title: Hypoxia-induced apoptosis in cultured glomerular endothelial cells: involvement of mitochondrial pathways.
abstract: BACKGROUND
  Glomerular endothelial cells (GENs) play a key role in the preservation and reconstruction of the glomerular capillary network following injury, thus maintaining the tissue oxygenation. Accumulating evidence has shown that failure to maintain the microcirculation leads to irreversible glomerular injury and glomerular sclerosis. In this regard, the behavior of endothelial cells in a hypoxic milieu is of interest.
  
  
  METHODS
  We exposed cultured GENs to hypoxia and observed apoptosis by annexin V assay. We examined mitochondrial signaling, focusing on Bcl2 and Bax by real-time polymerase chain reaction (PCR), immunocytochemistry, and immunoprecipitation. Furthermore, we examined the response to hypoxia in an overexpression model of Bcl2.
  
  
  RESULTS
  Hypoxic treatment induced apoptosis in 12.8%+/- 1.1% of GENs at 24 hours, and in 19.8%+/- 0.9% at 24 hours followed by 8 hours of reoxygenation. The expression of Bcl2 mRNA decreased to 0.45- +/- 0.15-fold at 24 hours, whereas that of Bax increased to 7.3- +/- 1.2-fold 1 hour after reoxygenation, accompanied by translocation from the cytosol to mitochondria. These changes were associated with a decrease in mitochondrial membrane potentials and an increase in caspase-9 activity. Both overexpression of Bcl2 and inhibition of Bax protected GENs from hypoxic injury.
  
  
  CONCLUSIONS
  We conclude that changes of quantity and localization of Bcl2 and Bax contribute to hypoxia-mediated apoptosis of GENs in vitro. Further investigation into glomerular endothelial cell injury and intracellular signaling in a hypoxic milieu is required to better understand and ultimately prevent progression of chronic kidney disease.
authors: Division of Nephrology and Endocrinology, University of Tokyo School of Medicine, Tokyo, Japan.
authors: Tetsuhiro
authors: T
authors: Tanaka
authors: Toshio
authors: T
authors: Miyata
authors: Reiko
authors: R
authors: Inagi
authors: Kiyoshi
authors: K
authors: Kurokawa
authors: Stephen
authors: S
authors: Adler
authors: Toshiro
authors: T
authors: Fujita
authors: Masaomi
authors: M
authors: Nangaku
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics; D001692:Biological Transport / Q000187:drug effects; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology*; D000860:Hypoxia / Q000503:physiopathology*; D007678:Kidney Glomerulus / Q000098:blood supply*; D008297:Male; D008564:Membrane Potentials; D008928:Mitochondria; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010100:Oxygen / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D051028:bcl-2-Associated X Protein
chemical_list: C494828:Bax protein, rat; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein; C506435:Casp9 protein, rat; D053453:Caspase 9; D020169:Caspases; D010100:Oxygen