miR-153-3p靶向调控CHI3L1对LPS所致人牙周膜干细胞炎症反应的影响

Effect of miR-153-3p Targeted Regulation of CHI3L1 on the Inflammatory Response of LPS-Induced Human Periodontal Ligament Stem Cells

目的:探索miR-153-3p是否能通过靶向调控几丁质酶3样蛋白1(CHI3L1)改善脂多糖(LPS)所致人牙周膜干细胞(hPDLSCs)炎症反应。方法:分离培养hPDLSCs,采用LPS处理建立炎症细胞模型,并进行miR-NC、miR-153-3p mimics、si-NC、si-CHI3L1以及miR-153-3p mimics+pc-NC、miR-153-3p mimics+pcDNA-CHI3L1转染。转染48h后qRT-PCR法检测各组细胞中miR-153-3p、CHI3L1 mRNA表达,ELISA法检测细胞上清液中IL-6、IL-1β、TNF-α水平,流式细胞术检测细胞凋亡情况,Western Blot法检测细胞中CHI3L1蛋白表达。双荧光素酶报告基因实验检测miR-153-3p与CHI3L1的靶向关系。结果:LPS处理后hPDLSCs中miR-153-3p表达下调(P<0.05),CHI3L1 mRNA与蛋白表达均显著上调(P<0.05);过表达miR-153-3p或抑制CHI3L1表达可降低LPS诱导的hPDLSCs中IL-6、IL-1β、TNF-α水平,下调CHI3L1 mRNA与蛋白表达,降低细胞凋亡率,差异均具有统计学意义(P<0.05)。双荧光素酶报告基因实验证实miR-153-3p可靶向调控CHI3L1表达(P<0.05)。此外,过表达CHI3L1可显著逆转过表达miR-153-3p对LPS诱导的hPDLSCs凋亡与炎症反应的抑制作用(P<0.05)。结论:miR-153-3p可通过靶向调控CHI3L1表达减轻LPS所致hPDLSCs炎症反应,减少细胞凋亡。

Objective:To explore whether miR-153-3p can improve the inflammatory response of human periodontal ligament stem cells(hPDLSCs)induced by lipopolysaccharide(LPS)by targeting regulates chitinase 3-like protein 1(CHI3L1).Methods:The hPDLSCs were isolated and cultured,and inflammatory cell models were established using LPS treatment and transfected with miR-NC,miR-153-3p mimics,si-NC,si-CHI3L1,and miR-153-3p mimics+pc-NC,miR-153-3p mimics+pcDNA-CHI3L1.48 h after transfection,the expressions of miR-153-3p and CHI3L1 mRNA in the cells of each group were detected by qRT-PCR,the levels of IL-6,IL-1βand TNF-αin the cell supernatant were detected by ELISA,cell apoptosis was detected by flow cytometry,the expression of CHI3L1 protein in cells was detected by Western Blot,the targeting relationship between miR-153-3p and CHI3L1 was detected by dual-luciferase reporter gene assay.Results:The expression of miR-153-3p was down-regulated in hPDLSCs after LPS treatment(P<0.05),while the expressions of CHI3L1 mRNA and protein were significantly up-regulated(P<0.05);over-expression of miR-153-3p or inhibition of CHI3L1 expression could reduce the levels of IL-6,IL-1βand TNF-αin LPS-induced hPDLSCs,down-regulate the expressions of CHI3L1 mRNA and protein,and reduce the apoptosis rate,and the differences were statistically significant(P<0.05).The dual-luciferase reporter gene assay confirmed that miR-153-3p could targeting regulates the expression of CHI3L1(P<0.05).In addition,the over-expression of CHI3L1 could significantly reverse the inhibitory effects of the over-expression of miR-153-3p on LPS-induced apoptosis and inflammatory response of hPDLSCs(P<0.05).Conclusion:MiR-153-3p can reduce the LPS-induced inflammatory response and apoptosis of hPDLSCs by targeting regulates CHI3L1 expression.