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Objective To investigate the impact of tanshinone Ⅱ A on the level of brain NMDAR1 protein in rats after cardiopulmonary resuscitation and its effects of brain function protection.Methods Seventy-eight SD male rats were randomly (random number) divided into three groups:group A (sham group,n =6),group B (control group,n =36) and group C (Tanshinone Ⅱ A intervention group,n =36).All animals were induced to be models of cardiac arrest by choking.The rats of group C received intravenous injection of Tanshinone Ⅱ A in dose of 15 mg/kg immediately at initiation of resuscitation,while rats of group B were intravenous injected same amount of normal saline instead.Brains tissues of all rats were taken at 1,6,12,24,48 and 72 h after the restoration of spontaneous circulation (ROSC).Immunohistochemical staining method was applied for measuring the levels of brain tissue NMDAR1 and Caspases-3,while water content of the brain was detected by wet and dry weight ratio.The experimentaldata were analyzed by using one-way ANOVA.Results (①)The level of brain NMDAR1 protein in group B increased at 1 h and reached its peak at 6 h after ROSC,then its level gradually declined and dropped below normal at 48 h,72 h,and there were significant difference in variation of NMDAR1 protein levels in comparison with the group A (P ＜ 0.05) ; the NMDAR1 protein levels at 1,6,12 h in group C were significantly lower than those in group B at the same intervals (P ＜ 0.01),but no significant differences were seen at 24,48,72 h (P ＞ 0.05).(②)The level of brain Caspases-3 in group B increased after ROSC,and reached its peak at 48 h after ROSC,then declined and maintained above normal at 72 h,and this variation was significantly different from that of the group A (P ＜ 0.01) ; while the levels of caspase-3 at 1,6,12,24 h in group C were significantly lower than those in group B (P ＜ 0.01),but thses differences at 48,72 h were still significant (P ＜ 0.05).(③)The water content of brain tissue in group B increased at 1 h and reached its peak at 24 h after ROSC,then gradually decreased from 48 h,but maintained above normal at 72 h,and this trend of variation was significantly different from that of group A (P ＜ 0.01 or P ＜ 0.05).Compared with group B,water content of brain tissue in group C decreased more significantly (P ＜ 0.01).Conclusions Tanshinone Ⅱ A down-regulates brain NMDAR1 protein level at early stage in rats as well as significantly inhibits the level of Caspases-3 thereby ameliorating brain edema after cardiopulmonary resuscitation.
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Objective To explore the role of caspase-3 activation and DNA fragmentation in later period of neonatal rat hypoxic-ischemic brain damage(HIBD). Methods DNA fragmentation,caspase-3 mRNA and caspase-3 protein were measured in 2 wks、3 wks and 4wks after setting HIBD animal model in newborn wistar rats by FCM, RT-PCR and Immunohistochemistry. Results (1) Apoptosis lasted 4ks after HIBD. This suggested a long lasting role of apoptosis in neonatal HIBD by TNNEL and EM.(2)Caspase-3 mRNA expression was estimated by semi-quantitative RT-PCR. It was higher in HIBD group(0.771?0.074) than in control group(0.620?0.038, P0.05. Average Avalue of Caspase-3 protein in HIBD group(0.374 ?0.038) at 3 wks was significantly higher than that in control group(0.356?0.020,P
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Objective To explore the relationship between the apoptosis of cerebellar granule cell and the changes of caspases-3 and Bcl-2 in rats with methylmercury intoxication.Methods We chronologically observed the pathological changes of cerebellum in rats given methylmercury[4 mg/(kg?d)] at day 11, 15, 18 and 19. The apoptosis of cerebellar granule cell was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL) method, and the relationship between the apoptosis and the expression of caspases-3 and Bcl-2 was evaluated.Results At day 18, some sparse TUNEL positive granular cells could be observed, mainly in deep lamina adjacent to the white matter. At day 19, the apoptotic cells markedly increased, while the number of granular cells decreased with a well preservation of Purkinje cells demonstrated by calvindine immunostaining. MRF-1 and glial fibrillary acidic protein(GFAP) dyeing demonstrated severe microglia reaction and astrocytosis. From day 11, caspasese-3 mRNA expression up-regulated, increased gradually and reached peak at day 18, then decreased markedly at day 19. In contrast, Bcl-2 mRNA expression down-regulated from day 11, and decreased obviously at day 19. Zic-1 as a marker of granule cell decreased gradually after methylmercury administration and indicated degeneration of granule cells.Conclusion Cerebellar granular cell degeneration induced by methylmercury is related to apoptosis mediated by caspases-3 and Bcl-2 down regulated.
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AIM: To investigate the effects of homocysteine (Hcy) on apoptosis and expression of caspases-3 in cultured human vascular smooth muscle cells (HVSMCs). METHODS: HVSMCs were cultured in vitro. The rate of apoptosis in HVSMCs was detected by flow cytometry. The expression of caspases-3 mRNA in HVSMCs was detected by reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: The rates of apoptosis in HVSMCs incubated with 500 ?mol/L Hcy for 0 h, 6 h, 12 h, 24 h and 48 h were 2.39%?0.47%, 2.45%?0.64%, 7.58%?1.02%, 13.37%?4.71% and 17.69%?3.13%, respectively. The ratio of the absorbance of caspases-3 mRNA/GAPDH was 0.24?0.08, 0.29?0.10, 0.89?0.26, 1.37?0.24 and 1.82?0.53,respectively, suggesting that Hcy induces the apoptosis and expression of caspases-3 mRNA in HVSMCs in a time-dependent manner. The rates of apoptosis in HVSMCs incubated with 0, 100, 200, 500 or 1 000 ?mol/L Hcy for 24 h were 2.68%?0.23%, 2.79%?0.12%, 8.45%?2.41%, 13.37%?4.71% and 23.75%?5.56%, respectively, revealing that Hcy induced the apoptosis in HVSMCs in a concentration-dependent manner. CONCLUSION: Hcy induces the apoptosis in HVSMCs by regulating the expression of caspases-3, which may be one of the mechanisms involved in atherosclerotic effects of Hcy.