摘要
目的:研究长链非编码RNA核富集转录体1(lncRNA NEAT1)对肺腺癌PC-9细胞增殖能力的影响,并初步探讨其作用机制。方法:qPCR检测人肺腺癌PC-9细胞与人胚肺二倍体2BS细胞中lncRNA NEAT1的表达水平;设计并合成lncRNA NEAT1的小干扰RNA(siRNA)序列,采用脂质体法转染PC-9细胞,通过qPCR检测转染前后PC-9细胞NEAT1的表达水平。MTT法、流式细胞术分别检测lncRNA NEAT1敲低对PC-9细胞增殖及细胞周期的影响;WB检测转染前后DNA损伤相关蛋白,即共济失调毛细血管扩张突变蛋白(ATM)和双链DNA损伤标志物γ-H2AX的表达水平。结果:与2BS细胞相比,PC-9细胞中lncRNA NEAT1呈高表达(P<0.05)。成功建立NEAT1敲低的PC-9细胞,转染siRNA 12 h后siNEAT1-1及siNEAT1-2干扰组细胞增殖能力较空白对照组及空转染组明显下降(P<0.05);干扰组细胞周期被阻滞在G1期[(88.97±2.64)%(,88.15±1.48)%vs(84.5±1.72)%,P<0.05]和G2/M期[(8.35±2.02)%,(8.11±1.36)%vs(4.28±1.28)%,P<0.05];干扰组细胞中DNA损伤相关蛋白ATM和γ-H2AX表达水平显著升高(均P<0.05)。结论:lncRNA NETA1在肺腺癌PC-9细胞呈高表达,其可通过抑制DNA损伤导致细胞周期G1/M期转变促进肺腺癌细胞PC-9的增殖能力。
Objective: To investigate the effects of long non-coding RNA nuclear enriched abundant transcript 1(lncRNA NEAT1) on the proliferation of lung adenocarcinoma PC-9 cells and to explore its mechanism. Methods: qPCR was used to detect the expression level of lncRNA NEAT1 in human lung adenocarcinoma PC-9 cells and human embryonic lung diploid 2 BS cells. The sequence of small interfering RNA(siRNA) targeting lncRNA NEAT1 gene was designed and synthesized, and then transfected into PC-9 cells by liposome method. The expression level of NEAT1 in PC-9 cells before and after transfection was detected by qPCR. MTT and flow cytometry were used to detect the effect of lncRNA NEAT1 knockdown on proliferation and cell cycle distribution of PC-9 cells, respectively. WB assay was used to detect the expressions of DNA damage-related proteins, namely, double-stranded DNA breaks(DSBs)biomarker γ-H2 AX and ataxia-telangiectasia mutated(ATM), before and after transfection. Results: Compared with 2 BS cells, lncRNA NEAT1 was highly expressed in PC-9 cells(P<0.05). The PC-9 cells with lncRNA NEAT1 knock-down were successfully established. After being transfected with siRNA for 12 h, the proliferation of PC-9 cells in siNEAT1 group and siNEAT2 group significantly decreased as compared with the blank control group and the empty transfection group(P<0.05). In the interference groups,cell cycle was arrested in G1 phase([88.97±2.64]%,[88.15±1.48]% vs [84.5±1.72]%, P<0.05) and G2/M phase([8.35±2.02]%,[8.11±1.36]% vs [4.28±1.28]%, P<0.05). The expression levels of DNA damage-related proteins ATM and γ-H2 AX in the interference groups were significantly increased(all P<0.05). Conclusion: lncRNA NEAT1 is highly expressed in lung adenocarcinoma PC-9 cells. lncRNA NEAT1 inhibits DNA damage and causes cell cycle at G1/M phase switch, and thus promotes the proliferation of lung adenocarcinoma cells.
作者
刘天旭
王梦杰
田聪
刘琰
吕微
贾云泷
刘丽华
LIU Tianxu;WANG Mengjie;TIAN Cong;LIU Yan;LYU Wei;JIA Yunlong;LIU Lihua(Department of Tumor Immunotherapy,the Fourth Hospital of Hebei Medical University,Shijiazhuang 050035,Hebei,China)
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2019年第8期845-849,共5页
Chinese Journal of Cancer Biotherapy
基金
国家自然科学基金资助项目(No.81871894)
河北省自然科学基金资助项目(No.H2018206318)~~
