Cadmium disrupts autophagic flux by inhibiting cytosolic Ca2+-dependent autophagosome-lysosome fusion in primary rat proximal tubular cells
Cadmium disrupts autophagic flux by inhibiting cytosolic Ca2+-dependent autophagosome-lysosome fusion in primary rat proximal tubular cells
复制标题
镉通过抑制原代大鼠近端肾小管细胞中胞浆 Ca2 依赖性自噬体-溶酶体融合来破坏自噬通量
DOI：
10.1016/j.tox.2017.03.016
复制
发表时间：
2017-05-15
期刊：
影响因子：
4.5
通讯作者：
Wang, Lin
中科院分区：
文献类型：
--
作者：
Liu, Fei;Wang, Xin-Yu;Wang, Lin
已有研究表明亚细胞内Ca ~（2+）再分布参与了镉诱导的大鼠原代近曲小管（rPT）细胞自噬抑制，但其机制尚不清楚。在这项研究中，自噬流量的状态监测GFP和RFP串联标记LC 3方法。用2-APB或BAPTA-AM抑制胞浆Ca ~（2+）浓度（[Ca ~（2+）]（c））可显著减轻Cd诱导的黄点形成，恢复Cd抑制的红点形成，而毒胡萝卜素（TG）则具有相反的调节作用，表明Cd诱导的[Ca ~（2+）]（c）升高可抑制rPT细胞的自噬通量。结果表明，2-APB、BAPTA-AM和TG对Cd诱导的自噬体积累有明显的调节作用。与此同时，自噬体-溶酶体融合的阻断和由镉暴露引起的Rab 7向自噬体的募集减少被2-APB或BAPTA-AM显著恢复，但与镉和TG的共处理进一步损害了镉诱导的自噬停滞。此外，镉诱导的氧化应激与胞质Ca 2+动员密切相关，N-乙酰半胱氨酸（NAC）显着拯救镉阻断的自噬体-溶酶体融合和招募Rab 7的自噬体在rPT细胞，这意味着镉诱导的自噬抑制是由于[Ca 2 +]（c）升高触发的氧化应激。总之，这些结果表明，镉介导的自噬抑制在rPT细胞是依赖于胞质Ca 2+超载。[Ca ~（2+）]（c）升高可抑制自噬体-溶酶体融合，阻断自噬体的降解，加重Cd诱导的rPT细胞毒性。(C)2017爱思唯尔B. V.保留所有权利。
Previous studies have shown that subcellular Ca2+ redistribution is involved in Cd-induced autophagy inhibition in primary rat proximal tubular (rPT) cells, but the mechanism remains unclear. In this study, the status of autophagic flux was monitored by the GFP and RFP tandemly tagged LC3 method. Pharmacological inhibition of cytosolic Ca2+ concentration ([Ca2+](c)) with 2-APB or BAPTA-AM significantly alleviated Cd-elevated yellow puncta formation and restored Cd-inhibited red puncta formation, while thapsigargin (TG) had the opposite regulatory effect, demonstrating that Cd-induced [Ca2+](c) elevation inhibited the autophagic flux in rPT cells. Resultantly, Cd-induced autophagosomes accumulation was obviously modulated by 2-APB, BAPTA-AM and TG, respectively. Meanwhile, blockage of autophagosome-lysosome fusion and decreased recruitment of Rab7 to autophagosomes by Cd exposure was noticeably restored by 2-APB or BAPTA-AM, but co-treatment with Cd and TG further impaired Cd-induced autophagy arrest. Moreover, Cd-induced oxidative stress intimately correlated with cytosolic Ca2+ mobilization, and N-acetylcysteine (NAC) markedly rescued Cd-blocked autophagosome-lysosome fusion and recruitment of Rab7 to autophagosomes in rPT cells, implying that Cd-induced autophagy inhibition was due to [Ca2+](c) elevation-triggered oxidative stress. In summary, these results suggest that Cd-mediated autophagy inhibition in rPT cells is dependent on cytosolic Ca2+ overload. Elevation of [Ca2+](c) inhibited the autophagosome-lysosome fusion to block the degradation of autophagosomes, which aggravated Cd-induced cytotoxicity in rPT cells. (C) 2017 Elsevier B.V. All rights reserved.