| Activation of polymoiphonuclear leukocytes and vascular endothelial cells in the early stage of burns was involved in early ischemic or hypoxic injuries by releasing many mediators such as reactive oxygen species (ROS), elastase and cytokines, which diffused into interstitial tissues and led to injures of organs including gut. ROS, one of the most important mediators released by activated polymoiphonuclear leukocytes and vascular endothelial cells, its role in the apoptosis was studied in in vitro model of intestinal epithelial cells.The key role of mitochondria in apoptosis has been well identified. But there is little study in the relationship between mitochondria and apoptosis of intestinal epithelial cells. We selected mitochondrion as main target for our studying. In vitro injurious model of intestinal epithelial cells induced by hydrogen peroxide was established to study the role of mitochondria in the injuries of intestinal epithelial cells, it would give birth to new ideas or clues for clinical treatments.Firstly, hydrogen peroxide-induced apoptosis of SW-480 cells, a kind of intestinal epithelial cells line, was established. We found: (1) Hydrogen peroxide led to apoptosis-like morphological changes of SW-480 cell such as condensed chromatin, karyopyknosis and apoptotic bodies. (2) Increasing numbers of mitochondria and mitochondria with different sizes, swelling intramitochondrial compartments, or reduced cristae or disappearing outmembrane could be observed in cells under transmission electron microscopy. (3) Consistent with the morphological results, hydrogen peroxide induced apoptosis of SW-480 cells and inhibition of cellular respiration. Our results indicated that hydrogenperoxide led to apoptosis of SW-480 cells related to morphological and functional changes of mitochondria.Secondly, The possible role of mitochondria in this in vitro model of hydrogen peroxide-induced apoptosis of SW-480 cells was studied by investigating expression of Bcl-2, Bax, ERK-1 and cytochrome c, changes of mitochondrial membrane potential, redistribution of mitochondria and expression of messenger RNA of AIF. We found: (1) Hydrogen peroxide resulted in decreasing expression of Bcl-2, increasing expression of Bax, especially in those wizened cells, and markedly reducing ratio of Bcl-2 to Bax.(2) Hydrogen peroxide led to inhibition of expression of ERK-1 in some degree.(3) Increasing expression of AIF mRNA and release of cytochrome c were also determined in hydrogen peroxide-treated SW-480 cells. (4) The spatial distribution of mitochondria evolved from an originally scattered bipolar or nearly symmetric distribution to an asymmetric, clustered distribution or even massive aggregation in SW-480 cell after treated with hydrogen peroxide. (5) Collapse of mitochondrial membrane potential occurred in hydrogen peroxide-treated SW-480 cells. Our results suggested the key role of mitochondria in the apoptosis of SW-480 cells treated by hydrogen peroxide.Thirdly, expression of mitochondrial genes encoding cytochrome c oxidase and ATPase was studied by RT-PCR and activities of them were investigated simultaneously in the same model. We found: (1) Hydrogen peroxide led to differential expression of mitochondrial genes with some genes up-regulated or down-regulated by dose dependent manner. (2) Difference was very obvious in expression of mitochondrial genes of cells treated with hydrogen peroxide in the concentration of 400umol/L or 4mmol/L. In generally, differential expression of mitochondrial genes was characterized with up-regulation of mitochondrial genes in the concentration of 400umol/L and down-regulation in the concentration of 4mmol/L. (3) Hydrogen peroxide resulted in decreasedzactivities of cytochrome c oxidase and ATPase. It was suggested that differential expression of mitochondrial genes encoding cytochrome c oxidase and ATPase be involved in apoptosis of intestinal epithelial cells by affecting activities of cytochorme c oxidase and ATPase.Finally, We studied expre...
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