Sertoli cells are the most important somatic cells contributing to the microenvironment (named niche) for spermatogonial stem cells (SSCs). They produce amounts of crucial growth factors and structure proteins that play essential roles in the complex processes of male SSCs survival, proliferation, and differentiation. It has been suggested that Sertoli cell abnormalities could result in spermatogenesis failure, eventually causing azoospermia in humans. However, to the end, the gene expression characteristics and protein functions of human Sertoli cells remained unknown. In this study, we aimed to evaluate the effect of fibroblast growth factor-5 (FGF5), a novel growth factor downregulated in Sertoli cells from Sertoli cell-only syndrome (SCOS) patients compared to Sertoli cells from obstructive azoospermia (OA) patients, on SSCs. We compared the transcriptome between Sertoli cell from SCOS and OA patients. Then, we evaluated the expression of FGF5, a growth factor which is downregulated in SCOS Sertoli cells, in human primary cultured Sertoli cells and testicular tissue. Also, the proliferation effect of FGF5 in mice SSCs was detected using EDU assay and CCK-8 assay. To investigate the mechanism of FGF5, Phospho Explorer Array was performed. And the results were verified using Western blot assay. Using RNA-Seq, we found 308 differentially expressed genes (DEGs) between Sertoli cells from SCOS and OA patients. We noted and verified that the expression of fibroblast growth factor-5 (FGF5) was higher in Sertoli cells of OA patients than that of SCOS patients at both transcriptional and translational levels. Proliferation assays showed that rFGF5 enhanced the proliferation of mouse SSCs line C18-4 in a time- and dose-dependent manner. Moreover, we demonstrated that ERK and AKT were activated and the expression of Cyclin A2 and Cyclin E1 was enhanced by rFGF5. The distinct RNA profiles between Sertoli cells from SCOS and OA patients were identified using RNA-Seq. Also, FGF5, a growth factor that downregulated in SCOS Sertoli cells, could promote SSCs proliferation via ERK and AKT activation.

Globozoospermia is a rare (prevalence of <0.1%) but severe male infertility condition. In our previous study, we found that robust KIFC1 immunostaining was detected in the human elongating/elongated spermatids during human acrosomogenesis. However, the relationship between the decreased expression of KIFC1 and human globozoospermia remains largely unknown. Testicular biopsies of 30 globozoospermia and 30 obstructive azoospermia patients who underwent infertility evaluation and treatment were utilized in this study. Reverse transcriptase polymerase chain reaction (RT-PCR), Western blots, immunohistochemistry, an in vivo model, and intratesticular injection of small inhibitory RNA (siRNA) against the Kifc1 gene were employed, and sperm abnormalities were evaluated by hematoxylin and eosin (H&E) staining and immunocytochemistry. We revealed that the testicular level of KIFC1 mRNA in globozoospermia was significantly reduced compared with that in obstructive azoospermia, and the KIFC1 protein was barely detectable in testicular specimens in 30% (9 of 30) of patients with globozoospermia. Furthermore, knockdown of the Kifc1 gene in mice increased the percentage of sperm with globozoospermic defects (26.5%). Decreased KIFC1 expression was mainly observed in the testes of patients with globozoospermia at the spermatid stage, which may be useful for counseling and management of such patients.

Aging has a significant negative impact on human testicular function; steroidogenesis is gradually impaired, and testosterone replacement therapy still has many risks. Low-intensity pulsed ultrasound (LIPUS) has been used as a novel non-invasive treatment for male erectile dysfunction and other fields, and has been shown to increase testosterone levels in animal models. Testosterone is synthesized and secreted by Leydig cells (LCs), and the serum testosterone level decreases after aging due to the LCs senescence. However, the effect of LIPUS on human senescent LCs has not been reported. In this study, human senescent LCs were isolated and stimulated with different energy intensities in vitro, and cell morphology, cell apoptosis, cell proliferation, cell senescence levels, lipid droplet number, testosterone and INSL3 secretion levels were tested and analyzed. Quantitative Polymerase Chain Reaction (QPCR) and Western Blot were performed to compare cell senescence characteristics and the expression profile of key pathways of testosterone secretion, and transcriptome analysis was performed to explore the signaling pathways of LCs alteration after LIPUS stimulation. It was safe and effective to stimulate LCs with the 75 mW/cm² energy of LIPUS in vitro, which not only improved the senescence phenotype, but also effectively enhanced the secretory function of LCs in vitro, and increased the expression of key pathways of the testosterone synthesis pathway. These results suggest that LIPUS could be used as a novel treatment to human senescent LCs with decreased testosterone secretion levels in vitro.

This study aimed to evaluate the efficacy and safety of vasal vessel-sparing modified single-armed 2-suture longitudinal intussusception vasoepididymostomy (SA-LIVE) to epididymal obstructive azoospermia patients. Forty consecutive epididymal obstructive azoospermia cases, who underwent microsurgical vasoepididymostomy in Shanghai General Hospital from January 2019 to October 2019, were included in this study. Twenty cases underwent SA-LIVE (group A), and 20 cases underwent vasal vessel-sparing SA-LIVE (group B). Until March 2021, the mean follow-up period was 16.9 ± 4.1 (12–23) months. The overall patency rate was 82.5%, and 80% and 85% for group A and group B respectively. The mean time to achieve patency was 4.11 ± 2.74 months. The overall natural pregnancy rate was 51.5%(17/33) at the mean follow-up of 16.9 months. The natural pregnancy rate was 50.0% for group A and 52.9% for group B (p > .05). At the time of 6 months post-operation, the patency rate was 70% for group A and 80% for group B (p = .465); the natural pregnancy rate was 0% for group A and 31.3% for group B (p = .022). Vasal vessel-sparing SA-LIVE is safe and effective to achieve favourable patency and pregnancy rates. Preserving vasal vessel would improve natural pregnancy rate at a very early stage.

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