title: Mitochondria-targeted antioxidant peptide SS31 prevents hypoxia/reoxygenation-induced apoptosis by down-regulating p66Shc in renal tubular epithelial cells.
abstract: OBJECTIVE
  Ischemia/reperfusion injury plays a crucial role in renal transplantation and represents a significant risk factor for acute kidney injury and delayed graft function. Mitochondria-targeted antioxidant peptide SS31 has been shown to attenuate ischemia/reperfusion injury by inhibiting oxidative stress. The present study was carried out to investigate whether the pretreatment of SS31 could reduce hypoxia/reoxygenation (H/R)-induced injury by inhibiting p66Shc.
  
  
  METHODS
  The cultured rat renal proximal tubular cell line NRK52E cells were exposed to 24 h hypoxia (5% CO2, 1% O2, 94% N2) followed by 6 h reoxygenation (5% CO2, 21% O2, 74% N2). SS31 was added to the culture medium 4 h prior to the treatment. Then the cell viability, apoptosis, and oxidative stress levels were determined. In addition, western blot analysis was performed to determine the expression of p66Shc, p-p66Shc, cytochrome c, and caspase-3.
  
  
  RESULTS
  H/R induced apoptotic cell death, accompanied with activation of total and p-p66Shc in NRK52E cells. Pretreatment with SS31 or overexpression of a dominantnegative Ser36 mutant p66Shc (p66Shc S36A) or p66Shc siRNA prevented cell death, whereas the protection effect of SS31 was completely blocked by overexpression of wild-type p66Shc. Furthermore, SS31 pretreatment reduced H/R-induced intracellular oxidative stress, cytochrome c translocation to the cytoplasm, and caspase-3 activation through inhibiting p66Shc.
  
  
  CONCLUSIONS
  This study revealed that SS31 pretreatment serves a protective role against H/R-induced apoptosis of human renal tubular epithelial cells, and the mechanism is related to suppression of p66Shc.
authors.affiliation: Organ Transplantation Institute of PLA, Changzheng Hospital, Second Military Medical University, Shanghai, China.
authors.forename: Wen-Yu
authors.initials: WY
authors.lastname: Zhao
authors.forename: Shu
authors.initials: S
authors.lastname: Han
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.forename: You-Hua
authors.initials: YH
authors.lastname: Zhu
authors.forename: Li-Ming
authors.initials: LM
authors.lastname: Wang
authors.forename: Li
authors.initials: L
authors.lastname: Zeng
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D015687:Cell Hypoxia; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology; D008315:Malondialdehyde / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects; D010766:Phosphorylation; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1
chemical_list: D000975:Antioxidants; D009842:Oligopeptides; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D055765:Shc Signaling Adaptor Proteins; C528035:Shc1 protein, rat; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; C506540:arginyl-2,'6'-dimethyltyrosyl-lysyl-phenylalaninamide; D008315:Malondialdehyde; D045304:Cytochromes c; D053148:Caspase 3