| Calcineurin is the only protein phosphatase that is modulated by eukaryotic Ca2+ and calmodulin-dependent serine/threonine. Calcineurin is widely distributed in mammalian tissues, with the highest levels found in brain. In addition, calcineurin has been observed in heart, skeletal muscle, T lymphocytes. CaN can be modulated by external signal that is mediated by Ca2+ and oxidation-reduction potential. In order to further investigate the function and mechanism of calcineufin, we examine the activity of calcineurin and ascertained the mechanism in case of different tissues, redox and in brain ischemia reperfusion.ObjectivePartâ… :Observing distribution of CaN and characteristic of calcium-dependence of calcineurin in different organs.Partâ…¡:To observe the effects of ischemic preconditioning on the activity of calcineurin in rat cerebral.Partâ…¢:Activities of calcineurin were compared among the control group, reactive oxygen species produces by xanthine/xanthine oxidase system, SOD, CAT, mannitol, and DTT group.MethodsPartâ… :Measuring the activity of calcineurin in different tissues, and the activity of calcineurin with different calcium concentration by UV spectrophotometer.Partâ…¡:1 We induced cerebral ischemia and cerebral ischemia preconditioning in normal rats and measured the activity of calcineurin in each group.2 The brain necrotic areas can be observed by the TTC staining. 3 We assayed the activities of calcineurin. The reaction temperature was 30℃, and the reaction was monitored in 405 nm and the interval of sampling was 30 second. The total reaction time was 5 minutes.Partâ…¢:We extract CaN from skeletal muscle. After add oxidize or antioxidize compound to each group, we measure the activity of CaN.ResultsPartâ… :1 Comparation of CaN activities in different tissues. The activity of calcineurin (nmol/min/mg protein) in brain is higher than that in other tissues (P＜0.01).2 The calcium-dependence of calcineurin in different tissues. With the increase of calcium concentration, the activity of calcineurin increased. When pCa is over 4.52, the activity of calcineurin decreased sharply.Partâ…¡:1 Comparation of the brain necrotic volume between the control group and that of the experimental group. After the brain reperfusion, the rat brain of the control group and the experimental group were stained by TTC. Brain necrotic areas appeared in both groups. The necrotic volume in the control group was larger than the experimenttal group (14.83±4.37 % vs. 5.82±2.98 %, Pï¼…0.05), which had significant discrimination.2 Comparation of the activities of CaN between the control group and that of the experimental group. The activity of calcineurin (nmol/min/mg protein) in ischemia preconditioning group is higher than that in control group (5.31±1.68 vs. 3.19±1.26, P＜0.05).Partâ…¢:1 Xanthine/xanthine oxidase system can reduce the activity of CaN. The activity of calcineurin (nmol/min/mg protein) in blank group is lower than that in control group (2.63±0.41 vs. 1.76±0.59, P＜0.01).2 Antioxidize compound can protect the activity of CaN. Comparing with control group, the activities of CaN (nmol/min/mg protein) in SOD group,catalase group and DTT group is higher obviously. SOD group and control group (2.05±0.38 vs.1.76±0.59, P＜0.05), catalase group and control group (2.40±0.48 vs.1.76±0.59, P＜0.01), DTT group and control group (2.22±0.40 vs.1.76±0.59, P＜0.01). But the mannitol group and control group (1.78±0.62 vs.1.76±0.59, P＞0.05) have no significant discrimination.Conclusions1 The results suggest that the activity of phosphatases in brain are higher than in other tissues, and the result of calcium dependence presents a change of a bell shape and it is same in different tissues.2 The ischemia preconditioning can reduce the brain necrotic volume after ischemia reperfusion, decrease the brain injury degree, and protect the ischemia brain.3 The result shows that SOD, catalase and DTr can protect the activity of CaN, but the discrimination between the control group and the mannitol group is not significant.4 The result of this study have approved the protective effect of the ischemia preconditioning on the ischemic rat brain, which can help to apply new ideas about prevention and cure the cerebral ischemia reperfusion injury.
|
PDF Full Text Request