title: Liposomal delivery of heat shock protein 72 into renal tubular cells blocks nuclear factor-kappaB activation, tumor necrosis factor-alpha production, and subsequent ischemia-induced apoptosis.
abstract: Heat shock protein 72 (HSP72) is a stress-inducible protein capable of protecting a variety of cells from toxins, thermal stress, and ischemic injury. The cytoprotective role and mechanism of action of HSP72 in renal cell ischemic injury remain unclear. To study this, HSP72 was introduced (liposomal transfer) or induced (thermal stress, 43 degrees Cx1 hour) in renal tubular cells (LLC-PK1) with Western blot confirmation. Cells were subjected to simulated ischemia 24 hours after liposomal HSP72 transfer or thermal stress, and the effect of HSP72 on nuclear factor-kappaB (NF-kappaB) activation (electrophoretic mobility shift assay and immunohistochemistry), IkappaBalpha production (Western blot), postischemic tumor necrosis factor-alpha (TNF-alpha) production (RT-PCR), and apoptosis (TUNEL assay) were determined. In separate experiments, the role of TNF-alpha in apoptosis was determined (anti-TNF-alpha neutralizing antibody). Results demonstrated that both liposomal transfer of HSP72 and thermal induction of HSP72 prevented NF-kappaB activation and translocation, TNF-alpha gene transcription, and subsequent ischemia-induced renal tubular cell apoptosis. Furthermore, TNF-alpha neutralization also inhibited ischemia-induced renal tubular cell apoptosis. These results indicate that liposomal delivery of HSP72 inhibits ischemia-induced renal tubular cell apoptosis by preventing NF-kappaB activation and subsequent TNF-alpha production. Further elucidation of the mechanisms of HSP-induced cytoprotection may result in therapeutic strategies that limit or prevent ischemia-induced renal damage.
authors.affiliation: Department of Urology, Indiana University Medical Center, Indianapolis, Ind 46202, USA. dmeldrum@iupui.edu
authors.forename: Kirstan K
authors.initials: KK
authors.lastname: Meldrum
authors.forename: Arthur L
authors.initials: AL
authors.lastname: Burnett
authors.forename: Xianzhong
authors.initials: X
authors.lastname: Meng
authors.forename: Rosalia
authors.initials: R
authors.lastname: Misseri
authors.forename: Matthew B K
authors.initials: MB
authors.lastname: Shaw
authors.forename: John P
authors.initials: JP
authors.lastname: Gearhart
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Meldrum
mesh_terms: D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D015153:Blotting, Western; D050884:HSP72 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism* / Q000494:pharmacology; D018869:Heat-Shock Response / Q000502:physiology; D006358:Hot Temperature; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D007511:Ischemia / Q000378:metabolism*; D007684:Kidney Tubules / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018374:LLC-PK1 Cells; D008081:Liposomes; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D013552:Swine; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000906:Antibodies; D050884:HSP72 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D020672:I-kappa B Proteins; D008081:Liposomes; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha