Heat shock protein-70 repairs proximal tubule structure after renal ischemia

Background Recent studies have suggested a role of heat shock protein (HSP) -70 in cytoskeletal repair during cellular recovery from renal ischemia. Th e aim of this study was to test the hypothesis that HSP-70 interacts in vit ro with cytoskeletal elements obtained from rat renal cortex during early r eflow after renal ischemia. Methods. Cellular proteins were fractionated into cytoskeletal pellets and noncytoskeletal supernatants by Triton X-100 extraction of rat renal cortex obtained after 15 minutes or IS hours of reflow after 45 minutes of renal ischemia, or from controls. Aliquots of isolated pellets were coincubated w ith aliquots of isolated supernatants in different combinations. A repeat T riton extraction was performed, and differential distribution of Na,K-ATPas e or HSP-70 was assessed by Western blots and densitometric analysis. Results. Coincubation of cytoskeletal pellets obtained during early reflow after renal ischemia (exhibiting severe injury of the cytoskeletal anchorag e of Na,K-ATPase) and noncytoskeletal supernatant obtained during later ref low (showing high HSP expression) resulted in specific translocation of HSP -70 from the supernatant into the pellet, functionally associated with dose -dependent stabilization of Na,K-ATPase within this cytoskeletal fraction. These effects could be reproduced by incubation with purified HSP-70 and we re abolished by the addition of anti-HSP-70 antibodies. Conclusion. These data support the hypothesis that HSP-70 interacts with cy toskeletal elements during the restoration of proximal tubule cell structur e and polarity after renal ischemia. This experimental approach represents a new in vitro assay to study further the role of HSP in cellular repair.