Myasthenia gravis genome-wide association study implicates AGRN as a risk locus.

BACKGROUND: Myasthenia gravis (MG) is a rare autoimmune disorder affecting the neuromuscular junction (NMJ). Here, we investigate the genetic architecture of MG via a genome-wide association study (GWAS) of the largest MG data set analysed to date. METHODS: We performed GWAS meta-analysis integrating three different data sets (total of 1401 cases and 3508 controls). We carried out human leucocyte antigen (HLA) fine-mapping, gene-based and tissue enrichment analyses and investigated genetic correlation with 13 other autoimmune disorders as well as pleiotropy across MG and correlated disorders. RESULTS: We confirmed the previously reported MG association with TNFRSF11A (rs4369774; p=1.09×10-13, OR=1.4). Furthermore, gene-based analysis revealed AGRN as a novel MG susceptibility gene. HLA fine-mapping pointed to two independent MG loci: HLA-DRB1 and HLA-B. MG onset-specific analysis reveals differences in the genetic architecture of early-onset MG (EOMG) versus late-onset MG (LOMG). Furthermore, we find MG to be genetically correlated with type 1 diabetes (T1D), rheumatoid arthritis (RA), late-onset vitiligo and autoimmune thyroid disease (ATD). Cross-disorder meta-analysis reveals multiple risk loci that appear pleiotropic across MG and correlated disorders. DISCUSSION: Our gene-based analysis identifies AGRN as a novel MG susceptibility gene, implicating for the first time a locus encoding a protein (agrin) that is directly relevant to NMJ activation. Mutations in AGRN have been found to underlie congenital myasthenic syndrome. Our results are also consistent with previous studies highlighting the role of HLA and TNFRSF11A in MG aetiology and the different risk genes in EOMG versus LOMG. Finally, we uncover the genetic correlation of MG with T1D, RA, ATD and late-onset vitiligo, pointing to shared underlying genetic mechanisms.

Novel genetic risk variants for pediatric celiac disease.

BACKGROUND: Celiac disease is a complex chronic immune-mediated disorder of the small intestine. Today, the pathobiology of the disease is unclear, perplexing differential diagnosis, patient stratification, and decision-making in the clinic. METHODS: Herein, we adopted a next-generation sequencing approach in a celiac disease trio of Greek descent to identify all genomic variants with the potential of celiac disease predisposition. RESULTS: Analysis revealed six genomic variants of prime interest: SLC9A4 c.1919G>A, KIAA1109 c.2933T>C and c.4268_4269delCCinsTA, HoxB6 c.668C>A, HoxD12 c.418G>A, and NCK2 c.745_746delAAinsG, from which NCK2 c.745_746delAAinsG is novel. Data validation in pediatric celiac disease patients of Greek (n = 109) and Serbian (n = 73) descent and their healthy counterparts (n = 111 and n = 32, respectively) indicated that HoxD12 c.418G>A is more prevalent in celiac disease patients in the Serbian population (P < 0.01), while NCK2 c.745_746delAAinsG is less prevalent in celiac disease patients rather than healthy individuals of Greek descent (P = 0.03). SLC9A4 c.1919G>A and KIAA1109 c.2933T>C and c.4268_4269delCCinsTA were more abundant in patients; nevertheless, they failed to show statistical significance. CONCLUSIONS: The next-generation sequencing-based family genomics approach described herein may serve as a paradigm towards the identification of novel functional variants with the aim of understanding complex disease pathobiology.

Introducing dAUTObase: a first step towards the global scale geoepidemiology of autoimmune syndromes and diseases.

MOTIVATION: An autoimmune disorder occurs when the immune system mistakenly attacks and destroys its own healthy body tissues. The initiation of a geoepidemiological database, for recording autoimmune incidents with a focus to clinical manifestations, demographic parameters and geographic background is crucial to detect correlations. RESULTS: The dAUTObase collects an ever increasing number of publications-currently counting 435-on autoimmune diseases' frequencies in various populations and ethnic groups. The respective data have been hosted by a web application developed for the task. It uses three data visualization tools: the PivotViewer, the Disease Treemap and the Disease World Map, to assist the effective data querying. AVAILABILITY AND IMPLEMENTATION: The dAUTObase 2.0 version (www.biodata.gr/dautobase) needs no registration for querying, but data entry and modification is reserved for registered users (curators-administrators). CONTACT: kpoulas@upatras.gr or tzimas@cti.gr.

Identification of cancer predisposition variants in apparently healthy individuals using a next-generation sequencing-based family genomics approach.

Cancer, like many common disorders, has a complex etiology, often with a strong genetic component and with multiple environmental factors contributing to susceptibility. A considerable number of genomic variants have been previously reported to be causative of, or associated with, an increased risk for various types of cancer. Here, we adopted a next-generation sequencing approach in 11 members of two families of Greek descent to identify all genomic variants with the potential to predispose family members to cancer. Cross-comparison with data from the Human Gene Mutation Database identified a total of 571 variants, from which 47 % were disease-associated polymorphisms, 26 % disease-associated polymorphisms with additional supporting functional evidence, 19 % functional polymorphisms with in vitro/laboratory or in vivo supporting evidence but no known disease association, 4 % putative disease-causing mutations but with some residual doubt as to their pathological significance, and 3 % disease-causing mutations. Subsequent analysis, focused on the latter variant class most likely to be involved in cancer predisposition, revealed two variants of prime interest, namely MSH2 c.2732T>A (p.L911R) and BRCA1 c.2955delC, the first of which is novel. KMT2D c.13895delC and c.1940C>A variants are additionally reported as incidental findings. The next-generation sequencing-based family genomics approach described herein has the potential to be applied to other types of complex genetic disorder in order to identify variants of potential pathological significance.

PheWAS and cross-disorder analysis reveal genetic architecture, pleiotropic loci and phenotypic correlations across 11 autoimmune disorders.

Introduction: Autoimmune disorders (ADs) are a group of about 80 disorders that occur when self-attacking autoantibodies are produced due to failure in the self-tolerance mechanisms. ADs are polygenic disorders and associations with genes both in the human leukocyte antigen (HLA) region and outside of it have been described. Previous studies have shown that they are highly comorbid with shared genetic risk factors, while epidemiological studies revealed associations between various lifestyle and health-related phenotypes and ADs. Methods: Here, for the first time, we performed a comparative polygenic risk score (PRS) - Phenome Wide Association Study (PheWAS) for 11 different ADs (Juvenile Idiopathic Arthritis, Primary Sclerosing Cholangitis, Celiac Disease, Multiple Sclerosis, Rheumatoid Arthritis, Psoriasis, Myasthenia Gravis, Type 1 Diabetes, Systemic Lupus Erythematosus, Vitiligo Late Onset, Vitiligo Early Onset) and 3,254 phenotypes available in the UK Biobank that include a wide range of socio-demographic, lifestyle and health-related outcomes. Additionally, we investigated the genetic relationships of the studied ADs, calculating their genetic correlation and conducting cross-disorder GWAS meta-analyses for the observed AD clusters. Results: In total, we identified 508 phenotypes significantly associated with at least one AD PRS. 272 phenotypes were significantly associated after excluding variants in the HLA region from the PRS estimation. Through genetic correlation and genetic factor analyses, we identified four genetic factors that run across studied ADs. Cross-trait meta-analyses within each factor revealed pleiotropic genome-wide significant loci. Discussion: Overall, our study confirms the association of different factors with genetic susceptibility for ADs and reveals novel observations that need to be further explored.

Genetics of myasthenia gravis: a case-control association study in the Hellenic population.

Myasthenia gravis (MG) is an heterogeneous autoimmune disease characterized by the production of autoantibodies against proteins of the postsynaptic membrane, in the neuromuscular junction. The contribution of genetic factors to MG susceptibility has been evaluated through family and twin studies however, the precise genetic background of the disease remains elusive. We conducted a case-control association study in 101 unrelated MG patients of Hellenic origin and 101 healthy volunteers in order to assess the involvement of common genetic variants in susceptibility to MG. We focused on three candidate genes which have been clearly associated with several autoimmune diseases, aiming to investigate their potential implication in MG pathogenesis. These are interferon regulatory factor 5 (IRF-5), TNFα-induced protein 3 (TNFAIP3), also known as A20, and interleukin-10 (IL-10), key molecules in the regulation of immune function. A statistical trend of association (P = 0.068) between IL-10 promoter single nucleotide polymorphisms (SNPs) and the subgroups of early and late-onset MG patients was revealed. No statistically significant differences were observed in the rest of the variants examined. As far as we are aware, this is the first worldwide attempt to address the possible association between IRF-5 and TNFAIP3 common genetic variants and the genetic basis of MG.

Prediction of Long-Term Survival after Coronary Artery Bypass with Bilateral Internal Thoracic Artery Grafting: External Validation of A Contemporary Nomogram.

Background: This study aimed to verify the external validation of a contemporary nomogram in predicting long-term survival after an isolated coronary artery bypass with bilateral internal thoracic artery grafting (CABG-BITA). Methods: Consecutive patients who underwent CABG-BITA at a single center were included in the study. All the predictors of the original risk score (age, diabetes mellitus, chronic obstructive pulmonary disease, congestive heart failure, chronic renal failure, old myocardial infarction, ejection fraction, intra-aortic balloon pump and peripheral arterial disease) were available for analysis. Results: Among the 2846 consecutive patients, a total of 1176 (41.3%) deaths were recorded during the 31,383 patient years of follow-up. The median EuroSCORE II was 2.35, and the median follow-up was 11.1 years. The risk score showed 72.7% overall discriminatory ability as measured by Harrell's concordance statistic. It showed satisfactory calibration at 10, 15 and 20 years of follow-up. The risk score showed a time-varying nonlinear effect, and accordingly, adjusted long-term survival predictions were calculated. There were subgroups (scores < 50 points) with favorable 20-year survival rates ranging from 77% to 60%. Higher risk subgroups (scores > 90 points) showed poor 20-year survival rates ranging from 22% to 4%. Conclusions: The validated risk score represents a useful algorithm for the detection of patients who could benefit after CABG-BITA with respect to long-term survival. Although further multi-center studies are required worldwide to reveal the usefulness of this score in the clinical setting, its wide adoption may act as a motivation for cardiac surgeons resulting in higher numbers of CABG-BITA procedures.

Microcurrent Stimulation Triggers MAPK Signaling and TGF-ß1 Release in Fibroblast and Osteoblast-Like Cell Lines.

Wound healing constitutes an essential process for all organisms and involves a sequence of three phases. The disruption or elongation of any of these phases can lead to a chronic or non-healing wound. Electrical stimulation accelerates wound healing by mimicking the current that is generated in the skin after any injury. Here, we sought to identify the molecular mechanisms involved in the healing process following in vitro microcurrent stimulation-a type of electrotherapy. Our results concluded that microcurrents promote cell proliferation and migration in an ERK 1/2- or p38-dependent way. Furthermore, microcurrents induce the secretion of transforming growth factor-beta-1 (TGF-ß1) in fibroblasts and osteoblast-like cells. Interestingly, transcriptomic analysis uncovered that microcurrents enhance the transcriptional activation of genes implicated in Hedgehog, TGF-ß1 and MAPK signaling pathways. Overall, our results demonstrate that microcurrents may enhance wound closure through a combination of signal transductions, via MAPK's phosphorylation, and the transcriptional activation of specific genes involved in the healing process. These mechanisms should be further examined in vivo, in order to verify the beneficial effects of microcurrents in wound or fracture healing.

Effects of Exposure to Tobacco Cigarette, Electronic Cigarette and Heated Tobacco Product on Adipocyte Survival and Differentiation In Vitro.

Cigarette smoking (CS) causes significant morbidity worldwide, attributed to the numerous toxicants generated by tobacco combustion. Electronic cigarettes (ECIG) and heated tobacco products (HTP) are considered alternative smoking/vaping products that deliver nicotine through an inhaled aerosol and emit fewer harmful constituents than CS. However, their long-term impacts on human health are not well established. Nicotine exposure has been linked to lipolysis and body weight loss, while smoking has been associated with insulin resistance and hyperinsulinemia. Enhanced function of beige (thermogenic) adipocytes has been proposed as a means to reduce obesity and metabolic disorders. In this study, we compared the effect of extract-enriched media via exposure of culture medium to CS, HTP aerosol, and ECIG aerosol on the viability and the differentiation of 3T3-L1 pre-adipocytes to beige adipocytes. Only CS extract caused a decrease in cell viability in a dose- and time-dependent manner. Furthermore, relative lipid accumulation and expression levels of the adipocyte markers Pgc-1α, Ppar-γ and Resistin were significantly decreased in cells exposed to CS extract. Our results demonstrate that CS extract, in contrast to HTP and ECIG extracts, significantly impairs differentiation of pre-adipocytes to beige adipocytes and may therefore impact significantly adipose tissue metabolic function.

Ocular conditions and dry eye due to traditional and new forms of smoking: A review.

Addiction to cigarette smoking has high prevalence rates recorded worldwide. Smoking has been linked to several life-threatening systemic conditions such as cancer, heart attack and stroke, in addition to a range of ocular pathologies. In recent years, electronic cigarettes (EC) have emerged as alternatives to smoking. ECs are nicotine delivery devices which produce an aerosol by heating, rather than combusting, a liquid which contains nicotine, flavours and preservatives. This review focuses on the association of traditional and new forms of smoking with dry eye disease, contact lens wear and four other common ocular diseases: cataract, age-related macular degeneration, glaucoma and Graves' ophthalmopathy. It is concluded that smoking and vaping appear as a risk factor for the aforementioned ocular conditions. An evidence-based, clear link between cigarette smoking, or EC vaping and ocular problems is yet to be discovered.

Wireless Direct Microampere Current in Wound Healing: Clinical and Immunohistological Data from Two Single Case Reports.

Chronic pressure ulcers are hard-to-heal wounds that decrease the patient's quality of life. Wireless Micro Current Stimulation (WMCS) is an innovative, non-invasive, similar to electrode-based electrostimulation (ES) technology, that generates and transfers ions that are negatively-charged to the injured tissue, using accessible air gases as a transfer medium. WMCS is capable of generating similar tissue potentials, as electrode-based ES, for injured tissue. Here, through immunohistochemistry, we intended to characterize the induced tissue healing biological mechanisms that occur during WMCS therapy. Two single cases of bedridden due to serious stroke white men with chronic non-healing pressure ulcers have been treated with WMCS technology. WMCS suppresses inflammatory responses by decreasing the aggregation of granulocytes, followed by stimulating myofibroblastic activity and a new formation of collagen fibers, as depicted by immunohistochemistry. As a result, WMCS provides a special adjunct or stand-alone therapy choice for chronic and non-healing injuries, similar to electrode-based ES, but with added (i.e., contactless) benefits towards its establishment as a routine clinical wound healing regime.

Real-Time Assessment of E-Cigarettes and Conventional Cigarettes Emissions: Aerosol Size Distributions, Mass and Number Concentrations.

Cigarette smoke is a complex mixture of chemical compounds which are emitted during the processes of tobacco combustion. Electronic cigarettes (e-cigs) are expected to produce less harmful compounds due to the absence of tobacco leaf combustion. However, potential risks of the passive exposure to the aerosol exhaled by e-cig users have been raised in the last decade. In this study, the aerosols with diameter less than 1 µm (PM1) produced by vaping of various e-cig liquids were compared to those generated by smoking conventional cigarettes in real time. The mass and number concentration along with the number size distribution were measured in a closed room of 35 m3 volume. Our results showed that aerosols emitted from e-cig liquids had a different profile compared to those from conventional cigarettes. Although e-cigs initially produced higher particle mass and number concentrations, their emissions had much shorter lifetime of approximately 10-20 s, in comparison with the conventional and hand-rolling cigarette particulate emissions which had a dissipation time of approximately 1.4 h in a 35 m3 room. E-cigs emitted aerosols which volatilized rapidly, as they probably consisted almost only of propylene glycol and/or vegetable glycerin.

Evidence for association of STAT4 and IL12RB2 variants with Myasthenia gravis susceptibility: What is the effect on gene expression in thymus?

Myasthenia gravis (MG) is an autoimmune disease mediated by the presence of autoantibodies that bind mainly to the acetylcholine receptor (AChR) in the neuromuscular junction. In our case-control association study, we analyzed common variants located in genes of the IL12/STAT4 and IL10/STAT3 signaling pathways. A total of 175 sporadic MG patients of Greek descent, positively detected with anti-AChR autoantibodies and 84 ethnically-matched, healthy volunteers were enrolled in the study. Thymus samples were obtained from 16 non-MG individuals for relative gene expression analysis. The strongest signals of association were observed in the cases of rs6679356 between the late-onset MG patients and controls and rs7574865 between early-onset MG and controls. Our investigation of the correlation between the MG-associated variants and the expression levels of each gene in thymus did not result in significant differences.

Tobacco-specific nitrosamines: A literature review.

Cigarette smoke is a complex mixture of chemicals, including several tobacco-specific nitrosamines (TSNA). Most TSNA are formed in tobacco during the post-harvest period, while a number are produced when a cigarette is burned. Considerable evidence supports the role of TSNA important causative factors for cancers of the lung, pancreas, esophagus, and oral cavity in people who use tobacco products. Of the known TSNA, nicotine-derived nitrosamine ketone (NNK) and N-nitrosonornicotine (NNN) are the most carcinogenic. Other TSNA include N'-nitrosoanatabine (NAT) and N-nitrosoanabasine (NAB). New tobacco products (e.g., e-cigarettes) designed to attract consumers who are concerned about the health effects of tobacco have been appearing on the market. Several studies have reported that certain TSNA have been detected in the replacement liquids and vapour of e-cigarettes, but the levels are generally considerably lower than in tobacco cigarettes. Additionally, the FDA recently announced its intention to regulate TSNA in e-cigarettes, cigar tobacco and pipe tobacco. With the rise of new technologies for reducing the use of tobacco products-such as e-cigarettes- to evaluate exposure levels to these harmful chemicals over time, researchers will be monitoring levels of TSNA in the body as a result of the use of these devices.

Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice.

Fibroblast growth factor 21 (Fgf21) is a hormone with emerging beneficial roles in glucose and lipid homeostasis. The interest in Fgf21 as a potential antidiabetic drug and the factors that regulate its production and secretion is growing. Statins are the most widely prescribed drug for the treatment of dyslipidemia. However, the function of statins is not limited to the lowering of cholesterol as they are associated with pleiotropic actions such as antioxidant, anti-inflammatory and cytoprotective effects. The recently described effect of statins on mitochondrial function and the induction of Fgf21 by mitochondrial stress prompted us to investigate the effect of statin treatment on Fgf21 expression in the liver. To this end, C57BL6J male mice and primary mouse hepatocytes were treated with simvastatin, and Fgf21 expression was subsequently assessed by immunoblotting and quantitative real-time PCR. Hepatic Fgf21 protein and mRNA and circulating levels of FGF21significantly decreased in mice that had received simvastatin in their food (0.1% w/w) for 1 week. This effect was also observed with simvastatin doses as low as 0.01% w/w for 1 week or following 2 intraperitoneal injections within a single day. The reduction in Fgf21 mRNA levels was further verified in primary mouse hepatocytes, indicating that the effect of simvastatin is cell autonomous. In conclusion, simvastatin treatment reduced the circulating and hepatic Fgf21 levels and this effect warrants further investigation with reference to its role in metabolism.

VAV1 and BAFF, via NFκB pathway, are genetic risk factors for myasthenia gravis.

OBJECTIVE: To identify novel genetic loci that predispose to early-onset myasthenia gravis (EOMG) applying a two-stage association study, exploration, and replication strategy. METHODS: Thirty-four loci and one confirmation loci, human leukocyte antigen (HLA)-DRA, were selected as candidate genes by team members of groups involved in different research aspects of MG. In the exploration step, these candidate genes were genotyped in 384 EOMG and 384 matched controls and significant difference in allele frequency were found in eight genes. In the replication step, eight candidate genes and one confirmation loci were genotyped in 1177 EOMG patients and 814 controls, from nine European centres. RESULTS: ALLELE FREQUENCY DIFFERENCES WERE FOUND IN FOUR NOVEL LOCI: CD86, AKAP12, VAV1, B-cell activating factor (BAFF), and tumor necrosis factor-alpha (TNF-α), and these differences were consistent in all nine cohorts. Haplotype trend test supported the differences in allele frequencies between cases and controls. In addition, allele frequency difference in female versus male patients at HLA-DRA and TNF-α loci were observed. INTERPRETATION: The genetic associations to EOMG outside the HLA complex are novel and of interest as VAV1 is a key signal transducer essential for T- and B-cell activation, and BAFF is a cytokine that plays important roles in the proliferation and differentiation of B-cells. Moreover, we noted striking epistasis between the predisposing VAV1 and BAFF haplotypes; they conferred a greater risk in combination than alone. These, and CD86, share the same signaling pathway, namely nuclear factor-kappaB (NFκB), thus implicating dysregulation of proinflammatory signaling in predisposition to EOMG.

Recent advances in genetic predisposition of myasthenia gravis.

Myasthenia gravis (MG) is an autoimmune disease mediated by the presence of autoantibodies that bind to components of the neuromuscular junction, causing the symptoms of muscular weakness and fatigability. Like most autoimmune disorders, MG is a multifactorial, noninherited disease, though with an established genetic constituent. The heterogeneity observed in MG perplexes genetic analysis even more, as it occurs in various levels, including diverse autoantigens, thymus histopathology, and age at onset. In this context of distinct subgroups, a plethora of association studies, discussed in this review, have assessed the involvement of various HLA and non-HLA related loci in MG susceptibility, over the past five years. As expected, certain HLA alleles were strongly associated with MG. Many of the non-HLA genes, such as PTPN22 and CTLA-4, have been previously studied in MG and other autoimmune diseases and their association with MG has been reevaluated in more cohesive groups of patients. Moreover, novel risk or protective loci have been revealed, as in the case of TNIP1 and FOXP3. Although the majority of these results have been derived from candidate gene studies, the focal point of all recent genetic studies is the first genome-wide association study (GWAS) conducted on early-onset MG patients.

Population-specific documentation of pharmacogenomic markers and their allelic frequencies in FINDbase.

AIMS: Population and ethnic group-specific allele frequencies of pharmacogenomic markers are poorly documented and not systematically collected in structured data repositories. We developed the Frequency of Inherited Disorders Pharmacogenomics database (FINDbase-PGx), a separate module of the FINDbase, aiming to systematically document pharmacogenomic allele frequencies in various populations and ethnic groups worldwide. MATERIALS & METHODS: We critically collected and curated 214 scientific articles reporting pharmacogenomic markers allele frequencies in various populations and ethnic groups worldwide. Subsequently, in order to host the curated data, support data visualization and data mining, we developed a website application, utilizing Microsoft™ PivotViewer software. RESULTS: Curated allelic frequency data pertaining to 144 pharmacogenomic markers across 14 genes, representing approximately 87,000 individuals from 150 populations worldwide, are currently included in FINDbase-PGx. A user-friendly query interface allows for easy data querying, based on numerous content criteria, such as population, ethnic group, geographical region, gene, drug and rare allele frequency. CONCLUSION: FINDbase-PGx is a comprehensive database, which, unlike other pharmacogenomic knowledgebases, fulfills the much needed requirement to systematically document pharmacogenomic allelic frequencies in various populations and ethnic groups worldwide.