State Key Laboratory of Cardiovascular Disease, FuWai Hospital, National Center for Cardiovascular Diseases, Peking Union Medical College & Chinese Academy of Medical Sciences, Beijing, China.
Xiamen Cardiovascular Hospital, Xiamen University, Xiamen, China.
Dis Markers. 2020 Nov 17;2020:8864322. doi: 10.1155/2020/8864322. eCollection 2020.
The endothelium is the first line of defence against harmful microenvironment risks, and microRNAs (miRNAs) involved in vascular inflammation may be promising therapeutic targets to modulate atherosclerosis progression. In this study, we aimed to investigate the mechanism by which microRNA-216a (miR-216a) modulated inflammation activation of endothelial cells. A replicative senescence model of human umbilical vein endothelial cells (HUVECs) was established, and population-doubling levels (PDLs) were defined during passages. PDL8 HUVECs were transfected with miR-216a mimics/inhibitor or small interfering RNA (siRNA) of SMAD family member 7 (Smad7). Real-time PCR and Western blot assays were performed to detect the regulatory role of miR-216a on Smad7 and NF-B inhibitor alpha (IB) expression. The effect of miR-216a on adhesive capability of HUVECs to THP-1 cells was examined. MiR-216a and Smad7 expression were measured using human carotid atherosclerotic plaques of the patients who underwent carotid endarterectomy ( = 41).
Luciferase assays showed that Smad7 was a direct target of miR-216a. Smad7 mRNA expression, negatively correlated with miR-216a during endothelial aging, was downregulated in senescent PDL44 cells, compared with young PDL8 HUVECs. MiR-216a markedly increased endothelial inflammation and adhesive capability to monocytes in PDL8 cells by promoting the phosphorylation and degradation of IB and then activating NF-B signalling pathway. The effect of miR-216a on endothelial cells was consistent with that blocked Smad7 by siRNAs. When inhibiting endogenous miR-216a, the Smad7/IB expression was rescued, which led to decreased endothelial inflammation and monocytes recruitment. In human carotid atherosclerotic plaques, Smad7 level was remarkably decreased in high miR-216a group compared with low miR-216a group. Moreover, miR-216a was negatively correlated with Smad7 and IB levels and positively correlated with interleukin 1 beta (IL1) expression .
In summary, our findings suggest a new mechanism of vascular endothelial inflammation involving Smad7/IB signalling pathway in atherosclerosis.
内皮细胞是抵御有害微环境风险的第一道防线，而参与血管炎症的 microRNAs（miRNAs）可能是调节动脉粥样硬化进展的有前途的治疗靶点。在这项研究中，我们旨在研究 microRNA-216a（miR-216a）调节内皮细胞炎症激活的机制。建立了人脐静脉内皮细胞（HUVEC）的复制性衰老模型，并在传代过程中定义了倍增水平（PDL）。将 miR-216a 模拟物/抑制剂或 SMAD 家族成员 7（Smad7）的小干扰 RNA（siRNA）转染至 PDL8 HUVECs。实时 PCR 和 Western blot 检测用于检测 miR-216a 对 Smad7 和 NF-B 抑制剂 alpha（IB）表达的调节作用。检测 miR-216a 对 HUVECs 与 THP-1 细胞黏附能力的影响。使用颈动脉内膜切除术（n=41）患者的人颈动脉粥样硬化斑块测量 miR-216a 和 Smad7 的表达。
荧光素酶测定表明 Smad7 是 miR-216a 的直接靶标。Smad7mRNA 表达在衰老过程中与 miR-216a 呈负相关，与年轻的 PDL8 HUVECs 相比，在衰老的 PDL44 细胞中下调。miR-216a 通过促进 IB 的磷酸化和降解，显著增加了 PDL8 细胞中的内皮炎症和单核细胞黏附能力，从而激活 NF-B 信号通路。miR-216a 对内皮细胞的作用与 siRNAs 阻断 Smad7 的作用一致。当抑制内源性 miR-216a 时，Smad7/IB 表达得到挽救，导致内皮炎症和单核细胞募集减少。在人颈动脉粥样硬化斑块中，高 miR-216a 组中 Smad7 水平明显低于低 miR-216a 组。此外，miR-216a 与 Smad7 和 IB 水平呈负相关，与白细胞介素 1 beta（IL1）表达呈正相关。
总之，我们的研究结果表明，在动脉粥样硬化中涉及 Smad7/IB 信号通路的血管内皮炎症的新机制。
