Autophagy blockade and lysosomal membrane permeabilization contribute to lead-induced nephrotoxicity in primary rat proximal tubular cells

Xiang-Bin Song,Lin Wang,Gang Liu,Zhen-Yong Wang,Zhen-Gui Yan,Zong-Ping Liu,Fei Liu

doi:10.1038/cddis.2017.262

Xiang-Bin Song, Lin Wang + Show 5 more

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https://doi.org/10.1038/cddis.2017.262

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Abstract

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.

Highlights

Lead (Pb) is a known nephrotoxicant that causes damage to proximal tubular cells
Protein levels of light chain 3 (LC3)-II in rat proximal tubular (rPT) cells treated with Pb (0.25, 0.5 and 1 μM) for 3 h (Figure 1a), 6 h (Figure 1b) and 12 h (Figure 1c) was detected to investigate the effect of Pb exposure on autophagy, respectively
Recent studies have suggested that autophagy acts as a protective mechanism to promote cell survival during acute kidney injury, and blockade of autophagic flux has a negative impact on the development of nephrotoxicity.[7,24,25,26,27]

Summary

Introduction

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. 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. Autophagy is a highly dynamic multi-step biological process which maintains cellular homeostasis via the degradation and recycling of damaged organelles, misfolded proteins, and long-lived macromolecules in lysosomes.[5] Previous studies have demonstrated that basal autophagy is vital for normal proximal tubule function, and genetic or pharmacologic blockade of autophagy strongly enhanced acute kidney injury induced by cisplatin or ischemia-reperfusion.[6,7,8,9] report by Lv et al.[10] and Sui et al.[11] showed that Pb promoted the autophagy in cultured osteoblasts and cardiofibroblasts, respectively. Our research group has recently found that 0.5 μM Pb treatment for 12 h blocked the autophagic flux in rPT cells,[12] while the underlying molecular mechanism of impaired autophagic flux during Pb exposure remains to be elucidated

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