6090Conditional ablation of SDF-1/CXCL12 in smooth muscle cells leads to severe cardiac hypertrophy and aortic valve stenosis

Abstract

Abstract Background Stromal cell derived factor-1 (SDF-1) and its corresponding receptors CXCR4 & CXCR7 have been shown to play pivotal roles during cardiovascular development, cardiac repair and tissue homeostasis after ischemia. Stabilization of the SDF-1/CXCR4+ axis has been shown to provide beneficial effects on myocardial repair. Nevertheless, detailed mechanisms of the cell specific role of SDF-1 are poorly understood. Since SDF-1-EGFP lineage tracking revealed high expression of SDF-1 in smooth muscle cells, we aimed to investigate the cell specific role by generating a smooth muscle cell specific SDF-1 (SM-SDF-1 KO) knockout mouse model. Methods SDF-1 expression was analyzed utilizing SDF-1-EGFP reporter mice. SM-SDF-1 KO mice were generated using Cre/LoxP technology (SM22a-Cre; SDF-1fl/fl). Morphology was analysed with immunohistochemistry and immunofluorescence. Cardiac function was assessed utilizing echocardiography and millar tip catheterization. Whole transcriptome analysis, qRT-PCR and western blotting were performed. Further, apoptotic index and cell proliferation were quantified by TUNEL assay and PH3 immunostaining, respectively. Results SDF-1-EGFP lineage tracking and immunofluorescence analysis revealed high expression of SDF-1 particularly in smooth muscle cells and less frequently in perivascular and endothelial cells. Conditional SM-SDF-1 KO mice showed a high pre- and perinatal mortality (50%). Immunohistochemistry in surviving adult SM-SDF-1 KO mice revealed a severe cardiac hypertrophy phenotype, associated with increased cardiac fibrosis and apoptotic cell death. SM-SDF-1 KO mice revealed very thin and dilated arteries. Echocardiography measurements confimed concentric hypertrophy, and decreased stroke volume reflecting restrictive hypertrophic cardiomyopathy. Immunohistochemistry confirmed pronounced hypertrophy of cardiomyocytes. Additionally, we found evidence for enhanced proliferation markers in cardiomyocytes of SM-SDF-1 KO mice. Transcriptome analyses from KO hearts vs. non-ablated littermates identified over 150 significantly up- and downregulated genes. Western blot analysis for HIF-1α, AKT and ERK cell-signalling pathways were significantly elevated, whereas Rho Kinase signalling was specifically downregulated in SM-SDF-1 KO mice. As a possible reason for the hypertrophic phenotype, SDF-1 mutants exhibited aortic stenosis due to aortic valve thickening associated with upregulation of the extracellular proteoglycan versican anddownregulation of the SDF-1 co-receptor CXCR7. We further noticed increased plasma levels of SDF-1 in aortic stenosis patients suggesting a cardioprotective role. Conclusion Our data suggest that smooth muscle cell specific expression of SDF-1 plays a prominent role in cardiovascular development leadingto cardiac hypertrophy in adult animals. Our data further suggest that SDF-1 is involved in maintaining the homeostasis of the aortic valve, possibly by regulating versican. Acknowledgement/Funding FWF Austria