Bidmon Bettina, Endemann Michaela, Arbeiter Klaus, Ruffingshofer Dagmar, Regele Heinz, Herkner Kurt, Eickelberg Oliver, Aufricht Christoph
Department of Pathology, Yale University, School of Medicine, New Haven Connecticut, USA.
Kidney Int. 2004 Dec;66(6):2300-7. doi: 10.1111/j.1523-1755.2004.66040.x.
Peritoneal dialysis is complicated by mesothelial cell injury due to low biocompatibility of peritoneal dialysis fluid (PDF). We have previously demonstrated that heat shock protein (HSP)-72 is potently up-regulated in response to PDF exposure of mesothelial cells in in vitro and in vivo models of peritoneal dialysis. The aim of this study was to evaluate potential cytoprotective effects of overexpression of HSP-72.
Cytoprotection was assessed by comparing cellular viability between pretreated versus nonpretreated human mesothelial cells (Met 5a; ATCC, Manassas, VA, USA, and primary cell cultures) subjected to extended, usually lethal PDF exposure times (120 min, CAPD2; Fresenius, Bad Homburg, Germany). Pretreatment was performed with exposure to PDF (60 min, CAPD2; Fresenius) or heat (15 min, 41.5 degrees C), and by transient transfection with HSP-72.
When mesothelial cells were pretreated by nonlethal exposure to PDF or heat, HSP-72 was markedly up-regulated (>5-fold, P < 0.01). Pretreated human mesothelial cells were significantly protected against subsequent "lethal" exposures to PDF, as assessed by dye exclusion (>50% reduction, P < 0.05) and lactate dehydrogenase (LDH) release (>30% reduction, P < 0.05). Comparable cytoprotection (50% reduction by dye exclusion) was indicated by overexpression of HSP-72 in cultered human mesothelial cells (>5-fold) after transient transfection with HSP-72. This cytoprotection was confirmed at a cellular basis by double staining techniques with HSP-72 and ApopTag (apoptosis detection kit).
Our study therefore shows that the mesothelial stress response confers cytoprotection in experimental peritoneal dialysis, mediated by the induction of HSP-72, and that the stimulus of the pretreatment does not have to be identical to the subsequent injury. These data offer the basis for an attractive novel therapeutic approach against PDF toxicity.
由于腹膜透析液（PDF）生物相容性低，腹膜透析会并发间皮细胞损伤。我们之前已经证明，在腹膜透析的体外和体内模型中，热休克蛋白（HSP）-72会因间皮细胞暴露于PDF而显著上调。本研究的目的是评估HSP-72过表达的潜在细胞保护作用。
通过比较经过预处理和未经过预处理的人间皮细胞（Met 5a；美国弗吉尼亚州马纳萨斯市美国典型培养物保藏中心以及原代细胞培养物）在延长的、通常具有致死性的PDF暴露时间（120分钟，CAPD2；德国巴特洪堡费森尤斯公司）下的细胞活力，来评估细胞保护作用。预处理通过暴露于PDF（60分钟，CAPD2；费森尤斯）或热（15分钟，41.5摄氏度），以及用HSP-72进行瞬时转染来进行。
当间皮细胞通过非致死性暴露于PDF或热进行预处理时，HSP-72显著上调（>5倍，P < 0.01）。通过染料排斥法（减少>50%，P < 0.05）和乳酸脱氢酶（LDH）释放（减少>30%，P < 0.05）评估，预处理后的人间皮细胞对随后的PDF“致死性”暴露具有显著的保护作用。在用HSP-72瞬时转染后，培养的人间皮细胞中HSP-72过表达（>5倍）显示出类似的细胞保护作用（通过染料排斥法减少50%）。通过HSP-72和ApopTag（凋亡检测试剂盒）的双重染色技术在细胞层面证实了这种细胞保护作用。
因此，我们的研究表明，间皮应激反应在实验性腹膜透析中通过诱导HSP-72介导细胞保护作用，并且预处理的刺激不必与随后的损伤相同。这些数据为针对PDF毒性的一种有吸引力的新型治疗方法提供了基础。
