miR-296-3p通过靶向ZBTB20抑制HSC-T6细胞的活化OA
miR-296-3p inhibits the avtivation of HSC-T6 cell by targeting ZBTB20
目的:探讨miR-296-3p通过靶向锌指和BTB结构域蛋白20(BTB domain-containing protein 20,ZBTB20)对大鼠肝星状细胞 HSC-T6活化的调控作用.方法:采用 qRT-PCR检测经过转化生长因子-β1(transforming growth factor-β1,TGF-β1)刺激的大鼠肝星状细胞HSC-T6中纤维化标志物和miR-296-3p的表达水平;利用脂质体转染法将miR-296-3p mimic/inhibitor及各自的对照转染到HSC-T6细胞中,分组为:mimic NC组、miR-296-3p mimic组、inhibitor组和miR-296-3p inhibitor组,并且通过qRT-PCR、Western Blot、CCK-8、集落形成和 Transwell实验检测转染 miR-296-3p mimic/inhibitor后对 HSC-T6细胞活化能力的影响;利用生物信息学方法预测 miR-296-3p的候选靶基因,并用双荧光素酶基因报告实验进行验证;qRT-PCR 和 Western blot实验检测 miR-296-3p与候选靶基因 ZBTB20之间的靶定关系;qRT-PCR、CCK-8及Transwell实验分析pcDNA3.1-ZBTB20能否逆转miR-296-3p+mimics对HSC-T6细胞活化能力的抑制作用.结果:HSC-T6细胞被 TGF-β1刺激后进一步活化,并且活化的细胞中的 miR-296-3p表达水平下调;miR-296-3p抑制HSC-T6的增殖、迁移,降低肝纤维化标志物I型胶原蛋白(Col1A1)和α-平滑肌肌动蛋白(α-SMA)的表达水平;通过生物信息学数据库预测以及双荧光素酶基因报告实验验证显示ZBTB20是miR-296-3p的功能靶基因,并且在HSC-T6中miR-296-3p负调控ZBTB20的表达;过表达ZBTB20可以部分逆转过表达miR-296-3p对HSC-T6活化的抑制作用.结论:miR-296-3p通过靶向抑制ZBTB20的表达从而抑制HSC-T6细胞的进一步活化.
Objective:To investigate the role of miR-296-3p in the regulation of HSC-T6 activation in rat hepatic stellate cells by targeting zinc finger and BTB domain-containing protein 20(ZBTB20).Methods:The expression levels of fibrosis markers and miR-296-3p in rat hepatic stellate cells HSC-T6 stimulated by transforming growth factor-β1(TGF-β1)were detected by qRT-PCR.The miR-296-3p mimic/inhibitor and their respective controls were transfected into HSC-T6 cells using liposome transfec-tion,respectively.There were mimic NC group,miR-296-3p mimic group,inhibitor group and miR-296-3p inhibitor group.The effects of miR-296-3p on the activation ability of HSC-T6 cells were detected by qRT-PCR,Western Blot,CCK-8,colony forma-tion,and Transwell assays.After transfection with miR-296-3p mimic/inhibitor,the candidate target genes of miR-296-3p were predicted by bioinformatics methods and validated with dual luciferase gene reporter assay.qRT-PCR and Western Blot experi-ments were performed to detect the binding relationship between miR-296-3p and the candidate target gene ZBTB20.qRT-PCR,CCK-8 and Transwell assays were performed to analyse whether pcDNA3.1-ZBTB20 could reverse the inhibitory effect of miR-296-3p+mimics on the activation ability of HSC-T6 cells.Results:HSC-T6 cells were further activated by TGF-β1 stimulation,and the expression level of miR-296-3p was down-regulated in activated cells.miR-296-3p inhibited the proliferation and migration of HSC-T6,and decreased the expression level of hepatic fibrosis markers,collagen type I(Col1A1)and α-smooth muscle actin(α-SMA).The bioinformatics database prediction and the verification of the double luciferase gene reporter assay showed that ZBTB20 was a functional target gene of miR-296-3p and that miR-296-3p negatively regulates ZBTB20 expression in HSC-T6.ZBTB20 could partially reverse the inhibitory effect of miR-296-3p on HSC-T6 activation.Conclusion:miR-296-3p inhibits fur-ther activation of HSC-T6 cells by targeting and inhibiting ZBTB20 expression.
周静;李以恒;李尹凡;崔笑妍;王梅梅;张荣花;刘志勇;章广玲
华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学基础医学院,河北省慢性疾病重点实验室,河北 唐山 063210华北理工大学临床医学院,河北省医工融合精准医疗重点实验室,河北 唐山 063000华北理工大学临床医学院,河北省医工融合精准医疗重点实验室,河北 唐山 063000
医药卫生
miR-296-3pZBTB20HSC-T6细胞肝纤维化增殖迁移
MiR-296-3pZBTB20HSC-T6 cellsHepatic fibrosisProliferationMigration
《海南医科大学学报》 2026 (9)
649-660,12
This study was supported by the Hebei Province Natural Science Foundation Project(H2023209047)Hebei Provincial Department of Education Funded Project for Cultivating Innovative Ability of Graduate Students(CXZZSS2024056) 河北省自然科学基金资助项目(H2023209047)河北省教育厅在读研究生创新能力培养资助项目(CXZZSS2024056)
评论