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Leydig cells (LCs) play crucial roles in producing testosterone, which is critical in the regulation of male reproduction and development. Low levels of testosterone will lead to male hypogonadism. LC transplantation is a promising alternative therapy for male hypogonadism. However, the source of LCs limits this strategy for clinical applications. Thus far, others have reported that LCs can be derived from stem cells by gene transfection, but the safe and effective induction method has not yet been reported. Here, we report that Leydig-like cells can be derived from human induced pluripotent stem cells (iPSCs) using a novel differentiation protocol based on molecular compounds. The iPSCs-derived Leydig-like cells (iPSC-LCs) acquired testosterone synthesis capabilities, had the similar gene expression profiles with LCs, and positively expressed Leydig cell lineage-specific protein markers LHCGR, STAR, SCARB1, SF-1, CYP11A1, HSD3B1, and HSD17B3 as well as negatively expressed iPSC-specific markers NANOG, OCT4, and SOX2. When iPSC-LCs labeled with lipophilic red dye (PKH26) were transplanted into rat testes that were selectively eliminated endogenous LCs using EDS (75 mg/kg), the transplanted iPSC-LCs could survive and function in the interstitium of testes, and accelerate the recovery of serum testosterone levels and testis weights. Collectively, these findings demonstrated that the iPSCs were able to be differentiated into Leydig-like cells by few defined molecular compounds, which may lay the safer groundwork for further clinical application of iPSC-LCs for hypogonadism.

Background: Polycystic ovary syndrome (PCOS) is the most common metabolic and endocrinopathies disorder in women of reproductive age and non-alcoholic fatty liver (NAFLD) is one of the most common liver diseases worldwide. Previous research has indicated potential associations between PCOS and NAFLD, but the underlying pathophysiology is still not clear. The present study aims to identify the differentially expressed genes (DEGs) between PCOS and NAFLD through the bioinformatics method, and explore the associated molecular mechanisms.Methods: The microarray datasets GSE34526 and GSE63067 were downloaded from Gene Expression Omnibus (GEO) database and analyzed to obtain the DEGs between PCOS and NAFLD with the GEO2R online tool. Next, the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis for the DEGs were performed. Then, the protein-protein interaction (PPI) network was constructed and the hub genes were identified using the STRING database and Cytoscape software. Finally, NetworkAnalyst was used to construct the network between the targeted microRNAs (miRNAs) and the hub genes.Results: A total of 52 genes were identified as DEGs in the above two datasets. GO and KEGG enrichment analysis indicated that DEGs are mostly enriched in immunity and inflammation related pathways. In addition, nine hub genes, including TREM1, S100A9, FPR1, NCF2, FCER1G, CCR1, S100A12, MMP9, and IL1RN were selected from the PPI network by using the cytoHubba and MCODE plug-in. Then, four miRNAs, including miR-20a-5p, miR-129-2-3p, miR-124-3p, and miR-101-3p, were predicted as possibly the key miRNAs through the miRNA-gene network construction.Conclusion: In summary, we firstly constructed a miRNA-gene regulatory network depicting interactions between the predicted miRNA and the hub genes in NAFLD and PCOS, which provides novel insights into the identification of potential biomarkers and valuable therapeutic leads for PCOS and NAFLD.

Perfluorododecanoic acid (PFDoA) has been used as a surfactant and may have reproductive toxicity. However, whether PFDoA influences Leydig cell development during prepuberty remains unknown. In the present study, 21-day-old male Sprague–Dawley rats were gavaged 0, 5, or 10 mg/kg PFDoA from postnatal day 21 to 35. PFDoA decreased the serum concentrations of testosterone, luteinizing hormone, and follicle-stimulating hormone at doses of 5 and 10 mg/kg without influencing Leydig cell number and proliferation. However, PFDoA down-regulated the expression of Leydig cell genes (Lhcgr, Scarb1, Star, Cyp11a1, Cyp17a1, and Hsd11b1) or their proteins. PFDoA dose-dependently reduced SIRT1 and PGC-1α levels. PFDoA did not affect AMPK and AKT2 levels but decreased their phosphorylation. We also treated primary progenitor Leydig cells purified from prepubertal rat testes with PFDoA for 24 h. It in vitro lowered viability and decreased mitochondrial membrane potential of progenitor Leydig cells, but it stimulated the generation of the intracellular reactive oxygen species and induced Leydig cell apoptosis at 10 μM. In conclusion, PFDoA blocks rat Leydig cell development during the prepubertal period possibly via targeting AMPK/SIRT1/PGC-1α and AKT2 signaling pathways.

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