Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a complex and disabling multi-system condition without a known cure. It is characterized by an abrupt or delayed onset of persistent and relapsing symptoms, notably severe fatigue that impedes daily activities. Despite ongoing research, the exact cause of ME/CFS remains elusive. Numerous studies have utilized peripheral blood mononuclear cells (PMBCs) from ME/CFS patients to identify features of autoimmune illness and mitochondrial dysfunction. Thus, there is a need to explore innovative approaches designed to better understand the contributions of immune and metabolic dysfunctions in ME/CFS, potentially paving the way for the development of targeted therapies. One such approach involves bone marrow mesenchymal stromal cell (BMMSC)-derived exosomes (exos), which are vesicles secreted by cells to facilitate effective cell-cell communication. Secreted exos deliver nucleic acids and proteins to adjacent cells, thereby modulating target cell signaling. Such paracrine factors, to a significant extent, capture the therapeutic effects of BMMSCs, an effect observed previously in multiple animal disease models; both RNAs and proteins from exos play roles in regulating processes such as cell survival, differentiation, and immunomodulation. Exos exhibit the ability to suppress the expansion and maturation of activated immune cells, inhibit their functional differentiation, and preserve regulatory T cells in vivo. This immuno-regulatory effect of exos is particularly useful in examining the contribution of immune cell dysfunction to the progression of ME/CFS. Additionally, numerous studies have highlighted the central role of exos in promoting mitochondrial function, leading to an increase in ATP production. These properties render exos an appealing probe for resetting cellular signaling in multi-system conditions such as ME/CFS. Thus, we propose the following two aims: Aim 1) Investigate the effects of exos on specific immune cell subsets using PBMCs isolated from individuals with ME/CFS. PBMCs obtained from ME/CFS patients will undergo co-culturing with exos, and the stimulation of T, B, and NK cells in their presence. Assessment will include surface markers indicative of cell subtype activation, along with the measurement of cell proliferation and apoptosis. Aim 2) Examine the impact of exos on selected mRNA expression and mitochondrial function of PBMCs. Leveraging previously conducted RNA sequencing of PBMCs from ME/CFS subjects, we will design a custom panel using NanoString nCounter technology for selected mRNA molecules. This panel will predominantly consist of genes associated with immune response and metabolism, as determined in our dataset. Additionally, we will assess the mitochondrial function of PBMCs in the presence of exos. Our research proposal is innovative, holding the potential to provide information regarding the underlying pathology of ME/CFS and unveil novel molecular and cellular therapeutic targets using exos as immunomodulators and ME/CFS patient samples.

NARRATIVE The availability of the bio-bank and data-bank from the longitudinal good day/bad day study of ME/CFS at the Nova Southeastern University and the collaboration with Nancy Klimas and her genomics group presents an opportunity to perform ex-vivo testing of a cell-based therapy on the some key drivers of the disease. The proposed project offers the opportunity to conduct a study of effects that mesenchymal stromal cell-derived exosomes may have on ME/CFS using viable, cryopreserved peripheral blood mononuclear cells. Both mesenchymal stromal cells and their exosomes have been shown to possess immunomodulatory and regenerative properties and we propose to utilize both cells and exosomes as probes to examine their effect(s) on aberrant inflammatory and metabolic responses observed in ME/CFS.