Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China.
Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China; Department of Urology, Hunan Cancer Hospital, The Affiliated Cancer Hospital of Xiangya, School of Medicine of Central South University, Changsha, Hunan, China.
Biomol Biomed. 2023 Sep 4;23(5):802-814. doi: 10.17305/bb.2023.8969.
A significant decrease in LINC00467 expression in testicular germ cell tumors (TGCTs) was found in our previous study in comparison to adjacent tissue. Interestingly, the expression of LINC00467 correlated with the pathological grade of the tumor in TGCT patients. The higher the expression of LINC00467 was, the worse the prognosis of the patients with TGCT was. Despite these findings, the exact role of LINC00467 in the development of TGCTs requires further investigation. LINC00467 expression was downregulated in the NCCIT and TCam-2 cell lines via small interfering RNA (siRNA) silencing. The levels of gene expression were validated using quantitative real-time polymerase chain reaction (qRT-PCR) analyses. Cell proliferation was evaluated by the MTT and Cell Counting Kit-8 (CCK8) assays, whereas flow cytometry was used to assess the effects on the cell cycle. Western blotting analysis was used to detect expression levels of protein. Additionally, RNA-sequencing and bioinformatics methods were used to investigate the mechanism of action of LINC00467 in TGCTs. The suppression of LINC00467 expression resulted in decreased cell proliferation and induced S-phase arrest. Furthermore, the suppression of LINC00467 downregulated proliferating cell nuclear antigen (PCNA), a protein related to cell cycle regulation, while it upregulated p21 expression. In other studies involving dihydrotestosterone (DHT) stimulation, it was observed that DHT could upregulate LINC00467 expression. In addition, silencing of the LINC00467 reversed the effect of testosterone on cell proliferation. The Gene Set Enrichment Analysis (GSEA) revealed that LINC00467 regulated the p53 pathway by modulating the expression of CCNG1. Our study found that LINC00467 regulates cell proliferation by inducing S-phase arrest through the cell cycle-related proteins PCNA and p21. These findings contribute to our understanding of non-coding RNAs mechanisms involved in the development of TGCTs.
在我们之前的研究中，与邻近组织相比，在睾丸生殖细胞瘤（TGCT）中发现 LINC00467 的表达显著降低。有趣的是，LINC00467 的表达与 TGCT 患者的肿瘤病理分级相关。LINC00467 的表达越高，TGCT 患者的预后越差。尽管有这些发现，但 LINC00467 在 TGCT 发展中的确切作用仍需要进一步研究。通过小干扰 RNA（siRNA）沉默下调 NCCIT 和 TCam-2 细胞系中的 LINC00467 表达。使用定量实时聚合酶链反应（qRT-PCR）分析验证基因表达水平。通过 MTT 和细胞计数试剂盒-8（CCK8）测定评估细胞增殖，而通过流式细胞术评估对细胞周期的影响。使用 Western blot 分析检测蛋白表达水平。此外，使用 RNA 测序和生物信息学方法研究 LINC00467 在 TGCT 中的作用机制。抑制 LINC00467 的表达导致细胞增殖减少并诱导 S 期停滞。此外，抑制 LINC00467 下调与细胞周期调控相关的增殖细胞核抗原（PCNA）的表达，同时上调 p21 的表达。在涉及二氢睾酮（DHT）刺激的其他研究中，观察到 DHT 可以上调 LINC00467 的表达。此外，沉默 LINC00467 逆转了睾酮对细胞增殖的影响。基因集富集分析（GSEA）表明，LINC00467 通过调节 CCNG1 的表达来调节 p53 途径。我们的研究发现，LINC00467 通过诱导与细胞周期相关的蛋白 PCNA 和 p21 引起 S 期停滞来调节细胞增殖。这些发现有助于我们理解非编码 RNA 参与 TGCT 发展的机制。
