摘要
目的探究LncRNA NORAD靶向miR-410-3p/HMGB1对胰腺癌细胞增殖、侵袭及肿瘤免疫微环境(TME)的影响.方法培养正常胰腺导管上皮细胞系HPDE6-C7和胰腺癌细胞系PANC-1、SW1990、AsPC-1,RT-qPCR检测NORAD/miR-410-3p/HMGB1轴在胰腺癌中的表达.将PANC-1细胞分为NC组、sh-NC(NORAD敲低阴性对照)组、sh-NORAD(NORAD敲低)组、sh-NORAD+anti-miR-410-3p(miR-410-3p抑制剂)组、sh-NORAD+oe-HMGB1(HMGB1过表达)组.RT-qPCR和West-ern blot检测各组PANC-1细胞转染效率.CCK-8、EdU染色、免疫荧光、Transwell法检测各组PANC-1细胞增殖和侵袭;流式细胞仪和ELISA检测各组PANC-1细胞TME情况;双荧光素酶报告基因和RIP实验检测miR-410-3p与NORAD或HMGB1靶向关系.结果与sh-NC组比较,sh-NORAD组NORAD、HMGB1、OD值、EdU阳性率、Ki-67阳性率、N-cadherin阳性率、侵袭数量、CD206^(+)/CD68^(+)、CD163含量、Arg-1含量均下调,miR-410-3p表达、E-cadherin阳性率、CD86^(+)/CD68^(+)、iNOS含量上调(P<0.05);与sh-NORAD组比较,sh-NORAD+anti-miR-410-3p组和sh-NORAD+oe-HMGB1组NORAD、HMGB1、OD值、EdU阳性率、Ki-67阳性率、N-cadherin阳性率、侵袭数量、CD206^(+)/CD68^(+)、CD163含量、Arg-1含量均上调,miR-410-3p表达、E-cadherin阳性率、CD86^(+)/CD68^(+)、iNOS含量下调(P<0.05).miR-410-3p与NORAD或HMGB1存在靶向调控关系(P<0.05).结论NORAD靶向miR-410-3p/HMGB1轴,促进胰腺癌细胞增殖、侵袭及M2型巨噬细胞极化,从而重塑TME.
Objective To explore the impacts of LncRNA NORAD on the proliferation,invasion and tumor immune microen-vironment(TME)of pancreatic cancer cells by targeting miR-410-3p/HMGB1.Methods The normal pancreatic duct epithelial cell line HPDE6-C7 and pancreatic cancer cell lines PANC-1,SW1990,AsPC-1 were cultured.RT-qPCR was used to detect the expres-sion of NORAD/miR-410-3p/HMGB1 axis in pancreatic cancer.PANC-1 cells were separated into NC group,sh-NC(NORAD knock-down negative control)group,sh-NORAD(NORAD knockdown)group,sh-NORAD+anti-miR-410-3p(miR-410-3p inhibitor)group,and sh-NORAD+oe-HMGB1(HMGB1 overexpression)group.RT-qPCR and Western blot were used to measure the transfec-tion efficiency of PANC-1 cells in each group.CCK-8,EdU staining,immunofluorescence,and Transwell assay were implemented to measure the proliferation and invasion of PANC-1 cells in each group.Flow cytometry and ELISA were implemented to measure the TME status of PANC-1 cells in each group.In addition,the dual luciferase reporter gene and RIP assay were used to detect the targe-ting relationship between miR-410-3p and NORAD or HMGB1.Results Compared with the sh-NC group,the sh-NORAD group showed downregulation of NORAD,HMGB1,OD value,EdU positivity rate,Ki-67 positivity rate,N-cadherin positivity rate,invasion quantity,CD206^(+)/CD68^(+),CD163 content,and Arg-1 content,while upregulation of miR-410-3p expression,E-cadherin positivity rate,CD86^(+)/CD68^(+),and iNOS content(P<0.05).Compared with the sh-NORAD group,the sh-NORAD+anti-miR-410-3p group and sh-NORAD+oe-HMGB1 group showed upregulation of NORAD,HMGB1,OD value,EdU positivity rate,Ki-67 positivity rate,N-cadherin positivity rate,invasion quantity,CD206^(+)/CD68^(+),CD163 content,and Arg-1 content,while downregulation of miR-410-3 p expression,E-cadherin positivity rate,CD86^(+)/CD68^(+),and iNOS content(P<0.05).Moreover,miR-410-3p had a targeted reg-ulatory relationship with NORAD or HMGB1(P<0.05).Conclusion NORAD targets the miR-410-3p/HMGB1 axis,promotes the proliferation and invasion of pancreatic cancer cells and its polarization to M2 macrophages,thus remodeling TME.
作者
王运良
王东华
贾喜花
WANG Yun-liang;WANG Dong-hua;JIA Xi-hua(Department of Oncology,TheFirst Central Hospital of Baoding City,Baoding 071000,China)
出处
《肝胆外科杂志》
2026年第1期65-71,共7页
Journal of Hepatobiliary Surgery
基金
河北省医学科学研究课题(20211822)。
