Lead (Pb) is a known nephrotoxicant that causes damage to proximal tubular cells. Autophagy has an important protective role in various renal injuries, but the role of autophagy in Pb-elicited nephrotoxicity remains largely unknown. In this study, Pb promoted the accumulation of autophagosomes in primary rat proximal tubular (rPT) cells, and subsequent findings revealed that this autophagosome accumulation was caused by the inhibition of autophagic flux. Moreover, Pb exposure did not affect the autophagosome–lysosome fusion in rPT cells. Next, we found that Pb caused lysosomal alkalinization, may be through suppression of two V-ATPase subunits. Simultaneously, Pb inhibited lysosomal degradation capacity by affecting the maturation of cathepsin B (CTSB) and cathepsin D (CTSD). Furthermore, translocation of CTSB and CTSD from lysosome to cytoplasm was observed in this study, suggesting that lysosomal membrane permeabilization (LMP) occurred in Pb-exposed rPT cells. Meanwhile, Pb-induced caspase-3 activation and apoptosis were significantly but not completely inhibited by CTSB inhibitor (CA 074) and CTSD inhibitor (pepstatin A), respectively, demonstrating that LMP-induced lysosomal enzyme release was involved in Pb-induced apoptosis in rPT cells. In conclusion, Pb-mediated autophagy blockade in rPT cells is attributed to the impairment of lysosomal function. Both inhibition of autophagic flux and LMP-mediated apoptosis contribute to Pb-induced nephrotoxicity in rPT cells.

Blood-testis barrier (BTB) creates a privileged niche indispensable for spermatogenesis. Glyphosate (GLY), the most commonly used herbicide worldwide, has been reported to decrease sperm quality. However, whether and how GLY destroys the BTB to affect sperm quality remains to be elucidated. Herein, this study was designed to investigate the influence of GLY on the BTB in vivo and in vitro experiments. The results showed that male rats exposed to GLY for 4 months exhibited a decrease in sperm quality and quantity, accompanied by BTB integrity disruption and testicular oxidative stress. Additionally, GLY-induced reactive oxygen species (ROS) contributed to the downregulation of BTB-related proteins in primary Sertoli cells (SCs). Intriguingly, we identified a marked upregulation of oxidative stress-related gene NOX1 in GLY-exposed testis based on transcriptome analysis. NOX1 knockdown blocked the GLY-induced oxidative stress, as well as prevented BTB-related protein decrease in SCs. Furthermore, the estrogen receptor (ER)-α was significantly upregulated in vivo and in vitro models. An ER-α inhibitor decreased the expression levels of both ER-α and NOX1. Mechanistically, GLY directly interacted with ER-α at the site of Pro39 and Lys401 to promote ER-α activation, which boosted NOX1 expression to trigger ROS accumulation. Collectively, these results demonstrate that long-term GLY exposure adversely affects BTB integrity, which disrupts spermatogenesis via activation of ER-α/NOX1 axis. This study presents a better understanding of the risk of long-term GLY exposure to male fertility.