, embolie pulmonară, terapie anticoagulantă, disfuncție endotelială

22

Fiziologia - Physiology • 2022.1(103)THE IMPACT OF LMWH TREATMENT ON IUGRINCIDENCE IN PREGNANCIES WITH INHERITED
THROMBOPHILIADIROESCU CIPRIAN2, ANGHEL-SAVCIU NICOLETA-LOREDANA1, PANA RAZVANCOSMIN1, CERNEA NICOLAE 1
1
Clinical Emergency County Hospital Craiova, Obstetrics and Gynecology Department Tabaci 1, Craiova 200642
2
Municipal Hospital Turnu Magurele, Obstetrics and Gynecology Department, Castanilor, Turnu Măgurele 145200

ABSTRACT

Thrombophilia is defined as a predisposition to the development of thrombosis being associated with an increased tendency
to develop venous thromboembolism. Numerous studies have shown an important association between hereditary
thrombophilia and intrauterine growth restriction or preeclampsia. There is still controversy regarding the treatment with low
molecular weight heparin of pregnant women with hereditary thrombophilia. The main points of contention are the selection of
the type of hereditary thrombophilia that requires anticoagulant treatment, the dose and frequency of administration, and thebenefits of LMWH anticoagulant treatment in these patients. Our aim was to evaluate the effects of LMWH treatment on oneof the most common fetal complications associated with thrombophilia pregnancies, namely intrauterine growth restriction.
Key words : LMWH, thrombophilia, intrauterine growth restriction

INTRODUCTION
Inherited thrombophilias are a group of genetic
disorders that interfere with the coagulation cascade
increasing the risk for venous thrombosis andthromboembolism. When pregnancy occurs in a woman
with hereditary thrombophilia the risk of thrombosis ispotentiated by the physiological state of hypercoagulability
existing in pregnancy. The most common inherited
thrombophilias are [1]:Factor V Leiden thrombophilia is the most common
inherited form of thrombophilia with a prevalence in theEuropean general population of 3-8% for one copy of thefactor V Leiden mutation;-Moderate protein S deficiency is estimated to affect 1:500
individuals. Severe deficiency is rare and its prevalence isunknown.
-Moderate protein C deficiency affects about 1:500
individuals.
- Prothrombin-related thrombophilia is the second mostcommon genetic form of thrombophilia, occurring in about
1.7-3% of the European general-Hereditary antithrombin III deficiency has a prevalence of
1:500-5000
inthegeneralpopulation.
-Plasminogen activator inhibitor 1 (PAI-1) polymorphisms

are associated with thrombotic disease. Prior studies haveshown a possible link with adverse pregnancy outcomes[2].
Many studies have shown different degrees ofassociation between hereditary thrombophilia and various
fetal and maternal complications during pregnancy [3] (eg.
recurrent pregnancy loss, foetal growth restriction, latemiscarriages, stillbirth and preeclampsia). However, there
is no consensus on therapeutic standards in addressing
hereditary thrombophilia in pregnancy. Also routine
screening for thrombophilic mutations is not considered
cost-efficient, as their prevalence is relatively low. [4]It has been long assumed that insufficient uterine,
placental, and fetal circulations result in adverse
pregnancy outcomes and that those abnormalities can be
defined by the use of Doppler ultrasonography. [5,6[.
Uteroplacental blood flow decreases in pregnancies thatare complicated by hypertension and intrauterine growth
restrictions (IUGRs). This decrease is associated with apathologic condition of spiral arteries, thought to arise
during placentation in the first trimester of pregnancy; thus,
it might be possible to predict the development of these
conditions by assessing uteroplacental blood flow early in
pregnancy with color Doppler [7,8]. The result is abnormal

Received 11th of March 2022. Accepted 19th of June 2022. Address for correspondence: Anghel Savciu Nicoleta Loredana,. Clinical Emergency
County Hospital Craiova, Obstetrics and Gynecology Department Tabaci 1, Craiova 200642
telephone: 004 0723181624 e-mail: lore.anghel@yahoo.commailto:chioreandianamaria@yahoo.com

Fiziologia - Physiology • 2022.1(103)23uteroplacental blood flow, and this has led to the idea ofusing Doppler assessment of uterine and umbilicalarteries velocity waveforms as a method of screening forthese antenatal complications. An abnormal test result isrepresented by an abnormal flow velocity ratio(systolic/diastolic (S/D) ratio), resistance index, pulsatility
index or the presence of an early diastolic notch [9].
Intrauterine growth restriction is one of the most
redoubtable complications in pregnancy and is the mostcommon indication for induction of labour before term.
Intrauterine growth restriction (IUGR) has been defined as
the rate of fetal growth that is below normal in light of thegrowth potential of a specific infant as per the race andgender of the fetus [10]. It has also been described as adeviation from or a reduction in an expected fetal growth
pattern and is usually the result of innate reduced growthpotential or because of multiple adverse effects on thefetus. The “normal” neonate is the one whose birth weight
is between the 10th and 90th percentile as per thegestational age, gender and race with no feature ofmalnutrition and growth retardation [11,12]. Possible
complications of foetuses with intrauterine growth
restriction include: perinatal asphyxia, hypothermia,
hypoglycemia, and polycythemia.
The choice of patients who would benefit mostfrom anticoagulation during pregnancy is made bycomparing the risk of bleeding resulting from theadministration of anticoagulant with the risk ofthromboembolism in the absence of anticoagulation.
Despite this and the lack of controlled trials, there has
been increased use of anticoagulants to prevent VTE(venous thromboembolism) and adverse pregnancy
outcomes. Although the effects of anticoagulants on thefetus and the mother may be considerable and there areno controlled studies with proven certainty of safe
administration, the use of low molecular weight heparine(LMWH) in pregnancy has been steadily increasing [13].
MATERIAL AND METHODS
We conducted a retrospective study at the Craiova
County Emergency Clinical Hospital in which we included
60 patients diagnosed with thrombophilia (various forms)
and who also gave birth in the hospital maternity ward.
Half of them opted for anticoagulant treatment of LMWH
( single dose per day s.c. ) and half refused
anticoagulation after being presented with the potential
risks and benefits of LMWH. The exclusion criterion was
the presence of a personal history of venous thrombosis
All the patients included in the study had been tested forhereditary thrombophilia using the same laboratory. In
addition to monitoring the incidence of intrauterine growth
restriction, we also monitored flows on the uterine arteries,
umbilical arteries and middle cerebral artery.
RESULTS
1. No anticoagulation
The incidence of intrauterine growth restriction
was recorded in approximately 50% of the patients without

24

anticoagulation. The mean age of patients in the nonanticoagulant group was 27.5 years. The weight of fetuseswith IUGR ranged from 1400 to 2600 grams and they were
all born prematurely. The most common variety ofthrombophilia in the IUGR group without anticoagulation
was homozygous PAI mutation. Over 80% of patients in
the group with IUGR and without anticoagulation had anotch on the uterine artery in the first trimester. Thischange persisted in almost half of these patients in thesecond trimester. In patients without anticoagulation andwith eutrophic fetuses, the most common variety ofthrombophilia was the MTHFR C677T gene mutation.
Uterine notch in this group in the first trimester was present
in approximately 70% of patients but the incidence
decreased to below 30% in the second trimester.
2. Anticoagulation
The mean age in the anticoagulant group was 29.4
years. The incidence of IUGR in this group was
approximately 25%. The weights of the fetuses with IUGR
in this group ranged from 1450 to 2770 grams. Most
varieties of thrombophilia in the group with IUGR andanticoagulation were represented by multiple concomitantmutations of MTHFRC677T, MTHFRA1298C and PAI
genes. Almost all patients in the IUGR and anticoagulationgroup had a notch on the uterine artery in the first trimesterand persisted at 50% in the second trimester. Similar to
the IUGR and no anticoagulation group, concomitantmutations in the MTHFR and PAI genes were present in
the IUGR and anticoagulation group. The frequency ofnotch on the uterine artery in the first trimester and its
persistence in the second trimester was relatively similar
to the IUGR and anticoagulation subgroup. If we exclude
the anticoagulation criterion IUGR is present inapproximately 25% of the studied patientsDISCUSSIONS
Although it is a small study group, thedifferences between anticoagulated and nonanticoagulated patients are significant in terms of IUGR
incidence. The literature offers percentages with a very
wide margin ranging from 3% to 35% in terms of IUGR
incidence in patients with thrombophilia. The evidence is
conflicting with respect to the presence and strength of theassociations between inherited thrombophilia andpregnancy complications other than VTE. Even if aheritable thrombophilic defect is found in a woman withrecurrent miscarriages or late pregnancy complications,no evidence exists to suggest that LMWH or otherinterventions, such as antiplatelet treatment, will provide
benefit, apart from some studies showing a beneficial
effect of low-dose aspirin to prevent preeclampsia in
women with a history of preeclampsia, regardless of thepresence of thrombophilia [13]If we restrict our search to higher quality
studies, there is currently insufficient evidence to support
LMWH use in patients at risk for placenta-mediatedpregnancy complications. Alternatively, in the
Fiziologia - Physiology • 2022.1(103)subpopulation of women with prior history of the more
severe placenta-mediated complications (early onset
preterm preeclampsia, major preeclampsia, small for
gestational age (SGA) ≤5th percentile or pregnancy
loss >20 weeks), there remains a possible benefit for
LMWH to prevent recurrent complications, however this isbased on lower quality evidence (single centre,
nonregistered trials) and further high quality research is
required to explore this higher risk subpopulation of
women.[14]Polymorphisms of the methylene tetrahydrofolate
reductase gene (C677T and A1298C) is known to be
implicated in the apparition of preeclampsia [15,16]. The
role of mutation in the MTHFR gene, as risk factors for
intrauterine growth restriction during pregnancy, is not very
well known so far; there are some studies but their results
are conflicting and ambigous. In our study group IUGR
was found mostly in case of association of multiple
mutations including the two most frequent MTHFR
mutations, C677T and A1298C.
Patients with plasminogen activator inhibitor 1 (PAI1) mutation were found in several studies to be at higher
risk of developing IUGR [17,18]. In our study PAI mutation
was present in the majority of cases which developed
IUGR and were not helped by the administration of LMWH.
Increased impedance to blood flow in the uterine
arteries in pregnancy , as determined by Doppler
ultrasound parameters, has been used as a screening
method for preeclampsia and the delivery of a SGA
neonate, but this screening method is associated with low
positive predictive values indicating that most patients witha positive test will not develop the disease [19]. Our study
showed the presence and persistence of uterine arterynotch in over 2 thirds of the patients which developed
IUGR, regardless of LMWH administration.
CONCLUSIONS
Our study shows a 25% incidence of IUGR
regardless of anticoagulation and a double percentage ofpregnancies with intrauterine growth restriction in patients
without LMWH compared to those who received LMWH. It
should be noted that the restriction was encounteredregardless of anticoagulation in patients with concomitantmutations on several genes (MTHFR, PAI) and that in thecase of patients without anticoagulation the most common
mutation is homozygosity of the PAI gene.
In view of the above, we believe that additional
stratified studies on subvariant mutations and mutation
combinations are needed to accurately determine which
patients benefit from LMWH administration.

2.

3.

4.
5.

6.

7.

8.

9.

10.

11.

12.

13.

14.

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preeclampsia.Chedraui P, Andrade ME, SalazarPousada D, Escobar GS, Hidalgo L, Ramirez C,
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10.7863/jum.2010.29.7.1103.

IMPACTUL TRATAMENTULUI CU HGMM ASUPRA INCIDENTEI RCIU IN SARCINILE CU
TROMBOFILIE EREDITARAREZUMAT

Trombofilia este definită ca o predispoziție la dezvoltarea de tromboză fiind asociată cu o tendință crescută de apariție atromboembolismului venos. Numeroase studii au arătat o asociere importantă între trombofilia ereditară și restricția de creștere
intrauterină sau preeclampsie. Există în continuare controverse în ceea ce privește tratamentul gravidelor cu trombofilie
ereditară cu heparine cu greutate moleculară mică. Principalele puncte de discuții contradictorii sunt reprezentate de selecția
tipului de trombofilie ereditară care necesită tratament anticoagulant, doza și frecvența administrării precum si beneficiile
tratamentului anticoagulant cu HGMM la aceste paciente. Obiectivul nostru a fost să evaluăm efectele tratamentului cu HGMM
asupra uneia dintre cele mai frecvente complicații fetale asociate sarcinilor cu trombofilie și anume restricția de creștere
intrauterină.
Cuvinte cheie: HGMM, trombofilie, restricție de creștere intrauterină

26

Fiziologia - Physiology • 2022.1(103)VASCULAR CHANGES IN THROMBOPHILIAPREGNANCIES. DOES ANTICOAGULATION MAKE A
DIFFERENCE?

ANGHEL-SAVCIU NICOLETA-LOREDANA1, PANA RAZVAN COSMIN1, CERNEANICOLAE1, DIROESCU CIPRIAN2
1
Clinical Emergency County Hospital Craiova, Obstetrics and Gynecology Department Tabaci 1, Craiova 200642
2
Municipal Hospital Turnu Magurele, Obstetrics and Gynecology Department, Castanilor, Turnu Măgurele 145200

ABSTRACT

Vascular and coagulation cascade changes that occur physiologically during pregnancy are well known. In women withhereditary thrombophilia associated with pregnancy, these changes are exacerbated and produce undesirable effects on
pregnancy. Although anticoagulant treatment during pregnancy seems to be the logical solution in these cases, however, manystudies show net benefits of using anticoagulants only in patients with previous adverse pregnancy outcomes (pregnancy loss,
preeclampsia, eclampsia, intrauterine growth restriction). In this study we aimed to observe the effects of HGMM treatment on
vascular flow in the uterine arteries, umbilical artery and middle cerebral artery.
Key words: LMWH, thrombophilia, intrauterine growth restriction, Doppler
INTRODUCTION
Doppler ultrasonography has been expanding day by day
thanks to its superiority in analyzing hemodynamics.
Changes in fetoplacental and uterine hemodynamics canbe identified without requiring an invasive procedure on
many sites, particularly umbilical artery, fetal middle
cerebral artery and uterine artery, for topics such as
intrauterine growth retardation, fetal anemia followup andmanagement, preeclampsia and even the prediction ofpoor obstetric outcomes. [1]Blood flow to theplacenta is provided by the uterine artery . Mostly Doppler
evaluation of uterine artery is being used for preeclampsia
prognostic [2]. By analyzing vascular placental resistence
using Doppler ultrasonography on uterine artery, serious
aid is given in the management of perinatal moratality fromintrauterine growth restriction [3]Another important parameter in evaluation of
palcental perfusion is umbilical artery flow whos
degradation is associated with adverse pregnancy
outcomes such as IUGR. Abnormal umbilical artery bloodflow has been implicated in pregnancy complications andfetal demise. Its relation to histopathological changes in
the placenta and to maternal or fetal thrombophilia is not
as well understood.[4] Vascular pulsatility index in fetal
middle cerebral artery (MCA) is very important for theantenatal follow-up and management of IUGR in termsof the brain sparing effect defined as centralization . It isalso very important to predict the poor obstetric outcomes
as a component of “cerebro-placental ratio” (CPR) which
has been investigated frequently in the recent years. [5].
Thrombophilia represents an important factor in maternalfetal complications during pregnancy and it is known toincrease
the risk of adverse pregnancy outcome.
According to current data, there is an extensive literaturedebate regarding the coagulation disorder in pregnant
patients, that may lead to various complications andincrease the morbidity and mortality of both mother andfetus, and lead to adverse pregnancy outcome, such as
spontaneous abortion, intrauterine growth restriction,
placental abruption, preeclampsia, stillbirth or venous
thromboembolism [6]. Thus an appropriate treatment
initiated at the right time can make the difference between
a good or bad pregnancy outcome for both mother andoffspring. Anticoagulation with low molecular weight
heparins (LMWHs) is a well-established antithrombotic
practice for primary and secondary thromboprophylaxis
during pregnancy. Due to their excellent safety record,
LMWHs have been offered to women at high risk of an
adverse pregnancy outcome in advance of scientificevidence [7]. The administration of LMWHs in the

Received 12th of May 2021. Accepted 11th of June 2021. Address for correspondence: Pana Razvan Cosmin, Clinical Emergency County
Hospital Craiova, Obstetrics and Gynecology Department Tabaci 1, Craiova 200642; Phone: 0040727344111, razvancosminpana@yahoo.com

Fiziologia - Physiology • 2022.1(103)27

prevention of pre-eclampsia and small for gestational age(SGA) fetuses is based on biological plausibility andextrapolation from antiphospholipid syndrome [8]MATERIAL AND METHODS
The study is a retrospective one on, patients
admitted at the Craiova County Emergency Clinical
Hospital. It was conducted on 60 patients with different
forms of thrombophilia (various gene mutations ) and who
were followed until the time of birth. 30 patients received
anticoagulant treatment ( single dose per day of LMWH
s.c. ) and 30 decided to refuse anticoagulation after being
presented with the potential risks and benefits of LMWH.
The exclusion criterion was the presence of apersonal history of venous thrombosis and adverse
pregnancy outcomes.
All the patients included in the study had been tested
for hereditary thrombophilia using the same laboratory. We
searched for the two most common mutations of MTHFR
gene, C677T and A1298C and also mutations of the PAI
gene.
In addition to monitoring the incidence of
intrauterine growth restriction, we also monitored flows on
the uterine arteries, umbilical arteries and middle cerebral
artery in the second and third trimester.
RESULTS
1. No anticoagulation
We had 30 patients who refused to be treated with LMWH.
We observed a natural decrease in uterine artery
pulsatility index from second to third trimester in all
patients without LMWH. Of the 30 patients without
anticoagulation, 7 (representing 23%) had an abnormallyhigh pulsatility index (PI) on the uterine artery in the 2ndtrimester and only 2 of these 7 had the same values in the
3rd trimester. IUGR pregnancies occurred in 5 of the 7patients with abnormally high PI in the 2nd trimester,
representing 16% of patients without anticoagulation and
71% of patients without anticoagulation and with high PI in
the 2nd trimester.
21 out of 30 patients had 2nd trimester uterine arterynotch which persisted only in 10 patients in the 3rdtrimester. More than 50% of the patients in this group had
a ratio of less than 1 for ACM/Umbilical Artery relation in
the 2nd trimester. In the third trimester ACM/Umbilical
Artery < 1 was found in over 75% of patients.
2. Anticoagulation
As in the previous group, a natural decrease in thepulsatility index was observed on the uterine artery in the
3rd trimester. High values of PI were found in 23% of thisgroup in the 2nd trimester with only 2 remaining the samein the 3rd trimester. IUGR occurred in 2 (6,6%) out of the 7patients with abnormally high PI in the 2nd trimester. 26 out
of 30 patients had a notch on the uterine artery in the 2ndtrimester with only 6 being present in the 3rd trimester.
ACM/Umbilical artery ratio was less than 1 in about 35%of the patients in this group in the 2nd trimester growing up
to 50% in the 3rd trimester.

28

DISCUSSIONSACM/UA pulsatility index ratio is a very goodpredictor of adverse outcome in the fetuses of women withpreeclampsia and gestational hypertension [9]. Early
studies (2003) showed some improvement in umbilicaland middle cerebral artery flow under thromboprophylaxis[10]. In our study the anticoagulated patients had asignificantly lower percentage of subunit ratio of
ACM/Umbilical artery. The use of LMWH to improveuterine artery indices has been studied and provenespecially in patients with gestational hypertension [12]and also in non-thrombophilic placental mediatedcomplications [13]. There are some studies that concluded
that the use of LMWH in thrombophilic pregnant women
do not add any benefits regarding flow indices on uterine
arteries. Our study showed no improvement in uterine
artery PI with LMWH administration, the percentage ofpatients with abnormally high PI was the same in both
anticoagulated and non-anticoagulated groups. Still, IUGR
occurred in a smaller percentage in the anticoagulatedgroup suggesting that uterine arteries flow indices might
not be implicated in the mechanism of IUGR emergence.
The effect of LMWH administration on uterine arterynotching in patients with thrombophilia is controversial andeven when the results show an improvement in blood flow
there is no significant effect on pregnancy outcomes[14].
Our study showed only a minor improvement in thepresence of uterine artery notching with insignificant
results on IUG occurrence crossed with uterine arterynotching.
CONCLUSIONS
Administration of LMWH was of little use, nomajor improvement being found regarding the vascular
indices on the uterine artery or umbilical/middle cerebral
artery ratio. Uterine artery notch improved from 2nd to 3rdtrimester irrespective of anticoagulation. IUGR incidence
was significantly lower in the group with anticoagulation
altough vascular indices were not improved by LMWH.
From our small study group we can conclude that
anticoagulation in patients with thrombophilia does reduce
the IUGR incidence but not by improving the Doppler
indices on uterine artery, umbilical artery or middle
cerebral artery. Thus different mechanisms of actions areprobably responsible of the poor pregnancy outcomes inwomen with thrombophilia.

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23. Li X, Zhang C. Comparative efficacy of nine
antidepressants in treating Chinese patients with poststroke depression: A network meta-analysis. J Affect Disord
2020 Apr 1;266:540-548. doi: 10.1016/j.jad.2020.02.005.

24. Kappelmann N, Lewis G, Dantzer R, Jones PB, Khandaker
GM. Antidepressant activity of anti-cytokine treatment: asystematic review and meta-analysis of clinical trials ofchronic inflammatory conditions. Mol Psychiatry. 2018
Feb;23(2):335-343. doi: 10.1038/mp.2016.167.
25. Wang SB, Wang YY, Zhang QE, Wu SL, Ng CH, Ungvari
GS, Chen L, Wang CX, Jia FJ, Xiang YT. Cognitive
behavioral therapy for post-stroke depression: A metaanalysis. J Affect Disord. 2018 Aug 1;235:589-596. doi:
10.1016/j.jad.2018.04.011.
26. Song K, Xiong F, Wang Y, Wang H, Huang A, Zhang H.
Complementary and alternative therapies for poststroke
depression: A protocol for systematic review and network
meta-analysis. Medicine (Baltimore). 2020 Sep
18;99(38):e21995. doi: 10.1097/MD.0000000000021995.

29

MODIFICARI VASCULARE IN SARCINILE CU TROMBOFILIE. ANTICOAGULAREAFACE DIFERENTA?

REZUMAT

Sunt binecunoscute modificarile vasculare si ale cascadei coagularii ce apar in mod fiziologic in timpul sarcinii. In cazul femeilor
cu trombofilie ereditara asociata sarcinii aceste modificari sunt accentuate si produc efecte nedorite asupra sarcinii. Desi
tratamentul anticoagulant pe perioada sarcinii pare sa fie solutia logica in aceste cazuri, totusi, multe studii arata beneficii netela folosirea anticoagulantelor doar la pacientele cu antecedente obstetricale nefavorabile (pierderi de sarcina, preeclampsie,eclampsie, restrictie de crestere intrauterina). In acest studiu ne-am propus sa observam efectele tratamentului cu HGMM
asupra fluxurilor vasculare la nivelul arterelor uterine , arterei ombilicale si arterei cerebrale medii.
Cuvinte cheie: HGMM, trombofilie, restricție de creștere intrauterină, Doppler

UA in second and third trimester
2.00
1.50
1.00
0.50
0.00

1 2 3 4 5

6 7 8 910 11 12 13 14

15 16 17 18

19 20 21 22

23 24 25 26

27 28 29 30

31 32

Fig 1. Uterine artery doppler evaluation in second and third trimester in the group without LMWH

UA in 2nd and 3rd trimester
2.00
1.00
0.00
1 2 3 4 5

6 7 8 910 11 12 13

14 15 16 17

18 19 20 21

22 23 24

25 26 27

28 29 30

Fig 2. Uterine artery doppler evaluation in 2nd and 3rd trimester in the group which received anticoagulation

30

Fiziologia - Physiology • 2022.1(103)CHALLENGES IN THE MANAGEMENT OF INVASIVE
BASAL CELL CARCINOMA OF DORSUM NASI

Ivan Vatamanesku2, Viorel Parasca1,2, Maria-Magdalena Smeu2, Dragos Popa4, Dan
Calota2, Tiberiu Tebeica3, Marius Eugen Ciurea4
1UMF Carol Davila Bucharest
2SCUCPRA Bucharest
3Leventer Center, Histopathology Department
4Department of Plastic Surgery, University of Medicine and Pharmacy of Craiova, Romania.

ABSTRACT

Basal cell carcinoma is the most frequent malignant tumor of the skin that usually appears on the sun exposed areas andmacroscopically looks like skin-colored bump with rolled edges or a patch with scaly-waxed surface that doesn’t heal with timeor local topical treatment.
We present the case of a 71-years old patient that was redirected to plastic surgery department by a dermatology specialist
with a presumed diagnosis of squamos cell carcinoma after punch-biopsy and dermatoscopy. The unusual macroscopicallyappearance pleaded to SCC due to large ulceration borders and center. Images of CT showed no regional adenopathy, but
local infiltration of alar-cartilages and superficial part of the septum.
We performed 3 stages excision surgeries with histopathology examination for a radical excision, followed by a 2 stage nosereconstruction with frontal flap based on supraorbital and supratrochlear arteries and it’s structure was based on osseous grafts,
cartilage grafts recruited from left concha and remaining septum and oral mucosal grafts for nasal cavity lining.
The particularity of this case report is the depth of the tumor invasion, the involvement of the alar cartilages that needed to be
excised, the multiple stages of tumor excision in order to obtain free-tumor edges and the multistep reconstruction of the nosein order to obtain a strong and breathable structure.
Keywords: nose reconstruction, basal cell carcinoma, multi stage excision, frontal flap, cartilage grafts

INTRODUCTION

Basal cell carcinoma is the most frequent
nonmelanotic skin cances, has a local invasive pattern,
but almost never metastasize. In Europe the incidenceof BCC (basal cell carcinoma) is 15/100 000 per year, andit had doubled in the last 15 years.[1,2] The estimatednumbers of new cases of cancer and deaths in 2021
cannot be compared with other previous years, due to thepandemic. Covid 19 pandemic already creates decreased
access and fewer resources for diagnosis and cancer
treatments, which will influence a higher mortality, a low

survival rate and a lower incidence of new cancer casesthat are more invasive and request multi-step
approach.[3,4,5,6].
MATERIAL AND METHODS
A 80-year-old man was reffered from a local
dermatology clinic to our department in July 2021, for askin tumor on dorsum nasi. The tumor was 2.5x3.5 cm(fig.1) , with an ulcerated and bleeding surface and it had
increased in size in the last 2 weeks with at least 15%.
anticoagulation after being presented with the

Received 12th of April 2022. Accepted 24th of June 2022. Address for correspondence: Vlad Petre Atanasescu M.D. 1,2, vladpetre.atanasescu@drd.umfcd.rotelephone: 004 0723181624 e-mail: lore.anghel@yahoo.commailto:chioreandianamaria@yahoo.com

Fiziologia - Physiology • 2022.1(103)31

Paraclinical investigations and pathological features
The punch biopsy revealed the suspicion of squamouscell carcinoma with atypical mitosis that invades the basal
membrane. Due to SCC suspicion, we recommended CT
scans and an oncology consult.
CT scan with iv contrast revealed discrete iodofile
modification of dorsum nasi and right paramedian in the
middle part of the nose, involvement of the right alar
cartilage and anterior portion of septum. No adenopathy.
No systemic visible metastasis.
Histopathology
Excision biopsy result revealed an ulcerated tumor
proliferation of the skin, organized in cords and spans ofhyper chromatic basal cell, with pleomorphic form, close
to the epidermis and diffused in the entire dermis, with theinvasion of muscular layer and of the perichondrium
beneath. Multiple centers of chondromalacia associatedwith important inflammation and fibrosis. Zones withsquamous cell metaplasia and intratumor keratinization.
As a conclusion the lesion represents an infiltrating basal
cell carcinoma with profound margin infiltrated (fig 2).
Treatment and Prognosis
The tumor was primarily excised with a 5 mm lateral
margins and in block with the right alar cartilage, anterior
septum, with a partial, medial left alar cartilage andsubjacent mucosa for a radical treatment (fig 3) and thedefect was covered with a tulle gras dressing until thehistopathology result came in (fig 3).
Histopathology result revealed not SCC (squamouscell carcinoma) like previous punch biopsy, but basal cell
carcinoma. The profound margin oriented in area 2-4
o’clock in the rapport with the cranial pole of the tumor
were tumor positive, which lead to the decision of reexcision in the noted areas.
The second histopathology result showed tumor freemargins (fig 4), but in order to maintain a radical surgicaltreatment we decided that in the same operation tominimally re-excise the border in a 3rd stage (fig 5) andperform the reconstruction of the nose.
We performed frontal flap for external coverage of thedorsum nasi (fig 6), conchal cartilage graft (fig 7) andinferior septum graft (fig 8) and bone graft to reconstruct
the alar cartilages and anterior septum, and a oral mucosal
graft (fig 9) to reconstruct the nasal cavity lining that was
excised.[7,8,9,10,11,12]
After putting in place all the cartilage grafts andsuturing to the remaining cartilage (fig. 10), the frontal flap
was created and rotated on top of the defect, suturing the
mucosal graft on the internal side of the nose (fig. 11), The
frontal flap will maintain its pedicle 3-4 weeks (fig. 12) for
an appropriate vascular integration and it will be trimmed
and divided after that in a second-stage-reconstruction
operation.[13,14,15,16,17]
The donor area on the forehead for the frontal flap was
primary closed, but was in tension and had suffered, andso a skin graft for the frontal area was planned for thesecond step surgery, as to pedicle-division and flap

32

trimming (fig. 13).[16,17]
Frontal flap reconstruction of the nose is the goldstandard procedure for large tridimensional nasal defects
and with time, the skin and subcutaneous tissue will
readjust and give a close to normal aspect of the operatedside (fig. 14). Due to reconstruction of the septal and alar
cartilage, the nasal cavity is not obstructed and nostrils
have a normal shape (fig. 15)[18,19,20,21,22,23]
The patient continued with the oncologist
recommendations and the adjuvant treatment(radiotherapy) after the final operation is healed. The
forehead flap brings tissue that is very well vascularised
into the defect area and is safe for adjuvant radiotherapy.
RESULTS
The patient prognosis is good, after complete andradical excision with reconstruction of nasal lesions andwith good follow-up and adjuvant procedures. The patientis very happy with the post operatory aspect and breathes
normally with his nose.
DISCUSSIONS
Late diagnosed and invasive skin cancers ofcomplex structures need aggressive treatments and multistep surgeries in order to obtain a radical excision and aclose to normal aspect or function. [24,25]
Punch biopsy result may not be the same as theexcision- biopsy due to polymorphic cells within the tumor
and interdisciplinary investigation need to be done before
deciding the treatment plan. [26,27,28]
Rare cases of mixed tumor histopathology need athorough investigation, a multi-stage radical treatment and,if necessary, post operatory adjuvant treatment,
followed by close follow-up exams in the years to come.[29]CONCLUSIONS
Our study shows a 25% incidence of IUGR
regardless of anticoagulation and a double percentage ofpregnancies with intrauterine growth restriction in patients
without LMWH compared to those who received LMWH. It
should be noted that the restriction was encounteredregardless of anticoagulation in patients with concomitantmutations on several genes (MTHFR, PAI) and that in thecase of patients without anticoagulation the most common
mutation is homozygosity of the PAI gene.
In view of the above, we believe that additional
stratified studies on subvariant mutations and mutation
combinations are needed to accurately determine which
patients benefit from LMWH administration.

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Ichinokawa,a,* Akiko Ohtuki,a Mariko Hattori,a Hiroko
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6. Delayed skin cancer diagnosis in 2020 because of the
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J AM ACAD DERMATOL VOLUME 85, NUMBER 3
7. Basal Cell Carcinoma of the Outer Nose: Overview on
Surgical Techniques and Analysis of 312 Patients, J Cutan
Aesthet Surg. 2014 Jul-Sep; 7(3): 143–150. doi: 10.4103/09742077.146660
8. Basal Cell Carcinoma of the Head and Neck, J Skin
Cancer. 2011; 2011: 496910. Published online 2010 Dec 15. doi:
10.1155/2011/496910
9. Red Dot Basal Cell Carcinoma: Report of Cases andReview of This Unique Presentation of Basal Cell Carcinoma.
Cureus Cohen P R (March 22, 2017)
9(3): e1110.
doi:10.7759/cureus.1110
10. Basal Cell Carcinoma of the Head and Neck Region: An
Analysis of 171 Cases", Omer Sefvan Janjua, Sana Mehmood
Qureshi, Journal of Skin Cancer, vol. 2012, Article ID 943472, 4
pages, 2012. https://doi.org/10.1155/2012/943472
11. Reconstruction algorithm for nasal basal cell carcinoma
with skin involvement only: analysis of 221 cases repaired byminor surgery, Y J Kim 1, H H Cho 2, S O Kim 3, J B Lee 1, S C
Lee 1, PMID: 25959078 DOI: 10.1111/ced.12676
12.
Complex nasal reconstruction: a case study:
reconstruction of full-thickness nasal defect , Amit D Bhrany,
PMID: 21112520 DOI: 10.1016/j.fsc.2010.10.007
13. The dorsal nasal flap for reconstruction of large nasal tip
defects, Marc S Zimbler, PMID: 18248465 DOI: 10.1111/j.15244725.2007.34101.x
14. Reconstructive rhinoplasty. Park SS, Cook TA. Facial
Plast Surg. 1997 Oct;13(4):309-16. doi: 10.1055/s-00281082430. PMID: 9656885
15. Aesthetic reconstruction of nasal defects using forehead
flaps. Konz B, Wörle B, Sander CA. Facial Plast Surg. 1997
Apr;13(2):111-7. doi: 10.1055/s-2008-1064473. PMID: 9243986

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16. Three-dimensional nasal reconstruction using aprefabricated forehead flap: case report. Alagöz MS, Işken T,
Sen C, Onyedi M, Izmirli H, Yücel E. Aesthetic Plast Surg. 2008
Jan;32(1):166-71. doi: 10.1007/s00266-007-9026-5. Epub 2007
Sep 1. PMID: 17763893
17. Midline forehead skin flap. , Millard DR. In: Berish S,
editor. Grabb's Encyclopedia of Flaps. Vol. 1. Philadelphia, Pa,
USA: Lippincott Williams & Wilkins; 2009. pp. 99–100.
18. Factors influencing the linear depth of invasion ofprimary basal cell carcinoma., Takenouchi T, Nomoto S, Ito M.
Dermatologic Surgery. 2001;27(4):393–396. [PubMed]
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20. [Reconstruction of nasal defect after tumor excision].
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2007 Jul;21(7):714-7. PMID: 17694661
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Zhang JD, Li L. Zhonghua Zheng Xing Wai Ke Za Zhi. 2007
Sep;23(5):378-80. PMID: 18161347
22. Nasal Reconstruction: An Overview and Nuances ,James F. Thornton, John R. Griffin, Fadi C. Constantine, Semin
Plast Surg. 2008 Nov; 22(4): 257–268. doi: 10.1055/s-00281095885, PMCID: PMC2884875
23. Multi-stage flap reconstruction of the nose: clinicalapplication and aesthetic refinement, M. Telfer, K. Abdel-Galil,
DOI:https://doi.org/10.1016/j.bjoms.2011.04.041
24. Aesthetic aspects of partial nose reconstruction , Jr. Clin
Plast Surg. 1981;8:177–191.
25. [Nasal reconstruction with a three-staged forehead flap:
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Guillou-Jamard MR, Rysanek B, Compère JF, Benateau H. Rev
Stomatol Chir Maxillofac. 2007 Feb;108(1):21-8; discussion 2830. doi: 10.1016/j.stomax.2005.12.001. Epub 2007 Jan 9. PMID:
17275048
26. Histopathological Study of Lesions of Nose andParanasal Sinuses, Aparna M. Kulkarni,corresponding author
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Ramteke, Indian J Otolaryngol Head Neck Surg. 2012 Sep; 64(3):275–279. Published online 2011 Sep 10. doi: 10.1007/s12070011-0286-7
27. Guidelines for the management of basal cell carcinoma,
N R Telfer 1, G B Colver, C A Morton, British Association of
Dermatologists, 2008 Jul;159(1):35-48. doi: 10.1111/j.13652133.2008.08666.x.
28. Dermatoscopy of basal cell carcinoma: Morphologic
variability of global and local features and accuracy of diagnosis
Altamura D., Menzies S. W., Argenziano G., et al.. Journal of theAmerican Academy of Dermatology. 2010;62(1):67–75. doi:
10.1016/j.jaad.2009.05.035
29. What causes basal cell carcinoma to be the commonest
cancer? ,Gilbody J. S., Aitken J., Green A. Australian Journal of
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33

PROVOCĂRI ÎN MANAGEMENTUL CARCINOMULUI BAZOCELULAR INVAZIV AL DORSUM
NASI
REZUMAT

Carcinomul bazocelular este cea mai frecventă tumoare malignă a pielii, care apare de obicei pe zonele expuse la soare și
macroscopic se prezintă ca o umflătură de culoarea pielii cu margini rulate sau un plasture cu suprafață solzoasă ceară carenu se vindecă cu timpul sau tratamentul local local. Prezentăm cazul unui pacient în vârstă de 71 de ani care a fost redirecționatcătre secția de chirurgie plastică de către un specialist dermatologie cu un presupus diagnostic de carcinom scuamos dupăpunch-biopsie și dermatoscopie. Aspectul macroscopic neobișnuit a pledat pentru SCC din cauza granițelor și centrului
ulcerației mari. Imaginile CT nu au arătat adenopatie regională, ci infiltrație locală a cartilajelor alare și a părții superficiale a
septului. Am efectuat intervenții chirurgicale de excizie în 3 etape cu examen histopatologic pentru o excizie radicală, urmată
de o reconstrucție a nasului în 2 etape cu lambou frontal bazat pe artere supraorbitale și supratrohleare și structura sa a fostbazată pe grefe osoase, grefe de cartilaj recrutate din conca stângă și sept și oral rămas. grefe de mucoase pentru căptușeala
cavității nazale. Particularitatea acestui raport de caz este profunzimea invaziei tumorale, implicarea cartilajelor alare caretrebuiau excizate, etapele multiple de excizie a tumorii pentru a obține margini tumorale libere și reconstrucția în mai multe
etape a nasului pentru a obține o structură puternică și respirabilă.
Cuvinte cheie: reconstrucție a nasului, carcinom bazocelular, excizie în mai multe etape, lambou frontal, grefe de cartilaj
FIGURES(fig1. Initial evaluation of tumor. Ulcerated and bleeding surface invading the beneath structures)(fig2. Histopathology aspect of lesion as an ulcerated basocellular carcinoma involving the cartilage)
34

Fiziologia - Physiology • 2022.1(103)(fig 3 Post tumor excision tulle gras dressing)(fig 4. Hp aspect of first re-excision- tumor free)(fig. 5. Second re-excision. Tumor free margins and specimen tissue. Radical treatment was achieved.)
Fiziologia - Physiology • 2022.1(103)35

(fig 6. Frontal flap design)(fig 7. Conchal graft to recreate

right alar cartilage)(fig 8. Inferior septal cartilage graft and bone graft from ethmoidal plate for reconstruction of the anterior septum)

36

Fiziologia - Physiology • 2022.1(103)(fig 9. oral mucosal graft)

fig. 10- allar cartilages and septum grafts)(fig. 11- oral mucosal graft inside the flap)

Fiziologia - Physiology • 2022.1(103)37

(fig. 12- frontal flap in place after 1st step reconstruction)(fig 13. Pedicle division and forehead skin graft

38

Fiziologia - Physiology • 2022.1(103)(fig 14. Post-op aspect at 2 months. Frontal view(fig15. Post-op aspect. Nostril shape close to normal, not deviated, not obstructed)

Fiziologia - Physiology • 2022.1(103)39

IDENTIFICATION OF CIS AND TRANS CONFIGURATIONS IN BIOACTIVE
LIPIDS USING MASS-ENERGY PROFILES*COJOCARIU CAROLINA1,2, *PASCARIU MIHAI-COSMIN1,3, *DINCA NICOLAE4, ONULOV ROBERT2, SISU EUGEN2,
PUSCASIU DANIELA2, SERB ALINA-FLORINA2, GEORGESCU MARIUS2
1 "Vasile Goldiș” Western University of Arad, Faculty of
Medicine/Faculty of Pharmacy, 86 Liviu Rebreanu, RO-310414 Arad,
Romania
2 “Victor Babes” University of Medicine and Pharmacy, Faculty of Medicine, 2 Eftimie Murgu Sq, RO-300041 Timisoara,
Romania
3 Renewable Energy and Electrochemistry Department, National Institute of R&D for Electrochemistry and Condensed Matter- INCEMC, 144 Aurel Păunescu-Podeanu, RO-300569, Timișoara, Romania
4 “Aurel Vlaicu” University, 81 Revolutiei Blvd., RO-310330 Arad, Romania* these authors have equal contribution

ABSTRACT
Lipids are important components of biomass and their molecular geometry has an essential role in ensuring their biologicalactivity. Knowing their correct structure is imperative for providing the proper diagnosis in medicine, or for designing efficient
drugs with minimal side effects. The cis and trans configurations of the C=C bond, responsible for the molecular geometry of
lipids, are difficult to establish due to the large number of diastereomers and their possible interconversion. This preliminary
study was performed in order to establish whether the quantum calculated mass – fragmentation energy profiles and mass
spectra of lipids can be useful in discriminating these configurations. The experimental validation, carried out with oleic andelaidic acids, shows that this type of analysis, based on the coupling of mass spectrometry with quantum-chemical methods,
could decode the structural information regarding the cis-trans configuration of the C=C bond in lipids.
Keywords: diastereomers recognition, mass spectrometry, mass-energy profiles, quantum chemical calculations
INTRODUCTION
The crucial role of lipids in cell, tissue, and organ physiology is demonstrated by a large number of genetic studies and by
the many human diseases that involve the disruption of lipid metabolic enzymes and pathways. Examples of such diseases
include cancer, diabetes, as well as neurodegenerative and infectious diseases [1]. Lipids are the main components of the cell
membrane, with a decisive role in cell activity through the selective permeability exhibited towards the chemical information[2]. Fatty acids are ubiquitous in the structure of sphingolipids from ceramides, cerebrosides and gangliosides, which arebioactive compounds with a complex role in the body and frequently used in diagnosis [3].
The cis configuration of the C=C bond predominates in fatty acids from natural lipids. This aspect is crucial in biochemistry,
as it is known and proven that the stereochemistry of an isomer is essential in ensuring its biological activity [4]. This is why
the content of trans fatty acids in raw alimentary materials and foodstuffs has long been the focus of the European Food Safety
Authority (EFSA) because numerous studies indicated them as risk factors for numerous diseases. Thus, the risk of death fromheart disease, the highest in the European Union, increases by 20-32% if more than 2% of the total daily diet is provided bytrans fats. Trans lipids accumulate in biomass due to food processing. Chromatography using chemical standards and IR
spectroscopy are the only accepted methods for their analysis [5,6]. Due to the fact that a single composition could correspondto lots of isomers, of which only some are biologically active, the difficulty of establishing the true structure inevitably arises
even when using these analytical methods. For example, a recent atlas includes for the lipidome no less than 117 lipid
subclasses and 8,051 lipids [7]. The linolenic acid group (18:3) alone contains 2912 possible positional and geometrical isomersof the C=C bond. For most of these lipid isomers no standards are available, and either they cannot be identified or a false
positive structural assignment is forced upon them. The correct structure is the only one that can be useful in medical diagnosis
or for designing efficient drugs with minimal side effects.
In this context, knowing the stereochemistry due to the C=C bond of lipids is essential because the cis configuration inlipids generates bends in the hydrocarbon chains, while the trans configuration does not change their linearity [6].
The current paper describes a preliminary study aimed at establishing whether recent strategies based on quantitative
structure–fragmentation relationship (QSFR) [8, 9] could be useful for this purpose. In these techniques, the mass –fragmentation energy profiles, quantum-chemically calculated for the candidate structures, are compared with thecorresponding mass-ion intensity profile from the experimental spectrum of the analyte. The true structure is considered to be
40

Fiziologia - Physiology • 2022.1(103)the one that gives the highest match score.
MATERIAL AND METHODS
The applied QSFR method used the mass-energy profiles calculated for two candidate cis-trans diastereomers, namely
oleic acid (18:9Z) and elaidic acid (18:9E) (Figs. 1 and 2). Method validation was done with positive ion mass spectra recorded
over time for standard oleic acids, available in spectral databases.
Mass-energy profiles
Oleic and elaidic acids have the same fragmentation pathways (Figures 1 and 2). The mass series chosen for the profiles
correspond to primary positive ions resulting from the ionization and cleavage of a single bond in the candidate structure: S 5,
S7, S8, S10, S11, OH cleavage, H2O loss and the molecular ion (Figures 1 and 2).
The enthalpy of molecule fragmentation, corresponding to each of these ions, was calculated by using the RM1 semiempirical method and the equation:ΔfHfrag = ΔfH(Ii+) + ΣΔfH(Fi) − ΔfH(M)(1)
where ΣΔfH(Fi) is the sum of the formation enthalpies of the accompanying fragments, ΔfH(Ii+) is the formation enthalpy of
the resulting ion and ΔfH(M) is the molecular enthalpy of the candidate structure. Both calculated profiles consist of 13 pairs ofcorresponding ion mass - enthalpy of fragmentation values. The experimental profiles are formed by ion mass – ion intensity
pairs.
Quantum calculations
The RM1 semi‐empirical method was chosen because it has provided good results in previous studies and is more
affordable [9]. All structures were initially modeled using the HyperChem 8.0.10 software. Neutral molecules, obtained afterMM+ pre‐optimization, were optimized using the RM1 method [10]. The radical cations were obtained from these structures
and were finally optimized using RM1. As for “spin pairing”, RHF operators were used for neutral molecules and cations while
UHF operators were employed for radicals and radical cations. For geometry optimization and ΔfH calculation, the “Polak–
Ribière (conjugate gradient)” algorithm was selected with an RMS gradient of 0.01 kcal/(Å∙mol). The molecules wereconsidered to be placed in vacuum.
Matching profiles
Energy profiles were aligned by using the order of increasing ionic masses (Table 1: B, C and D columns). The match score(P) of the candidate structures’ profiles with the experimental profile was calculated with the MS Excel software by using
equation (2) in which the linear correlation coefficient R appears.
P (%) = 100(1-R)/2(2)
Their match is one of complementarity [9] since the ionic intensity is inversely proportional to the fragmentation enthalpy,
a fundamental principle in chemical kinetics. Relation (2) is advantageous (Table 1, field f(x)) because, when the intensities ofsome ions in the spectrum are missing, the linear correlation function processes only the existing values.
S8
139
S7

183
169

129
101
O
HO

181
S5

143 113
153

265

S10

99 S

11

Oleic acid
C18H34O2
M=282

Fig. 1. Ion formation pathways used for the mass-energy profile of oleic acid, the cis isomer.

Fiziologia - Physiology • 2022.1(103)

41

O

101

S8

S10

143

113
S11

S7

153

99

HO
129

265

183
S5 181

139 169
Elaidic acid
C18H34O2
M=282

Fig. 2. Ion formation pathways used for the mass-energy profile ofoleic acid, the trans isomer (elaidic acid).

RESULTS
Table 1 shows the energy values calculated with RM1 and equation (1) (columns B and C), the masses of the ionsconsidered for profiling (column D), the intensities of the respective ions from standard spectra (columns E-M) and the resulting
matching scores for oleic acid (row 17) and elaidic acid (row 18).
DISCUSSION

The fragmentations S7 and S11 are allylic (Figures 1 and 2), and the resulting allylic ions (m/z 99, 129, 153, 183) arestabilized by conjugation. Quantum calculations show that the enthalpies of allylic fragmentation for oleic acid are higher thanthose for elaidic acid (Tab. 1), in agreement with the stabilization of allylic ions, greater in the trans than in the cis configuration.
In the case of the molecular ion (m/z 282) the situation is also similar. The higher scores obtained for oleic acid (formatted
green in Table 1) for eight out of the nine standards, indicate the true structure, thus validating this method of discrimination for
cis-trans diastereomers based on mass-energy profiles calculated with RM1. Its further optimization can improve its accuracy
and selectivity.

Table 1. MS Excel page for calculating the matching score (rows 17 and 18) between the calculated mass-energy profiles for oleic and elaidic
acids (columns B and C), and the mass-ion intensity profiles from the reference mass spectra (columns E-M)

42

Fiziologia - Physiology • 2022.1(103)CONCLUSION
The relativity of the primary ions’ intensities in the mass spectra of fatty acids contains structural information regarding the cistrans configuration of the C=C bond. Mass – fragmentation energy profiles, calculated with the quantum method RM1, canserve to decode this structural information. This QSFR method can be further developed for its use in the study of the chemical
structure-biological activity relationship, in which lipid chain geometry is involved.

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METODĂ DE IDENTIFICARE A CONFIGURAŢIILOR CIS ŞI TRANS ÎN LIPIDELE BIOACTIVE UTILIZÂND PROFILURI
MASĂ-ENERGIEREZUMAT
Lipidele sunt componente importante ale biomasei iar geometria lor moleculară are un rol esențial în asigurarea activității biologice.
Cunoașterea structurii lor corecte este singura care poate fi utilă în diagnostic sau proiectarea de medicamente eficiente cu efecte secundare
minime. Implicate în geometria moleculară a lipidelor, configurațiile cis și trans ale legăturii C=C se stabilesc anevoios din cauza număruluimare de diastereomeri și interconversiei posibile a acestora. Lucrarea noastră face un studiu preliminar pentru a stabili dacă profilurile masăenergie de fragmentare calculate cuantic și spectrele de masă ale lipidelor pot fi utile la discriminarea acestor configurații. Validarea
experimentală, realizată cu acizii oleic și elaidic, arată că acest tip de analiză, bazat pe cuplarea spectrometriei de masă cu metodele cuantochimice, ar putea decodifica informația structurală referitoare la configurația cis-trans a legăturii C=C din lipide.

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