摘要
目的探讨转录因子c-Myc的R-loops结构及其对胶质瘤细胞恶性生物学行为的调控作用。方法采用DNA-RNA免疫沉淀PCR(DNA-RNA immunoprecipitation PCR,DRIP-PCR)法检测胶质瘤细胞(U251和U87-MG)中c-Myc的R-loops结构。分别将空载质粒、c-Myc过表达质粒、RNASEH1过表达质粒转染至U251和U87-MG细胞,设立对照组(CTRL组)、核糖核酸内切酶H1(RNASEH1)过表达组(RNASEH1-OE组)、c-Myc过表达组(c-Myc-OE组)及共转染c-Myc与RNASEH1共转染组(c-Myc-OE+RNASEH1-OE组)。采用细胞计数试剂盒-8(CCK-8)、5-乙炔基-2'-脱氧尿苷(EdU)实验检测细胞增殖能力;通过细胞划痕实验和Transwell实验评估细胞迁移和侵袭水平;利用蛋白印迹法(Western blot)检测RNASEH1和c-Myc的蛋白表达水平。结果转录因子c-Myc在胶质瘤细胞中存在R-loops结构(P<0.05)。过表达RNASEH1可清除c-Myc依赖的R-loops结构,从而抑制胶质瘤细胞的增殖、侵袭和迁移(P<0.05)。c-Myc的过表达可提高核糖核酸内切酶H1(RNASEH1)的表达,从而挽救其R-loops结构,促进胶质瘤细胞侵袭和迁移能力的影响(P<0.05)。结论转录因子c-Myc可能通过其R-loops结构促进胶质瘤细胞的增殖、侵袭和迁移等恶性生物学行为。
Objective To explore the c-Myc-dependent R-loop structure and its regulatory effect on the proliferation,invasion and migration of glioma cells.Methods DRIP-PCR was used to detect the R-loop structure of c-Myc in glioma cells(U251 and U87-MG).U251 and U87-MG were transfected into empty plasmids,c-Myc overexpression plasmids,and RNASEH1 overexpression plasmids respectively,which were used respectively to establish control group(CTRL group),RNASEH1 overexpression group(RNASEH1-OE group),c-Myc overexpression group(c-Myc-OE group),and co-transfected c-Myc and RNASEH1 overexpression group(c-Myc-OE+RNASEH1-OE group).CCK-8 assay,EdU assay,scratch assay,and Transwell assay were used to detect the proliferation,migration,and invasion levels of glioma cells.The Western blot was used to detect the expression levels of RNASEH1 and c-Myc proteins expressions in glioma cells.Results c-Myc had an R-loop structure in glioma cells(P<0.05).Overexpression of RNASEH1 could knock out the c-Myc-dependent R-loop structure,further reducing glioma cell proliferation,invasion,and migration(P<0.05).Overexpression of c-Myc could upregulate the expression of knockdown RNASEH1,thereby rescuing the c-Myc-dependent R-loop structure and enhancing the invasion and migration abilities of glioma cells(P<0.05).Conclusions c-Myc-dependent R-loop structure can promote the proliferation,invasion and migration of glioma cells.
作者
林家汉
陈彩燕
陈申波
叶富跃
杨堃
LIN Jiahan;CHEN Caiyan;CHEN Shenbo;YE Fuyue;YANG Kun(Department of Neurology,First Affiliated Hospital of Hainan Medical University,Haikou,Hainan 570000,China;Department of Critical Care Medicine,First Affiliated Hospital of Hainan Medical University,Haikou,Hainan 570000,China)
出处
《中华神经外科疾病研究杂志》
2026年第2期16-22,共7页
Chinese Journal of Neurosurgical Disease Research
基金
国家自然科学基金地区项目(82060456)
海南省自然科学基金青年项目(823QN350)
海南省“南海新星”科技创新人才平台项目(NHXXRCXM202351)
海南医科大学第一附属医院青年培育基金(HYYFYPY202303)。
