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. The aim of this study was to test the hypothesis that HSP-70 interacts in vitro with cytoskeletal elements obtained from rat renal cortex during early reflow 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 18 hours of reflow after 45 minutes of renal ischemia, or from controls. Aliquots of isolated pellets were coincubated with aliquots of isolated supernatants in different combinations. A repeat Triton extraction was performed, and differential distribution of Na, K-ATPase 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 anchorage of Na,K-ATPase) and noncytoskeletal supernatant obtained during later reflow (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 were abolished by the addition of anti-HSP-70 antibodies.


CONCLUSIONS
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.
Bidmon
B
B
Kinderdialyse, and Ludwig Boltzmann Institut, Department of Pediatrics, Universitatsklinik für Kinder- und Jugendheilkunde, AKH Wien, Wien, Austria.
Endemann
M
M
Müller
T
T
Arbeiter
K
K
Herkner
K
K
Aufricht
C
C
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D015153:Blotting, Western; D002458:Cell Fractionation / Q000379:methods; D003599:Cytoskeleton / Q000378:metabolism; D003902:Detergents; D004305:Dose-Response Relationship, Drug; D018840:HSP70 Heat-Shock Proteins / Q000276:immunology / Q000494:pharmacology*; D007511:Ischemia / Q000188:drug therapy* / Q000473:pathology*; D007672:Kidney Cortex / Q000098:blood supply / Q000201:enzymology / Q000473:pathology; D007687:Kidney Tubules, Proximal / Q000098:blood supply / Q000201:enzymology / Q000473:pathology*; D008297:Male; D017830:Octoxynol; D051381:Rats; D017207:Rats, Sprague-Dawley; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis / Q000378:metabolism
D000906:Antibodies; D003902:Detergents; D018840:HSP70 Heat-Shock Proteins; D017830:Octoxynol; D000254:Sodium-Potassium-Exchanging ATPase