Project description:Chronic Fatigue Syndrome (CFS/ME) is a complex multisystem disease, with a pathogenesis that is undetermined. A large cohort of genes demonstrating altered expression in CFS/ME implicates the role of translational regulatory molecules, microRNA (miRNA), in the pathogenesis of this disease. We aimed to define the changes in microRNA expression in peripheral blood mononuclear cell (PBMC) samples in CFS/ME patients. miRNA expression was analysed in PBMC samples taken from CFS/ME patients and healthy controls, using the Ambion Bioarray V1. miRNA demonstrating differential expression were validated by qRT-PCR and analysed in an independent patient cohort in fractionated blood cell populations. The targets of miRNA hsa-miR-99b and hsa-miR-330-3p were then identified by gene expression analysis after transfection into primary NK cells.Microarray analysis identified differential expression of 34 miRNA, all of which were up-regulated. Four of the 34 miRNA had confirmed expression changes by qRT-PCR. Fractionating PBMC samples by cell type from an independent patient cohort identified changes in miRNA expression in NK-cells, B-cells and monocytes with the most significant abnormalities occurring in NK cells. Transfecting primary NK cells with hsa-miR-99b and hsa-miR-330-3p, respectively, resulted in gene expression changes consistent with NK cell activation and diminished cytotoxicity.This study demonstrates altered microRNA expression in the peripheral blood mononuclear cells of CFS/ME patients, which are potential diagnostic biomarkers. The greatest degree of miRNA deregulation was identified in NK cells with targets consistent with cellular activation and altered effector function.
Project description:We extended the mathematical models of measuring biodiversity to estimate DNA methylation heterogeneity in a cell population. We propose a model-based approach (abundance-based, phylogeny-based and pairwise similarity-based heterogeneity) and consider similarity in DNA methylation patterns from individual cells to evaluate heterogeneity that overcomes biases due to missing data. We also applied commonly used non-model based method (methylation entropy) and other reported methods of estimating methylation heterogeneity such as single-cell based approach to evaluate methylation heterogeineity.
Project description:The gene expression in vascular endothelial cells (VECs) and circulating fibrocytes (CFs) was tested either culturing alone or co-cultured. Our previous study showed that CFs inhibit both proliferation and apoptosis of VECs. In this present study, we co-cultured CFs and VECs in Transwell and tested the gene expression in CFs and VECs in order to delight the mechanism under which CFs affect the proliferation and apoptosis of VECs.
Project description:Examination of DNA methylome patterns in a larger cohort of ME/CFS samples using the Illumina Infinium HumanMethylation450 Beadchip Array
Project description:This study compared whole blood gene expression in CFS adolescent and healthy controls, and explored associations between gene expression and neuroendocrine markers, immune markers and clinical markers within the CFS group.
Project description:Autoimmunity is an established clinical feature in both Myalgic encephalomyelitis / chronic fatigue syndrome (ME/CFS) and Post-Acute Sequelae of COVID (PASC). Passive transfer of immunoglobulins from patients' sera into mice induces some clinical features of PASC. IgG-induced transfer of disease phenotypes has long been appreciated, yet the exact mechanism of disease development remains largely elusive. Here, we show that IgG isolated from ME/CFS patients selectively induces mitochondrial fragmentation in primary human endothelial cells and alters mitochondrial energetics. Blocking IgG entry into cells using an Fc blocker inhibits the mitochondrial fragmentation phenotype. Both the digested Fab and Fc regions of IgG separately entered the endothelial cells. However, only the Fab fragment alone was able to alter the mitochondrial energetics, similar to native IgG. Proteomics analysis of IgG-bound immune complex revealed significant and specific changes within the immune complex of ME/CFS and PASC patients compared to healthy controls and other disease controls. We demonstrate that IgGs from ME/CFS patients carry a chronic protective response signal that promotes mitochondrial adaptation to stress through mitochondrial fragmentation, without altering mitochondrial ATP generation capacity in endothelial cells. Our results suggest that chronic and pathogenic IgG-induced mitophagy and metabolic alterations to stress-dependent ATP production may lead to cellular saturation, ultimately affecting mitochondrial health. These results provide an opportunity to develop alternative therapeutic intervention strategies.
Project description:Post-infectious Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a chronic disease with a yet unresolved pathophysiology and sparse diagnostic options. Extracellular vesicles (EVs) serve as carriers of disease-specific protein and microRNA (miRNA) signatures. We aim to identify novel miRNA markers as potential biomarker in isolated plasma EVs from female post-COVID-19 ME/CFS patients in comparison to healthy controls (HCs). EVs were isolated from plasma of post-infectious ME/CFS patients and HCs by size exclusion chromatography, and characterized for their number, size, morphology and surface marker expression. Small RNA sequencing identifed four small RNAs, which are significantly differential expressed between ME/CFS patients and healthy donors. Subsequently performed qPCR studies revealed significant downregulation exclusively for the hsa-let-7b-5p miRNA in EVs from ME/CFS patients versus HCs. Notably, reduced hsa-let-7b-5p expression correlated with impaired physical functioning as well as increased fatigue, pain, and immune activation scores. Our data indicate, that hsa-let-7b-5p could serve as a promising candidate for development as diagnostic and stratification marker of ME/CFS.