Assessment of Immune and Molecular Changes in T Lymphocytes, Natural Killer Cells and Vasoactive Neuropeptides in Chronic Fatigue Syndrome

Abstract

Immune dysregulation due to infection, inflammation or compromises to immune and molecular mechanisms can have detrimental effects on normal physiological functions. Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is a disorder of unknown mechanism where diagnosis is often delayed and evidence-based effective treatments are lacking. CFS/ME is characterised by severe fatigue, flu-like symptoms, pain and cognitive disturbances. To date a mechanism explaining the relationship between these symptoms and the physiological irregularities presented by patients remains obscure. As such, CFS/ME patients spend a considerable amount of money on health service cost in pursuit of the most effective treatment to alleviate their symptoms. An important initiative towards better management of CFS/ME is the development of accurate diagnosis. The principal aim of this research is to identify feasible biomarkers that can be used in effective diagnosis of CFS/ME. This research examines the status of lymphocytes in CFS/ME with a specific focus on their cellular function including apoptosis, protein secretion, receptor expression and gene expression. It is hypothesized that irregularities in these cellular processes contribute to the mechanism of CFS/ME and thus may form a suite of diagnostic markers for assessing CFS/ME patients. Participants for the study comprised 95 CFS/ME patients and 50 non-fatigued controls at baseline. Data for 50 CFS/ME and 27 non-fatigued controls was available for the follow up study at 6 and 12 months. Participants were aged between 25-65 years. The criteria for inclusion were based on the Centre for Disease Prevention and Control (CDC) 1994 clinical diagnostic criteria. Whole blood was collected from each participant at baseline, at 6 months and at 12 months. Flow cytometric protocols were employed in examining measures of cytotoxic activity in CD8+T and Natural Killer (NK) cells, levels of CD56brightCD16negative NK cells CD56dimCD16positive NK phenotypes, CD4+T helper cytokine secretion and levels of Foxp3 and vasoactive neuropeptide receptor (VPACR2). The expression pattern of cytotoxic related genes including granzyme A (GZMA), granzyme K (GZMK), perforin (PRF1) and interferon-gamma (IFN-G) and the expression of microRNA (miRNA) molecules in NK and CD8+T cells were investigated. At baseline, compared to the non-fatigued controls, CFS/ME patients exhibited significant decreases in cytotoxic activity of NK and CD8+T cells, levels of CD56brightCD16negative NK cells and repression of IFN-G, GZMA and GZMK gene expression. At the same time, significant increases in cytokines, IL-10, IFN-γ and TNF-α, FOXP3, VPACR2 and PRF1expression were noticed in the CFS/ME patients compared to controls. At 6 months follow up, significant reductions in NK activity, CD56brightCD16negative NK cells, IL-10 and IL-17A were observed in the CFS/ME patients compared to the non-fatigued controls. At 12 months, in contrast to the nonfatigued controls, CFS/ME patients continued to demonstrate significant decreases in NK activity and significant increases in only IL-2. Assessment of miRNA expression revealed a significant down-regulation of a number of miRNAs in CFS/ME patients in comparison to the non-fatigued controls, specifically, miR-21, miR-146a, miR-223, miR-17-5p, miR-103, miR-106, miR-10a, miR-191 and miR-152 were significantly down-regulated mainly in the CFS/ME patients in comparison to the controls. In conclusion, the results from this study have elucidated the extent of decreases in cytotoxic activity in CFS/ME. In addition, this study has identified unique immune related processes and molecules that are compromised in CFS/ME patients. These novel parameters may have important implications in the development of biomarkers for CFS/ME. Moreover the consistent decrease in cytotoxic activity and NK phenotypes over the 12 month period strongly supports their usefulness as biomarkers for diagnosing CFS/ME. These biomarkers if implemented in the clinical setting could potentially assist in improving diagnosis and demonstrating a clear pathomechanism for CFS/ME. Eventually, this may result in the development of better therapeutic and treatment strategies for managing CFS/ME.

Date of Award	9 Jun 2012
Original language	English
Supervisor	Sonya Marshall-Gradisnik (Supervisor) & Kevin John Ashton (Supervisor)