孕酮预处理对大鼠局灶性脑缺血再灌注损伤的保护作用及机制

Neuronal protection of progesterone against ischemic brain injury and underlying molecular mechanisms

目的探讨孕酮预处理对局灶性脑缺血再灌注损伤（focalcerebralischemicandreperfusioninjury，fCIRI）的作用及机制。方法在建立大鼠fCIRI模型前96、48、1h分别给予孕酮（8mg／kg）1次性腹腔注射。建立大鼠fCIRI模型，在fCIRI后第3～8天，检查大鼠的空间记忆功能和缺血侧海马CA，区存活神经元数量。检测孕酮预处理后1—96h的海马细胞外信号调节激酶1／2（ERK1／2）磷酸化水平和活化ERK1／2核转移。结果造模前48、1h经孕酮预处理的大鼠海马CA，区神经元存活数量（mm“）较造模后明显增多（164．3±11．0、218．5±9．1与142．7±12．1，F=29．45，P〈0．01）；造模前48、1h经孕酮预处理的大鼠能明显缩短脑缺血导致的水迷宫逃避潜伏期（S）延长（32．4±14．3、10．3±11．1与19．2±9．6；F=35．84，P〈0．01）。海马CA，区ERK1／2磷酸化水平在孕酮给药后的12～48h明显增强，96h恢复基线。核内ERK1／2磷酸化水平在孕酮给药后2h增加，并持续到48h。孕酮增加胞质ERK1／2及胞核内ERK1／2磷酸化的水平能被RU486阻断。结论孕酮预处理对缺血引起的海马CA，区神经元死亡有保护作用，并能改善空间记忆功能；孕酮预处理的神经保护作用有48h以上的有效时间窗；在孕酮的神经保护作用依赖于孕酮受体介导的ERK1／2信号通路。

Objective To investigate the effect of progesterone pretreatment of focal cerebral ischemie and reperfusion injury （fCIRI） and underlying molecular mechanisms. Methods A single intraperitoneal injection of progesterone （8 mg/kg） given 1 h, 48 h and 96 h before fCIRI was established in male Sprague-Dawley rats. The number of survival of neurons in hippocampal CAl region of the ischemia- side, as well as spatial memory function, was detected on days 3--8 after fCIRI. Extracellular-signal- regulated kinase 1/2 phosphorylation （p-ERK1/2） and nuclear translocation of p-ERK1/2 in hippoeampal CAj region were examined using western blot. Results The number of survival of neuronal cells was significantly increased in ischemic groups treated with progesterone at 1 h and 48 h pre-fCIRI （ 164. 3 ± 11.0, 218. 5 ±9. 1 and 142. 7 ±12. 1 ,F =29. 4,P 〈0. 01 ） compared with fCIRI group treated with vehicle. Likewise, the escape-latency to reach the hidden-platform recorded in day 5 of Morris water maze test was reduced markedly in fCIRI-treatment groups compared with the vehicle group（ 10. 3 ±11.1, 19. 2 ±9. 6 and 32. 4 ±14. 3 ;F = 35.8, P 〈 0. 01 ）. The level of p-ERK1/2 was elevated notably during 24 h to 48 h post- progesterone by western blot, while restored to the baseline at 96 h post-progesterone. Improved nuclear translocation of p-ERK1/2 was observed from 2 h to 48 h post-progesterone. The progesterone receptor antagonist RU486 blocked the exaltation of either intracellular level or nuclear translocation of p-ERK1/2, which was induced by progesterone. Conclusions The pretreatment with progesterone exerts a neuroprotective effect against the ischemia-induced neuronal death arid ameliorates the deficits in spatial memory through enhancing the activation of ERK1/2. The neuroprotection derived from pretreatment with progesterone achieves a time window of not less than 48 h, which is progesterone receptor-mediated ERK1/2 signaling pathway-dependent.