肿瘤相关成纤维细胞上调hsa-miR-18b-5p靶向FBXL3促进前列腺癌的增殖及转移

扫码查看

原文链接

国家科技期刊平台
NETL
NSTL
万方数据

中文摘要：目的探讨受肿瘤相关成纤维细胞(CAFs)调控的hsa-miR-18b-5p在前列腺癌(PCa)中的表达水平及作用，并研究对其在PCa发生发展过程中的分子机制。方法利用生物信息学技术分析在PCa中高表达的miRNA，构建肿瘤相关成纤维细胞并与PCa细胞系共培养，验证CAFs对PCa细胞增殖和迁移的影响及对hsa-miR-18b-5p的调控作用;RT-qPCR验证20例患者癌组织及癌旁正常组织、前列腺正常上皮增生细胞及PCa各细胞系中hsa-miR-18b-5p的表达水平;通过脂质体法将阴性对照及hsa-miR-18b-5p抑制物分别转染入PCa细胞C4-2、LNCAP中，分为NC inhibitor组和hsa-miR-18b-5p inhibitor组;采用细胞集落形成、CCK-8实验、划痕愈合、Transwell、IC50实验、流式细胞术分别检测C4-2、LNCAP细胞增殖、迁移、侵袭、耐药能力、凋亡和周期;建立PCa裸鼠移植瘤模型，定期测量移植瘤的质量和体积，Kaplan-Meier生存曲线分析裸鼠生存情况;利用靶基因预测分析网站(Targetscan、Mirtarbase、miRDB、miRDIP)预测has-miR-18b-5p的靶基因，并通过双荧光素酶报告基因分析验证hsa-miR-18b-5p与靶基因的靶向关系，RT-qPCR、Western blotting检测各组细胞靶基因的表达水平。结果生物信息学分析结果显示，在PCa中高表达的miRNA有17个，结合差异表达程度以及相关文献筛选出拟研究的基因:miR-148a、miR-17、miR-18b-5p、miR-770、miR-297-3p。CAFs与PCa细胞系共培养后hsa-miR-18b-5p的表达水平升高(P<0。01)，且促进PCa细胞的增殖与迁移(P<0。01)，敲除has-miR-18b-5p可抵消CAFs对PCa细胞增殖及迁移的影响，确定has-miR-18b-5p为研究基因。与正常前列腺上皮细胞或肿瘤旁组织相比，PCa细胞或肿瘤组织中hsa-miR-18b-5p的表达升高(P<0。05);敲除hsa-miR-18b-5p可抑制C4-2、LNCAP细胞的增殖、迁移、侵袭以及耐药(P<0。05);敲除hsa-miR-18b-5p可抑制裸鼠移植瘤质量和体积的增长，增加裸鼠的生存时间(P<0。05)。靶基因预测分析网站显示，FBXL3是hsa-miR-18b-5p的一个潜在作用靶点，双荧光素酶报告基因证实has-miR-18b-5p与FBXL3基因间存在结合位点，且敲除hsa-miR-18b-5p可增加C4-2、LNCAP细胞中FBXL3的蛋白表达(P<0。05)。结论 CAFs上调前列腺癌细胞hsa-miR-18b-5p的表达水平，后者可能通过靶向调控FBXL3基因的表达促进PCa细胞的增殖和转移。

外文标题：Tumor-associated fibroblasts promotes proliferation and migration of prostate cancer cells by suppressing FBXL3 via upregulating hsa-miR-18b-5p

外文摘要：Objective To explore the mechanism of tumor-associated fibroblasts(CAFs)for regulating proliferation and migration of prostate cancer(PCa)cells.Methods We conducted a bioinformatics analysis to identify miRNAs with high expression in PCa.The proliferation,migration and hsa-miR-18b-5p expression levels were observed in PCa cells co-cultured with CAFs.We further examined hsa-miR-18b-5p expression level in 20 pairs of PCa and adjacent tissue samples and in different PCa cell lines and normal epithelial cells using RT-qPCR.In PCa cell lines C4-2 and LNCAPNC,the effects of transfection with a hsa-miR-18b-5p inhibitor on cell proliferation,migration,invasion,drug resistance,apoptosis and cell cycle were evaluated,and the effects of has-miR-18b-5p knockdown on C4-2 cell xenograft growth and mouse survival were observed in nude mice.Dual luciferase reporter gene assay was used to validate the targeting relationship between hsa-miR-18b-5p and its target genes,whose expressions were detected in PCa cells using RT-qPCR and Western blotting.Results The expression of hsa-miR-18b-5p was significantly increased in the co-culture of CAFs and PCa cell lines,which exhibited significantly enhanced proliferation and migration abilities.Transfection with has-miR-18b-5p inhibitor strongly attenuated the effect of CAFs for promoting proliferation and migration of PCa cells,and in C4-2 and LNCAP cells cultured alone,inhibition of hsa-miR-18b-5p obviously suppressed cell proliferation,migration,invasion,and drug resistance.In the tumor-bearing mice,hsa-miR-18b-5p knockdown in the transplanted cells significantly inhibited xenograft growth and increased the survival time of the mice.Target gene prediction suggested that FBXL3 was a potential target of hsa-miR-18b-5p,and dual luciferase reporter gene confirmed a binding site between them.In C4-2 and LNCAP cells,hsa-miR-18b-5p knockdown resulted in significantly increased expression levels of FBXL3.Conclusion CAFs promotes proliferation and migration of PCa cells by up-regulating hsa-miR-18b-5p to suppress FBXL3 expression.

外文关键词：

hsa-miR-18b-5ptumor-associated fibroblastsFBXL3prostate cancerpromoting cancer

作者：

骆金光、陶怀祥、闻志远、陈龙、胡昊、关翰

展开 >

作者单位：

蚌埠医科大学第一附属医院泌尿外科,安徽蚌埠 233004

蚌埠医科大学慢性疾病免疫学基础与临床安徽省重点实验室,安徽蚌埠 233030

关键词：

hsa-miR-18b-5p 肿瘤相关成纤维细胞 FBXL3 前列腺癌促癌

基金：

安徽省自然科学基金安徽省高等学校自然科学研究重点项目蚌埠医学院2023年度研究生科研创新计划项目慢性疾病免疫学基础与临床安徽省重点实验室开放课题基金

项目编号：

2008085QH3582023AH051942Byycx23114AHIAI2022K01

出版年：

2024

DOI：

10.12122/j.issn.1673-4254.2024.07.08

南方医科大学学报

南方医科大学

南方医科大学学报

CSTPCD北大核心

影响因子：1.654

ISSN：1673-4254

年,卷(期)：2024.44(7)