Characterization of human proximal tubular cells after hypoxic preconditioning: constitutive and hypoxia-induced expression of heat shock proteins HSP70 (A, B, and C), HSC70, and HSP90.
In animal models of cardiac or cerebral ischemic preconditioning, induction of heat shock proteins (HSPs), especially HSP70, correlates with protection from subsequent injury. The extent of HSP70 induction after stress correlates inversely with initial HSP70 levels. Primate cells, unlike nonprimate cells, express high basal levels of HSP70; thus, primate cells may respond differently to preconditioning than nonprimate cells. We have demonstrated that exposing cultured human proximal tubular epithelial cells (PTEC) to 12 h of hypoxia followed by a 24-h recovery period (hypoxic preconditioning) induces resistance to subsequent hypoxic injury. Herein, we characterize the expression of HSP70, HSP90, and heat shock cognate-70 (HSC70) in PTEC under basal conditions and after hypoxic preconditioning. By Northern blot analysis, we demonstrate that hypoxic preconditioning of PTEC increases mRNA for HSP70 > HSP90 > HSC70. With reverse transcription and polymerase chain reaction, mRNA transcripts from three different HSP70 genes (HSP70 A, B, and C) were detected in unstressed PTEC. Transcripts from these genes were also detected in freshly isolated human renal cortex, indicating that all three genes are expressed in vivo. By Western blot analysis, we demonstrate that PTEC express high basal levels of HSP70, HSC70, and HSP90. Hypoxic preconditioning did not lead to a significant increase in protein content of any of these HSPs, despite increased mRNA levels. This suggests that HSP accumulation cannot account for the development of cytoresistance after hypoxic preconditioning in PTEC. However, high basal expression of HSP70 in human PTEC may contribute to their innate resistance for hypoxia.
Turman
M A
MA
Department of Pediatrics, Ohio State University, Columbus 43205, USA.
Kahn
D A
DA
Rosenfeld
S L
SL
Apple
C A
CA
Bates
C M
CM
D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D015687:Cell Hypoxia; D002478:Cells, Cultured; D017931:DNA Primers; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D018869:Heat-Shock Response; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism*; D008969:Molecular Sequence Data; D010100:Oxygen; D012333:RNA, Messenger
D002352:Carrier Proteins; D017931:DNA Primers; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; C494371:HSPA8 protein, human; D012333:RNA, Messenger; D010100:Oxygen