Abstract 2040: Rapamycin Extends Life Of Matrix Gla Protein Knockout Mice Independent Of Vascular Calcification Content.

Abstract

Background: Peripheral artery disease (PAD) is characterized by the narrowing and blockage of the vessels in lower extremities. While atherosclerosis has been the assumed culprit of PAD recent data now shows that medial arterial calcification (MAC) is the cause of below-the knee ischemia. MAC promotes thrombosis via inward remodeling, reduced elasticity, and arterial stiffness. The mechanisms driving MAC pathogenesis are unclear. Matrix Gla Protein (MGP) is an inhibitor of vascular calcification and present in arterial vessel walls. MGP -/- mice exhibit extensive MAC throughout their vasculature and die within 5-7 weeks due to aortic rupture or heart failure. The drug rapamycin functions as an inhibitor of cell proliferation and an activator of autophagy. Hypothesis: That rapamycin reduces MAC in MGP -/- mice via inhibiting inward remodeling and activating autophagy. Methods: MGP +/+ and MGP -/- mice treated with vehicle or rapamycin (5mg/kg, 3x weekly) starting at 10 days old for 2 weeks. Calcification was quantified via micro-computed tomography (uCT). Collagen structure and elastin quality were assessed histological staining. Markers of proliferation (Ki-67), cell death (Caspase 3), autophagy (LC3), and calcification (Runx2) were quantified via immunofluorescence staining. Western blot was used to assess autophagy flux and proliferation in smooth muscle cells (SMCs) isolated from MGP +/+ and MGP -/- mice. Results: While rapamycin treatment doubled the life-expectancy of MGP -/- mice, the calcification content was unchanged compared to the vehicle. However, rapamycin caused a significant decline in mineral density. Higher collagen content and improved elastin structure were observed in MGP -/- treated with rapamycin compared to vehicle. Runx2 and Ki-67 were elevated in MGP -/- mice compared to MGP +/+ , but there was no difference in response to rapamycin treatment. No difference was observed in caspase 3 levels. Under rapamycin treatment, MGP -/- SMCs exhibited higher autophagy flux than MGP +/+ SMCs. Conclusion: While rapamycin did not reduce MAC, our data suggest that rapamycin potentially preserves the integrity of the aortic vessel wall by inducing autophagy, which requires further investigation.