Abstract 19733: Shear-Dependent Aortic Valve Endothelial Inflammation and Calcification Are Regulated by O-GlcNAc Modification of MEF2C Transcription Factor
Introduction: Aortic valve (AV) disease is a significant contributor to cardiovascular mortality. AV calcification occurs preferentially on the fibrosa side, where endothelial cells (ECs) are subjected to characteristic oscillatory shear stress (OS), whereas the ventricularis ECs experience stable unidirectional laminar shear stress (LS). OS and LS differently regulate endothelial function via gene expression and protein modification. While the post-translational β-N-acetylglucosamine modification of amino acids (O-GlcNAcylation) has been shown to be important in the function of various cell types, its role in AV disease is unknown. We hypothesized that OS impairs EC O-GlcNAcylation, leading to AV inflammation and calcification.
Methods and Results: Immunostaining analysis showed that O-GlcNAcylation was decreased in fibrosa endothelium compared to ventricularis in human and porcine AVs, and human AV ECs (HAVECs) exhibited increased O-GlcNAcylation by LS (20 dyn/cm2) and decreased by OS (+5 dyn/cm2) for 24h. LS increased expression of the O-GlcNAc transferase (OGT), while reducing monocyte adhesion and increasing expression of the shear-sensitive, anti-inflammatory gene Klf2. This LS-induced anti-inflammatory effect was inhibited by siRNA-medicated OGT knockdown in HAVECs. Immunoprecipitation studies using MEF2C and O-GlcNAc antibodies showed that LS increased O-GlcNAcylation of MEF2C, a well-known regulator of Klf2. Mass spectrometry analysis revealed three novel O-GlcNAcylation sites of MEF2C at T9, S219, and S269. Adenoviral expression of each MEF2C mutant (T9A, S219A, and S269A) in HAVECs decreased MEF2C O-GlcNAcylation, Klf2 promoter activity, and Klf2 mRNA expression by LS. In ex vivo porcine AV studies, we found that overexpression of wild-type MEF2C, S219A or S269A mutant increased Klf2 expression and inhibited valve calcification, while MEF2C-T9A mutant failed to increase Klf2 expression and to prevent valve calcification.
Conclusion: Our studies identified OGT-mediated O-GlcNAcylation of MEF2C as a novel shear-sensitive post-translational modification pathway regulating AV inflammation and calcification. These novel studies may lead to new non-surgical therapies for AV disease.
Author Disclosures: J. Heath: None. H. Kim: None. J. Fernandez Esmerats: None. R. Simmons: None. S. Kumar: None. J. Cho: None. H. Jo: None.
- © 2016 by American Heart Association, Inc.