Exploring potential biomarkers for ME/CFS

Abstract

Myalgic Encephalomyelitis (ME) is a complex disease which affects many different body systems. Disturbance of the immune system, brain and central nervous systems cardiac function, dysregulation of metabolism generally exhibit as post exertional malaise are just a few of the many examples of how this disease can affect an individual. So far, molecular biomarkers have been identified then can diagnose a patient with accuracy. The two purposes of this study are: (1) to investigate differences in cytokines in two different independent cohorts of ME/CFS patients and gender-and-aged-matched controls as a measure of the disturbance of the immune system, any difference shown would be potential hypotheses that could be followed up in larger cohorts. (2) to develop tools and test them in a model cell culture system mimicking ME/CFS. These tools can then be applied to ME/CFS patient samples. Two independent cohorts in New Zealand were studied. The first was 10 ME/CFS patients and 10 gender-and-aged controls in a Dunedin pilot study. The second was a Palmerston North cohort targeting post exertional malaise of 9 ME/CFS patients 9 gender-and-aged-matched controls and 5 Multiple sclerosis (MS) patients. Results of the cytokine analysis of patients and controls in the initial Dunedin Pilot indicated two statistically significant downregulated cytokines IL-13 and IL-7 in ME/CFS patients compared with controls and IP-10 and VEGF were found to be near to significance. The Palmerston North exercise study four cytokines were found to be differentially regulated in ME/CFS patients compared with controls IL-4, IL-9, IL-13 and PDGFBB. The two cohorts were then combined for analysis to give greater numbers to the study, and IL-7, IL-13, IP-10 and VEGF were found to be significant, closely mimicking the initial Dunedin pilot study. Analysing the aspect of post exertional malaise was done through a paired T test before and after exercise and 6 cytokines in controls showed significant change. Neither disease ME/CFS and MS followed this trend. The connection between cytokine change and immune dysfunction was tested with tools developed to measure change in the phosphorylation status of a key protein eIF2α that determines the fate of the immune cell. A cell culture model was set up to model this, and many variations of transfections were performed to test for over expression. Antibodies against eIF2α were produced and purified by both negative and positive selection to detect phosphorylation in Western blots. The antibody was effective in detecting increased eIF2α phosphorylation seen in both neutrophils and lymphocytes in protein extractions from a ME/CFS patient and gender-and-aged matched control. The outcomes from both pilot studies, showed evidence of dysregulation in some cytokines between ME/CFS patients and their gender-age-matched controls. When an exercise component is introduced ME/CFS patient’s cytokines act very differently to that seen in controls. In summary, disturbance in plasma cytokines, were measured directly and inhibited of protein synthesis in immune cell individually by increased phosphorylation of eIF2α. These observations should be followed up with a larger cohort of patients and focussed study design.