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There is increasing attention focussed on the risks associated with mobile phones possibly serving as ‘Trojan Horse’ fomites for microbial transmission in healthcare settings. However, little is reported on the presence of microbes on community derived mobile phones which in 2021, numbered in the billions in circulation with majority being used on a daily basis. Identify viable microbial organisms swabbed from smartphones on a university campus. Entire surfaces of 5 mobile phones were swabbed and examined for their microbial content using pre-agar-based growths followed by downstream DNA metagenomic next-generation sequencing analysis. All phones were contaminated with viable microbes. 173 bacteria, 8 fungi, 8 protists, 53 bacteriophages, 317 virulence factor genes and 41 distinct antibiotic resistant genes were identified. While this research represents a pilot study, the snapshot metagenomic analysis of samples collected from the surface of mobile phones has revealed the presence of a large population of viable microbes and an array of antimicrobial resistant factors. With billions of phones in circulation, these devices might be responsible for the rise of community acquired infections. These pilot results highlight the importance of public health authorities considering mobile phones as ‘Trojan Horse’ devices for microbial transmission and ensure appropriate decontamination campaigns are implemented.

Syrine Boucherabine,1,* Rania Nassar,1,2,* Lobna Mohamed,1 Matthew Olsen,3 Fatma Alqutami,1 Shroque Zaher,1 Mahmood Hachim,1 Abdulmajeed Alkhajeh,4 Simon McKirdy,5 Rashed Alghafri,3,5– 7 Lotti Tajouri,3,5,7 Abiola Senok1 1College of Medicine, Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates; 2Oral and Biomedical Sciences, School of Dentistry, College of Biomedical and Life Sciences, Cardiff University, Cardiff, UK; 3Faculty of Health Sciences and Medicine, Bond University, Robina, QLD, Australia; 4Dubai Health Authority, Dubai, United Arab Emirates; 5Harry Butler Institute, Murdoch University, Murdoch, WA, Australia; 6General Department of Forensic Sciences and Criminology, Dubai Police, Dubai, United Arab Emirates; 7Dubai Police Scientists Council, Dubai, United Arab Emirates*These authors contributed equally to this workCorrespondence: Abiola Senok, Mohammed Bin Rashid University of Medicine and Health Sciences, Building 14, Dubai Healthcare City, P. O. Box 505055, Dubai, United Arab Emirates, Tel +97143838717, Email abiola.senok@mbru.ac.aeBackground: As high touch wearable devices, the potential for microbial contamination of smart watches is high. In this study, microbial contamination of smart watches of healthcare workers (HCWs) was assessed and compared to the individual’s mobile phone and hands.Methods: This study was part of a larger point prevalence survey of microbial contamination of mobile phones of HCWs at the emergency unit of a tertiary care facility. Swabs from smart watches, mobile phones and hands were obtained from four HCWs with dual ownership of these digital devices. Bacterial culture was carried out for all samples and those from smart watches and mobile phones were further assessed using shotgun metagenomic sequencing.Results: Majority of the participants were females (n/N = 3/4; 75%). Although they all use their digital devices at work and believe that these devices could harbour microbes, cleaning in the preceding 24 hours was reported by one individual. Predominant organisms identified on bacterial culture were multidrug resistant Staphylococcus hominis and Staphylococcus epidermidis. At least one organism identified from the hands was also detected on all mobile phones and two smart watches. Shotgun metagenomics analysis demonstrated greater microbial number and diversity on mobile phones compared to smart watches. All devices had high signatures of Pseudomonas aeruginosa and associated bacteriophages and antibiotic resistance genes. Almost half of the antibiotic resistance genes (n/N = 35/75;46.6%) were present on all devices and majority were related to efflux pumps. Of the 201 virulence factor genes (VFG) identified, majority (n/N = 148/201;73%) were associated with P. aeruginosa with 96% (n/N = 142/148) present on smart watches and mobile phones.Conclusion: This first report on microbial contamination of smart watches using metagenomics next generation sequencing showed similar pattern of contamination with microbes, VFG and antibiotic resistance genes across digital devices. Further studies on microbial contamination of wearable digital devices are urgently needed.Keywords: smart watches, mobile phones, microbial contamination, shotgun sequencing

Backgrounds\udIn 2022, smartphone use continues to expand with the number of smartphone subscriptions surpassing 6 billion and forecasted to grow to 7.5 billion by 2026. The necessity of these ‘high touch’ devices as essential tools in professional healthcare settings carries great risks of cross-contamination between mobile phones and hands. Current research emphasises mobile phones as fomites enhancing the risk of nosocomial disease dissemination as phone sanitisation is often overlooked. To assess and report via a large-scale E-survey the handling practices and the use of phones by healthcare workers.\ud \ud Methods\ud A total of 377 healthcare workers (HCWs) participated in this study to fill in an E-survey online consisting of 14 questions (including categorical, ordinal, and numerical data). Analysis of categorical data used non-parametric techniques such as Pearson's chi-squared test.\ud \ud Results\ud During an 8-h shift, 92.8% (n/N = 350/377) use their phone at work with 84.6% (n/N = 319/377) considering mobile phones as an essential tool for their job. Almost all HCWs who participated in this survey believe their mobile phones could potentially harbour microorganisms (97.1%; n/N = 366/377). Fifty-seven respondents (15.1%) indicated that they use their phones while wearing gloves and 10.3% (n/N = 39/377) have never cleaned their phones. The majority of respondents (89.3%; n/N = 337/377) agreed that contaminated mobile phones could contribute to dissemination of SARS-CoV-2.\ud \ud Conclusion\ud Mobile phone use is now almost universal and indispensable in healthcare. Medical staff believe mobile phones can act as fomites with a potential risk for dissemination of microbes including SARS-COV-2. There is an urgent call for the incorporation of mobile phone sanitisation in infection prevention protocol. Studies on the use of ultraviolet-C based phone sanitation devices in health care settings are needed.

Background Multiple sclerosis is a demyelinating autoimmune disease, for which there is no blood‐borne biomarker. Erythrocytes may provide a source of such biomarkers as they contain microRNAs. MicroRNAs regulate protein translation through complementary binding to messenger RNA. As erythrocytes are transcriptionally inactive, their microRNA profiles may be less susceptible to variation. The aim of this study was to assess the biomarker potential of erythrocyte microRNAs for multiple sclerosis and assess the potential contribution of erythrocyte‐derived extracellular vesicle microRNAs to pathology. Methods Erythrocytes were isolated from whole blood by density gradient centrifugation. Erythrocyte microRNAs of a discovery cohort (23 multiple sclerosis patients and 22 healthy controls) were sequenced. Increased expression of miR‐183 cluster microRNAs (hsa‐miR‐96‐5p, hsa‐miR‐182‐5p and hsa‐miR‐183‐5p) was validated in an independent cohort of 42 patients and 45 healthy and pathological (migraine) controls. Erythrocyte‐derived extracellular vesicles were created ex vivo and their microRNAs were sequenced. Targets of microRNAs were predicted using miRDIP. Results Hsa‐miR‐182‐5p and hsa‐miR‐183‐5p were able to discriminate relapsing multiple sclerosis patients from migraine patients and/or healthy controls with 89‐94% accuracy and around 90% specificity. Hsa‐miR‐182‐5p and hsa‐miR‐183‐5p expression correlated with measures of physical disability and hsa‐miR‐96‐5p expression correlated with measures of cognitive disability in multiple sclerosis. Erythrocytes were found to selectively package microRNAs into extracellular vesicles and 34 microRNAs were found to be differentially packaged between healthy controls and multiple sclerosis patients. Several gene targets of differentially expressed and packaged erythrocyte microRNAs overlapped with multiple sclerosis susceptibility genes. Gene enrichment analysis indicated involvement in nervous system development and histone H3‐K27 demethylation. Conclusions Erythrocyte miR‐183 cluster members may be developed into specific multiple sclerosis biomarkers that could assist with diagnosis and disability monitoring. Erythrocyte and their extracellular microRNAs were shown to target multiple sclerosis susceptibility genes and may be contributing to the pathophysiology via previously identified routes.

Introduction. Advancements in technology and communication have revolutionised the 21st century with the introduction of mobile phones and smartphones. These phones are known to be platforms harbouring microbes with recent research shedding light on the abundance and broad spectrum of organisms they harbour. Mobile phone use in the community and in professional sectors including health care settings is a potential source of microbial dissemination.Aim. To identify the diversity of microbial genetic signature present on mobile phones owned by hospital medical staff.Methods. Twenty-six mobile phones of health care staff were swabbed. DNA extraction for downstream next generation sequencing shotgun metagenomic microbial profiling was performed. Survey questionnaires were handed to the staff to collect information on mobile phone usage and users’ behaviours.Results. A total of 11259 organisms derived from 26 phones were found with 2096 genes coding for antibiotic resistance and virulent factors. These organisms corresponded to 5717 bacteria, 675 fungi, 93 protists, 320 viruses, 4456 bacteriophages. The survey of medical staff showed that 46% (12/26) of the participants used their mobile phones in the bathroom.Discussion/conclusion. Mobile phones are vectors of microbes and can contribute to microbial dissemination and nosocomial diseases worldwide. As fomites, mobile phones that are not decontaminated may pose serious risks for public health and biosecurity.

INTRODUCTION: Mobile phones are used the world over, including in healthcare settings. This study aimed to investigate the viable microbial colonisation of mobile phones used by healthcare personnel. METHODS: Swabs collected on the same day from 30 mobile phones belonging to healthcare workers from three separate paediatric wards of an Australian hospital were cultured on five types of agar plate, then colonies from each phone were pooled, extracted and sequenced by shotgun metagenomics. Questionnaires completed by staff whose phones were sampled assisted in the analysis and interpretation of results. RESULTS AND DISCUSSION: All phones sampled cultured viable bacteria. Overall, 399 bacterial operational taxonomic units were identified from 30 phones, with 1432 cumulative hits. Among these were 58 recognised human pathogenic and commensal bacteria (37 Gram-negative, 21 Gram-positive). The total number of virulence factor genes detected was 347, with 1,258 cumulative hits. Antibiotic resistance genes (ARGs) were detected on all sampled phones and overall, 133 ARGs were detected with 520 cumulative hits. The most important classes of ARGs detected encoded resistance to beta-lactam, aminoglycoside and macrolide antibiotics and efflux pump mediated resistance mechanisms. CONCLUSION: Mobile phones carry viable bacterial pathogens and may act as fomites by contaminating the hands of their users and indirectly providing a transmission pathway for hospital-acquired infections and dissemination of antibiotic resistance. Further research is needed, but meanwhile adding touching mobile phones to the five moments of hand hygiene is a simple infection control strategy worth considering in hospital and community settings. Additionally, the implementation of practical and effective guidelines to decontaminate mobile phone devices would likely be beneficial to the hospital population and community at large.

Mobile phones have become an integral part of modern society. As possible breeding grounds for microbial organisms, these constitute a potential global public health risk for microbial transmission.Scoping review of literature examining microbial's presence on mobile phones in both health care (HC) and community settings.A search (PubMedGoogleScholar) was conducted from January 2005-December 2019 to identify English language studies. Studies were included if samples from mobile phones were tested for bacteria, fungi, and/or viruses; and if the sampling was carried out in any HC setting, and/or within the general community. Any other studies exploring mobile phones that did not identify specific microorganisms were excluded.A total of 56 studies were included (from 24 countries). Most studies identified the presence of bacteria (54/56), while 16 studies reported the presence of fungi. One study focused solely on RNA viruses. Staphylococcus aureus, and Coagulase-Negative Staphylococci were the most numerous identified organisms present on mobile phones. These two species and Escherichia coli were present in over a third of studies both in HC and community samples. Methicillin-resistant S. aureus, Acinetobacter sp., and Bacillus sp. were present in over a third of the studies in HC settings.While this scoping review of literature regarding microbial identification on mobile phones in HC and community settings did not directly address the issue of SARS-CoV-2 responsible for COVID-19, this work exposes the possible role of mobile phones as a 'Trojan horse' contributing to the transmission of microbial infections in epidemics and pandemics.

Background: Variations in genotype may contribute to heterogeneity in functional adaptations to exercise. Methods: A systematic search of eight databases was conducted, and 9,696 citations were screened. Results: Eight citations from seven studies measuring 10 single-nucleotide polymorphisms and nine different functional performance test outcomes were included in the review. There was one observational study of physical activity and six experimental studies of aerobic or resistance training. The ACE (D) allele, ACTN3 (RR) genotype, UCP2 (GG) genotype, IL-6-174 (GG) genotype, TNF-α-308 (GG) genotype, and IL-10-1082 (GG) genotype all predicted significantly superior adaptations in at least one functional outcome in older men and women after prescribed exercise or in those with higher levels of physical activity. Conclusion: There is a small amount of evidence that older adults may have better functional outcomes after exercise/physical activity if they have specific alleles related to musculoskeletal function or inflammation. However, more robust trials are needed.

Purpose: The present study examined the effect of low-dose thumb exertion on lymphocyte subpopulation trafficking. The potential role of blood lactate in mediating lymphocyte redistribution was also investigated. Methods: 27 male participants (18 weightlifting-trained; 9 untrained) were separated into 3 groups of 9 (Weightlifting and Untrained Experimental: WLEXP, UTEXP; Weightlifting Placebo: WLPLA). WLEXP and UTEXP performed 4x60 second isometric thumb intervals separated by 60 second rest intervals in a single-blinded placebo-controlled study. Participants were assessed over a 60 minute post-intervention recovery period for pain, blood lactate and lymphocyte subpopulation counts. Results: WLPLA did not change for any measured variable (p>0.05). The two experimentalgroups increased significantly (p0.05). No differences in cell count were seen for CD56+/CD16+ lymphocytes across time for any group (p>0.05). UTEXP showed an early significant increase (20 min post-intervention) in CD4+CD3+ (20.78%, p0.05). Conversely, WLEXP group showed no early increase followed by a delayed increase in cell count evident at the final time-point; CD4+CD3+ (19.06%, p

Multiple Sclerosis (MS) is an inflammatory and neurodegenerative disease of the central nervous system. The inflammatory process in MS is driven by both T and B cells and current therapies are targeted to each of these cell types. Epigenetic mechanisms may provide a valuable link between genes and environment. DNA methylation is the best studied epigenetic mechanism and is recognized as a potential contributor to MS risk. The objective of this study was to identify DNA methylation changes associated with MS in CD19+ B-cells. We performed an epigenome-wide association analysis of DNA methylation in the CD19+ B-cells from 24 patients with relapsing-remitting MS on various treatments and 24 healthy controls using Illumina 450 K arrays. A large differentially methylated region (DMR) was observed at the lymphotoxin alpha (LTA) locus. This region was hypermethylated and contains 19 differentially methylated positions (DMPs) spanning 860 bp, all of which are located within the transcriptional start site. We also observed smaller DMRs at 4 MS-associated genes: SLC44A2, LTBR, CARD11 and CXCR5. These preliminary findings suggest that B-cell specific DNA-methylation may be associated with MS risk or response to therapy, specifically at the LTA locus. Development of B-cell specific epigenetic therapies is an attractive new avenue of research in MS treatment. Further studies are now required to validate these findings and understand their functional significance.

Background There is a paucity of knowledge concerning erythrocytes in the aetiology of Multiple Sclerosis (MS) despite their potential to contribute to disease through impaired antioxidant capacity and altered haemorheological features. Several studies have identified an abundance of erythrocyte miRNAs and variable profiles associated with disease states, such as sickle cell disease and malaria. The aim of this study was to compare the erythrocyte miRNA profile of relapsing-remitting MS (RRMS) patients to healthy sex- and age-matched controls. Methods Erythrocytes were purified by density-gradient centrifugation and RNA was extracted. Following library preparation, samples were run on a HiSeq4000 Illumina instrument (paired-end 100 bp sequencing). Sequenced erythrocyte miRNA profiles (9 patients and 9 controls) were analysed by DESeq2. Differentially expressed miRNAs were validated by RT-qPCR using miR-152-3p as an endogenous control and replicated in a larger cohort (20 patients and 18 controls). After logarithmic transformation, differential expression was determined by two-tailed unpaired t-tests. Logistic regression analysis was carried out and receiver operating characteristic (ROC) curves were generated to determine biomarker potential. Results A total of 236 erythrocyte miRNAs were identified. Of twelve differentially expressed miRNAs in RRMS two showed increased expression (adj. p

This study investigated whether the putative physiological benefits induced by growth hormone (GH) and insulin-like growth factor-1 (IGF-1) are countered at supra-physiological concentrations because of an augmentation in the production of mitochondrial-derived free radicals with a subsequent increase in oxidative damage, compromising mitochondrial function. To test this hypothesis, peripheral blood mononuclear cells were incubated for 4 h with either recombinant human GH (rhGH) (range = 0.25–100 μg/L) or recombinant IGF-1 (rIGF-1) (range = 100–600 μg/L) and along with control samples were subsequently analyzed by flow cytometry for the determination of cellular viability, mitochondrial membrane potential (Δψm), mitochondrial superoxide (O2–) generation, and mitochondrial permeability transition pore (mtPTP) activity. Results showed levels of mitochondrial O2– generation to be significantly reduced compared with control samples (lymphocytes: 21.5 ± 1.6 AU; monocytes: 230.2 ± 9.8 AU) following rhGH treatment at both concentrations of 5 μg/L (13.5 ± 1.3 AU, P ≤ 0.05) and 10 μg/L (12.3 ± 1.5 AU, P ≤ 0.05) in lymphocytes and at 10 μg/L (153.4 ± 11.4 AU, P ≤ 0.05) in monocytes. However, no significant effect was found at either higher rhGH concentrations or following treatment with any concentration of rIGF-1. In addition, neither of the 2 hormones had any significant effect on Δψm, mtPTP activity, or on cellular viability. In conclusion, physiological concentrations of rhGH elicited a protective cellular effect through the reduction of oxidative free radicals within mitochondria. This antioxidant effect was diminished at supra-physiological concentrations but not to a level that would elicit disruption of mitochondrial function.

Background Immunoactivation is less evident in secondary progressive MS (SPMS) compared to relapsing-remitting disease. MicroRNA (miRNA) expression is integral to the regulation of gene expression; determining their impact on immune-related cell functions, especially CD4+ T cells, during disease progression will advance our understanding of MS pathophysiology. This study aimed to compare miRNA profiles of CD4+ T cells from SPMS patients to healthy controls (HC) using whole miRNA transcriptome next-generation sequencing (NGS). Total RNA was extracted from CD4+ T cells and miRNA expression patterns analyzed using Illumina-based small-RNA NGS in 12 SPMS and 12 HC samples. Results were validated in a further cohort of 12 SPMS and 10 HC by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Results The ten most dysregulated miRNAs identified by NGS were selected for qPCR confirmation; five (miR-21-5p, miR-26b-5p, miR-29b-3p, miR-142-3p, and miR-155-5p) were confirmed to be down-regulated in SPMS (p

Multiple sclerosis (MS) is an autoimmune disease characterised by lymphocytic infiltration of the central nervous system and subsequent destruction of myelin and axons. On the background of a genetic predisposition to autoimmunity, environmental triggers are assumed to initiate the disease. The majority of MS research has focused on the pathological involvement of lymphocytes and other immune cells, yet a paucity of attention has been given to erythrocytes, which may play an important role in MS pathology. The following review briefly summarises how erythrocytes may contribute to MS pathology through impaired antioxidant capacity and altered haemorheological features. The effect of disease-modifying therapies on erythrocytes is also reviewed. It may be important to further investigate erythrocytes in MS, as this could broaden the understanding of the pathological mechanisms of the disease, as well as potentially lead to the discovery of novel and innovative targets for future therapies.

Background The purpose of this study was to investigate the mitochondrial effects exerted by physiological and supra-physiological concentrations of recombinant human growth hormone (rhGH) and recombinant insulin-like growth factor-1 (rIGF-1) under conditions of substrate saturation in peripheral blood mononuclear cells (PBMCs). Methods PBMCs from healthy male subjects were treated with either rhGH, at concentrations of 0.5, 5 and 50 μg/L, or rIGF-1 at concentrations of 100, 300 and 500 μg/L for 4 h. Mitochondrial membrane potential (Δψm) and mitochondrial levels of highly reactive oxygen species (hROS) were subsequently analysed. This analysis was performed by flow cytometry in digitonin permeabilized cells, following treatment with saturating concentrations of various respiratory substrate combinations and the use of specific electron transport chain (ETC.) complex inhibitors, enabling control over both the sites of electron entry into the ETC. at complexes I and II and the entry of electrons from reduced carriers involved in β-oxidation at the level of ubiquinol. Results Neither rhGH nor rIGF-1 exerted any significant effect on Δψm or the rate of hROS production in either lymphocyte or monocyte sub-populations under any of the respiratory conditions analysed. Conclusion That neither hormone was capable of attenuating levels of oxidative stress mediated via either complex I linked respiration or lipid-derived respiration could have serious health implications for the use of rhGH in healthy individuals, which is frequently associated with significant increases in the bioavailability of free fatty acids (FFA). Such elevated supplies of lipid-derived substrates to the mitochondria could lead to oxidative damage which would negatively impact mitochondrial function.

Growth hormone (GH) is a commonly used drug aimed at improving sport performance. The aim of this study is to evaluate the immunomodulatory effects of short-term administration of recombinant GH (rhGH) in healthy young males. NK cell number, activity and phenotype, T cell number, CD4(+) (Th1/Th2) cytokine production of IL2, IL4, IL6, IL10, TNF-α and IFN-γ and CD4(+)/CD8(+) ratio with particular attention to the possible correlation to IGF-I production were investigated. 30 males (27 ± 9 years) were randomly assigned to placebo (n = 15) or drug (rhGH) 1 mg/day groups (n = 15) with daily injection for 7 days. IGF-I plasma concentration and flow cytometry data were generated at baseline and days 8, 15, 22 and 29 post injection. Data analysis used General Linear Model with repeated measures, Bonferroni correction factor and significance at p ≤ 0.05. Serum IGF-I levels (ng/mL) increased significantly (p ≤ 0.01) on day 8 (0.48 ± 0.78) after injections compared to baseline (0.31 ± 0.07) and days 15 (0.33 ± 0.06), 22 (0.29 ± 0.05) and 29 (0.29 ± 0.06). A significant time effect was noted in IL10 secretion (pg/mL) from day 15 (P = 35.14 ± 19.93, rhGH = 26.63 ± 16.39) to days 22 (P = 61.32 ± 20.41, rhGH = 74.99 ± 46.91) and 29 (P = 101.98 ± 67.25, rhGH = 107.74; ± 122.58). There was no correlation between IGF-I and NK activity, phenotype or number along with T lymphocyte number, CD4(+)/CD8(+) ratio or Th1 and Th2 cytokine production. In conclusion, cytokine secretion spectrum was not affected by short-term rhGH administration in young males.

To investigate the migraine locus around the C19p13 region through analysis of the NOTCH3 gene (C19p13.2-p13.1), previously shown to be a gene involved in CADASIL and the TNFSF7 gene (C19p13), homologous to the ligands of TNF-alpha and TNF-beta, genes that have previously been associated with migraine. The NOTCH3 gene was analysed by sequencing all exons with known CADASIL mutations in a typical (non-familial hemiplegic) migraine family (MF1) that has previously been shown to be linked to C19p13. The TNFSF7 gene was investigated through SNP association analysis using a matched case-control migraine population. NOTCH3 gene sequencing results for affected members of MF1 proved to be negative for all known sequence variants giving rise to mutations for CADASIL. TNFSF7 gene chi-square results showed non-significant P values across all populations tested against controls, except for the MO subgroup which displayed a possible association with the TNFSF7 SNP (genotype, allele analysis P = 0.036, P = 0.017 respectively). Our results suggest that common migraine is not caused by any known CADASIL mutations in the NOTCH3 gene of interest. However, the TNFSF7 gene displayed signs of involvement in a MO affected population and indicates that further independent studies of this marker are warranted.

Multiple sclerosis (MS) is a serious neurological disorder affecting young Caucasian individuals, usually with an age of onset at 18 to 40 years old. Females account for approximately 60x of MS cases and the manifestation and course of the disease is highly variable from patient to patient. The disorder is characterised by the development of plaques within the central nervous system (CNS). Many gene expression studies have been undertaken to look at the specific patterns of gene transcript levels in MS. Human tissues and experimental mice were used in these gene-profiling studies and a very valuable and interesting set of data has resulted from these various expression studies. In general, genes showing variable expression include mainly immunological and inflammatory genes, stress and antioxidant genes, as well as metabolic and central nervous system markers. Of particular interest are a number of genes localised to susceptible loci previously shown to be in linkage with MS. However due to the clinical complexity of the disease, the heterogeneity of the tissues used in expression studies, as well as the variable DNA chips/membranes used for the gene profiling, it is difficult to interpret the available information. Although this information is essential for the understanding of the pathogenesis of MS, it is difficult to decipher and define the gene pathways involved in the disorder. Experiments in gene expression profiling in MS have been numerous and lists of candidates are now available for analysis. Researchers have investigated gene expression in peripheral mononuclear white blood cells (PBMCs), in MS animal models Experimental Allergic Encephalomyelitis (EAE) and post mortem MS brain tissues. This review will focus on the results of these studies.

Multiple sclerosis (MS) is a complex neurological disease that affects the central nervous system (CNS) resulting in debilitating neuropathology. Pathogenesis is primarily defined by CNS inflammation and demyelination of nerve axons. Methionine synthase reductase (MTRR) is an enzyme that catalyzes the remethylation of homocysteine (Hcy) to methionine via cobalamin and folate dependant reactions. Cobalamin acts as an intermediate methyl carrier between methylenetetrahydrofolate reductase (MTHFR) and Hcy. MTRR plays a critical role in maintaining cobalamin in an active form and is consequently an important determinant of total plasma Hcy (pHcy) concentrations. Elevated intracellular pHcy levels have been suggested to play a role in CNS dysfunction, neurodegenerative, and cerebrovascular diseases. Our investigation entailed the genotyping of a cohort of 140 cases and matched controls for MTRR and MTHFR, by restriction length polymorphism (RFLP) techniques. Two polymorphisms: MTRR A66G and MTHFR A1298C were investigated in an Australian age and gender matched case-control study. No significant allelic frequency difference was observed between cases and controls at the alpha = 0.05 level (MTRR chi2 = 0.005, P = 0.95, MTHFR chi2 = 1.15, P = 0.28). Our preliminary findings suggest no association between the MTRR A66G and MTHFR A1298C polymorphisms and MS.

Background Multiple sclerosis (MS) is thought to be a T cell-mediated autoimmune disorder. MS pathogenesis is likely due to a genetic predisposition triggered by a variety of environmental factors. Epigenetics, particularly DNA methylation, provide a logical interface for environmental factors to influence the genome. In this study we aim to identify DNA methylation changes associated with MS in CD8+ T cells in 30 relapsing remitting MS patients and 28 healthy blood donors using Illumina 450K methylation arrays. Findings Seventy-nine differentially methylated CpGs were associated with MS. The methylation profile of CD8+ T cells was distinctive from our previously published data on CD4+ T cells in the same cohort. Most notably, there was no major CpG effect at the MS risk gene HLA-DRB1 locus in the CD8+ T cells. Conclusion CD8+ T cells and CD4+ T cells have distinct DNA methylation profiles. This case–control study highlights the importance of distinctive cell subtypes when investigating epigenetic changes in MS and other complex diseases.

The purpose of this study was to determine whether recombinant human growth hormone (rhGH) would show any significant effects on the expression of apoptosis regulating proteins in peripheral blood mononuclear cells (PBMCs). Additionally, the potential for post-transcriptional regulation of gene expression by miRNA was assessed in two cellular compartments, the cytosol and the mitochondria. Ten male subjects were subcutaneously injected with either rhGH (1 mg) or saline (0.9%) for seven consecutive days in a double-blinded fashion. Blood sampling was undertaken prior to treatment administration and over a period of three weeks following treatment cessation. Bcl-2 and Bak gene and protein expression levels were measured in PBMCs, while attention was also directed to the expression of miR-181a and miR-125b, known translational inhibitors of Bcl-2 and Bak respectively. Results showed that rhGH significantly decreased Bak protein concentrations compared to placebo samples for up to 8 days post treatment. While cytosolic miRNA expression was not found to be significantly affected by rhGH, measurement of the expression of miR-125b in mitochondrial fractions showed a significant down-regulation eight days post-rhGH administration. These findings suggest that rhGH induces short-term anti-apoptotic effects which may be partially mediated through a novel pathway that alters the concentration of mitochondrially-associated miRNAs.

International audience; Multiple sclerosis (MS) is a chronic relapsing-remitting inflammatory disease of the central nervous system characterized by oligodendrocyte damage, demyelination and neuronal death. Genetic association studies have shown a 2-fold or greater prevalence of the HLA-DRB1*1501 allele in the MS population compared with normal Caucasians. In discovery cohorts of Australasian patients with MS (total 2941 patients and 3008 controls), we examined the associations of 12 functional polymorphisms of P2X7, a microglial/macrophage receptor with proinflammatory effects when activated by extracellular adenosine triphosphate (ATP). In discovery cohorts, rs28360457, coding for Arg307Gln was associated with MS and combined analysis showed a 2-fold lower minor allele frequency compared with controls (1.11% for MS and 2.15% for controls, P = 0.0000071). Replication analysis of four independent European MS case-control cohorts (total 2140 cases and 2634 controls) confirmed this association [odds ratio (OR) = 0.69, P = 0.026]. A meta-analysis of all Australasian and European cohorts indicated that Arg307Gln confers a 1.8-fold protective effect on MS risk (OR = 0.57, P = 0.0000024). Fresh human monocytes heterozygous for Arg307Gln have >85% loss of 'pore' function of the P2X7 receptor measured by ATP-induced ethidium uptake. Analysis shows Arg307Gln always occurred with 270His suggesting a single 307Gln-270His haplotype that confers dominant negative effects on P2X7 function and protection against MS. Modeling based on the homologous zP2X4 receptor showed Arg307 is located in a region rich in basic residues located only 12 Å from the ligand binding site. Our data show the protective effect against MS of a rare genetic variant of P2RX7 with heterozygotes showing near absent proinflammatory 'pore' function.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) affecting most commonly the Caucasian population. Nitric oxide (NO) is a biological signaling and effector molecule and is especially important during inflammation. Inducible nitric oxide synthase (iNOS) is one of the three enzymes responsible for generating NO. It has been reported that there is an excessive production of NO in MS concordant with an increased expression of iNOS in MS lesions. This study investigated the role of a bi-allelic tetranucleotide polymorphism located in the promoter region of the human iNOS (NOS2A) gene in MS susceptibility. A group of MS patients (n = 101) were genotyped and compared to an age- and sex-matched group of healthy controls (n = 101). The MS group was subdivided into three subtypes, namely relapsing-remitting MS (RR-MS), secondary-progressive MS (SP-MS) and primary-progressive MS (PP-MS). Results of a chi-squared analysis and a Fisher's exact test revealed that allele and genotype distributions between cases and controls were not significantly different for the total population (chi(2) = 3.4, P(genotype) = 0.15; chi(2) = 3.4, P(allele) = 0.082) and for each subtype of MS (P > 0.05). This suggests that there is no direct association of this iNOS gene variant with MS susceptibility.

Multiple sclerosis (MS) is a complex autoimmune disorder of the CNS with both genetic and environmental contributing factors. Clinical symptoms are broadly characterized by initial onset, and progressive debilitating neurological impairment. In this study, RNA from MS chronic active and MS acute lesions was extracted, and compared with patient matched normal white matter by fluorescent cDNA microarray hybridization analysis. This resulted in the identification of 139 genes that were differentially regulated in MS plaque tissue compared to normal tissue. Of these, 69 genes showed a common pattern of expression in the chronic active and acute plaque tissues investigated (Pvalue0.0001, rho=0.73, by Spearman's rho analysis); while 70 transcripts were uniquely differentially expressed (or = 1.5-fold) in either acute or chronic active tissues. These results included known markers of MS such as the myelin basic protein (MBP) and glutathione S-transferase (GST) M1, nerve growth factors, such as nerve injury-induced protein 1 (NINJ1), X-ray and excision DNA repair factors (XRCC9 and ERCC5) and X-linked genes such as the ribosomal protein, RPS4X. Primers were then designed for seven array-selected genes, including transferrin (TF), superoxide dismutase 1 (SOD1), glutathione peroxidase 1 (GPX1), GSTP1, crystallin, alpha-B (CRYAB), phosphomannomutase 1 (PMM1) and tubulin beta-5 (TBB5), and real time quantitative (Q)-PCR analysis was performed. The results of comparative Q-PCR analysis correlated significantly with those obtained by array analysis (r=0.75, Pvalue0.01, by Pearson's bivariate correlation). Both chronic active and acute plaques shared the majority of factors identified suggesting that quantitative, rather than gross qualitative differences in gene expression pattern may define the progression from acute to chronic active plaques in MS.

Background: Multiple sclerosis (MS) is thought to be caused by T-cell mediated autoimmune dysfunction. Risk of developing MS is influenced by environmental and genetic factors. Modifiable differences in DNA methylation are recognized as epigenetic contributors to MS risk and may provide a valuable link between environmental exposure and inherited genetic systems. Objectives and methods: To identify methylation changes associated with MS, we performed a genome-wide DNA methylation analysis of CD4+ T cells from 30 patients with relapsing–remitting MS and 28 healthy controls using Illumina 450K methylation arrays. Results: A striking differential methylation signal was observed at chr. 6p21, with a peak signal at HLA-DRB1. After prioritisation, we identified a panel of 74 CpGs associated with MS in this cohort. Most notably we found evidence of a major effect CpG island in DRB1 in MS cases ( pFDR < 3 × 10−3). In addition, we found 55 non-HLA CpGs that exhibited differential methylation, many of which localise to genes previously linked to MS. Conclusions: Our findings provide the first evidence for association of DNA methylation at HLA-DRB1 in relation to MS risk. Further studies are now warranted to validate and understand how these findings are involved in MS pathology.

Autoimmune disorders are known to affect a substantial number of people worldwide and in some cases may be fatal. They occur in the presence of unregulated inflammatory re‐ sponses including failure in self-tolerance. Some unexplained disorders with immune com‐ promises may demonstrate certain characteristics that suggest an autoimmune disorder including Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME). CFS/ME re‐ mains an unsolved disorder with multiple symptoms and no single causative factor. These symptoms may include but are not limited to incapacitating fatigue, weakened short term memory or attentiveness, sore throat, tender cervical or axillary lymph nodes, muscle pain, severe headaches, impaired sleep and postexertional malaise. To date succinct and concise mechanisms that underlie this disorder have not yet being identified although, many hy‐ potheses have been put forward. CFS/ME often occurs as a consequence of a post-infectious episode accompanied by compromises in the immune, endocrine and nervous systems. The sequences of these events have not being clearly identified. Importantly, immune deteriora‐ tion in CFS/ME is related to heightened or suppressed cell function, differential gene expres‐ sion, equivocal levels of immune cell numbers and protein secretion promoting adverse inflammatory activation. Both innate and adaptive immune system perturbations persist in CFS/ME. These characteristics are in many respects similar to mechanisms of disease in au‐ toimmune disorders suggesting that the changes in immune response may develop from cellular and molecular changes in immune cells and proteins. We propose here that as the mechanism of CFS/ME may involve certain immunological factors that have been shown to be compromised in other autoimmune diseases, CFS/ME may in some cases have an autoim‐

Multiple sclerosis (MS) is a serious neurological disorder affecting young Caucasian individ‐ uals, usually with an age of onset at 18 to 40 years old. Females account for approximately 60% of MS cases and the manifestation and course of the disease is highly variable from pa‐ tient to patient. The disorder is characterised by the development of plaques within the cen‐ tral nervous system (CNS). MS remains the most frequent cause of neurological disability, with the exception of trauma, for young adults. Investigations on twins show higher con‐ cordance rates of MS in monozygotic compared to dizygotic twins. In addition, familial sus‐ ceptibility studies show that around 15% of MS patients have an affected relative. Familial risk for MS is thus very high compared to the lifetime prevalence in the general population of approximately 0.2%. Genome wide screens for MS have provided potential data for find‐ ing specific chromosomal loci involved in MS susceptibility. A series of whole genome screens for linkage to MS have been undertaken and resulted in the discovery of significant chromosomal susceptibility loci in the genome. These data have triggered a lot of interest in the regions found associated with MS and interestingly there are a number of genes that may plausibly be involved in the aetiology and pathophysiology of MS. These candidate genes have been implicated in a variety of approaches but usually involve immunological and/or genetic studies. One of the most consistent findings has been an association of specif‐ ic major histocompatibility molecules which genes are located in the chromosome 6p21. However, other significant Non HLA regions pinpoint the involvement of several candidate genes that are currently under investigation at the sequencing and proteomic levels. Many gene expression studies have been undertaken to look at the specific patterns of gene tran‐ script levels in MS. Human tissues and experimental mice were used in these gene-profiling

Objective To investigate the role of adrenoceptor subtypes in regulating the spontaneous contractile activity of the inner lining of the urinary bladder (urothelium/lamina propria). Materials and Methods The responses of isolated strips of porcine urothelium/lamina propria to noradrenaline, phenylephrine, and isoprenaline were obtained in the absence and presence of receptor subtype-selective antagonists. Quantitative reverse-transcriptase polymerase chain reaction was undertaken to assess the expression of adrenoceptor genes. Results The tissues expressed all α 1 - and β -adrenoceptor subtypes, with the α 1A -, α 1B -, and β 2 -adrenoceptors the predominant receptors at the messenger RNA level. In the functional experiments, the rate of phasic contractions and the basal tension were increased by the α 1 -adrenoceptor agonists phenylephrine (100 μ M) and A61603 (10 μ M). The rate and tension responses to phenylephrine were reduced by low concentrations of tamsulosin (3 nM) and RS100329 (10 nM) but were unaffected by BMY7378 (100 nM), prazosin (10 nM), and RS17053 (1 μ M). In contrast, isoprenaline and salbutamol (both 1 μ M) induced a relaxation of tissues and slowing of phasic contractions. The rate and tension responses to isoprenaline were inhibited by propranolol (100 nM) or a combination of CGP20712A (30 nM) and ICI118551 (70 nM). The rate responses were also significantly inhibited by ICI118551 alone (70 nM). Conclusion Although all α 1 - and β -adrenoceptor subtypes were expressed in the pig urothelium/lamina propria, the α 1A/L -adrenoceptor appeared to mediate increases in the contractile rate and tension. The β -adrenoceptor induced inhibition of spontaneous contractile activity appears to be predominately mediated by β 2 -adrenoceptors, with β 1 - and β 2 -adrenoceptors possibly involved in the tension responses.

Heat shock proteins (HSPs) are important molecules required for ideal protein function. Extensive research on the functional properties of HSPs indicates that HSPs may be implicated in a wide range of physiological functions including immune function. In the immune system, HSPs are involved in cell proliferation, differentiation, cytokine release, and apoptosis. Therefore, the ability of the immune system, in particular immune cells, to function optimally and in unison with other physiological systems is in part dependent on signaling transduction processes, including bidirectional communication with HSPs. Regulatory T cells (Tregs) are important T cells with suppressive functions and impairments in their function have been associated with a number of autoimmune disorders. The purpose of this paper is to examine the relationship between HSPs and Tregs. The interrelationship between cells and proteins may be important in cellular functions necessary for cell survival and expansion during diseased state.

Background Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is an etiologically unexplained disorder characterised by irregularities in various aspects of the immunological function. Presently, it is unknown whether these immunological changes remain consistent over time. This study investigates Natural Killer (NK) cell cytotoxic activity, NK cell subsets (CD56brightCD16- and CD56dimCD16+) and cytokines, over the course of a12 month period in patients with CFS/ME. Methods The participants in the study comprised 65 (47.2 ± 11.5 years) CFS/ME participants and 21 (45.2 ±9.3 years) non-fatigued controls. Flow cytometry protocols were used to assess NK subsets and NK cytotoxic activity at various time points that included baseline (T1), 6 (T2) and 12 months (T3). Cytokine secretions were measured following mitogenic stimulation of peripheral blood mononuclear cells. Results NK cytotoxic activity was significantly decreased in the CFS/ME patients at T1, T2 and T3 compared to the non-fatigued group. Additionally, in comparison to the non-fatigued controls, the CFS/ME group had significantly lower numbers of CD56brightCD16- NK cells at both T1 and T2. Interestingly, following mitogenic stimulation, cytokine secretion revealed significant increases in IL-10, IFN-γ and TNF-α at T1 in the CFS/ME group. A significant decrease was observed at T2 in the CFS/ME group for IL-10 and IL-17A while at T3, IL-2 was increased in the CFS/ME group in comparison to the non-fatigued controls. Overall cytotoxic activity was significantly decreased at T3 compared to T1 and T2. CD56brightCD16- NK cells were much lower at T2 compared to T1 and T3. IL-10 and IL-17A secretion was elevated at T2 in comparison to T1 and T3. Conclusion These results confirm decreases in immune function in CFS/ME patients, suggesting an increased susceptibility to viral and other infections. Furthermore, NK cytotoxic activity may be a suitable biomarker for diagnosing CFS/ME as it was consistently decreased during the course of the 12 months study.

Introduction: It has been shown that growth hormone (GH) exerts regulatory effects on hemorheology and other metabolic functions. GH stimulates the production of insulin-like growth factor I (IGF-I) and GH-IGF-I system has profound effects on body fluid status. There are speculations that GH has become widely used as a performance enhancing drug among athletes of various sports. The present study evaluated the possible hemorheological effects of short term administration of human recombinant growth hormone (rhGH) in healthy young males. Methodology: Thirty young healthy males (27 ± 9) participated in a 29 days study where it was administered either 0.9% sodium chloride or 1 mg of human rhGH from day 1 to day 7. The participants were randomly assigned into either placebo (C) n = 15 or rhGH 1 mg/day (rhGH) group n = 15. This study evaluated plasma fibrinogen levels, red blood cell (RBC) aggregation, deformability and serum IGF-I levels between and within the groups along 29 days. Results: There was a significant increase in erythrocytes aggregation index post injection (day 8), in accordance to an increase in serum IGF-I.

Multiple sclerosis (MS) is a common cause of neurological disability in young adults. The disease generally manifests in early to middle adulthood and causes various neurological deficits. Autoreactive T lymphocytes and their associated antigens have long been presumed important features of MS pathogenesis. The Protein tyrosine phosphatase receptor type C gene (PTPRC) encodes the T-cell receptor CD45. Variations within PTPRC have been previously associated with diseases of autoimmune origin such as type 1 diabetes mellitus and Graves' disease. We set out to investigate two variants within the PTPRC gene, C77G and C772T in subjects with MS and matched healthy controls to determine whether significant differences exist in these markers in an Australian population. We employed high resolution melt analysis (HRM) and restriction length polymorphism (RFLP) techniques to determine genotypic and allelic frequencies. Our study found no significant difference between frequencies for PTPRC C77G by either genotype (Χ2 = 0.65, P = 0.72) or allele (Χ2 = 0.48, P = 0.49). Similarly, we did not find evidence to suggest an association between PTPRC C772T by genotype (Χ2 = 1.06, P = 0.59) or allele (Χ2 = 0.20, P = 0.66). Linkage disequilibrium (LD) analysis showed strong linkage disequilibrium between the two tested markers (D' = 0.9970, SD = 0.0385). This study reveals no evidence to suggest that these markers are associated with MS in the tested Australian Caucasian population. Although the PTPRC gene has a significant role in regulating CD4+ and CD8+ autoreactive T-cells, interferon-beta responsiveness, and potentially other important processes, our study does not support a role for the two tested variants of this gene in MS susceptibility in the Australian population.

The aims of the study were: (i) to extend our linkage analysis of chromosome 1q microsatellite markers in predominantly migraine with aura pedigrees and (ii) to test the novel FHM-2 ATP1A2 gene for involvement in these migraine affected pedigrees and a previous pedigree (MF14) showing evidence of linkage of markers to C1q31.A chromosome 1 scan (31 markers) was performed in 21 multiplex pedigrees affected predominantly with migraine with aura (MA). The known FHM-2 ATP1A2 gene mutations were tested, by sequencing, for the involvement in MA and migraine without aura (MO) in these pedigrees. Sequencing was performed in the coding areas of the ATP1A2 gene through three MA individuals from MF14.Evidence for linkage was obtained at C1q23 to markers spanning the ATP1A2 gene. However, testing of the known ATP1A2 gene mutations (for FHM) in common migraine probands of pedigrees showing excess allele sharing was negative. Sequencing of the entire coding areas of the gene through all the three MA affected from MF14 was also negative for mutations.Microsatellite markers on chromosome 1q23 show evidence of excess allele sharing in MA and some MO pedigrees, suggesting linkage to the common forms of migraine and the presence of a susceptibility gene in this region. The FHM-2 (ATP1A2 gene) does not seem to be involved in the common types of migraine. Despite certain clinical characteristics, the genetic correlation between FHM and familial typical migraine remains unclear. Several candidate genes lie within the C1q23 and C1q31 cytogenetic regions; therefore, further studies are needed.

Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS) resulting in accumulating neurological disability. The disorder is more prevalent at higher latitudes. To investigate VDR gene variation using three intragenic restriction fragment length polymorphisms (Apa I, Taq I and Fok I) in an Australian MS case-control population. One hundred and four Australian MS patients were studied with patients classified clinically as Relapsing Remitting MS (RR-MS), Secondary Progressive MS (SP-MS) or Primary Progressive MS (PP-MS). Also, 104 age-, sex-, and ethnicity-matched controls were investigated as a comparative group. Our results show a significant difference of genotype distribution frequency between the case and control groups for the functional exon 9 VDR marker Taq I (p(Gen) = 0.016) and interestingly, a stronger difference for the allelic frequency (p(All) = 0.0072). The Apa I alleles were also found to be associated with MS (p(All) = 0.04) but genotype frequencies were not significantly different from controls (p(Gen) = 0.1). The Taq and Apa variants are in very strong and significant linkage disequilibrium (D' = 0.96, P < 0.0001). The genotypic associations are strongest for the progressive forms of MS (SP-MS and PP-MS). Our results support a role for the VDR gene increasing the risk of developing multiple sclerosis, particularly the progressive clinical subtypes of MS.