Tle6 (Transducin-like enhancer of split 6) is a member of the Tle co-repressor superfamily, which is expressed in various tissues of invertebrates and vertebrates and participates in the developmental process. However, the current research has only found that the TLE6 mutation is related to infertility, and the key regulatory mechanism of TLE6 remains to be explored. In this study, we combined Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9 and the Tet-on system to construct mouse spermatogonia cell lines that induced TLE6 protein knockout (KO), and studied the effect of Tle6 on mouse spermatogonia proliferation and the cell cycle. The results showed that, after drug induction, the Tle6 gene in mouse spermatogonia was successfully knocked out at the genome and protein levels, and the Tle6 gene knockout efficiency was confirmed to be 87.5% with gene-cloning technology. At the same time, we also found that the mouse spermatogonia proliferated slowly after the Tle6 knockout. Using flow cytometry, we found that the cells did not undergo significant apoptosis, and the number of cells in the S phase decreased. After real-time quantity PCR (qRT-PCR) analysis, we found that the expression of cell-proliferation-related genes, CCAAT enhancer-binding protein α(C/ebp α), granulocyte-colony stimulating factor(G-csf), cyclin-dependent kinases 4(Cdk 4), Cyclin E, proliferating cell nuclear antigen(Pcna), and S-phase kinase-associated protein 2 (Skp2) was significantly reduced, which further affected cell growth. In summary, Tle6 can regulate spermatogonia cell proliferation and the cell cycle and provide a scientific basis for studying the role of TLE6 on spermatogenesis.
Testicular seminoma is one of the most common tumours in the field of urology, and its aetiology is still unclear. The aim of the present study was to identify the factors responsible for the development of testicular cancer and to investigate whether mutations in these genes were primarily congenital or acquired. To identify the key genes and miRNAs linked to testicular seminoma, as well as their potential molecular mechanisms, the GSE15220, GSE1818 and GSE59520 microarray datasets were analysed. A total of 5,195 and 1,163 differentially expressed genes (DEGs) were identified after analysing the GSE15220 and GSE1818 datasets, respectively. Among them, 287 genes were common between the two datasets. Of these, 110 were upregulated and 177 were downregulated. Five differentially expressed microRNAs (miRs; DEMs) that were downregulated in seminoma were identified after analysing the GSE59520 dataset. Following protein-protein interaction network and Gene Ontology analysis, the five nodes with the highest degrees were screened as hub genes. Among them, the high expression of hub genes, such as protein tyrosine phosphatase receptor type C (PTPRC), was associated with worse overall survival. We also predicted the potential target genes of the DEMs. DNA topoisomerase II α (TOP2A), marker of proliferation Ki-67 (MKI67), PTPRC and ubiquitin conjugating enzyme E2 C were associated with the PI3K/AKT and Wnt/β-catenin signalling pathways. In addition, hsa-miR-650 and hsa-miR-665 were associated with the PI3K/AKT and Wnt/β-catenin signalling pathways. Additionally, TOP2A and MKI67 were strongly associated with the target genes hsa-miR-650 and hsa-miR-665, respectively. We proposed that the hub genes reported in the present study may have a certain impact on cellular proliferation and migration in testicular seminoma. The roles of these hub genes in seminoma may provide novel insight to improve the diagnosis and treatment of patients with seminoma.
<b><i>Introduction:</i></b> The concentration of 25-hydroxycholecalciferol (25OHD<sub>3</sub>) in the serum of obese people is low and often accompanied by symptoms of low fertility. Therefore, vitamin D is recommended as a potential treatment option. However, after clinical trials, it was found that vitamin D cannot effectively increase the concentration of 25OHD<sub>3</sub> in the serum of obese people. How obesity causes low 25OHD<sub>3</sub> concentration and low fertility is unclear. <b><i>Methods:</i></b> We analyzed the physiological and pathological changes in obese mice induced by a high-fat diet (HFD) and the changes in mice after supplementing with 25OHD<sub>3</sub>. <b><i>Results:</i></b> The concentration of 25OHD<sub>3</sub> in the serum of obese mice induced by HFD was significantly reduced, and these mice showed liver hypertrophy accompanied by abnormal liver injury, testicular hypertrophy, low testosterone levels, high leptin levels, and low sperm motility. The mRNA and protein expression of CYP2R1 of hydroxylated vitamin D<sub>3</sub> was significantly reduced; CYP11A1 and CYP11A2, which synthesize testosterone, were significantly reduced. After supplementing with 25OHD<sub>3</sub>, there was an increase in serum 25OHD<sub>3</sub> concentration, testosterone level, and sperm motility, but it cannot improve the degree of obesity, CYP2R1 expression, and liver damage. <b><i>Conclusion:</i></b> Our research shows that there is a metabolic interference mediated by 25OHD<sub>3</sub> and testosterone between obesity and low sperm motility. The results of this study can provide a scientific basis for studying the mechanism of 25OHD<sub>3</sub> and hormone regulation and treating obese people with low sperm motility.
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