Departments of coronary heart disease, First Affiliated Hospital of Xinjiang Medical University, Urumqi City, China.
Departments of coronary heart disease, Xinjiang Medical University, Urumqi City, China.
Cardiovasc Drugs Ther. 2024 Apr;38(2):263-277. doi: 10.1007/s10557-022-07416-x. Epub 2022 Dec 31.
BACKGROUND: Circular RNAs (circRNAs) have shown important regulatory roles in cardiovascular diseases, including atherosclerosis (AS). However, the role and mechanism of circ_0026218 in AS remain unclear. METHODS: The cell model of AS in vitro was established by stimulating human umbilical vein endothelial cells (HUVECs) with oxidized low-density lipoprotein (ox-LDL). In addition, circ_0026218, microRNA-188-3p (miR-188-3p), and toll-like receptor 4 (TLR4) expression was determined via real-time quantitative polymerase chain reaction (RT-qPCR) in serum samples from AS patients and healthy volunteers. Cell proliferation was assessed using Cell Counting Kit-8 (CCK-8) assay and 5-ethynyl-2'-deoxyuridine (EdU) assay. Cell apoptosis was measured using flow cytometry. The inflammatory response was assessed using enzyme-linked immunosorbent assay (ELISA). Oxidative stress level was assessed using corresponding kits. Nitric oxide (NO) level was examined using NO detection assay. The interaction between miR-188-3p and circ_0026218 or TLR4 was determined via dual-luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays. Exosomes were observed using transmission electron microscopy (TEM). The size distribution of exosomes was analyzed using nanoparticle tracking analysis (NTA). RESULTS: Ox-LDL treatment caused HUVEC dysfunction by inhibiting cell proliferation and promoting apoptosis, inflammation, and oxidative stress. Circ_0026218 was upregulated in AS serum samples and ox-LDL-treated HUVECs. Knockdown of circ_0026218 attenuated ox-LDL-induced dysfunction in HUVECs. MiR-188-3p acted as a target of circ_0026218, and miR-188-3p downregulation reversed the suppression role of circ_0026218 knockdown on ox-LDL-induced HUVEC disorder. TLR4 was a target of miR-188-3p, and miR-188-3p overexpression alleviated ox-LDL-induced dysfunction in HUVECs by targeting TLR4. Circ_0026218 could deregulate the TLR4/NF-κB pathway by sponging the miR-188-3p. Importantly, circ_0026218 was overexpressed in exosomes from ox-LDL-treated HUVECs and could be delivered via exosomes. CONCLUSION: Circ_0026218 knockdown attenuated ox-LDL-induced dysfunction in HUVECs via regulating miR-188-3p/TLR4/NF-κB pathway.
背景：环状 RNA（circRNAs）在心血管疾病（包括动脉粥样硬化（AS））中表现出重要的调节作用。然而，circ_0026218 在 AS 中的作用和机制尚不清楚。
方法：通过用氧化低密度脂蛋白（ox-LDL）刺激人脐静脉内皮细胞（HUVEC）在体外建立 AS 细胞模型。此外，通过实时定量聚合酶链反应（RT-qPCR）测定 AS 患者和健康志愿者血清样本中的 circ_0026218、微小 RNA-188-3p（miR-188-3p）和 Toll 样受体 4（TLR4）的表达。使用细胞计数试剂盒-8（CCK-8）测定和 5-乙炔基-2'-脱氧尿苷（EdU）测定评估细胞增殖。通过流式细胞术测量细胞凋亡。通过酶联免疫吸附测定（ELISA）评估炎症反应。通过相应试剂盒评估氧化应激水平。通过一氧化氮（NO）检测试剂盒检测 NO 水平。通过双荧光素酶报告、RNA 免疫沉淀（RIP）和 RNA 下拉测定确定 miR-188-3p 与 circ_0026218 或 TLR4 之间的相互作用。使用透射电子显微镜（TEM）观察外泌体。通过纳米颗粒跟踪分析（NTA）分析外泌体的大小分布。
结果：ox-LDL 处理通过抑制细胞增殖和促进细胞凋亡、炎症和氧化应激来导致 HUVEC 功能障碍。circ_0026218 在 AS 血清样本和 ox-LDL 处理的 HUVEC 中上调。circ_0026218 敲低减轻了 ox-LDL 诱导的 HUVEC 功能障碍。miR-188-3p 作为 circ_0026218 的靶标，miR-188-3p 的下调逆转了 circ_0026218 敲低对 ox-LDL 诱导的 HUVEC 紊乱的抑制作用。TLR4 是 miR-188-3p 的靶标，miR-188-3p 通过靶向 TLR4 减轻 ox-LDL 诱导的 HUVEC 功能障碍。circ_0026218 可以通过海绵吸附 miR-188-3p 来调节 TLR4/NF-κB 通路。重要的是，circ_0026218 在 ox-LDL 处理的 HUVEC 来源的外泌体中过表达，并可通过外泌体传递。
结论：circ_0026218 敲低通过调节 miR-188-3p/TLR4/NF-κB 通路减轻 ox-LDL 诱导的 HUVEC 功能障碍。
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