长链非编码RNA MALAT1调控星形胶质细胞增殖和凋亡及对MAPK/ERK1,2信号通路的影响OACSTPCD
Long non-coding RNA MALAT1 regulates astrocyte proliferation and apoptosis and affects MAPK/ERK1,2 signaling pathway
目的 探讨肺腺癌转移相关转录本1(MALAT1)的表达对星形胶质细胞(C8-D1A)增殖和凋亡,以及丝裂原活化蛋白激酶(MAPK)/细胞外信号调节激酶(ERK1,2)通路的影响.方法 通过慢病毒载体和质粒载体在C8-D1A细胞中分别敲低及过表达MALAT1基因,用实时荧光定量PCR(qPCR)法检测敲低及过表达效率、细胞增殖相关标志物Ki67、微染色体维持蛋白2(MCM2)、细胞核增殖抗原(PCNA)、凋亡相关蛋白含半胱氨酸的天冬氨酸蛋白水解酶3(Caspase-3)、B细胞淋巴瘤2(Bcl-2)、Bcl-2相关X蛋白(Bax)的表达变化,CCK-8、流式细胞术检测敲低及过表达MALAT1后对C8-D1A细胞增殖和凋亡的影响.免疫荧光染色法检测Caspase-3和Ki67蛋白表达,Western印迹法检测细胞凋亡相关蛋白Caspase-3、Bax、Bcl-2及ERK1/2、p-ERK 1/2、p38MAPK、p-p38MAPK蛋白表达变化情况.结果 与对照组相比,过表达MALAT1抑制细胞增殖并诱导细胞凋亡,其G0/G1期、G2/M期比例升高,S期比例降低,Caspase-3蛋白表达升高,p-ERK 1/2蛋白表达降低,Ki67、PCNA mRNA表达量降低;而敲低MALAT1后细胞增殖显著增加并抑制细胞凋亡,其S期比例增高、G,/M期比例降低,Caspase-3、Bax蛋白表达降低,p-ERK1/2、p-p38MAPK蛋白表达升高,Ki67、MCM2、PCNA mRNA表达量增加(P＜0.05).结论 MALAT1基因可能通过调控MAPK/ERK1,2信号通路进而促进星形胶质细胞凋亡并抑制其增殖.
Objective To investigate the effect of MALAT1 expressions on cell proliferation and apoptosis in astrocytes by regulating mitogen-activated protein kinase(MAPK)/extracellular signal-regulated kinase(ERK1,2)pathway.Methods The MALAT1 gene was knocked down and over-expressed in C8-D1A cells by lentiviral and plasmid vectors,respectively.The expressions of MALAT1,cell proliferation-related markers(Ki67,MCM2,PCNA)and apoptosis-related proteins(Caspase-3,Bax,Bcl-2)were detected by quantitative real-time polymerase chain reaction(qPCR).CCK-8 assay and flow cytometry were used for cell proliferation and apoptosis in C8-D1A cells.Immunofluorescence was adopted to detect the protein expressions of Caspase-3 and Ki67.Western blotting was used to detect the protein expressions of Caspase-3,Bax,Bcl-2,ERK1/2,p-ERK1/2,p38MAPK and p-p38MAPK.Results Compared with the control group,over-expressed MALAT1 inhibited cell proliferation and induced cell apoptosis in C8-D1A cells while the knockdown of MALAT1 significantly enhanced cell proliferation and anti-apoptotic ability in C8-D1A cells.The proportion of C8-D1A cells in G0/G1-phase and G2/M-phase was higher than in the control group as evidenced by flow cytometry,but was lower in S-phase.Meanwhile,data showed that Caspase-3 was increased while p-ERK1/2 was decreased in terms of protein levels.The mRNA expressions of Ki67 and PCNA were decreased.After knockdown of MALAT1,the proportion of C8-D1A cells in S-phase was higher,but was lower in G2/M-phase.The protein expressions of Caspase-3 and Bax decreased while those of p-ERK1/2 and p-p38MAPK increased.The mRNA expressions of Ki67,MCM2 and PCNA were increased.The differences were all statistically significant(P＜0.05).Conclusion MALAT1 promotes astrocyte apoptosis and inhibits proliferation by regulating the MAPK/ERK1,2 signaling pathway.
胡辉;钱令嘉;王雪;吴雨涵;董华丰;张玲;韦爱君;谢方;赵云;孙兆炜
军事科学院军事医学研究院,北京 100850
临床医学
肺腺癌转移相关转录本1星形胶质细胞丝裂原活化蛋白激酶细胞外信号调节激酶细胞凋亡细胞增殖
MALAT1astrocytesmitogen-activated protein kinaseextracellular signal-regulated kinasecell apoptosiscell proliferation
《军事医学》 2024 (005)
347-354 / 8
国家自然科学基金项目(82271553),北京市自然科学基金项目(5222033)
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