OBJECTIVE: To investigate the effect of recombinant adenovirus-mediated heat shock protein 70 (HSP70) on energy metabolism of mitochondria in intestinal epithelial cells (IEC-6) after hypoxia/reoxygenation injury . METHODS: IEC-6 cells were transfected with HSP70 recombinant adenovirus vectors (Ad-HSP70) and empty adenovirus vectors. The expression of HSP70 protein was detected by Western blotting. Cultured IEC-6 cells were divided into: control group (without treatment), hypoxia/reoxygenation group (with challenge of hypoxia/reoxygenation) and Ad-HSP70 transfection group (with challenge of hypoxia/reoxygenation after Ad-HSP70 transfection). The activity of mitochondrial dehydrogenase was assessed by MTf method. The contents of cellular ATP, ADP , AMP and energy charge (EC)were determined by high-performance liquid chromatography (HPLC). RESULTS: The expression of HSP70 protein in IEC-6 cells was significantly upregulated after Ad-HSP70 transfection compared with empty adenovirus vector transfection. Compared with that in control group, the activity of mitochondrial dehydrogenase was significantly lowered in IEC-6 cells in hypoxia/reoxygenation group (P < 0.01). The activity of mitochondrial dehydrogenase in Ad-HSP70 transfection group was significantly greater than that in hypoxia/reoxygenation group (P < 0.01). Compared with those in control group,the content of cellular ATP was significantly decreased in hypoxia/reoxygenation group, the contents of cellular ADP and AMP were significantly increased. The above cell energy indices in Ad-HSP70 transfection group was similar to those in control group (P > 0.05), which were ameliorated compared with those in hypoxia/reoxygenation group (P < 0.050 or P < 0.01). The cellular EC in hypoxia/reoxygenation group (0.615 +/- 0.060) was significantly lower than that in control group (0.748 +/- 0.012, P < 0.01) and Ad-HSP70 transfection group (0.736 +/- 0.028, P < 0.01). CONCLUSION: Ad-HSP70 transfection in IEC-6 cells can upregulate the expression of HSP70, the content of cellular ATP and EC after hypoxia/reoxygenation, and protect mitochondrial function. Mitochondria may be one of main target organelles for HSP70 in protection of IEC against hypoxia/reoxygenation injury.
OBJECTIVE: To investigate the effect of recombinant adenovirus-mediated heat shock protein 70 (HSP70) on energy metabolism of mitochondria in intestinal epithelial cells (IEC-6) after hypoxia/reoxygenation injury . METHODS: IEC-6 cells were transfected with HSP70 recombinant adenovirus vectors (Ad-HSP70) and empty adenovirus vectors. The expression of HSP70 protein was detected by Western blotting. Cultured IEC-6 cells were divided into: control group (without treatment), hypoxia/reoxygenation group (with challenge of hypoxia/reoxygenation) and Ad-HSP70 transfection group (with challenge of hypoxia/reoxygenation after Ad-HSP70 transfection). The activity of mitochondrial dehydrogenase was assessed by MTf method. The contents of cellular ATP, ADP , AMP and energy charge (EC)were determined by high-performance liquid chromatography (HPLC). RESULTS: The expression of HSP70 protein in IEC-6 cells was significantly upregulated after Ad-HSP70 transfection compared with empty adenovirus vector transfection. Compared with that in control group, the activity of mitochondrial dehydrogenase was significantly lowered in IEC-6 cells in hypoxia/reoxygenation group (P < 0.01). The activity of mitochondrial dehydrogenase in Ad-HSP70 transfection group was significantly greater than that in hypoxia/reoxygenation group (P < 0.01). Compared with those in control group,the content of cellular ATP was significantly decreased in hypoxia/reoxygenation group, the contents of cellular ADP and AMP were significantly increased. The above cell energy indices in Ad-HSP70 transfection group was similar to those in control group (P > 0.05), which were ameliorated compared with those in hypoxia/reoxygenation group (P < 0.050 or P < 0.01). The cellular EC in hypoxia/reoxygenation group (0.615 +/- 0.060) was significantly lower than that in control group (0.748 +/- 0.012, P < 0.01) and Ad-HSP70 transfection group (0.736 +/- 0.028, P < 0.01). CONCLUSION: Ad-HSP70 transfection in IEC-6 cells can upregulate the expression of HSP70, the content of cellular ATP and EC after hypoxia/reoxygenation, and protect mitochondrial function. Mitochondria may be one of main target organelles for HSP70 in protection of IEC against hypoxia/reoxygenation injury.