Abstract 36: Application of “Omics” to Assess the Inter Passage Variability in the Immunoprivilege of Mesenchymal Stem Cells

Abstract

Introduction: Bone marrow derived allogeneic mesenchymal stem cells (MSCs) from young healthy donors are immunoprivileged. However recent data from preclinical and clinical studies indicate that allogeneic MSCs provoke immune response in the recipient heart and are rejected after transplantation. We recently observed that stem cell surface oxidized phospholipids (ox-PCs) modulate allogeneic immune cell response. In most of the allogeneic MSCs based clinical trials in cardiac patients, cells from a single donor were expanded in cultures and transplanted. However, the effect of increase in the passage number on stem cell membrane lipidome and its association with immunogenicity of MSCs has not been investigated yet, the present study was carried out to address this Methods: MSCs isolated from Wistar rats were allowed to grow for different passages namely Passage 3, 5 and 7 (P 3, 5 and 7). The immunoprivilege of MSCs was assessed by mixed leukocyte reaction assay. Briefly, MSCs were co-cultured with mixed leukocytes derived from SD rats for 72 h. Leukocyte mediated cytotoxicity (lactate dehydrogenase release) and apoptosis (Bax/Bcl-xl ratio) in MSCs was assessed after the co-culture. Furthermore, MSCs at different passages were analyzed by LC/MS/MS -“lipidomic platform” for cell surface ox-PCs levels and “proteomic platform” for intracellular proteins. Results and conclusions: We observed a decrease in lymphocyte mediated cytotoxicity and Bax/ Bcl-xl ratio in MSCs at P5 and7 compared to P3. The lipidome analysis showed that the levels of immunomodulatory ox-PCs increased in MSCs at P5 and P7. Inhibition of ox-PCs was associated with the loss of immunoprivilege of MSCs. To find if the ox-PCs were acting in tandem with downstream intracellular protein alterations, we performed proteome analysis. There were significant changes in proteins related to intracellular immune response, lipid metabolism and mitochondrial dynamics. In depth analysis of the ‘omics’ data is now going on to pinpoint associations between cell surface ox-PCs and the candidate intracellular proteins involved in immune modulation. To the best of our knowledge this is the first study integrating membrane lipidomic with intracellular proteins to assess immunogenicity of MSCs.