Chuan'ai Chen, Haolong Li, Pengpeng An, Yang Yu, Chunyan Liu, Yumiao Yang
Department of Quality Management, Tianjin Blood Center, Tianjin, China.
Nephrology (Carlton). 2025 Feb;30(2):e70000. doi: 10.1111/nep.70000.
To study the effect and elucidate the underlying mechanisms of VDAC1-ΔC on autophagy in renal tubular epithelial cells injured by hypoxia/reoxygenation.
C57/BL6 mice were randomly divided into groups: sham operation group, IRI 1d group and IRI 2d group. The inner canthal blood of mice was collected to detect the levels of serum creatinine and urea nitrogen and kidney tissues were sampled, and sections were stained with Periodic acid-Schiff for morphological evaluation. The expression of VDAC1 in kidney tissue was detected by Western blot. An immortalised human proximal tubular epithelial cell line, HK-2 cells, were subjected to hypoxia/reoxygenation treatment. HK-2 cells were incubated under hypoxia for 6 h, followed by 6 and 24 h of reoxygenation, cells were divided into four groups: H6/R0 group, H6/R6 group, H6/R24 group and control group. The release of LDH and cytosolic ROS were assessed, the expression of autophagy-related proteins LC3 and p62 was detected by Western blot, autophagy flux was monitored by transfecting mRFP-GFP-LC3 lentivirus in HK2 cells, and cells were pretreated with bafilomycin A1 to further monitor the autophagy flux. VDAC1-cleavage-defective mutant in HK-2 cells silencing VDAC1 was established to examine the effect of VDAC1 cleavage on autophagy and hypoxia/reoxygenation injury.
In vivo, IRI 1d/2d promoted the disorder of renal tubular structure and the cleavage of VDAC1 in kidney tissue; in vitro, hypoxia/reoxygenation promoted cytosolic ROS accumulation, LDH release, VDAC1 cleavage and induced autophagy and autophagic flux; reduced VDAC1 cleavage inhibited autophagy; and decreased cytosolic ROS accumulation and LDH release, thus alleviated cell injury.
In renal tubular epithelial cells injured by H/R, VDAC1 cleavage was increased, triggering an autophagic response, and VDAC1 cleavage promoted autophagy to regulate cell injury.
研究电压依赖性阴离子通道1截短体（VDAC1-ΔC）对缺氧/复氧损伤的肾小管上皮细胞自噬的影响并阐明其潜在机制。
将C57/BL6小鼠随机分为假手术组、缺血再灌注损伤1天组（IRI 1d组）和缺血再灌注损伤2天组（IRI 2d组）。采集小鼠内眦血以检测血清肌酐和尿素氮水平，并取肾脏组织进行采样，切片用高碘酸-希夫染色进行形态学评估。通过蛋白质免疫印迹法检测肾脏组织中VDAC1的表达。将永生化的人近端肾小管上皮细胞系HK-2细胞进行缺氧/复氧处理。HK-2细胞在缺氧条件下孵育6小时，然后复氧6小时和24小时，细胞分为四组：H6/R0组、H6/R6组、H6/R2组和对照组。评估乳酸脱氢酶（LDH）释放和胞质活性氧（ROS）水平，通过蛋白质免疫印迹法检测自噬相关蛋白LC3和p62的表达，在HK2细胞中通过转染mRFP-GFP-LC3慢病毒监测自噬流，并用巴佛洛霉素A1预处理细胞以进一步监测自噬流。构建沉默VDAC1的HK-2细胞中VDAC1切割缺陷突变体，以研究VDAC1切割对自噬和缺氧/复氧损伤的影响。
在体内，IRI 1d/2d促进肾小管结构紊乱和肾脏组织中VDAC1的切割；在体外，缺氧/复氧促进胞质ROS积累、LDH释放、VDAC1切割并诱导自噬和自噬流；减少VDAC1切割可抑制自噬；并减少胞质ROS积累和LDH释放，从而减轻细胞损伤。
在缺氧/复氧损伤的肾小管上皮细胞中，VDAC1切割增加，引发自噬反应，且VDAC1切割促进自噬以调节细胞损伤。
