摘要
目的结合生物信息学分析,探讨趋化因子CXc配体L12(CXCL12)、白细胞介素6(IL-6)和吲哚胺-2,3-双加氧酶(IDO)在去势抵抗性前列腺癌细胞中的表达水平。方法利用激素敏感性前列腺癌细胞株LNCaP和去势抵抗性前列腺癌细胞株C4-2细胞的基因表达谱,采用R3.6.1软件对2组的表达数据进行差异基因分析,并根据差异基因进行基因本体(GO)功能富集分析,筛选差异基因富集的京都基因和基因组百科全书(KEGG)信号通路,最终选定CXCL12、IL-6和IDO三个差异基因;将LNCaP和C4-2细胞分为control组(常规处理)、si-NC组(将si-NC转染入细胞内)、si-IL-6组(将si-IL-6转染入细胞内)和si-CXCL12组(将si-CXCL12转染乳细胞内);采用实时荧光定量PCR(RT-PCR)及Western blot法检测LNCaP和C4-2细胞株中CXCL12、IL-6和IDO各自mRNA及蛋白表达水平;使用si-IL-6、si-CXCL12敲低IL-6、CXCL12在LNCaP、C4-2细胞中的表达,采用qRT-PCR法检测细胞中IDO、CXCL12和IL-6基因表达,WB检测相关蛋白表达,MTT法检测LNCaP或C 4-2细胞增殖、流式凋亡术检测细胞凋亡、Transwell实验检测细胞侵袭能力、划痕恢复实验检测细胞迁移能力。结果生物信息学分析结果显示,细胞株LNCaP和细胞株C4-2之间共有3505个差异基因,其中上调基因1695个、下调的基因1810个;RT-PCR法和WB检测显示,与C4-2细胞比较,LNCaP细胞中IL-6和IDOmRNA及蛋白表达量均明显增高,CXCL12 mRNA及蛋白量显著降低,差异有统计学意义(P<0.05);将si-IL-6转染入LNCaP和C4-2细胞后,与si-NC组比较,LNCaP细胞的si-IL-6组中细胞增殖、侵袭和迁移能力受到显著抑制,细胞凋亡率显著增加,差异有统计学意义(P<0.01);与si-NC组比较,si-IL-6组中IL-6和IDO蛋白和mRNA表达水平显著降低,差异有统计学意义(P<0.01);将si-ICXCL12转染入LNCaP或C4-2细胞后,与正常C4-2细胞比较,si-CXCL12组中C4-2细胞增殖、侵袭和迁移能力受到显著抑制,同时细胞凋亡率显著增加,差异有�
Objective To explore the expression levels of chemokine CXC ligand L12(CXCL12),interleukin-6(IL-6),and indoleamine 2,3-dioxygenase(IDO)in castration-resistant prostate cancer cells,combining with bioinformatics analysis.Methods R3.6.1 software was run on gene expression profiles of hormone-sensitive prostate cancer cell line LNCaP and castration-resistant prostate cancer cell line C4-2 to analyze differentially expressed genes in above two cell lines.Gene ontology(GO)functional enrichment analysis and Kyoto genes and the Genome Encyclopedia(KEGG)signaling pathway enrichment were performed on the differentially expressed genes.Three differentially expressed genes were ultimately selected:CXCL12,IL-6,and IDO.Real time fluorescence quantitative PCR(RT-qPCR)and Western blot were used to detect the mRNA and protein expression levels of CXCL12,IL-6,and IDO in LNCaP and C4-2 cell lines.si-IL-6 and si-CXCL12 were used to knockdown the expressions of IL-6 and CXCL12 in LNCaP and C4-2 cells,respectively.RT-qPCR and Western blot were used to detect mRNA and protein expressions of IDO,CXCL12,and IL-6.MTT assay was used to measure cell proliferation of LNCaP or C4-2 cells.Flow cytometry was used to examine cell apoptosis.Transwell assay was performed to examine cell invasion capacity.Wound healing assay was applied to examine cell migration capacity.Results Bioinformatics analysis showed that there were 3505 differentially expressed genes shared by cell lines LNCaP and C4-2,including 1695 upregulated genes and 1810 downregulated genes.Divide LNCaP and C4-2 cells into control group(conventional treatment),si-NC group(transfection of si-NC into cells),si-IL-6 group(transfection of si-IL-6 into cells),and si-CXCL12 group(transfection of si-CXCL12 into milk cells);RT-qPCR and Western blot results showed that when compared with C4-2 cells,the mRNA and protein expression levels of IL-6 and IDO in LNCaP cells were significantly increased,while the mRNA and protein expression levels of CXCL12 were significantly decreased(P<0.05).Afte
作者
金宏亮
宣睿
许立军
JIN Hongliang;XUAN Rui;XU Lijun(Department of Urologic Surgery,the Second Affiliated Hospital of Soochow University,Suzhou 215004,Jiangsu,China)
出处
《贵州医科大学学报》
CAS
2023年第10期1156-1165,共10页
Journal of Guizhou Medical University
基金
江苏省自然基金面上项目(BK20191170)。
