1511-P: Identifying How Endothelial Protease-Activated Receptors Control Insulin Signaling—Implications for Diabetes

Endothelial cell (EC) dysfunction occurs in diabetes. Thrombin, a circulating serine protease with increased activity in diabetics, signals through protease-activated receptors 1 and 4 (PAR1/PAR4). To determine EC-specific roles of PARs in diabetes, we generated Par1^fl//fl; Par4^fl/fl; iCdh5 (PAC)-Cre^ERT2 (Par1/4^dECko) mice and challenged them with streptozotocin (STZ) to induce hyperglycemia. STZ-treated Par1/4^dECko mice had reduced blood glucose levels, weighed significantly more, and lost less abdominal fat than control littermates. Neither Par4^ECko nor Par1^ECko mice phenocopied Par1/4^ECko mutants, suggesting that loss of both thrombin receptors is required to drive these phenotypes. Notably, STZ-treated Par1/4^dECko mice showed comparable insulin levels to their STZ-treated littermates, suggesting mutant mice had heightened insulin sensitivity, which was verified by insulin/glucose tolerance testing. Using primary human ECs that express PAR1 but not PAR4, we found that PAR1/4 depletion increased basal insulin receptor (IR) activity as measured by the uptake of a glucose-analog. PAR1/4-depleted ECs also showed increased insulin receptor Type-A (IR-A) spliceform expression, which has high insulin-independent basal activity. Conversely, treatment of ECs with a PAR1 ligand upregulated the insulin-dependent IR Type-B (IR-B) spliceform, suggesting that endothelial PAR1/4 can modulate IR splicing/activity. Additionally, we found that ECs in many wildtype mouse organs, including skeletal muscle, predominantly express IR-B in vivo. Therefore, a switch to IR-A expression in Par1/4^dECko mice may enhance endothelial IR activity. IR activity in ECs regulates insulin transport from the bloodstream to the parenchyma. We propose that loss of EC PAR1/4 elevates whole-body insulin sensitivity by increasing IR splicing/activity and subsequent transcytosis to the parenchyma. Thus, EC PAR1/4 modulation may hold promise for treating diabetes.