Nili Avidan

Genetic basis of myasthenia gravis – A comprehensive review

Myasthenia gravis (MG) is a rare autoimmune disease characterized by the production of autoantibodies against proteins of the postsynaptic membrane in the neuromuscular junction. The estimated number of MG patients is steadily increasing, and it had more than doubled in the last 20 years. Monozygotic MG twin concordance is estimated to be about 35% supporting the central role of environmental factors in MG etiology. Epigenetics, presume to be the mechanistic link between environmental and genetic risk factors in disease development, provides support for specific microRNAs associated with MG. Genetic studies have mainly pointed at specific HLA alleles implicated in MG susceptibility, however recently both TNFAIP3-interacting protein 1 (TNIP1) and tyrosine phosphatase non-receptor 22 (PTPN22) were indicated to be associated with MG in a GWAS study. A gender bias was observed for SNPs in the HLA-locus, suggesting female-specific alleles have an increase risk for MG. Moreover, sex hormones play a pivotal role in the gender bias in autoimmunity in general and in MG in particular. Hence the genetic basis of gender bias might be highly pertinent to MG and deserves further characterization. Pathway-based analyses that combine information across multiple genes into a limited number of molecular networks have been found to be a powerful approach. Both regulatory T-cell (Treg) differentiation and NF-κB signaling pathway have been shown to have relevance to MG pathophysiology. Hence studies centered around two pathways might be a fruitful approach to identify additional polymorphisms associated with myasthenia gravis.

Insulin-Like Growth Factor-Binding Protein 7 Regulates Keratinocyte Proliferation, Differentiation and Apoptosis

Insulin-like growth factor (IGF)-binding protein 7 (IGFBP7) belongs to the IGFBP superfamily, which is involved in the regulation of IGF and insulin signaling. Recently, a global gene expression study revealed that IGFBP7 is downregulated in the psoriatic epidermis, with UVB phototherapy restoring its expression to normal. In the present study, we confirmed that IGFBP7 expression is decreased in psoriatic lesions. Given the previous data suggesting a role for IGFBP7 in the control of cancer cell growth, we investigated its involvement in the regulation of keratinocyte (KC) proliferation and differentiation, which are abnormal in psoriasis. To model IGFBP7 downregulation in vitro, we used IGFBP7-specific small interfering RNA or small hairpin RNA-expressing lentiviral vectors in HaCaT cells or primary human KCs. Downregulation of IGFBP7 was found to markedly enhance KC proliferation in both systems, was associated with a significant decrease in KC susceptibility to tumor necrosis factor-alpha-induced apoptosis, but did not affect senescence. Downregulation of IGFBP7 was also shown to block expression of genes associated with calcium-induced differentiation of human KCs. Finally, recombinant IGFBP7 was found to inhibit KC proliferation and enhanced their apoptosis. These data position IGFBP7 as a regulator of KC proliferation and differentiation, suggesting a potential role for this protein in the pathophysiology and treatment of hyperproliferative dermatoses such as psoriasis.

Interferon-beta induces distinct gene expression response patterns in human monocytes versus T cells.

Background Monocytes, which are key players in innate immunity, are outnumbered by neutrophils and lymphocytes among peripheral white blood cells. The cytokine interferon-β (IFN-β) is widely used as an immunomodulatory drug for multiple sclerosis and its functional pathways in peripheral blood mononuclear cells (PBMCs) have been previously described. The aim of the present study was to identify novel, cell-specific IFN-β functions and pathways in tumor necrosis factor (TNF)-α-activated monocytes that may have been missed in studies using PBMCs. Methodology/Principal Findings Whole genome gene expression profiles of human monocytes and T cells were compared following in vitro priming to TNF-α and overnight exposure to IFN-β. Statistical analyses of the gene expression data revealed a cell-type-specific change of 699 transcripts, 667 monocyte-specific transcripts, 21 T cell-specific transcripts and 11 transcripts with either a difference in the response direction or a difference in the magnitude of response. RT-PCR revealed a set of differentially expressed genes (DEGs), exhibiting responses to IFN-β that are modulated by TNF-α in monocytes, such as RIPK2 and CD83, but not in T cells or PBMCs. Known IFN-β promoter response elements, such as ISRE, were enriched in T cell DEGs but not in monocyte DEGs. The overall directionality of the gene expression regulation by IFN-β was different in T cells and monocytes, with up-regulation more prevalent in T cells, and a similar extent of up and down-regulation recorded in monocytes. Conclusions By focusing on the response of distinct cell types and by evaluating the combined effects of two cytokines with pro and anti-inflammatory activities, we were able to present two new findings First, new IFN-β response pathways and genes, some of which were monocytes specific; second, a cell-specific modulation of the IFN-β response transcriptome by TNF-α.

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

To identify novel genetic loci that predispose to early‐onset myasthenia gravis (EOMG) applying a two‐stage association study, exploration, and replication strategy.

Integrative analysis of methylome and transcriptome in human blood identifies extensive sex- and immune cell-specific differentially methylated regions

The relationship between DNA methylation and gene expression is complex and elusive. To further elucidate these relations, we performed an integrative analysis of the methylome and transcriptome of 4 circulating immune cell subsets (B cells, monocytes, CD4+, and CD8+ T cells) from healthy females. Additionally, in light of the known sex bias in the prevalence of several immune-mediated diseases, the female datasets were compared with similar public available male data sets. Immune cell-specific differentially methylated regions (DMRs) were found to be highly similar between sexes, with an average correlation coefficient of 0.82; however, numerous sex-specific DMRs, shared by the cell subsets, were identified, mainly on autosomal chromosomes. This provides a list of highly interesting candidate genes to be studied in disorders with sexual dimorphism, such as autoimmune diseases. Immune cell-specific DMRs were mainly located in the gene body and intergenic region, distant from CpG islands but overlapping with enhancer elements, indicating that distal regulatory elements are important in immune cell specificity. In contrast, sex-specific DMRs were overrepresented in CpG islands, suggesting that the epigenetic regulatory mechanisms of sex and immune cell specificity may differ. Both positive and, more frequently, negative correlations between subset-specific expression and methylation were observed, and cell-specific DMRs of both interactions were associated with similar biological pathways, while sex-specific DMRs were linked to networks of early development or estrogen receptor and immune-related molecules. Our findings of immune cell- and sex-specific methylome and transcriptome profiles provide novel insight on their complex regulatory interactions and may particularly contribute to research of immune-mediated diseases.

Methylome and transcriptome profiling in Myasthenia Gravis monozygotic twins

OBJECTIVE To identify novel genetic and epigenetic factors associated with Myasthenia gravis (MG) using an identical twins experimental study design. METHODS The transcriptome and methylome of peripheral monocytes were compared between monozygotic (MZ) twins discordant and concordant for MG, as well as with MG singletons and healthy controls, all females. Sets of differentially expressed genes and differentially methylated CpGs were validated using RT-PCR for expression and target bisulfite sequencing for methylation on additional samples. RESULTS >100 differentially expressed genes and ∼1800 differentially methylated CpGs were detected in peripheral monocytes between MG patients and controls. Several transcripts associated with immune homeostasis and inflammation resolution were reduced in MG patients. Only a relatively few genes differed between the discordant healthy and MG co-twins, and both their expression and methylation profiles demonstrated very high similarity. INTERPRETATION This is the first study to characterize the DNA methylation profile in MG, and the expression profile of immune cells in MZ twins with MG. Results suggest that numerous small changes in gene expression or methylation might together contribute to disease. Impaired monocyte function in MG and decreased expression of genes associated with inflammation resolution could contribute to the chronicity of the disease. Findings may serve as potential new predictive biomarkers for disease and disease activity, as well as potential future targets for therapy development. The high similarity between the healthy and the MG discordant twins, suggests that a molecular signature might precede a clinical phenotype, and that genetic predisposition may have a stronger contribution to disease than previously assumed.

Personalized Medicine and Theranostics: Applications to Multiple Sclerosis

Tailored therapeutics based on diagnostic tests—theranostics—is shaping future health care. We describe recent progress in the search for biomarkers for multiple sclerosis (MS) related to drug response. Robust and clinically validated predictive biomarkers for support of treatment optimization are of increased need, especially as the treatment options increase, to improve efficacy and safety as part of tailored therapeutics. Pharmacogenomic studies in MS were mainly focused on biomarkers for interferon beta response, and none of the markers has translated into clinical practice. The antibody-based tests for John Cunningham virus have been clinically validated and are implemented in clinical practice to aid care for natalizumab-treated patients. Participatory medicine empowers patients to be actively involved in their care by tailored medical information, according to their needs and expectations. It can increase adherence and, together with the biomarkers in development, contribute to improved health care of the patient with MS.Abstract Tailored therapeutics based on diagnostic tests—theranostics—is shaping future health care. We describe recent progress in the search for biomarkers for multiple sclerosis (MS) related to drug response. Robust and clinically validated predictive biomarkers for support of treatment optimization are of increased need, especially as the treatment options increase, to improve efficacy and safety as part of tailored therapeutics. Pharmacogenomic studies in MS were mainly focused on biomarkers for interferon beta response, and none of the markers has translated into clinical practice. The antibody-based tests for John Cunningham virus have been clinically validated and are implemented in clinical practice to aid care for natalizumab-treated patients. Participatory medicine empowers patients to be actively involved in their care by tailored medical information, according to their needs and expectations. It can increase adherence and, together with the biomarkers in development, contribute to improved health care of the patient with MS.

Integrative analysis of DNA methylation and gene expression identifies distinct profiles among immune cells subsets

The immune network is a complex system for host defenses comprising various types of inflammatory and regulatory cells. Autoimmune diseases occur because of dysregulation in homeostasis caused by imbalance of these cells. The pattern of imbalance depends on the disease. Most autoimmune diseases are chronic, and the mechanism underlying this chronic nature is yet to be determined. Monoclonal antibody therapy is highly specific to the molecules targeted and is therefore highly effective. However, this therapy cannot be applied to all autoimmune diseases, and even the most effective therapy is incapable of completely inhibiting disease activity. Antigen-specific therapies have the ability to inhibit disease activity; however, their application is limited because of the presence of various disease-specific antigens. Regulatory cell therapies also have potential, but pose a plasticity problem. By focusing our research on experimental autoimmune encephalomyelitis (EAE), which is a multiple sclerosis (MS) model and normally initially has a relapse or remission course, followed by a progressive course, we can develop alternative therapies for the treatment of autoimmune diseases by exploring the mechanism of relapse and remission. From the phenomena that immunizing peptide itself determines the difference between monophasic-EAE and relapsingEAE, we found out that two-correlated non-genetic factors can shut down the reactivation in the EAE. One is kinetics and the component of CD4CD25 regulatory T cells (Treg), which are characterized by their later expansion after emerging of encephalitogenic T cells and by their subset expressing both CD69 and CD103 (=DP-Treg). DP-Treg is the most efficacious subset showing the highest Treg-compatible signatures with high antigen-specificity exerting stability due to downregulation of IL-6R expression, nevertheless with activated effector phenotype. The other is a hierarchy of encephalitogenic peptide itself to be immunized, which means that the more dominant peptide can develop acute EAE, the less relapse and re-induction of EAE occurs. This superiority of dominant peptide is characterized by specificity to itself, that is, it can suppress response to other peptides broadly and establish permanent remission. Peptide tolerance induced orally or intravenously could not get such wide suppression. Taken together, efficient induction of DP-Treg is correlated with superiority of self-peptide itself, suggesting ‘immunological homunculus’. This can be applied for treatment such as tissue-specific inverse vaccination for autoimmune diseases.