Hypoxia‐induced apoptosis of mouse spermatocytes is mediated by HIF‐1α through a death receptor pathway and a mitochondrial pathway

Abstract

Hypoxia in vivo induces oligozoospermia, azoospermia, and degeneration of the germinal epithelium, but the underlying molecular mechanism of this induction is not fully clarified. The aim of this study was to investigate the role of the death receptor pathway and the mitochondrial pathway in hypoxia-induced apoptosis of mouse GC-2spd (GC-2) cells and the relationship between HIF-1α and apoptosis of GC-2 cells induced by hypoxia. GC-2 cells were subjected to 1% oxygen for 48 hr. Apoptosis was detected by flow cytometry, TUNEL staining, LDH, caspase-3/8/9 in the absence and presence of HIF-1α siRNA. The protein levels of apoptosis-related markers were determined by Western blot in the presence and absence of HIF-1α siRNA. Mitochondrial transmembrane potential change was observed by in situ JC-1 staining. Cell viability was assessed upon treatment of caspase-8 and 9 inhibitors. The results indicated that hypoxia at 1% oxygen for 48 hr induced apoptosis of GC-2 cells. A prolonged exposure of GC-2 cells to hypoxic conditions caused downregulation of c-FLIP, D R and Bcl-2 and upregulation of DR , TRAIL, Fas, p53, and Bax, with an overproduction of caspase-3/8/9. Moreover, hypoxia at this level had an effect on mitochondrial depolarization. In addition, specific inhibitors of caspase-8/9 partially suppressed hypoxia-induced GC-2 cell apoptosis, and the anti-apoptotic effects of the caspase inhibitors were additive. Of note, HIF-1α knockdown attenuated hypoxia and induced apoptosis of GC-2 cells. In conclusion, our data suggest that the death receptor pathway and mitochondrial pathway, which are likely mediated by HIF-1α, contribute to hypoxia-induced GC-2 cell apoptosis.

Elevation of autophagy rescues spermatogenesis by inhibiting apoptosis of mouse spermatocytes

“…In our previous study, we demonstrated the apoptosis of GC-2 cells was increased in a time-dependent manner under hypoxic conditions, which was evaluated by using TUNEL staining and flow cytometry (Yin et al 2017). In this study, we further determined whether different hypoxic conditions induced the apoptosis of GC-2 cells through the mitochondrial pathway and/or death receptor pathway.…”

“…The underlying mechanisms include the activation of caspases and clearance of the Bruce protein (an anti-apoptotic protein, a member of the inhibitors of apoptosis proteins family) (Nezis et al , Young et al 2012. Taking all the above into consideration, the interaction between autophagy and apoptosis in spermatocytes under hypoxia treatment remains unclear.In our previous study, we established an apoptosis model of mouse spermatocytes under hypoxia treatment and found that caspase-8 is involved in apoptosis of mouse spermatocytes (Yin et al 2017). In the present study, we further explore whether and how hypoxia affects the autophagy of mouse spermatocytes and how autophagy interacts with apoptosis mutually in vitro and in vivo, which ultimately determines the fate of spermatocytes.…”

“…In our previous study, we established an apoptosis model of mouse spermatocytes under hypoxia treatment and found that caspase-8 is involved in apoptosis of mouse spermatocytes (Yin et al 2017). In the present study, we further explore whether and how hypoxia affects the autophagy of mouse spermatocytes and how autophagy interacts with apoptosis mutually in vitro and in vivo, which ultimately determines the fate of spermatocytes.…”

Hypoxia induces apoptosis of mouse spermatocyte GC‐2 cells through activation of autophagy

“…HIF‐1 is one of the key mediators in asthenozoospermia by influencing sperm motility (Salvolini et al, ). In addition, it is reported that hypoxia‐induced GC‐2 cell apoptosis mediated by HIF‐1α (Yin et al, ). In hypoxic GC‐2 cells, we found that HIF‐1α silencing promoted Bcl‐2 and p62 protein expression, but suppressed caspase‐3, Bax, LC3‐II, and Beclin‐1 protein expression.…”

“…Several previous studies have indicated that hypoxia‐induced cell damage is closely related to apoptosis in various cell types (Li et al, ; Ma et al, ; Olcina et al, ). And more notably, a recent study demonstrated the treatment of hypoxia can induce the apoptosis of spermatogenic cells (GC‐2 cells) in mice (Yin et al, ). Accordingly, in a male infertility model treated with hypobaric hypoxia find that hypoxia induced the apoptosis of the spermatocytes (Liao et al, ).…”