Overexpression of HSP-72 confers cytoprotection in experimental peritoneal dialysis

Abstract

Background: 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.

Methods: 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.

Results: 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).

Conclusion: 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.