Background Heat shock protein family A member 1 like (Hspa1l) is a member of the 70kD heat shock protein (Hsp70) family. HSPA1L is an ancient, evolutionarily conserved gene with a highly conserved domain structure. The gene is highly abundant and constitutively expressed in the mice testes. However, the role of Hspa1l in the testes has still not been elucidated. Methods Hspa1l-mutant mice were generated using the CRISPR/Cas9 system. Histological and immunofluorescence staining were used to analyze the phenotypes of testis and epididymis. Apoptotic cells were detected through TUNEL assays. Fertility and sperm motilities were also tested. Quantitative RT-PCR was used for analyzing of candidate genes expression. Heat treatment was used to induce heat stress of the testis. Results We successfully generated Hspa1l knockout mice. Hspa1l-/- mice exhibited normal development and fertility. Further, Hspa1l-/- mice shown no significant difference in spermatogenesis, the number of apoptotic cells in testes epididymal histology, sperm count and sperm motility from Hspa1l+/+ mice. Moreover, heat stress does not exacerbate the cell apoptosis in Hspa1l-/- testes. These results revealed that HSPA1L is not essential for physiological spermatogenesis, nor is it involved in heat-induced stress responses, which provides a basis for further studies.

Mebendazole (MBZ) is a synthetic benzimidazole known for its antiparasitic properties. In recent years, growing evidence showed that MBZ was also used as an anti-tumor agent. However, whether (and to what extent) this drug treatment affected the male reproductive system was not well-understood. In this study, male C57BL/6 mice were injected with 40 mg/kg/day of MBZ. The treatment was for 3 and 7 days. Our results showed that the injected mice exhibited an abnormal spermatogenic phase with a significant decrease in sperm. We further detected microtubule disruption and transient functional destruction of the blood–testes barrier (BTB) in the MBZ-injected mice testes (BTB). Our data confirmed that MBZ suppressed the expression of the BTB junction-associated proteins and disrupted the Sertoli cells’ function in vivo. Moreover, MBZ-treated mice demonstrated an aberrant caspase-3 signalling pathway, which resulted in the apoptosis of the germ cells. Here, we present our data, indicating that MBZ impairs BTB by reducing the expression of the microtubules’ and BTB junction-associated proteins. The last leads to activating the caspase-3 pathway, which triggers extensive germ cell apoptosis.

Peroxisomes are organelles enclosed by a single membrane and are present in various species. The abruption of peroxisomes is correlated with peroxisome biogenesis disorders and single peroxisomal enzyme deficiencies that induce diverse diseases in different organs. However, little is known about the protein compositions and corresponding roles of heterogeneous peroxisomes in various organs. Through transcriptomic and proteomic analyses, we observed heterogenous peroxisomal components among different organs, as well as between testicular somatic cells and different developmental stages of germ cells. As Pex3 is expressed in both germ cells and Sertoli cells, we generated Pex3 germ cell- and Sertoli cell-specific knockout mice. While Pex3 deletion in Sertoli cells did not affect spermatogenesis, the deletion in germ cells resulted in male sterility, manifested as the destruction of intercellular bridges between spermatids and the formation of multinucleated giant cells. Proteomic analysis of the Pex3 -deleted spermatids revealed defective expressions of peroxisomal proteins and spermiogenesis-related proteins. These findings provide new insights that PEX3-dependent peroxisomes are essential for germ cells undergoing spermiogenesis, but not for Sertoli cells.