The Effects of Cardiolipin on Vascular Smooth Muscle Cell Dedifferentiation and Migration

Abstract

PURPOSEThe initial phases of atherosclerosis involve the transition of smooth muscle cells (SMCs) to a synthetic, dedifferentiated phenotype, which is associated with alterations in receptor, ion channel and contractile filament composition. The late, irreversible atherosclerosis stages are characterized by SMC migration and subsequent formation of a ‘neointima.’ This leads to blood flow obstruction which, when destined to the cardiomyocytes, induces hypoxic conditions and myocardial infarction. Apoptotic cardiomyocytes release a phospholipid called cardiolipin (CL) into the systemic circulation. Previously, we have shown that physiological concentrations of CL inhibit endothelial cell migration. The objective of this project was to investigate the impact of CL on SMCs, specifically addressing cellular dedifferentiation and migration.METHODSFor this study, we used adult, male C57Bl/6 mice. Aortas from these mice were extracted and incubated in Dulbecco's modification of Eagle medium containing physiological CL concentrations (1 μM and 10 μM) at 37°C for 48 hours. We subsequently quantified contractile proteins (such as smooth muscle α‐actin and calponin) using immunoblotting to determine whether CL affects SMC dedifferentiation. These concentrations were also used to investigate the migration of SMCs using a migration assay. For this, we mimicked wounds by scraping off the cell culture plates' diameters to create cell‐free gaps. The plates were then incubated at 37°C for six days, during which the width of the gaps was measured every 48 hours under a light microscope.RESULTSThe immunoblotting results demonstrated a significant drop in calponin in a dose‐dependent manner with increasing CL concentrations, while the other de/differentiation markers remained unchanged. The migration assay revealed that the gap width was significantly narrower in control plates compared to CL‐treated plates.CONCLUSIONSOur results suggest that the effect of physiological concentrations of CL on SMCs is calponin‐specific and does not initiate dedifferentiation within 48 hours. The significantly wider gaps observed in the CL‐treated plates compared to the control plates indicate the potent effect CL has on inhibiting SMC migration. Since SMC migration is considered the key step in the atherogenic progression, our migration assay data suggest that, in a physiological context, CL has the potential to arrest the progression of the disease into its irreversible stages. Thus, we hypothesize the potential involvement of CL in developing anti‐atherosclerosis therapeutics.Support or Funding InformationCanadian Institute of Health Research (CIHR)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.