Silencing of LINC00467 inhibits cell proliferation in testicular germ cell tumors cells

Fang Zhu; Zhizhong  Liu; Qianyin Zhou; Dai Zhou; Jingyu Fan; Hao Bo; Liqing Fan

doi:10.17305/bb.2023.8969

Authors

Fang Zhu Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China https://orcid.org/0000-0002-8289-7362
Zhizhong Liu 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
Qianyin Zhou Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China
Dai Zhou Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China; Clinical Research Center for Reproduction and Genetics in Hunan Province, Reproductive and Genetic Hospital of CITIC-Xiangya, Changsha, China
Jingyu Fan Department of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United States
Hao Bo Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China; Clinical Research Center for Reproduction and Genetics in Hunan Province, Reproductive and Genetic Hospital of CITIC-Xiangya, Changsha, China
Liqing Fan Institute of Reproductive and Stem Cell Engineering, School of Basic Medical Science, Central South University, Changsha, China; Clinical Research Center for Reproduction and Genetics in Hunan Province, Reproductive and Genetic Hospital of CITIC-Xiangya, Changsha, China https://orcid.org/0000-0002-8289-7362

DOI:

https://doi.org/10.17305/bb.2023.8969

Keywords:

Testicular germ cell tumors (TGCT), LINC0046, dihydrotestosterone (DHT), cell proliferation, cell cycle

Abstract

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.