Abstract 169: Lipoproteins and their Modified Forms Regulate Smooth Muscle Cell Calcification

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Background: Vascular calcification (VC), alongside atherogenic lipoprotein profiles, have been correlated with poor cardiovascular outcome, however there is a paucity of literature exploring the relationship between the two regarding VC progression. We therefore aim to examine the roles of lipoprotein species and their oxidised forms on both medial and intimal VC. Methods: Human aortic smooth muscle cells (HAoSMC) were pre-treated with 200 μg/ml high (HDL), low (LDL) and very low (VLDL) density lipoproteins for 24 hours before treating cells with a calcification medium (CM; Ca 2.7mM, PO 4 2.0 mM). Cells were harvested using an alizarin red (ARS) calcification assay, or at various time points for qPCR analysis. In parallel, in vivo studies using apolipoprotein E knock-out mice were fed an atherogenic diet for 40 weeks and received reconstituted HDL (rHDL) infusions containing apoA-I (20mg/kg) and 1-palmitoyl-2-linoleoyl phosphatidylcholine during the final 4 weeks of the study. Tissues were harvested and stained for plaque assessment (H&E) and calcium deposits (ARS). Results: Pre-treatment of HAoSMC with rHDL inhibited calcification (43.9%, p<0.001), whereas ox-rHDL removed its protective effects. Likewise, ox-LDL (77.1%, p<0.05) also upregulated calcium deposition and interestingly ox-VLDL significantly decreased calcification (70.5%, p<0.05) with their native counterparts having no effects. PCR measures of calcification markers Runx2, RANKL and alkaline phosphatase show a time-dependent increase in expression as calcification occurs. In animal studies, no change in weight gain, cholesterol or triglyceride levels were observed with treatment. In addition, rHDL infusions did not alter plaque size however ARS staining of the brachiocephalic artery demonstrated a significant reduction (6.58%, p<0.05) in calcification present in the atherosclerotic plaque. Conclusions: This study is the first to demonstrate the effects of lipoproteins on VC in vitro and the effects of rHDL on in vivo VC. Somewhat in accordance to the roles of lipoproteins in atherosclerosis, HDL and ox-VLDL show a reduction of calcification, where-as ox-LDL enhances calcification of HAoSMC.