Hypoxia/re-oxygenation injury induces apoptosis of LLC-PK1 cells by activation of caspase-2.
Hypoxia/re-oxygenation injury induces apoptosis in renal tubule cells, but its underlying molecular pathways are not fully elucidated. Activation of caspase-2 has recently been proposed as a novel mechanism of apoptosis in fibroblasts. In this study we examined whether hypoxia/re-oxygenation injury induces apoptosis in proximal tubule cells by activation of caspase-2. Porcine proximal tubule (LLC-PK1) cells were subjected to hypoxia/re-oxygenation injury in the presence or absence of caspase inhibitors. Apoptosis was detected by DNA laddering, flow cytometry, and immunocytochemistry for Bax and cytochrome c. The activity of caspases-2, 8 and 9 was measured. Apoptosis was evident after hypoxia/re-oxygenation and was best prevented by pretreatment with caspase-2 inhibitor. Hypoxia/re-oxygenation resulted in a dramatic increase in caspase-2 activity (32-fold, in comparison with a 16-fold increase in caspase-8 activity and a tenfold increase in caspase-9 activity). Immunocytochemistry revealed Bax activation and translocation to mitochondria and cytochrome c release into the cytosol following hypoxia/re-oxygenation, both of which were significantly suppressed by pretreatment with caspase-2 inhibitor. These results indicate that hypoxia/re-oxygenation injury in cultured proximal tubule cells induced apoptosis by activation of caspase-2, which is required for the mitochondrial translocation of Bax.
Park
Moon Soo
MS
Pediatrics, Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul, South Korea. parkms2512@yahoo.co.kr
Kim
Beom-Su
BS
Devarajan
Prasad
P
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053143:Caspase 2 / Q000378:metabolism*; D015687:Cell Hypoxia / Q000502:physiology*; D002460:Cell Line; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D007687:Kidney Tubules, Proximal / Q000378:metabolism / Q000473:pathology*; D008928:Mitochondria / Q000378:metabolism; D010100:Oxygen / Q000502:physiology*; D013552:Swine; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; D053143:Caspase 2; D010100:Oxygen