| Goal and purpose of the research:Despite advances in surgery and chemotherapy over the past few decades,gastric cancer(GC)remains the third most common cause of cancer-related mortality worldwide,accounting for 8.8%of all cancer-related deaths.The lack of biomarkers in early detection and peritoneal metastasis are the main reasons for the failure of gastric cancer treatment.Therefore,finding molecular biomarkers that can predict disease progression may promote drug development and treatment strategies for gastric cancer.Research method:High-throughput mRNA sequencing(RNA-seq)revealed that collagen type X alpha 1(COL10A1)is a disease progression-associated gene.QPCR analysis of tissue samples from 103 patients with gastric cancer showed that high COL10A1 mRNA expression was associated with gastric cancer metastasis and reduced survival.We analyzed the COL10A1 promoter using the UCSC genome website and JASPAR database,and we found potential SOX9 binding site.The cancer tissues and corresponding normal tissues of gastric cancer patients were collected,and the localization and quantitative analysis of SOX9 and COL10A1 expression were performed by the technique of immunohistochemical.At the same time,the clinical data of the examined tissues and related information on the survival prognosis were collected.The clinical relevance of SOX9 and COL10A1 was then verified by QPCR and Western blot at the transcriptional and translational levels,respectively.The stably transfected cell line overexpressing COL10A1 gene and the stable cell line of COL10A1 knockdown were constructed,and the gastric cancer cell lines were stimulated by TGF-?1 recombinant factor.TGF-? signal,tumor-associated EMT markers and metal matrix enzyme family proteins were detected by Western blot.We verified the relationship between COL10A1 and TGF-? signaling pathways.Research content:In this study,we demonstrated that both SOX9 and COL10A1 are up-regulated in gastric cancer tissues.We observed a positive correlation between the expression patterns of SOX9 and COL10A1 in gastric cancer cell lines and tissues.The results of electrophoretic mobility shift assay(EMSA),chromatin immunoprecipitation(ChIP)analysis and promoter reporter indicate that SOX9 can directly bind to the COL10A1 gene promoter and activate its transcription.Biological function experiments showed that COL10A1 regulates the migration and invasion of gastric cancer cells.Knockdown of COL10A1 inhibited lung and abdominal cavity metastasis in nude mouse metastasis models.Moreover,transforming growth factor-?1(TGF-?1)treatment up-regulated the phosphorylation of Smad2 and increased SOX9 and COL10A1 expression.COL10A1 was confirmed to be a potential inducer of epithelial-to-mesenchymal transition(EMT).SOX9 was essential for COL10A1-mediated EMT,and cell migration,invasion and metastasis.Co-expression of SOX9 and COL10A1 was associated with tumor progression and was strongly predictive of overall survival in GC patients.In summary,this study elucidated the mechanistic link between COL10A1 and the TGF-?1-SOX9 axis.Main conclusion:Our mRNA expression profiling analyses have indicated that COL10A1 is significantly elevated during the development and progression of gastric cancer.The expression of COL10A1 is markedly overexpressed in gastric cancer patients and enhances malignant and metastatic ability in vivo.Increased mRNA and protein expression of COL10A1 are associated with shorter survival in gastric cancer patients.Moreover,COL10A1 promotes cell migration and invasion through positive transcriptional regulation of SOX9 and the involvement of the TGF-? signaling pathway.Therefore,this study not only elucidates the precise mechanism of COL10A1 in the progression of gastric cancer but also provides molecular insight into cancer-related ECM and may serve as a predictor of clinical outcome in gastric cancer. |
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