MiR-15a regulates bicuspid aortic valve calcification via TGF-β signaling pathway via inhibiting Smad7
MiR-15a regulates bicuspid aortic valve calcification via TGF-β signaling pathway via inhibiting Smad7
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DOI：
10.1016/j.jksus.2020.101278
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发表时间：
2020-12
期刊：
影响因子：
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通讯作者：
Jiankang Xu;Hao Liu;Rui Zheng;Minyan Dang;Y. Shao;Junjie Du
中科院分区：
文献类型：
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作者：
Jiankang Xu;Hao Liu;Rui Zheng;Minyan Dang;Y. Shao;Junjie Du
背景：二尖瓣主动脉瓣是一种遗传性心脏瓣膜疾病，与三尖瓣主动脉瓣相比，其特点是瓣膜钙化和狭窄早发，进展迅速。强有力的证据表明，许多mirna及其靶基因参与BAV钙化的发生。本研究旨在探讨miR-15a和Smad7在BAV中的表达及其调节钙化的机制。方法采用免疫组化和western blotting检测Smad7和Runx2在人主动脉瓣组织中的表达水平。RT-PCR检测miR-15a、Smad7、Runx2的表达。通过双荧光素酶测定评估miR-15a与Smad7之间的关系。在培养的猪瓣膜间质细胞（PVICs）中过表达MiR-15a，通过RT-PCR检测Smad7和Runx2 mRNA的变化，以及Smad7和Runx2蛋白的变化。采用茜素红染色验证miR-15a在钙化诱导培养PVICs中的抑制钙化作用。结果与钙化TAV相比，钙化BAV中Smad7和Runx2的表达显著升高，miR-15a的表达显著降低，Smad7和Runx2 mRNA的表达显著升高，miR-15a的表达与Smad7 mRNA水平呈负相关。miR-15a过表达的PVICs显示miR-15a上调，Smad7和Runx2表达下调。在钙化诱导下，mir -15a过表达组PVICs的钙化程度明显降低。结论BAV中miR-15a的低表达导致Smad7的高表达，从而降低Smad2/3-Smad4复合物对runx2的拮抗作用，促进BAV钙化进程。
BackgroundBicuspid aortic valve is a heritable heart valve disease with characteristic early onset of valve calcification and stenosis with rapid progression compared with tricuspid aortic valve. Strong evidence indicates that many miRNAs and their target genes are involved in the development of BAV calcification. This study was designed to investigate miR-15a and Smad7 expressions in BAV and their mechanism of regulating calcification.MethodsImmunohistochemistry and western blotting were used to explore the expression levels of Smad7 and Runx2 in human aortic valve tissue. The expression of miR-15a, Smad7 and Runx2 were detected by RT-PCR. The relationship between miR-15a and Smad7 was assessed by dual-luciferase assay. MiR-15a was overexpressed in cultured porcine valve interstitial cells (PVICs) and the variations of Smad7 and Runx2 mRNA were evaluated by RT-PCR, as well as the changes of Smad7 and Runx2 protein. Alizarin reds staining was used to verify the calcification inhibition effect of miR-15a in cultured PVICs under calcification induction.ResultsCompared with calcified TAV, remarkable higher expression of Smad7 and Runx2 were found in calcified BAV, significantly lower expressions of miR-15a and higher expressions of Smad7 and Runx2 mRNA, along with negative correlation between expressions of miR-15a and mRNA level of Smad7. MiR-15a-overexpressed PVICs shown upregulation of miR-15a, downregulation of Smad7 and Runx2 expression. PVICs under calcification induction shown significantly reduced calcification in miR-15a-overexpressed group.ConclusionsThe lower miR-15a expression in BAV results in the high expression of Smad7, thereby reducing antagonism of Smad2/3–Smad4 complex to runx2 and promoting the progress of BAV calcification.