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miR-769-5p/类Krüppel因子6轴下调circ_0097010对脂多糖刺激的牙周膜细胞的抗凋亡和抗炎活性

Anti-apoptosis and anti-inflammation activity of circ_0097010 downregulation in lipopolysaccharide-stimulated periodontal ligament cells by miR-769-5p/Krüppel like factor 6 axis.

作者信息

Sun Dan-Dan, Wu Xue, Lin Shi-Chen, Duan Shao-Yu

机构信息

Department of Stomatology, Electric Power Teaching Hospital, Capital Medical University, Beijing, China.

出版信息

J Dent Sci. 2023 Jan;18(1):310-321. doi: 10.1016/j.jds.2022.04.024. Epub 2022 Jun 4.

Abstract

BACKGROUND/PURPOSE: Periodontitis is a prevalent infectious inflammatory disease. Growing evidence has revealed important roles for circular RNAs (circRNAs) and circRNA sponge activity in periodontitis. Here, we elucidated the precise part of circ_0097010 in periodontitis pathogenesis.

MATERIALS AND METHODS

Human periodontal ligament cells (hPDLCs) were exposed to lipopolysaccharide (LPS). Cell viability, proliferation and apoptosis were evaluated by CCK-8 assay, EdU incorporation assay and flow cytometry, respectively. Circ_0097010, microRNA (miR)-769-5p and Krüppel like factor 6 (KLF6) were quantified by qRT-PCR and Western blot. Interleukin 6 (IL-6) level, tumor necrosis factor-α (TNF-α) secretion, superoxide dismutase (SOD) activity and malondialdehyde (MDA) level were detected by enzyme-linked immunosorbent assay (ELISA). Dual-luciferase reporter, RNA immunoprecipitation (RIP) and RNA pull-down assays were used to confirm the direct relationship between miR-769-5p and circ_0097010 or KLF6.

RESULTS

Our data showed that LPS repressed cell proliferation and induced cell apoptosis and inflammation in hPDLCs. Circ_0097010 was upregulated in periodontitis samples and LPS-exposed hPDLCs. Downregulation of circ_0097010 exerted anti-apoptosis and anti-inflammation functions in LPS-exposed hPDLCs. Mechanistically, circ_0097010 acted as a miR-769-5p sponge, and reduced abundance of miR-769-5p reversed the anti-apoptosis and anti-inflammation effects of circ_0097010 suppression. KLF6 was a direct miR-769-5p target, and miR-769-5p-mediated inhibition of KLF6 possessed anti-apoptosis and anti-inflammation functions in LPS-induced hPDLCs. Moreover, circ_0097010 controlled KLF6 expression by miR-769-5p.

CONCLUSION

These data identify circ_0097010 as a key regulator of LPS-induced inflammation and apoptosis in hPDLCs and highlight a novel mechanism of circ_0097010 regulation through miR-769-5p/KLF6 axis.

摘要

背景/目的:牙周炎是一种常见的感染性炎症疾病。越来越多的证据表明环状RNA(circRNA)及其海绵活性在牙周炎中发挥着重要作用。在此,我们阐明了circ_0097010在牙周炎发病机制中的具体作用。

材料与方法

将人牙周膜细胞(hPDLCs)暴露于脂多糖(LPS)中。分别通过CCK-8法、EdU掺入法和流式细胞术评估细胞活力、增殖和凋亡。通过qRT-PCR和蛋白质免疫印迹法对circ_0097010、微小RNA(miR)-769-5p和Krüppel样因子6(KLF6)进行定量分析。采用酶联免疫吸附测定(ELISA)检测白细胞介素6(IL-6)水平、肿瘤坏死因子-α(TNF-α)分泌、超氧化物歧化酶(SOD)活性和丙二醛(MDA)水平。利用双荧光素酶报告基因、RNA免疫沉淀(RIP)和RNA下拉实验来确认miR-769-5p与circ_0097010或KLF6之间的直接关系。

结果

我们的数据显示,LPS抑制hPDLCs的细胞增殖并诱导细胞凋亡和炎症反应。circ_0097010在牙周炎样本和LPS处理的hPDLCs中表达上调。下调circ_0097010对LPS处理的hPDLCs具有抗凋亡和抗炎作用。机制上,circ_0097010作为miR-769-5p的海绵,降低miR-769-5p的丰度可逆转circ_0097010抑制的抗凋亡和抗炎作用。KLF6是miR-769-5p的直接靶标,miR-769-5p介导的对KLF6的抑制在LPS诱导的hPDLCs中具有抗凋亡和抗炎作用。此外,circ_0097010通过miR-769-5p调控KLF6的表达。

结论

这些数据表明circ_0097010是LPS诱导的hPDLCs炎症和凋亡的关键调节因子,并揭示了circ_0097010通过miR-769-5p/KLF6轴调控的新机制。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b749/9831795/d20d70c28ffa/gr1.jpg

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