RACK1对人宫颈癌细胞增殖和凋亡的影响及其机制  

Effect of RACK1 on the proliferation and apoptosis of human cervical cancer cells

作　　者：李金秋 徐丽秀[1] 朱佳瑜 夏依达·吐尔逊 阿仙姑·哈斯木[1] Li Jin-Qiu;Xu Li-Xiu;Zhu Jia-Yu;Xia Yida·Turxun;Ashamgul·Hasim(Basic Medicine College of Xinjiang Medical University/Xinjiang Key Laboratory of Molecular Biology of Endemic Diseases,Urumqi 836001,China)

机构地区：[1]新疆医科大学基础医学院/新疆地方病分子生物学重点实验室,乌鲁木齐836001

出　　处：《解放军医学杂志》2022年第5期442-450,共9页Medical Journal of Chinese People's Liberation Army

基　　金：新疆维吾尔自治区自然科学基金(2022D01D13);新疆维吾尔自治区天山创新团队项目(2021D14002)。

摘　　要：目的探讨激活性蛋白激酶C受体1(RACK1)对人宫颈癌细胞增殖和凋亡的影响及其潜在的机制。方法收集2020年12月-2021年12月新疆医科大学第一附属医院24例患者的宫颈鳞癌(CSCC)组织及24例无宫颈病变的正常宫颈(NC)组织,采用qRT-PCR检测CSCC和NC组织中RACK1 mRNA的表达水平,并检测CSCC组织中增殖和凋亡相关基因c-myc、caspase-3、caspase-9、Bax、Bcl-2 mRNA的表达水平;采用Spearman秩相关分析CSCC组织中RACK1 mRNA与增殖和凋亡相关基因mRNA表达的相关性。采用qRT-PCR及Western blotting检测RACK1在宫颈癌细胞C33a、SiHa和正常宫颈内皮细胞H8中的表达情况。通过慢病毒转染C33a、SiHa细胞以沉默RACK1的表达,根据不同处理分为正常对照组(NC)、空载组(sh-NON)和沉默组1(shRACK1-1)、沉默组2(shRACK1-2),并验证细胞转染效率,采用MTT法及平板克隆实验检测细胞增殖能力,流式细胞术检测细胞凋亡水平,qRT-PCR检测RACK1沉默后各组细胞中c-myc、caspase-3、caspase-9、Bax、Bcl-2 mRNA的表达水平,Western blotting检测RACK1沉默后各组细胞中c-myc、caspase-3、caspase-9、Bax、Bcl-2以及磷酸化Janus激酶2(p-JAK2)、JAK2、磷酸化细胞信号传导与转录活化因子3(p-STAT3)、STAT3蛋白的表达水平。结果CSCC组织RACK1 mRNA表达水平明显高于NC组织(P<0.001)。Spearman秩相关分析结果显示,宫颈癌组织中RACK1 mRNA的表达水平与caspase-3(r=–0.679,P<0.001)、caspase-9(r=–0.735,P<0.001)、Bax(r=–0.691,P<0.001)mRNA表达水平呈明显负相关,而与c-myc(r=0.713,P<0.001)、Bcl-2(r=0.846,P<0.001)mRNA表达水平呈明显正相关。qRT-PCR与Western blotting检测结果显示,与H8细胞比较,C33a、SiHa细胞中RACK1 mRNA和蛋白表达水平均明显升高(P<0.001)。RACK1沉默后,与sh-NON组及NC组比较,shRACK1-1组及shRACK1-2组RACK1 mRNA和蛋白表达水平明显降低(P<0.001)。与sh-NON组比较,shRACK1-1组及shRACK1-2组细胞增殖和集落形成能力也明显降低(P<0.001Objective To investigate the effect of activated protein kinase C receptor 1(RACK1)on the proliferation and apoptosis of human cervical cancer cells and its potential mechanism.Methods The cervical cancer tissues of 24 patients diagnosed as cervical squamous cell carcinoma(CSCC),admitted in the First Affiliated Hospital of Xinjiang Medical University from Dec.2020 to Dec.2021,were collected as the experimental group(CSCC group),and 24 normal cervical(NC)tissues were collected as the control group(NC group).The expression levels of RACK1 mRNA in CSCC tissues and NC tissues and in proliferation and apoptosis related genes c-myc,caspase-3,caspase-9,Bax and Bcl-2 in CSCC tissue samples were detected by qRT-PCR,and the correlation between RACK1 mRNA expression and apoptosis-related genes was analyzed by Spearman rank correlation.qRT-PCR and Western blotting were used to detect the expression of RACK1 in cervical cancer cells C33a and SiHa and normal cervical epithelial cells H8.C33a and SiHa cells were transfected with lentivirus to silence the expression of RACK1.According to different treatments,they were divided into:normal control(NC)group,sh-NON(RACK1 silent expression no-load group),shRACK1-1(RACK1 silent expression group 1),shRACK1-2(RACK1 silent expression group 2),and the transfection efficiency was verified.MTT assay and plate cloning assay were used to detect cell proliferation,and flow cytometry was performed to detect cell apoptosis.After RACK1 was silenced,qRT-PCR was used to detect the expression levels of c-myc,caspase-3,caspase-9,Bax and Bcl-2.Western blotting was used to detect the expression levels of c-myc,caspase-3,caspase-9,Bax,Bcl-2,phosphorylated Janus kinase 2(p-JAK2),JAK2,phosphorylated cell signal transduction and transcription activating factor 3(p-STAT3)and STAT3.Results The expression level of RACK1 mRNA was significantly higher in CSCC tissue than that in NC tissue(P<0.001).Spearman rank correlation analysis showed that the expression level of RACK1 mRNA in cervical cancer tissues was si

关 键 词：宫颈癌 激活性蛋白激酶C受体1 细胞增殖 细胞凋亡 

分 类 号：R737.33[医药卫生—肿瘤]