bluuebunny/crossref_metadata_2025 huggingface dataset:
Endocervical swabs transported in first void urine as combined specimens in the detection of Mycoplasma genitalium by real-time PCR
Authors: Edberg Andreas , Aronsson Fredrik , Johansson Eva , Wikander Elisabeth , Ahlqvist Thomas , Fredlund Hans
Published: December 2008

The aim of this study was to determine whether a patient's endocervical swab specimen can be transported in first void urine (FVU) as combined specimens for the detection of Mycoplasma genitalium by real-time PCR. The study also compared two different DNA extraction methods for observation of possible PCR inhibition. Three specimens, one endocervical swab specimen transported in 2-SP medium, one endocervical swab specimen transported in FVU and a FVU specimen, were collected from 329 women. All sample types underwent manual DNA extraction whereas in the DNA extraction study, 329 endocervical swab specimens transported in FVU were subjected to both manual Chelex and automated BioRobot M48 DNA extraction. A total of 100 endocervical swab specimens transported in FVU from patients PCR-negative for M. genitalium in the study were used in the PCR inhibition analysis. M. genitalium was detected in 25/329 (7.6 %) women. The endocervical swab specimens transported in 2-SP medium and transported in FVU were positive for M. genitalium in 17/25 (68 %) and 24/25 (96 %) women, respectively. The FVU specimens alone were positive for M. genitalium in 22/25 (88 %) women. In the DNA extraction study, M. genitalium DNA was detected in 24/329 (7.3 %) and 28/329 (8.5 %) of endocervical swab specimens transported in FVU subjected to manual Chelex extraction and automated BioRobot M48 extraction, respectively. Partial PCR inhibition was detected in 6 % of samples subjected to manual Chelex extraction whereas no inhibition was detected with the automated BioRobot M48 extraction. Thus endocervical swab specimens transported in FVU demonstrate higher sensitivity than FVU specimens only and have considerably increased sensitivity compared with endocervical swab specimens transported in 2-SP medium for detection of M. genitalium DNA. Moreover, automated BioRobot M48 extraction was shown to be superior to a crude manual Chelex extraction, leaving no PCR inhibition and giving a slightly higher DNA yield and/or better sensitivity.
Evaluation of clinical and socio-demographic risk factors for antibacterial resistance of Helicobacter pylori in Bulgaria
Authors: Boyanova Lyudmila , Ilieva Juliana , Gergova Galina , Spassova Zoya , Nikolov Rossen , Davidkov Lubomir , Evstatiev Ivailo , Kamburov Victor , Katsarov Nikolai , Mitov Ivan
Published: December 2008

The aim of this study was to assess the clinical and socio-demographic risk factors for primaryHelicobacter pyloriantibacterial resistance. In total, 266 consecutiveH. pyloristrains, from untreated symptomatic adult patients who answered a questionnaire, were evaluated. Strain susceptibility to amoxicillin, metronidazole, clarithromycin and tetracycline was tested by a breakpoint susceptibility test. Metronidazole resistance was found in fewer (17.0 %) peptic ulcer patients than in non-ulcer subjects (28.3 %,P=0.037), as well as in fewer patients born in villages (12.7 %) than in those born in towns (27.6 %,P=0.016). Clarithromycin resistance varied from 8.8 to 23.4 % (P=0.009) within the hospital centres. The highest clarithromycin resistance rate was found in hospital centre A (23.4 %) compared to other centres (12.9 %,P=0.041). The factors sex, age, symptom duration, non-steroidal anti-inflammatory drug use, diabetes, type of profession and educational level were not associated withH. pyloriresistance. Logistic regression revealed that the risk factors for metronidazole resistance were non-ulcer disease [odds ratio (OR) 1.95, 95 % confidence interval (95 % CI) 1.04–3.65] and a birthplace of a town (OR 2.64, 95 % CI 1.18–5.93). The hospital centre may be a risk factor (OR 2.07, 95 % CI 1.02–4.21) for clarithromycin resistance but further studies are required to verify this suggestion. In conclusion, the knowledge of the risk factors forH. pyloriresistance to antibacterials could facilitate the treatment choice forH. pylorieradication.
Multilocus genetic analysis reveals that the Australian strains of Vibrio cholerae O1 are similar to the pre-seventh pandemic strains of the El Tor biotype
Authors: Safa Ashrafus , Bhuiyan Nurul A. , Murphy Denise , Bates John , Nusrin Suraia , Kong Richard Y. C. , Chongsanguan Manas , Chaicumpa Wanpen , Nair G. Balakrish
Published: December 2008

Episodes of cholera stemming from indigenous Vibrio cholerae strains in Australia are mainly associated with environmental sources. In the present study, 10 V. cholerae O1 strains of Australian origin were characterized. All of the strains were serogroup O1 and their conventional phenotypic traits categorized them as belonging to the El Tor biotype. Genetic screening of 12 genomic regions that are associated with virulence in V. cholerae showed variable results. Analysis of the ctxAB gene showed that the Australian environmental reservoir contains both toxigenic and non-toxigenic V. cholerae strains. DNA sequencing revealed that all of the toxigenic V. cholerae strains examined were of ctxB genotype 2. Whole genome PFGE analysis revealed that the environmental toxigenic V. cholerae O1 strains were more diverse than the non-toxigenic environmental O1 strains, and the absence of genes that make up the Vibrio seventh pandemic island-I and -II in all of the strains indicates their pre-seventh pandemic ancestry.
Anaerobiospirillum succiniciproducens bacteraemia in a patient with acute lymphoblastic leukaemia
Authors: Fadzilah M. N. , Faizatul L. J. , Hasibah M. S. , Sam I.-C. , Bador M. Kahar , Gan G. G. , AbuBakar S.
Published: December 2008

A 17-year-old man with acute lymphoblastic leukaemia had fever and diarrhoea during a febrile neutropenic episode. A spiral-shaped, Gram-negative anaerobic bacterium was isolated from blood, and confirmed as Anaerobiospirillum succiniciproducens by 16S rRNA sequencing. The patient responded to imipenem.
Vibrio cholerae O139 requires neither capsule nor LPS O side chain to grow inside Acanthamoeba castellanii
Authors: Abd Hadi , Saeed Amir , Weintraub Andrej , Sandström Gunnar
Published: December 2008

Vibrio cholerae, the causative agent of cholera, has the ability to grow and survive in the aquatic free-living amoebaAcanthamoeba castellanii. The aim of the present study was to examine the ability of the clinical isolateV. choleraeO139 MO10 to grow inA. castellaniiand to determine the effect of the bacterial capsule and LPS O side chain on intracellular growth. Results from co-cultivation, viable counts, a gentamicin assay, electron microscopy and statistical analysis showed that the association ofV. choleraeO139 MO10 withA. castellaniidid not inhibit growth of the amoeba, and enhanced growth and survival ofV. choleraeO139 MO10 occurred. The wild-typeV. choleraeO139 MO10 and a capsule mutant or capsule/LPS double mutant grew insideA. castellanii. Neither the capsule nor the LPS O side chain ofV. choleraeO139 was found to play an important role in the interaction withA. castellanii, disclosing the ability ofV. choleraeto multiply and survive insideA. castellanii, as well as the role ofA. castellaniias an environmental host forV. cholerae.
Burkholderia cepacia complex: epithelial cell–pathogen confrontations and potential for therapeutic intervention
Authors: McClean Siobhán , Callaghan Máire
Published: December 2008

Burkholderia cepaciacomplex (Bcc) is an important and virulent pathogen in cystic fibrosis patients. The interactions between this pathogen and the host lung epithelium are being widely investigated but remain to be elucidated. The complex is very versatile and its interactions with the lung epithelial cells are many and varied. The first steps in the interaction are penetration of the mucosal blanket and subsequent adherence to the epithelial cell surface. A range of epithelial receptors have been reported to bind to Bcc. The next step in pathogenesis is the invasion of the lung epithelial cell and also translocation across the epithelium to the serosal side. Furthermore, pathogenesis is mediated by a range of virulence factors that elicit their effects on the epithelial cells. This review outlines these interactions and examines the therapeutic implications of understanding the mechanisms of pathogenesis of this difficult, antibiotic-resistant, opportunistic pathogen.
Post-processing intensity measurements at favourable dose values
Authors: Diederichs Kay , Junk Michael
Published: December 2008

In a macromolecular X-ray experiment, many sets of intensity measurements are collected, with each measurement corresponding to the intensity of a unique reflection at a different X-ray dose. The computational correction of radiation damage, which occurs as a function of dose during the experiment, is a concept suggesting the approximation of each set of measured intensities with a smooth function. The value of the approximating function at a user-defined point common to all unique reflections is then used as an interpolated snapshot of the true intensities at this specific dose. It is shown here that, under realistic assumptions, interpolation with a linear function has the smallest amount of error at or near two well defined points in the dose interval. This result is a special case from a mathematical analysis of polynomial approximations which proves that the points of minimum error in the approximation of a polynomial of ordernby a polynomial of ordern − 1 are independent of the function values. Conditions are formulated under which better intensities are obtained from linear interpolation than from the usual averaging of observations.
Instrument-independent specification of the diffraction geometry and polarization state of the incident X-ray beam
Authors: Schiltz Marc , Bricogne Gérard
Published: December 2008

This work augments the proposal of Schwarzenbach & Flack [J. Appl. Cryst.(1989),22, 601–605], who have advocated the use of a diffractometer-independent definition of the azimuthal angle ψ to specify the diffraction geometry of a Bragg reflection. It is here proposed that one additional angle ξ, which is also based on a diffractometer-independent definition, is needed to encode the direction of linear polarization for those experiments where this quantity is of importance. This definition is then extended to the cases of partially and/or elliptically polarized X-ray beams, and the use of three normalized Stokes parameters,P1,P2andP3, together with ξ, is advocated in order to characterize exhaustively the polarization state of the incident beam. The conventions proposed here present a general, unambiguous and economical means of encoding the information about the diffraction geometry, without the need to record any further information about the instrument, crystal orientation matrix and goniometer angles. Data-processing software using these definitions to analyse polarization-dependent phenomena becomes instrument-independent and completely general. These methods have been implemented in the macromolecular phasing programSHARPfor exploiting the polarization anisotropy of anomalous scattering in protein crystals.
Disordered materials studied using neutron polarization analysis on the multi-detector spectrometer, D7
Authors: Stewart J. R. , Deen P. P. , Andersen K. H. , Schober H. , Barthélémy J.-F. , Hillier J. M. , Murani A. P. , Hayes T. , Lindenau B.
Published: December 2008

The technique of longitudinal neutron polarization analysis on a multi-detector neutron spectrometer (so-called `xyz'-polarization analysis) is presented. This technique allows the simultaneous and unambiguous determination of the nuclear, magnetic and nuclear spin-incoherent scattering cross sections as a function of both momentum transfer and energy transfer. The implementation ofxyz-polarization analysis on the recently upgraded D7 spectrometer at the Institut Laue–Langevin in Grenoble is described. Several examples of neutron polarization analysis studies of disordered systems on D7 are given, illustrating the valuable information which can be retrieved from the analysis of neutron diffraction patterns between the Bragg peaks.
Incarceration and drug use patterns among a cohort of injection drug users
Authors: DeBeck Kora , Kerr Thomas , Li Kathy , Milloy M.‐J. , Montaner Julio , Wood Evan
Published: December 2008

Aims  Drug law enforcement remains the dominant response to drug‐related harm. However, the impact of incarceration on deterring drug use remains under‐evaluated. We sought to explore the relationship between incarceration and patterns of drug use among people who inject drugs (IDU).

Design  Using generalized estimating equations (GEE), we examined the prevalence and correlates of injection cessation among participants in the Vancouver Injection Drug User Study followed over 9 years. In subanalyses, we used McNemar's tests and linear growth curve analyses to assess changes in drug use patterns before and after a period of incarceration among participants reporting incarceration and those not incarcerated.

Findings  Among 1603 IDU, 842 (53%) reported injection cessation for at least 6 months at some point during follow‐up. In multivariate GEE analyses, recent incarceration was associated negatively with injection cessation [adjusted odds ratio (AOR) = 0.43, 95% confidence interval (CI) 0.37–0.50], whereas the use of methadone was associated positively with cessation (AOR = 1.38, 95% CI 1.22–1.56). In subanalyses assessing longitudinal patterns of drug use among incarcerated individuals and those not incarcerated over the study period, linear growth curve analyses indicated that there were no statistically significant differences in patterns of drug use between the two groups (all P > 0.05).

Conclusions  These observational data suggest that incarceration does not reduce drug use among IDU. Incarceration may inhibit access to mechanisms that promote injection cessation among IDU. In contrast, results indicate that methadone use is associated positively with injection cessation, independent of previous frequency of drug use.
Multiple genetic tests for susceptibility to smoking do not outperform simple family history
Authors: Gartner Coral E. , Barendregt Jan J. , Hall Wayne D.
Published: December 2008

Aims  To evaluate the utility of using predictive genetic screening of the population for susceptibility to smoking.

Methods  The results of meta‐analyses of genetic association studies of smoking behaviour were used to create simulated data sets using Monte Carlo methods. The ability of the genetic tests to screen for smoking was assessed using receiver operator characteristic curve analysis. The result was compared to prediction using simple family history information. To identify the circumstances in which predictive genetic testing would potentially justify screening we simulated tests using larger numbers of alleles (10, 15 and 20) that varied in prevalence from 10 to 50% and in strength of association [relative risks (RRs) of 1.2–2.1].

Results  A test based on the RRs and prevalence of five susceptibility alleles derived from meta‐analyses of genetic association studies of smoking performed similarly to chance and no better than the prediction based on simple family history. Increasing the number of alleles from five to 20 improved the predictive ability of genetic screening only modestly when using genes with the effect sizes reported to date.

Conclusions  This panel of genetic tests would be unsuitable for population screening. This situation is unlikely to be improved upon by screening based on more genetic tests. Given the similarity with associations found for other polygenic conditions, our results also suggest that using multiple genes to screen the general population for genetic susceptibility to polygenic disorders will be of limited utility.
The relative efficacy of two levels of a primary care intervention for family members affected by the addiction problem of a close relative: a randomized trial
Authors: Copello Alex , Templeton Lorna , Orford Jim , Velleman Richard , Patel Asmita , Moore Laurence , MacLeod John , Godfrey Christine
Published: December 2008

Objectives  A randomized trial to compare two levels of an intervention (full versus brief) for use by primary health‐care professionals with family members affected by the problematic drug or alcohol use of a close relative.

Design  A prospective cluster randomized comparative trial of the two interventions.

Setting  A total of 136 primary care practices in two study areas within the West Midlands and the South West regions of England.

Participants  A total of 143 family members affected by the alcohol or drug problem of a relative were recruited into the study by primary health‐care professionals. All recruited family members were seen on at least one occasion by the professional delivering the intervention and 129 (90 %) were followed‐up at 12 weeks.

Main outcome measures  Two validated and standardized self‐completion questionnaires measuring physical and psychological symptoms of stress (Symptom Rating Test) and behavioural coping (Coping Questionnaire) experienced by the family members. It was predicted that the full intervention would show increased reduction in both symptoms and coping when compared to the brief intervention.

Results  The primary analysis adjusted for clustering, baseline symptoms and stratifying variables (location and professional group) showed that there were no significant differences between the two trial arms. The symptom score at follow‐up was 0.23 [95% confidence interval (CI): −3.65, +4.06] higher in the full intervention arm than in the brief intervention arm, and the coping score at follow‐up was 0.12 (95% CI: −5.12, +5.36) higher in the full intervention arm than in the brief intervention arm.

Conclusions  A well‐constructed self‐help manual delivered by a primary care professional may be as effective for family members as several face‐to‐face sessions with the professional.
Randomized controlled trial of a brief intervention for problematic prescription drug use in non‐treatment‐seeking patients
Authors: Zahradnik Anne , Otto Christiane , Crackau Brit , Löhrmann Ira , Bischof Gallus , John Ulrich , Rumpf Hans‐Jürgen
Published: December 2008

Aims  Dependence on or problematic use of prescription drugs (PD) is estimated to be between 1 and 2% in the general population. In contrast, the proportion of substance‐specific treatment in PD use disorders at 0.5% is comparatively low. With an estimated prevalence of 4.7%, PD‐specific disorders are widespread in general hospitals compared to the general population. Brief intervention delivered in general hospitals might be useful to promote discontinuation or reduction of problematic prescription drug use.

Design  A randomized, controlled clinical trial.

Setting  Internal, surgical and gynaecological wards of a general and a university hospital.

Participants  One hundred and twenty‐six patients fulfilling criteria for either regular use of PD (more than 60 days within the last 3 months) or dependence on or abuse of PD, respectively, were allocated randomly to two conditions.

Intervention  Subjects received two counselling sessions based on Motivational Interviewing plus an individualized written feedback (intervention group, IG) or a booklet on health behaviour (control group, CG).

Measurements  The outcome was measured as reduction (>25%) and discontinuation of PD intake in terms of defined daily dosages (DDD).

Findings  After 3 months, more participants in the IG reduced their DDD compared to the participants in the CG (51.8% versus 30%; χ2 = 6.17; P = 0.017). In the IG 17.9%, in the CG 8.6% discontinued use of PD (χ2 = 2.42; P = 0.17).

Conclusions  Brief intervention based on Motivational Interviewing is effective in reducing PD intake in non‐treatment‐seeking patients.
Does sports participation during adolescence prevent later alcohol, tobacco and cannabis use?
Authors: Wichstrøm Tove , Wichstrøm Lars
Published: December 2008

Aims  To study whether participation in organized sports during adolescence predicts increased smoking of tobacco, alcohol intoxication and cannabis use from late adolescence to adulthood when controlling for potential confounders. Moreover, to study whether such increased drug use varies according to type of sport (team versus individual), main skills needed (endurance, strength or technical) and level of competition.

Design, setting and participants  Survey of national sample of Norwegian high school students (aged 13–19 years) in 1992 (T1) followed‐up in 1994 (T2), 1999 (T3) and 2006 (T4) (n = 3251).

Measurements  Outcome measures included smoking of tobacco and 12‐month prevalences of alcohol intoxication and cannabis use, respectively. Confounders included pubertal timing, friends' drug use, perceived social acceptance, grades and parental socio‐economic status.

Findings  Latent growth curve analyses showed that initial level of participation in organized sports predicted growth in alcohol intoxication. Those involved initially in team sports had greater growth in alcohol intoxication, but lower growth in tobacco use and cannabis use, during the adolescent and early adult years compared to those involved in technical or strength sports. Practising endurance sports, as opposed to technical or strength sports, predicted reduced growth in alcohol intoxication and tobacco use.

Conclusions  Sports participation in adolescence, and participation in team sports in particular, may increase the growth in alcohol intoxication during late adolescent and early adult years, whereas participation in team sports and endurance sports may reduce later increase in tobacco and cannabis use.
Saccharomyces cerevisiae Med9 comprises two functionally distinct domains that play different roles in transcriptional regulation
Authors: Takahashi Hiroyuki , Kasahara Koji , Kokubo Tetsuro
Published: December 2008

Mediator is one of the most important co‐activators that function in eukaryotic transcriptional regulation. In Saccharomyces cerevisiae, Mediator is comprised of 25 subunits belonging to four structurally distinct modules: Head, Middle, Tail, and Cyc‐C. Although each module plays a critical role in the regulation of a distinct set of genes, the precise molecular mechanisms remain unclear. To gain new insight into the role of the less‐characterized Middle module, we analyzed the function of Med9 by constructing a set of mutants and subjecting them to a range of in vivo and in vitro assays. Our results demonstrate that Med9 has two functional domains. The species‐specific amino‐terminal half (aa 1–63) plays a regulatory role in transcriptional regulation in vivo and in vitro. In contrast, the well‐conserved carboxy‐terminal half (aa 64–149) has a more fundamental function involved in direct binding to the amino‐terminal portions of Med4 and Med7 and the assembly of Med9 into the Middle module. Importantly, activator‐dependent recruitment of TBP and Taf11 to the promoter is differentially affected in med9 extracts and in extracts lacking Mediator. Add‐back experiments indicate that some unidentified factor(s) in med9 extracts may impact the binding of TFIID to the promoter.
Identification and characterization of a novel Tre‐2/Bub2/Cdc16 (TBC) protein that possesses Rab3A‐GAP activity
Authors: Ishibashi Koutaro , Kanno Eiko , Itoh Takashi , Fukuda Mitsunori
Published: December 2008

The Tre‐2/Bub2/Cdc16 (TBC) domain is a conserved protein motif that consists of approximately 200 amino acids and is thought to function as a specific Rab‐GAP domain. Although more than 40 distinct TBC domain‐containing proteins have been identified in humans, the GAP activity and specificity of most TBC proteins have never been determined. In this study we developed a novel method of screening for Rab3A‐GAP and identified two TBC proteins (FLJ13130 and RN‐tre) whose expression in PC12 cells was associated with exclusion of endogenous Rab3A molecules from dense‐core vesicles. As expression of RN‐tre caused fragmentation of the Golgi, which presumably resulted in the loss of dense‐core vesicles themselves, we further characterized FLJ13130 as a candidate Rab3A‐GAP. The results showed that expression of FLJ13130, but not of its catalytically inactive R134K mutant, greatly reduced the amount of GTP‐Rab3A in living cells and promoted the GTPase activity of Rab3A in vitro. Unexpectedly, however, FLJ13130 also promoted the GTPase activity of Rab22A, Rab27A, and Rab35, but not of Rab2A or Rab6A. Based on these results, we propose that FLJ13130 is a novel type of Rab‐GAP that exhibits broad GAP specificity and inactivates several distinct Rab isoforms, including Rab3A, just near the plasma membrane.
Genome‐wide histone methylation profile for heart failure
Authors: Kaneda Ruri , Takada Shuji , Yamashita Yoshihiro , Choi Young Lim , Nonaka‐Sarukawa Mutsuko , Soda Manabu , Misawa Yoshio , Isomura Tadashi , Shimada Kazuyuki , Mano Hiroyuki
Published: December 2008

Epigenetic alterations are implicated in the development of cardiac hypertrophy and heart failure, but little is known of which epigenetic changes in which regions of the genome play such a role. We now show that trimethylation of histone H3 on lysine‐4 (K4TM) or lysine‐9 (K9TM) is markedly affected in cardiomyocytes in association with the development of heart failure in a rat disease model. High‐throughput pyrosequencing performed with ChIP products for K4TM or K9TM prepared from human left ventricular tissue with retained or damaged function also revealed that protein‐coding genes located in the vicinity of K4TM marks differ between functional and disabled myocytes, yet both sets of genes encode proteins that function in the same signal transduction pathways for cardiac function, indicative of differential K4TM marking during the development of heart failure. However, K9TM mark‐profile was less dependent on the disease status compared to that of K4TM. Our data collectively reveal global epigenetic changes in cardiac myocytes associated with heart failure.
Consanguinity and reproductive wastage in the Palestinian Territories
Authors: Assaf Shireen , Khawaja Marwan , DeJong Jocelyn , Mahfoud Ziad , Yunis Khalid
Published: December 2008

Many studies have found that consanguinity poses a threat to child mortality and health and can also pose a threat to offspring survival before birth. However, there are conflicting findings with some studies having found no increased risk on offspring survival associated with consanguinity. Data from a population‐based survey conducted in 2004 in the Palestinian Territories was used to assess the risk of consanguinity on offspring survival. The analysis was conducted on 4418 women aged 15–49 who were asked whether or not they had experienced a stillbirth or a spontaneous abortion. These two outcomes were combined together for the analysis of reproductive wastage. Multivariable negative binomial regression was conducted to calculate the incidence risk ratios (IRR) for each region in the Palestinian Territories separately.

The strongest risk factors for reproductive wastage, after controlling for other variables, were found to be consanguinity, age and parity with age presenting the highest IRRs. Standard of living, locality type, education level, women's employment and past intrauterine device use were not found to be significant risk factors for reproductive wastage. In the West Bank only first cousin level of consanguinity was found to be significant and ‘hamola’ level (or from same family clan) lost its significance after adjusting for other variables. In the Gaza Strip both the first cousin and ‘hamola’ levels of consanguinity were significant and presented almost equal IRRs of 1.3. In conclusion, consanguinity was found to be a significant risk factor for reproductive wastage.
Laminin receptor 1 is differentially expressed in thoracic and limb wounds in the horse
Authors: Miragliotta Vincenzo , Lussier Jacques G. , Theoret Christine L.
Published: December 2008

Healing of wounds located on the distal limbs of horses is often complicated by retarded epithelialization and the development of exuberant granulation tissue (proud flesh). Treatments that definitively resolve this pathological process are still unavailable. Molecular studies of the repair mechanism might contribute to the development of new therapeutic strategies. The study presented herein aimed to clone the full length cDNA and to study the spatio‐temporal expression profile of mRNA and protein for LAMR1, previously attributed a role in wound epithelialization, during the repair of body and limb wounds in the horse. Cloning was achieved by screening a cDNA library previously derived from 7‐day wound biopsies. Expression was studied in unwounded skin and in samples from 1‐, 2‐, 3‐, 4‐ and 6‐week‐old wounds of the body and limb. Temporal gene expression was determined by real time polymerase chain reaction (RT‐PCR) while protein expression was mapped immunohistochemically. Full‐length cDNA for equine LAMR1 was shown to be highly similar to that of other species. The mRNA expression of LAMR1 was significantly up‐regulated only in thoracic wounds, 4 and 6 weeks following wounding (upon epithelialization). Cutaneous wounding induced protein expression at both locations. Our data suggest that up‐regulation of LAMR1 protein might favour epithelialization during wound healing. However, its interaction with ligands other than laminin complicates data interpretation. Future studies should quantitatively verify the temporal expression of this protein in order to provide the basis for targeted therapies that might enhance epithelialization.
PRECLINICAL STUDY: Pavlovian drug discrimination with bupropion as a feature positive occasion setter: substitution by methamphetamine and nicotine, but not cocaine
Authors: Wilkinson Jamie L. , Li Chia , Bevins Rick A.
Published: December 2008

Bupropion can serve as a discriminative stimulus (SD) in an operant drug discrimination task, and a variety of stimulants substitute for the bupropion SD. There are no reports, however, of bupropion functioning as a Pavlovian occasion setter (i.e. feature positive modulator). The present experiment seeks to fill this gap in the literature by training bupropion in rats as a feature positive modulator that disambiguates when a light will be paired with sucrose. Specifically, on bupropion (10 mg/kg intraperitoneal) sessions, offset of 15‐second cue lights were followed by brief delivery of liquid sucrose; saline sessions were similar except no sucrose was available. Rats readily acquired the discrimination with more conditioned responding to the light on bupropion sessions. Bupropion is approved for use as a smoking cessation aid, and more recently has drawn attention as a potential pharmacotherapy for cocaine and methamphetamine abuse. Accordingly, after discrimination training, we tested the ability of cocaine (1–10 mg/kg), methamphetamine (0.1 to 1 mg/kg) and nicotine (0.00625 to 0.2 mg/kg) to substitute for the bupropion feature. Nicotine (0.05 mg/kg) and methamphetamine (0.3 mg/kg) substituted fully for bupropion; cocaine did not substitute. These results extend previous research on shared stimulus properties between bupropion and other stimulants to a Pavlovian occasion setting function. Further, this is the first report of nicotine and methamphetamine substitution for bupropion. The overlap in stimulus properties might explain the effectiveness of bupropion as a smoking cessation aid and highlight the possible utility of bupropion for treatment of stimulant use disorder.
Plasma protein profiles in early asthmatic responses to inhalation allergen challenge
Authors: Rhim T. , Choi Y.‐S. , Nam B.‐Y. , Uh S. T. , Park J. S. , Kim Y.‐H. , Paik Y.‐K. , Park C.‐S.
Published: December 2008

Although mediators, such as lipids, cytokines, and chemokines, are related to the appearance of an IPR, there has been no reliable indicator to predict conditions for the appearance of an IPR.

In this study, we adopted a proteomic approach to investigate the pathogenesis at the level of the plasma proteins and to develop plasma markers to predict the appearance of an IPR following an inhalation challenge with Dermatophagoides pteronyssinus (D.p.). Sixteen mild asthmatics were recruited. Plasma was obtained before challenge and when a decline in forced expiratory volume in 1 s (FEV1) values greater than 20% from the phosphate‐buffered saline value was achieved during D.p. allergen challenge (positive responders), or at 60 min after the highest concentration of D.p. allergen was inhaled (negative responders). After comparing normalized volumes of the spots in the two groups, differentially expressed spots were identified using intra‐gel digestion and mass spectrometric analysis. Before D.p. antigen challenge, four spots of gamma fibrinogen and its isoforms were significantly decreased and two spots of complement C3 fragments were significantly increased in the positive responders compared to the negative responders. After D.p. antigen challenge, complement C3 fragment was persistently higher, while gamma fibrinogen was lower in the positive responders than in the negative responders. A validation study using Western blotting showed that gamma fibrinogen expression in the IPR‐positive asthmatics was significantly decreased compared to the average of the IPR‐negative asthmatic control group. These results indicate that alterations in the complement cascade and fibrinogen may predispose patients to the appearance of an immediate response to D.p. allergen challenge and may provide plasma markers to predict the appearance of an IPR.
Gene targeting demonstrates that inducible nitric oxide synthase is not essential for resistance to oral candidiasis in mice, or for killing of Candida albicans by macrophages in vitro
Authors: Farah C. S. , Saunus J. M. , Hu Y. , Kazoullis A. , Ashman R. B.
Published: December 2008

Introduction:  Oral candidiasis is caused by opportunistic infections with the yeast Candida albicans. Previous studies have demonstrated important roles for innate immunity and T helper type 1‐mediated inflammatory reactions in recovery from infection, with macrophages and neutrophils as key effector cells. Both effector cell types use the inducible isoform of nitric oxide synthase (iNOS) to generate candidacidal molecules, but it is not clear whether nitric oxide (NO) is an absolute requirement for candidacidal effector activity.

Methods:  In this study we directly investigated the role of iNOS‐derived NO in resistance to murine experimental oral candidiasis, using iNOS knockout mice.

Results:  Knockout mice were no more susceptible to oral candidiasis than wild‐type controls. Bone marrow‐derived macrophages from the knockout mice killed C. albicans yeasts efficiently in vitro, and were still able to produce nitrites in an iNOS‐independent manner, albeit less efficiently than wild‐type controls. There were no significant differences in local mucosal production of interleukins 6, 12, 17A, or 23, interferon‐γ, or transforming growth factor‐β 24 h after oral challenge with C. albicans.

Conclusion:  These data suggest that iNOS‐derived NO is not required for resistance to oral candidiasis in vivo, and that bone marrow‐derived macrophages may have iNOS‐independent means of generating reactive nitrogen species.
Evidence of the presence of T helper type 17 cells in chronic lesions of human periodontal disease
Authors: Cardoso C. R. , Garlet G. P. , Crippa G. E. , Rosa A. L. , Júnior W. M. , Rossi M. A. , Silva J. S.
Published: December 2008

Introduction:  Periodontal disease is a chronic inflammation of the attachment structures of the teeth, triggered by potentially hazardous microorganisms and the consequent immune‐inflammatory responses. In humans, the T helper type 17 (Th17) lineage, characterized by interleukin‐17 (IL‐17) production, develops under transforming growth factor‐β (TGF‐β), IL‐1β, and IL‐6 signaling, while its pool is maintained by IL‐23. Although this subset of cells has been implicated in various autoimmune, inflammatory, and bone‐destructive conditions, the exact role of T lymphocytes in chronic periodontitis is still controversial. Therefore, in this study we investigated the presence of Th17 cells in human periodontal disease.

Methods:  Gingival and alveolar bone samples from healthy patients and patients with chronic periodontitis were collected and used for the subsequent assays. The messenger RNA expression for the cytokines IL‐17, TGF‐β, IL‐1β, IL‐6, and IL‐23 in gingiva or IL‐17 and receptor activator for nuclear factor‐κB ligand in alveolar bone was evaluated by real‐time polymerase chain reaction. The production of IL‐17, TGF‐β, IL‐1β, IL‐6, and IL‐23 proteins was evaluated by immunohistochemistry and the presence of Th17 cells in the inflamed gingiva was confirmed by immunofluorescence confocal microscopy for CD4 and IL‐17 colocalization.

Results:  Our data demonstrated elevated levels of IL‐17, TGF‐β, IL‐1β, IL‐6, and IL‐23 messenger RNA and protein in diseased tissues as well as the presence of Th17 cells in gingiva from patients with periodontitis. Moreover, IL‐17 and the bone resorption factor RANKL were abundantly expressed in the alveolar bone of diseased patients, in contrast to low detection in controls.

Conclusion:  These results provided strong evidence for the presence of Th17 cells in the sites of chronic inflammation in human periodontal disease.
Cytokines in gingival crevicular fluid of adolescents and young adults
Authors: Kamma J. , Mombelli A. , Tsinidou K. , Vasdekis V. , Giannopoulou C.
Published: December 2008

Background/aim:  The purpose of this study was to compare the levels of the cytokines interleukin‐1β (IL‐1β), IL‐4, and IL‐8 in the gingival crevicular fluid (GCF) of adolescents and young adults.

Methods:  Twenty‐five adolescents aged between 14 and 16 years (Group A) and 20 periodontally healthy young adults aged between 25 and 35 years (Group B) were selected from two private dental clinics limited to pedodontics and periodontics respectively in Piraeus Greece. All subjects were systemically healthy. Clinical examination included probing pocket depth (PPD), presence or absence of plaque, and bleeding on probing (BOP). GCF was collected from four sites per subject. IL‐1β, IL‐4, and IL‐8, measured as total amounts (pg/30 s), were evaluated in 180 samples using a commercially available sandwich enzyme‐linked immunosorbent assay.

Results:  IL‐1β mean levels of Groups A and B were adjusted for BOP and PPD. Differences of IL‐1β mean levels between the two age groups were statistically significant (F = 50.245, P < 0.001) in favour of Group A. Adolescents showed statistically significantly lower mean levels of IL‐4 than young adults in the presence of BOP (F = 10.690, P = 0.001). There was no statistically significant difference between adolescents and adults for the means of IL‐8 adjusted for BOP and plaque presence (F = 2.032, P = 0.161).

Conclusions:  Within the limits of this study the differences reported in mean levels of IL‐1β and IL‐4 may be attributed to the different age status.
The host cytokine response to Porphyromonas gingivalis is modified by gingipains
Authors: Stathopoulou P. G. , Benakanakere M. R. , Galicia J. C. , Kinane D. F.
Published: December 2008

Background/aims:  Clinical studies indicate that primary proinflammatory cytokines, such as interleukin‐1β (IL‐1β) are elevated in the gingival crevice around teeth with periodontitis but the secondary cytokines and chemokines, IL‐6 and IL‐8, are not. The human gingival epithelial cells (HGECs) lining the gingival sulcus respond to perturbation by microbes of dental plaque by releasing a wide range of cytokines. Porphyromonas gingivalis, a putative periodontal pathogen, possesses numerous virulence factors some of which directly impact on the host response. In the present study, we sought to determine how P. gingivalis influences the inflammatory cytokine responses.

Methods:  HGECs were challenged with P. gingivalis and other putative periodontal pathogens, and the resultant production of IL‐1β, IL‐6, and IL‐8 was assayed by enzyme‐linked immunosorbent assay (ELISA). Culture supernatants and recombinant human cytokines were challenged with live P. gingivalis wild‐type and gingipain‐deficient strains and the resultant cytokine profile was assessed by ELISA and Western blot.

Results:  We show here that primary HGECs challenged with live P. gingivalis result in high levels of IL‐1β but not the related secondary cytokines IL‐6 and IL‐8. We further demonstrate that cytokine response differences are the result of the action of P. gingivalis proteases, with lysine gingipain being the most effective.

Conclusion:  We conclude that P. gingivalis, through lysine gingipain, can subvert the protective host proinflammatory response by direct cytokine degradation. Changes in the crevicular cytokine profile have consequences in periodontal disease pathogenesis that should be considered in the development of diagnostic and therapeutic modalities.
Multiple components contribute to ability of saliva to inhibit influenza viruses
Authors: White M. R. , Helmerhorst E. J. , Ligtenberg A. , Karpel M. , Tecle T. , Siqueira W. L. , Oppenheim F. G. , Hartshorn K. L.
Published: December 2008

Introduction:  Saliva is a potentially important barrier against respiratory viral infection but its mechanism of action is not well studied.

Methods:  We tested the antiviral activities of whole saliva, specific salivary gland secretions, and purified salivary proteins against strains of influenza A virus (IAV) in vitro.

Results:  Whole saliva or parotid or submandibular/sublingual secretions from healthy donors inhibited IAV based on hemagglutination inhibition and neutralization assays. This differs from human immunodeficiency virus (HIV), for which only submandibular/sublingual secretions are reported to be inhibitory. Among purified salivary proteins, MUC5B, scavenger receptor cysteine‐rich glycoprotein 340 (salivary gp‐340), histatins, and human neutrophil defensins (HNPs) inhibited IAV at the concentrations present in whole saliva. In contrast, some abundant salivary proteins (acidic proline‐rich proteins and amylase) had no activity, nor did several other less abundant salivary proteins with known activity against HIV (e.g. thrombospondin or serum leukocyte protease inhibitor). Whole saliva and MUC5B did not inhibit neuraminidase activity of IAV and viral neutralizing and aggregating activity of MUC5B was potentiated by the neuraminidase inhibitor oseltamivir. Hence, MUC5B inhibits IAV by presenting a sialic acid ligand for the viral hemagglutinin. The mechanism of action of histatins requires further study.

Conclusions:  These findings indicate that saliva represents an important initial barrier to IAV infection and underline the complexity of host defense activity of oral secretions. Of interest, antiviral activity of saliva against IAV and HIV differs in terms of specific glandular secretions and proteins that are inhibitory.
Factors involved in the T helper type 1 and type 2 cell commitment and osteoclast regulation in inflammatory apical diseases
Authors: Fukada S. Y. , Silva T. A. , Garlet G. P. , Rosa A. L. , Da Silva J. S. , Cunha F. Q.
Published: December 2008

Introduction:  Periapical chronic lesion formation involves activation of the immune response and alveolar bone resorption around the tooth apex. However, the overall roles of T helper type 1 (Th1), Th2, and T‐regulatory cell (Treg) responses and osteoclast regulatory factors in periapical cysts and granulomas have not been fully determined. This study aimed to investigate whether different forms of apical periodontitis, namely cysts and granulomas, show different balances of Th1, Th2 regulators, Treg markers, and factors involved in osteoclast chemotaxis and activation.

Methods:  Gene expression of these factors was assessed using quantitative real‐time polymerase chain reaction, in samples obtained from healthy gingiva (n = 8), periapical granulomas (n = 20), and cysts (n = 10).

Results:  Periapical cysts exhibited a greater expression of GATA‐3, while a greater expression of T‐bet, Foxp3, and interleukin‐10 (IL‐10) was seen in granulomas. The expression of interferon‐γ, IL‐4, and transforming growth factor‐β was similar in both lesions. Regarding osteoclastic factors, while the expression of SDF‐1α/CXCL12 and CCR1 was higher in cysts, the expression of RANKL was significantly higher in granulomas. Both lesions exhibited similar expression of CXCR4, CKβ8/CCL23, and osteoprotegerin, which were significantly higher than in control.

Conclusion:  Our results showed a predominance of osteoclast activity in granulomas that was correlated with the Th1 response. The concomitant expression of Treg cell markers suggests a possible suppression of the Th1 response in granulomas. On the other hand, in cysts the Th2 activity is augmented. The mechanisms of periradicular lesion development are still not fully understood but the imbalance of immune and osteoclastic cell activity in cysts and granulomas seems to be critically regulated by Treg cells.
Association between Bifidobacteriaceae and the clinical severity of root caries lesions
Authors: Mantzourani M. , Fenlon M. , Beighton D.
Published: December 2008

Background/aims:  The isolation of members of the family Bifidobacteriaceae (bifids) from oral samples has been sporadic and a recent cloning study has suggested that they are not detectable in root caries lesions.

Methods:  To better understand the presence of bifids in root caries we obtained clinical samples (15 of each) from sound exposed root surfaces, leathery remineralizing root lesions, and soft active root lesions. We investigated each for the presence of bifids using a mupirocin‐containing selective medium and identified the isolates using 16S recombinant RNA sequencing.

Results:  The proportion of bifids, as a percentage of the total anaerobic count, was significantly related to the clinical status of the sites sampled, being 7.88 ± 1.93 in the infected dentine from soft lesions, 1.61 ± 0.91 in leathery lesions, and 0.05 ± 0.39 in plaque from sound exposed root surfaces. Bifids were isolated from all soft lesions, 13 of 15 leathery lesions, and five of the plaque samples. Bifidobacterium dentium was isolated from four of the plaque samples, from 13 samples from leathery lesions, and from 12 of the 15 samples of infected dentine from the soft active lesions. Parascardovia denticolens and Scardovia genomospecies C1 were each isolated from samples associated with all three clinical conditions whereas Scardovia inopicata and Bifidobacterium subtile were both isolated from the infected dentine of the leathery and soft lesions. Bifidobacterium breve was isolated from the infected dentine of soft root caries lesions.

Conclusion:  Bifids may be routinely isolated from root caries lesions using appropriate cultural methods.