טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentSucher Mamrut Shimrat
SubjectEpigenetic Mechanisms in Autoimmune Diseases
DepartmentDepartment of Medicine
Supervisor Professor Ariel Miller
Full Thesis textFull thesis text - English Version


Abstract

Autoimmune diseases are the world’s second leading cause of chronic illness. Their complex etiology arises from both genetic and environmental factors. Epigenetic changes, such as DNA methylation, are considered to link genetics and environment, thus methylation changes could mark disease-altered gene function. Monozygotic twins, discordant and concordant for a disease, provide a unique opportunity to identify genetic and epigenetic disease- related factors.

We hypothesized that analyzing the methylome and transcriptome of immune cell-subsets from monozygotic twins, will identify genes involved in Myasthenia Gravis (MG), a rare autoimmune disease. The project includes two parts: A- Establishing methylome and transcriptome reference maps of B cells, monocytes, CD4 and CD8 T cells from healthy controls. B- MG-associated methylatome and transcriptome patterns in immune cells from MG discordant and concordant monozygotic twins.

In part A, methylome and transcriptome of 4 immune cell-subsets from healthy females were characterized. Due to known methylation gender bias and autoimmune disease prevalence, we compared our female datasets with public available male datasets. Subset-specific DNA methylated regions (DMRs) were identified, highly similar between genders, mainly located distant from CpG-islands. Many subset-specific DMRs overlapped enhancer elements, suggesting their role in immune cell regulation. Gender-specific DMRs shared among subsets were identified, many on autosomal chromosomes, over-represented in CpG-islands, pointing different regulatory mechanisms of gender versus subset-specific methylation.

In part B, transcriptome and methylome of 5 immune cell-subsets from 7 monozygotic twins discordant and concordant for MG, MG singletons and healthy controls were analyzed. While very few differentially expressed genes (DEGs) and DMRs were detected between the discordant twins, 134 DEGs and over 1800 DMRs were identified between controls and MG patients in monocytes, a major innate immune system player. 60% of selected DEGs and 42% of selected DMRs were successfully validated by different methods between MG patients and controls. Pathway analysis revealed that 6 validated DEGs are connected to the myeloid cells triggering receptor (TREM-1), an important innate inflammation mediator, suggesting monocytes have impaired function in MG. Cluster analysis of monocyte MG-associated DMRs and DEGs placed the healthy discordant twins within the MG patients group, which includes also unrelated MG patients, indicating that MG-associated genetic profile is present, despite no clinical phenotype.

Transcriptome analysis revealed also numerous B cell and Treg DEGs between MG patients and controls, some also differing between the discordant twins. Due to the recognized role of these subsets in MG, these DEGs represent a highly interesting candidates list for further disease cause studies.

We also identified MG-associated DEGs shared by B cells, CD4 and CD8 T cells. We successfully validated 20% of the selected shared DEGs using PBMC samples, which are the most accessible material for disease biomarker detection. These genes may be valuable for future disease diagnosis.

In summary, this study is the first report characterizing DNA methylation profiles in MG using a unique twins group. Novel MG associated genomic and methylomic profiles were identified and a possible predisposition to MG among discordant twins was detected. These findings provide potential disease biomarkers, and future MG therapy targets.