Th130 - SH2B3 Risk Allotype Is Associated with Increased Vascular Inflammation in Induced Pluripotent Stem Cells from Type 1 Diabetes Donors

Todd Brusko – Professor, Pathology, Immunology, and Laboratory Medicine, University of Florida; Clayton Mathews – Professor, Pathology, Immunology, and Laboratory Medicine, University of Florida; Amy Meacham – University of Florida

Abstract Text: Genome wide association studies have identified a single nucleotide polymorphism within exon 3 of SH2B3 (rs3184504 (C784T)) as a genetic element associated with increased risk for Type 1 Diabetes (T1D). SH2B3 encodes lymphocyte adaptor protein (LNK), a potent negative regulator of inflammatory signaling in leukocytes and endothelial cells. rs3184504 results in a non-synonymous change, R262W, with LNK262W associated with elevated disease risk. We hypothesize that LNK262W fails to restrain inflammatory signaling leading to increased vascular inflammation contributing to the development of T1D. Induced pluripotent stem cells (iPSC) from donors with T1D (n=4) were edited using CRIPSR/Cas9 at rs3184504 (to 784C (LNK262R) or 784T (LNK262W risk)) and differentiated into endothelial cells (iEC). A single donor gene-edited iEC was treated with TNFα in static culture and the expression of adhesion molecules assessed by flow cytometry. At 500U/mL for 4 hours, LNK262W (risk) iEC exhibited elevated adhesion molecule expression necessary for T cell transendothelial migration compared to common allotype (LNK262R) expressing HUVEC (CD54 = 3.88 fold-change; CD106 = 3.19 fold-change p=0.0032) and iEC (CD54 & CD106 = 1.65 fold-changes each). These results demonstrate that LNK262W fails to restrain inflammatory signaling in EC, which may contribute to the resulting mobilization of autoreactive T cells into the islets of Langerhans to promote beta cell destruction. Follow-up studies involve longer more physiologically relevant treatment times, and results from these studies will contribute to our understanding of how the risk allotype of LNK262W influences mechanistic disease progression and clinical outcomes by regulating vascular inflammatory functions and phenotypes.