Ⅺ型胶原α1蛋白调节蛋白激酶通路促进人肺腺癌原代细胞迁移和侵袭

COL11A1 regulates PI3K/Akt/GSK-3βpathway and promotes human lung adenocarcinoma primary cell migration and invasion

目的探讨Ⅺ型胶原α1蛋白(COL11A1)在肺腺癌迁移和侵袭中的作用和机制。方法采用2020年9至11月贵州医科大学附属医院收治的4例肺腺癌患者手术病理组织,经免疫组织化学方法鉴定肺腺癌组织及癌旁组织并行转录组测序,使用TCGA和GTEx数据库行单基因预后分析。用Western blot法检测肺腺癌组织和癌旁组织中COL11A1基因表达水平。提取及培养人肺腺癌原代细胞。COL11A1 siRNA转染人肺腺癌原代细胞后转录组测序,差异基因做KEGG富集分析,分析差异基因富集的通路,Western blot法检测该通路各关键位点蛋白表达及磷酸化水平变化,划痕愈合实验检测细胞迁移、CCK8法检测细胞增殖、Transwell检测侵袭能力。结果肺腺癌组织转录测序筛选差异表达较高的10个基因,单基因预后分析发现COL11A1基因表达水平与生存率相关(P<0.001);Western blot法检发现COL11A1在肺腺癌组织中表达较癌旁组织高(P<0.001);COL11A1 siRNA转染人肺腺癌原代细胞后转录组测序发现差异基因集中在PI3K-akt通路上,Western blot检测发现抑癌基因PTEN在siRNA转染组中表达显著高于对照组和阴性转染组,而Akt、p-Akt 473、p-Akt 308、p-PTEN、p-PDK1、p-c-Raf、p-GSK-3β基因表达均显著下调(均P<0.05),siRNA转染组相较于阴性对照组人肺腺癌原代细胞迁移、增殖、侵袭能力均降低(均P<0.05)。结论COL11A1调节PI3K/Akt/GSK-3β通路促进人肺腺癌原代细胞迁移和侵袭。

Objective To investigate the role and mechanism of COL11A1 in lung adenocarcinoma migration and invasion.Methods Surgical pathological tissues of 4 patients with lung adenocarcinoma admitted to the Affiliated Hospital of Guizhou Medical University from September to November 2020 were used.Immunohistochemical methods were used to identify lung adenocarcinoma tissues,para-cancerous tissues and parallel transcriptome sequencing.Genetic prognostic analysis was conducted by TCGA and GTEx databases.The expression level of COL11A1 gene in lung adenocarcinoma and adjacent tissues was detected by Western blotting.The primary human lung adenocarcinoma cells cultured.The COL11A1 siRNA was transfected into primary human lung adenocarcinoma cells,then the transcriptome sequencing of differential genes was performed,and KEGG enrichment analysis of differential gene enrichment pathway was conducted.Protein expression and phosphorylation were detected by Western blot method.Cell migration was detected by scratch healing test.Cell proliferation was detected by CCK8 method and invasion ability was detected by Transwell method.Results Ten differentially expressed genes were screened by transcription sequencing in lung adenocarcinoma.Prognostic analysis of single gene showed that COL11A1 gene expression level was correlated with survival rate(P<0.001).The expression of COL11A1 in lung adenocarcinoma was higher than that in adjacent tissues by Western blot(P<0.001).Transcriptome sequencing of COL11A1 siRNA transfection into primary human lung adenocarcinoma cells showed that differential genes were concentrated in PI3K-akt pathway.The expression of tumor suppressor gene PTEN in siRNA transfection group was significantly higher than that in control group and negative transfection group by Western blot.The expression of Aktp-Akt 473 p-Akt 308 p-PTENp-PDK1p-c-Rafp-GSK-3βwas down-regulated(all P<0.05).Compared with the negative control group,the ability of migration,proliferation and invasion of primary human lung adenocarcinoma cells in siRNA transfection group decreased(all P<0.05).COL11A1 regulates PI3K/Akt/GSK-3βpathway to promote migration and invasion of primary human lung adenocarcinoma cells.Conclusion COL11A1 regulates PI3K/Akt/GSK-3βpathway to promote migration and invasion of primary human lung adenocarcinoma cells.