These data support the hypothesis that HSP-70 interacts with cytoskeletal elements during the restoration of proximal tubule cell structure and polarity after renal ischemia. This experimental approach represents a new in vitro assay to study further the role of HSP in cellular repair.

Analysis of HSP expression represents a novel tool to assess biocompatibility of PDF. Among the HSP investigated, HSP-72 is the most predictive and accurate parameter to assess mesothelial cell injury in the early phase of exposure to PDF.

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.

Low biocompatibility of peritoneal dialysis fluid (PDF) injures mesothelial cells and activates their stress response. In this study, we investigated the role of heat shock proteins (HSP), the main cytoprotective effectors of the stress response, in cytoskeletal stabilization of mesothelial cells in experimental peritoneal dialysis. In cultured human mesothelial cells, cytoskeletal integrity was assessed by detergent extractability of marker proteins following in vitro PDF exposure. Effects of HSP on stabilization of ezrin were evaluated by a conditioning protocol (PDF pretreatment) and repair assay, based on coincubation of cytoskeletal protein fractions with recombinant HSP-72 or HSP-72 antibodies. In the rat model, detachment of mesothelial cells from their peritoneal monolayer during in vivo PDF exposure was assessed with and without overexpression of HSP-72 (by heat conditioning). In vitro, cytoskeletal disruption on sublethal PDF exposure was demonstrated by significantly altered detergent extractability of ezrin and ZO-1. Restoration was associated with significant induction and cytoskeletal redistribution of HSP during recovery. Both the conditioning protocol and in vitro repair assay provided evidence for HSP-72-mediated cytoskeletal stabilization. In the rat model, overexpression of HSP-72 following heat conditioning resulted in significantly reduced detachment of mesothelial cells on in vivo exposure to PDF. Our results establish an essential role of HSP in repair and cytoprotection of cytoskeletal integrity in mesothelial cells following acute in vitro and in vivo exposure to PDF. Repeated exposure to PDF, as is the rule in the clinical setting, may not only cause repeat injury to mesothelial cells but rather represents a kind of inadvertent conditioning treatment.

Long-term peritoneal dialysis results in continuous injury to the mesothelial cell layer. In order to assess degrees of peritoneal dialysis fluid (PDF)-induced cellular injury, morphologic and functional parameters have been investigated in leukocytes (from the blood or peritoneal cavity), cultured mesothelial cells, or fibroblasts (1,2). Whereas most investigations focused on the assessment of cell viability or function after PDF exposure, little is known about early cellular events activated upon the cells' sensing of injury.In this respect, the heat-shock proteins (HSP) are the major class of proteins involved in cytoprotection against injury. Heat-shock proteins are found throughout the cell and the amount of all HSP isoforms can constitute up to 5% of total cellular proteins. Heat-shock proteins are rapidly induced by a variety of cellular stressors such as heat, UV light, or cytotoxic agents. Increased HSP expression is preceded by activation of the respective transcription factor, heat-shock factor (HSF), which has multiple binding sites in the promoter region of most inducible HSP genes (3,4). Although staining for HSP has recently been described in peritoneal biopsy samples of continuous ambulatory peritoneal dialysis (CAPD) patients, there are no data available on the mesothelial stress response after exposure to PDF (5).In this study, we hypothesized that commercially available PDF induce HSP expression in cultured human mesothelial cells. MATERIALS AND METHODS