内皮-间质转化在钙化性主动脉瓣疾病发病过程中的作用研究

Endothelial-to-mesenchymal transition (EndMT) was identified as a very important process in the development of cardiac valve. However, in the valve disease, the opinions on the role of EndMT were not consistent. Studies focus on the markers of late-stage EndMT revealed that upregulation of these markers was a key process of the pathogenesis of valve; inversely, studies focused on the early-stage of EndMT demonstrated that transcript factors and upstream molecules were downregulated in calcific aortic valve disease (CAVD) samples. Another single-cell sequencing study conducted by us suggested that early-stage transcript factors were upregulated in the normal valve. Thus, we put on a hypothesis that EndMT had different effect on different stage of CAVD, and the early-stage EndMT had the potential to suppress the process of CAVD. So we designed the study, trying to use human valve samples, mouse CAVD models and porcine valve endothelial cells and valve interstitial cells to prove our hypothesis.

内皮-间质转化（EndMT）在瓣膜发育中有重要意义，但既往对EndMT在瓣膜疾病中的研究结论并不一致，对EndMT在瓣膜病发生中的作用缺乏全面系统的认识。申请人课题组前期针对瓣膜病的单细胞测序提示，人类正常瓣膜中EndMT早期转录因子表达水平较之CAVD显著升高。基于以上背景，我们假设不同阶段的CAVD中，EndMT的程度及作用并不相同，而早期EndMT对于CAVD的进展具有抑制作用。本研究拟利用人类瓣膜组织标本、CAVD小鼠模型研究CAVD各阶段EndMT的现象，全面描述EndMT在CAVD各阶段的表现和作用；另外，本研究拟利用猪瓣膜内皮与间质细胞的共培养模型，通过干预EndMT早期标志物Snai1在内皮细胞的表达水平，检测间质细胞的钙化水平，证明早期EndMT对CAVD的抑制作用。研究将通过细胞、组织和动物三个层面，首次全面阐述EndMT在CAVD的作用，为早期干预瓣膜病提供新思路。

内皮-间质转化（EndoMT）在主动脉瓣的发育中起着重要作用，但其在钙化性主动脉瓣疾病（CAVD）发生发展过程中的作用仍不清楚。我们在不同的动物模型（高脂+维生素D喂养的兔模型、高脂喂养的APOE-/-小鼠模型及导丝损伤主动脉瓣的小鼠模型）证实，CAVD中存在一群内皮细胞同时表达一系列间质细胞的特异性标记，换言之，EndoMT现象存在于不同的主动脉瓣钙化模型中。进一步的体外模型证实，EndoMT是由TGF-β信号驱动的，来自人类和动物模型的CAVD样本显示PDK4的高表达。我们进一步证实了PDK4在体外促进瓣膜内皮细胞向间质细胞进行转换。此外，PDK4抑制剂二氯乙酸盐明显缓解了主动脉瓣钙化小鼠模型的主动脉瓣钙化程度，表现为钙沉积减少，超声心动图参数改善，主动脉瓣的间质和成骨分化标志物减少。这些结果表明，PKD4可以通过介导EndoMT促进CAVD的发生和发展，这可能是CAVD的一个新的治疗目标。