Li Nan, Liu Ming, Cao Xiaohui, Li Wei, Li Yunfang, Zhao Zongmao
Department of Radiotherapy and Oncology, The Third Hospital of Hebei Medical University, Shijiazhuang, Hebei 050051, P.R. China.
Department of Neurosurgery, The Second Hospital of Hebei Medical University, Shijiazhuang, Hebei 050000, P.R. China.
Oncol Lett. 2021 Oct;22(4):693. doi: 10.3892/ol.2021.12954. Epub 2021 Aug 1.
Non-small cell lung cancer (NSCLC) is a major cause of cancer-associated mortality worldwide, and bone metastasis is the most prevalent event observed in patients with advanced NSCLC. However, the pathogenesis of bone metastases has not been fully elucidated. In the present study, differentially expressed genes (DEGs) were identified by gene expression microarray analysis of NSCLC tissue samples with or without bone metastases. Subsequently, collagen type 6A1 (COL6A1) was chosen as the target gene through Ingenuity Pathway Analysis and reverse transcription-quantitative (RT-q) PCR validation of the top eight DEGs. COL6A1 was overexpressed or knocked down, and the proliferation and invasion of NSCLC cells was assessed using Cell Counting Kit-8, colony formation and Transwell invasion assays. Additionally, the osteogenic capacity of HOB and hES-MP 002.5 cells was assessed using RT-qPCR, western blotting, Alizarin Red and alkaline phosphatase staining. A total of 364 DEGs were identified in NSCLC tissues with bone metastases compared with NSCLC tissues without bone metastases, including 140 upregulated and 224 downregulated genes. Gene Ontology analysis results demonstrated that the upregulated and downregulated genes were primarily enriched in 'cellular process', 'metabolic process' and 'biological regulation'. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis revealed that the upregulated genes were primarily enriched in 'cysteine and methionine metabolism', 'oxidative phosphorylation' and 'ribosome', whereas the downregulated genes were primarily enriched in the 'transcriptional misregulation in cancer', 'ribosome' and 'mitophagy-animal' pathways. COL6A1 was highly expressed in NSCLC tissue samples with bone metastases. Functionally, COL6A1 overexpression induced the proliferation and invasion of HARA cells, and its knockdown inhibited the proliferation and invasion of HARA-B4 cells. Finally, it was demonstrated that HOB and hES-MP 002.5 cells exhibited osteogenic capacity, and overexpression of COL6A1 in HARA cells increased the adhesion of these cells to the osteoblasts, whereas knockdown of COL6A1 in HARA-B4 cells reduced their adhesive ability. In conclusion, COL6A1 may serve as a potential diagnostic marker and therapeutic target for bone metastasis in NSCLC.
非小细胞肺癌（NSCLC）是全球癌症相关死亡的主要原因，骨转移是晚期NSCLC患者中最常见的事件。然而，骨转移的发病机制尚未完全阐明。在本研究中，通过对有或无骨转移的NSCLC组织样本进行基因表达微阵列分析，鉴定出差异表达基因（DEG）。随后，通过Ingenuity Pathway Analysis和对前八个DEG进行逆转录定量（RT-q）PCR验证，选择6A1型胶原蛋白（COL6A1）作为靶基因。对COL6A1进行过表达或敲低，并使用细胞计数试剂盒-8、集落形成和Transwell侵袭试验评估NSCLC细胞的增殖和侵袭。此外，使用RT-qPCR、蛋白质印迹、茜素红和碱性磷酸酶染色评估HOB和hES-MP 002.5细胞的成骨能力。与无骨转移的NSCLC组织相比，在有骨转移的NSCLC组织中共鉴定出364个DEG，包括140个上调基因和224个下调基因。基因本体分析结果表明，上调和下调基因主要富集在“细胞过程”、“代谢过程”和“生物调节”中。京都基因与基因组百科全书通路富集分析显示，上调基因主要富集在“半胱氨酸和甲硫氨酸代谢”、“氧化磷酸化”和“核糖体”中，而下调基因主要富集在“癌症中的转录失调”、“核糖体”和“线粒体自噬-动物”通路中。COL6A1在有骨转移的NSCLC组织样本中高表达。在功能上，COL6A1过表达诱导HARA细胞的增殖和侵袭，其敲低抑制HARA-B4细胞的增殖和侵袭。最后，证明HOB和hES-MP 002.5细胞具有成骨能力，HARA细胞中COL6A1的过表达增加了这些细胞与成骨细胞 的粘附，而HARA-B4细胞中COL6A1的敲低降低了它们的粘附能力。总之，COL6A1可能作为NSCLC骨转移的潜在诊断标志物和治疗靶点。
