Levels of Leydig cell autophagy regulate the fertility of male naked mole-rats

Fertility is abolished in nonbreeding males in colonies of natal naked mole-rats (NMRs). Although spermatogenesis occurs in both breeding and nonbreeding male NMRs, the mechanisms underlying the differences in fertility between breeders and nonbreeders remain unexplored. In this study, a significant decrease in autophagy was observed in Leydig cells of the testis from nonbreeding male NMRs. This alteration was visualised as a significant decrease in the levels of autophagy-related gene 7 (Atg7), Atg5, microtubule-associated protein 1A/B light chain 3 (LC3-II/I) and the number of autophagosomes and an increase in P62 levels using Western blotting analyses. Furthermore, monodansylcadaverine (MDC) staining and Western blot analyses revealed that testosterone production decreased in nonbreeding male NMR Leydig cells, this decrease was associated with a reduction in autophagy. Primary Leydig cells from breeding and nonbreeding male NMRs were processed to investigate the effect of an autophagy inhibitor (3-MA, 3-methyladenine) or an autophagy activator (rapamycin) on testosterone production. Rapamycin induced an increase in testosterone production in NMR Leydig cells, whereas 3-MA had the opposite effect. Consequently, spermatogenesis, the weight of the testis, and androgen levels were dramatically reduced in nonbreeding male NMRs. While rapamycin treatment restored the fertility of nonbreeding male NMRs. Based on these results, inadequate autophagy correlates with a decrease in steroid production in nonbreeding male NMR Leydig cells, which may ultimately influence the spermatogenesis and fertilities of these animals.