Abstract
Abstract IL-17A secreting T cells represent a distinct lineage of CD4+ T effector cells (Th17) that have been suggested to play a role in the pathogenesis of inflammatory/autoimmune diseases such as Crohn’s disease. Precision for Medicine cryopreserves primary Th17 cells (AccuCell®) to provide an easy alternative to isolating fresh cells, for use in screening assays and functional immunological studies. The human MDR1 gene encodes a membrane protein, P glycoprotein (Pgp), whose function is to export toxic substances or metabolites from the cell. MDR1 expresssion has been demonstrated in tissues of Crohn’s disease patients and on their circulating Th17 cells. Using AccuCell® Th17 cells from healthy and diseased patients we investigate how MDR1 expression differs in the disease state and highlight the challenges of screening drugs using healthy cells as opposed to the targeted disease state. First we evaluated our cryopreserved Th17 cells to fresh isolations for surface marker, chemokines, intracellular cytokines (IL-17A, IFNγ, IL-22), and RORγt. Full retention of the tested markers were observed in the recovered cryopreserved cells. On our disease state PBMCs phenotypic and functional evaluation of MDR1+ Th17/17.1 cells included activation markers CD5, CD69, CD2, human leukocyte antigen-DR (HLA-DR) and co-stimulatory receptor CD28, compared to normal healthy donors. T-cell receptor dependent proliferation in MDR1+ Th17 cells was compared to healthy donor demographic matched Th17 cells. Utilizing AccuCell® cryopreserved Th17 cells which are fully functional empowered these studies to elucidate mechanisms as discussed above; thus eliminating fresh isolations so researchers can focus on studying the pathogenesis of these cells.
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