Vesey David A, Cheung Catherine, Pat Betty, Endre Zoltan, Gobé Glenda, Johnson David W
Department of Renal Medicine, University of Queensland at Princess Alexandra Hospital, Brisbane, Australia.
Nephrol Dial Transplant. 2004 Feb;19(2):348-55. doi: 10.1093/ndt/gfg547.
Erythropoietin (EPO) has recently been shown to exert important cytoprotective and anti-apoptotic effects in experimental brain injury and cisplatin-induced nephrotoxicity. The aim of the present study was to determine whether EPO administration is also renoprotective in both in vitro and in vivo models of ischaemic acute renal failure.
Primary cultures of human proximal tubule cells (PTCs) were exposed to either vehicle or EPO (6.25-400 IU/ml) in the presence of hypoxia (1% O(2)), normoxia (21% O(2)) or hypoxia followed by normoxia for up to 24 h. The end-points evaluated included cell apoptosis (morphology and in situ end labelling [ISEL], viability [lactate dehydrogenase (LDH release)], cell proliferation [proliferating cell nuclear antigen (PCNA)] and DNA synthesis (thymidine incorporation). The effects of EPO pre-treatment (5000 U/kg) on renal morphology and function were also studied in rat models of unilateral and bilateral ischaemia-reperfusion (IR) injury.
In the in vitro model, hypoxia (1% O(2)) induced a significant degree of PTC apoptosis, which was substantially reduced by co-incubation with EPO at 24 h (vehicle 2.5+/-0.5% vs 25 IU/ml EPO 1.8+/-0.4% vs 200 IU/ml EPO 0.9+/-0.2%, n = 9, P<0.05). At high concentrations (400 IU/ml), EPO also stimulated thymidine incorporation in cells exposed to hypoxia with or without subsequent normoxia. LDH release was not significantly affected. In the unilateral IR model, EPO pre-treatment significantly attenuated outer medullary thick ascending limb (TAL) apoptosis (EPO 2.2+/-1.0% of cells vs vehicle 6.5+/-2.2%, P<0.05, n = 5) and potentiated mitosis (EPO 1.1+/-0.3% vs vehicle 0.5+/-0.3%, respectively, P<0.05) within 24 h. EPO-treated rats exhibited enhanced PCNA staining within the proximal straight tubule (6.9+/-0.7% vs vehicle 2.4+/-0.5% vs sham 0.3+/-0.2%, P<0.05), proximal convoluted tubule (2.3+/-0.6% vs vehicle 1.1+/-0.3% vs sham 1.2+/-0.3%, P<0.05) and TAL (4.7+/-0.9% vs vehicle 0.6+/-0.3% vs sham 0.3+/-0.2%, P<0.05). The frequency of tubular profiles with luminal cast material was also reduced (32.0+/-1.6 vs vehicle 37.0+/-1.3%, P = 0.05). EPO-treated rats subjected to bilateral IR injury exhibited similar histological improvements to the unilateral IR injury model, as well as significantly lower peak plasma creatinine concentrations than their vehicle-treated controls (0.04+/-0.01 vs 0.21+/-0.08 mmol/l, respectively, P<0.05). EPO had no effect on renal function in sham-operated controls.
The results suggest that, in addition to its well-known erythropoietic effects, EPO inhibits apoptotic cell death, enhances tubular epithelial regeneration and promotes renal functional recovery in hypoxic or ischaemic acute renal injury.
最近研究表明，促红细胞生成素（EPO）在实验性脑损伤和顺铂诱导的肾毒性中发挥重要的细胞保护和抗凋亡作用。本研究旨在确定在缺血性急性肾衰竭的体外和体内模型中，给予EPO是否也具有肾脏保护作用。
将人近端肾小管细胞（PTCs）原代培养物在缺氧（1% O₂）、常氧（21% O₂）或缺氧后再常氧条件下，分别加入溶剂或EPO（6.25 - 400 IU/ml），培养长达24小时。评估的终点指标包括细胞凋亡（形态学和原位末端标记法[ISEL]）、细胞活力（乳酸脱氢酶[LDH]释放）、细胞增殖（增殖细胞核抗原[PCNA]）和DNA合成（胸腺嘧啶核苷掺入）。还在单侧和双侧缺血再灌注（IR）损伤的大鼠模型中研究了EPO预处理（5000 U/kg）对肾脏形态和功能的影响。
在体外模型中，缺氧（1% O₂）诱导了显著程度的PTC凋亡，在24小时时与EPO共同孵育可使凋亡显著减少（溶剂组2.5±0.5% vs 25 IU/ml EPO组1.8±0.4% vs 200 IU/ml EPO组0.9±0.2%，n = 9，P<0.05）。在高浓度（400 IU/ml）时，EPO还刺激了暴露于缺氧且无论随后是否常氧的细胞中的胸腺嘧啶核苷掺入。LDH释放未受到显著影响。在单侧IR模型中，EPO预处理在24小时内显著减轻了外髓质厚升支（TAL）凋亡（EPO组细胞凋亡率为2.2±1.0% vs 溶剂组6.5±2.2%，P<0.05，n = 5）并增强了有丝分裂（EPO组分别为1.1±0.3% vs 溶剂组0.5±0.3%，P<0.05）。EPO处理的大鼠在近端直小管（6.9±0.7% vs 溶剂组2.4±0.5% vs 假手术组0.3±0.2%，P<0.05）、近端曲小管（2.3±0.6% vs 溶剂组1.1±0.3% vs 假手术组1.2±0.3%，P<0.05）和TAL（4.7±0.9% vs 溶剂组0.6±0.3% vs 假手术组0.3±0.2%，P<0.05）中PCNA染色增强。管腔内有铸型物质的肾小管轮廓频率也降低（32.0±1.6 vs 溶剂组37.0±1.3%，P = 0.05）。接受双侧IR损伤的EPO处理大鼠表现出与单侧IR损伤模型相似的组织学改善，并且其血浆肌酐峰值浓度显著低于溶剂处理的对照组（分别为0.04±0.01 vs 0.21±0.08 mmol/l，P<0.05）。EPO对假手术对照组的肾功能无影响。
结果表明，除了其众所周知的促红细胞生成作用外，EPO还可抑制缺氧或缺血性急性肾损伤中的凋亡性细胞死亡，增强肾小管上皮再生并促进肾功能恢复。
