Liu Zongtao, Dong Nianguo, Hui Haipeng, Wang Yixuan, Liu Fayun, Xu Li, Liu Ming, Rao Zhenqi, Yuan Zhen, Shang Yuqiang, Feng Jun, Cai Zhejun, Li Fei
Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1277 Jiefang Ave, Wuhan 430022, Hubei, China.
Department of Cardiology, the Second Medical Center & National Clinical Research Center for Geriatric Diseases, Chinese PLA General Hospital, 28# Fuxing Road, Beijing 100853, China.
Eur Heart J. 2022 May 1;43(17):1652-1664. doi: 10.1093/eurheartj/ehac037.
Tetrahydrobiopterin (BH4) is a critical determinant of the biological function of endothelial nitric oxide synthase. The present study was to investigate the role of valvular endothelial cell (VEC)-derived BH4 in aortic valve calcification.
Plasma and aortic valve BH4 concentrations and the BH4:BH2 ratio were significantly lower in calcific aortic valve disease patients than in controls. There was a significant decrease of the two key enzymes of BH4 biosynthesis, guanosine 5'-triphosphate cyclohydrolase I (GCH1) and dihydrofolate reductase (DHFR), in calcified aortic valves compared with the normal ones. Endothelial cell-specific deficiency of Gch1 in Apoe-/- (Apoe-/-Gch1fl/flTie2Cre) mice showed a marked increase in transvalvular peak jet velocity, calcium deposition, runt-related transcription factor 2 (Runx2), dihydroethidium (DHE), and 3-nitrotyrosine (3-NT) levels in aortic valve leaflets compared with Apoe-/-Gch1fl/fl mice after a 24-week western diet (WD) challenge. Oxidized LDL (ox-LDL) induced osteoblastic differentiation of valvular interstitial cells (VICs) co-cultured with either si-GCH1- or si-DHFR-transfected VECs, while the effects could be abolished by BH4 supplementation. Deficiency of BH4 in VECs caused peroxynitrite formation increase and 3-NT protein increase under ox-LDL stimulation in VICs. SIN-1, the peroxynitrite generator, significantly up-regulated alkaline phosphatase (ALP) and Runx2 expression in VICs via tyrosine nitration of dynamin-related protein 1 (DRP1) at Y628. Finally, folic acid (FA) significantly attenuated aortic valve calcification in WD-fed Apoe-/- mice through increasing DHFR and salvaging BH4 biosynthesis.
The reduction in endothelial-dependent BH4 levels promoted peroxynitrite formation, which subsequently resulted in DRP1 tyrosine nitration and osteoblastic differentiation of VICs, thereby leading to aortic valve calcification. Supplementation of FA in diet attenuated hypercholesterolaemia-induced aortic valve calcification by salvaging BH4 bioavailability.
四氢生物蝶呤（BH4）是内皮型一氧化氮合酶生物学功能的关键决定因素。本研究旨在探讨瓣膜内皮细胞（VEC）衍生的BH4在主动脉瓣钙化中的作用。
钙化性主动脉瓣疾病患者血浆和主动脉瓣BH4浓度以及BH4:BH2比值显著低于对照组。与正常主动脉瓣相比，钙化主动脉瓣中BH4生物合成的两种关键酶，即鸟苷5'-三磷酸环化水解酶I（GCH1）和二氢叶酸还原酶（DHFR）显著减少。在24周西方饮食（WD）挑战后，Apoe-/-（Apoe-/-Gch1fl/flTie2Cre）小鼠内皮细胞特异性Gch1缺乏与Apoe-/-Gch1fl/fl小鼠相比，主动脉瓣叶跨瓣峰值射流速度、钙沉积、 runt相关转录因子2（Runx2）、二氢乙锭（DHE）和3-硝基酪氨酸（3-NT）水平显著增加。氧化型低密度脂蛋白（ox-LDL）诱导与si-GCH1或si-DHFR转染的VEC共培养的瓣膜间质细胞（VIC）成骨分化，而补充BH4可消除这种作用。VEC中BH4缺乏导致VIC在ox-LDL刺激下过氧亚硝酸根形成增加和3-NT蛋白增加。过氧亚硝酸根生成剂SIN-1通过在Y628位点对动力相关蛋白1（DRP1）进行酪氨酸硝化，显著上调VIC中碱性磷酸酶（ALP）和Runx2表达。最后，叶酸（FA）通过增加DHFR和挽救BH4生物合成，显著减轻WD喂养的Apoe-/-小鼠的主动脉瓣钙化。
内皮依赖性BH4水平降低促进过氧亚硝酸根形成，随后导致DRP1酪氨酸硝化和VIC成骨分化，从而导致主动脉瓣钙化。饮食中补充FA通过挽救BH4生物利用度减轻高胆固醇血症诱导的主动脉瓣钙化。
