Liu Bo, Cai Zhiyuan, Wang Yan, Liu Xinye, Zhang Bin, Zheng Qian, Li Jingye, Li Cien, Cui Yuanbo, Lv Pengju, Yang Dongwei
Department of Cardiology, Zhengzhou Central Hospital, Zhengzhou University, Zhengzhou, Henan, China.
The First Department of Ocular Fundus Diseases, Zhengzhou Second Hospital, Zhengzhou, Henan, China.
Front Pharmacol. 2024 Nov 25;15:1393534. doi: 10.3389/fphar.2024.1393534. eCollection 2024.
Endothelial cell (EC) calcification is an important marker of atherosclerotic calcification. ECs play a critical role not only in atherogenesis but also in intimal calcification, as they have been postulated to serve as a source of osteoprogenitor cells that initiate this process. While the role of transglutaminase 2 (TG2) in cellular differentiation, survival, apoptosis, autophagy, and cell adhesion is well established, the mechanism underlying the TG2-mediated regulation of EC calcification is yet to be fully elucidated.
The TG2 gene was overexpressed or silenced by using siRNA and recombinant adenovirus. RT-PCR and WB were used to analyze the relative expression of target genes and proteins. 5-BP method analyzed TG2 activity. mCherry-eGFP-LC3 adenovirus and transmission electron microscopy analyzed EC autophagy level. Calcium concentrations were measured by using a calcium colorimetric assay kit. Alizarin red S staining assay analyzed EC calcification level. Elisa analyzed IL-6 level. Establishing EC calcification model by using a calcification medium (CM).
Our findings demonstrated that CM increased TG2 activity and expression, which activated the NF-κB signaling pathway, and induced IL-6 autocrine signaling in ECs. Furthermore, IL-6 activated the JAK2/STAT3 signaling pathway to suppress cell autophagy and promoted ECs calcification.
ECs are not only critical for atherogenesis but also believed to be a source of osteoprogenitor cells that initiate intimal calcification. Previous research has shown that TG2 plays an important role in the development of VC, but the mechanism by which it exerts this effect is not yet fully understood. Our results demonstrated that TG2 forms complexes with NF-κB components inhibition of autophagy promoted endothelial cell calcification through EndMT. Therefore, our research investigated the molecular mechanism of EC calcification, which can provide new insights into the pathogenesis of atherosclerosis.
内皮细胞（EC）钙化是动脉粥样硬化钙化的一个重要标志。内皮细胞不仅在动脉粥样硬化形成过程中起关键作用，而且在内膜钙化中也发挥着重要作用，因为它们被认为是启动这一过程的骨祖细胞来源。虽然转谷氨酰胺酶2（TG2）在细胞分化、存活、凋亡、自噬和细胞黏附中的作用已得到充分证实，但TG2介导的内皮细胞钙化调节机制仍有待充分阐明。
通过使用小干扰RNA（siRNA）和重组腺病毒使TG2基因过表达或沉默。采用逆转录-聚合酶链反应（RT-PCR）和蛋白质免疫印迹法（WB）分析靶基因和蛋白质的相对表达。采用5-溴吡啶法分析TG2活性。利用mCherry-eGFP-LC3腺病毒和透射电子显微镜分析内皮细胞自噬水平。使用钙比色测定试剂盒测量钙浓度。茜素红S染色试验分析内皮细胞钙化水平。酶联免疫吸附测定（ELISA）分析白细胞介素-6（IL-6）水平。使用钙化培养基（CM）建立内皮细胞钙化模型。
我们的研究结果表明，CM增加了TG2活性和表达，激活了核因子κB（NF-κB）信号通路，并在内皮细胞中诱导了IL-6自分泌信号。此外，IL-6激活Janus激酶2/信号转导子和转录激活子3（JAK2/STAT3）信号通路以抑制细胞自噬并促进内皮细胞钙化。
内皮细胞不仅对动脉粥样硬化形成至关重要，而且被认为是启动内膜钙化的骨祖细胞来源。先前的研究表明，TG2在血管钙化发展中起重要作用，但其发挥这种作用的机制尚未完全了解。我们的结果表明，TG2与NF-κB成分形成复合物，通过内皮-间质转化（EndMT）抑制自噬促进内皮细胞钙化。因此，我们的研究探讨了内皮细胞钙化的分子机制，可为动脉粥样硬化的发病机制提供新的见解。
