Zhang Hui-Ling, Gu Zhen-Lun, Savitz Sean I, Han Feng, Fukunaga Kohji, Qin Zheng-Hong
Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Soochow University School of Medicine, Suzhou, China.
J Neurosci Res. 2008 Apr;86(5):1132-41. doi: 10.1002/jnr.21569.
We have previously reported that prostaglandin A(1) (PGA(1)) reduces infarct size in rodent models of focal ischemia. This study seeks to elucidate the possible molecular mechanisms underlying PGA(1)'s neuroprotective effects against ischemic injury. Rats were subjected to permanent middle cerebral artery occlusion (pMCAO) by intraluminal suture blockade. PGA(1) was injected intracerebroventricularly (icv) immediately after ischemic onset. Western blot analysis was employed to determine alterations in IkappaBalpha, pIKKalpha, and peroxisome proliferator-activated receptor-gamma (PPAR-gamma). Immunohistochemistry was used to confirm the nuclear translocation of nuclear factor-kappaB (NF-kappaB) p65 and the expression of PPAR-gamma. RT-PCR was used to detect levels of c-Myc mRNA. The contribution of PPAR-gamma to PGA(1)'s neuroprotection was evaluated by pretreatment with the PPAR-gamma irreversible antagonist GW9662. A brief increase in pIKKalpha levels and rapid reduction in IkappaBalpha were observed after ischemia. PGA(1) blocked ischemia-induced increases in pIKKalpha levels and reversed the decline in IkappaBalpha levels. Ischemia-induced nuclear translocation of NF-kappaB p65 was attenuated by PGA(1). PGA(1) also repressed the ischemia-induced increase in expression of NF-kappaB target gene c-Myc mRNA. Immunohistochemistry demonstrated an increase in PPAR-gamma immunoreactivity in the nucleus of striatal cells at 3 hr after pMCAO. Western blot analysis revealed that the expression of PPAR-gamma protein significantly increased at 12 hr and peaked at 24 hr. PGA(1) enhanced the ischemia-triggered induction of PPAR-gamma protein. Pretreatment with the irreversible PPAR-gamma antagonist GW9662 attenuated PGA(1)'s neuroprotection against ischemia. These findings suggest that PGA(1)-mediated neuroprotective effect against ischemia appears to be associated with blocking NF-kappaB activation and likely with up-regulating PPAR-gamma expression.
我们之前曾报道，前列腺素A(1)（PGA(1)）可减小局灶性缺血啮齿动物模型的梗死面积。本研究旨在阐明PGA(1)对缺血性损伤的神经保护作用背后可能的分子机制。通过腔内缝线阻塞法对大鼠进行永久性大脑中动脉闭塞（pMCAO）。在缺血发作后立即经脑室内（icv）注射PGA(1)。采用蛋白质免疫印迹分析来确定IκBα、磷酸化IκKα和过氧化物酶体增殖物激活受体γ（PPAR-γ）的变化。免疫组织化学用于确认核因子κB（NF-κB）p65的核转位以及PPAR-γ的表达。逆转录聚合酶链反应（RT-PCR）用于检测c-Myc mRNA水平。通过用PPAR-γ不可逆拮抗剂GW9662进行预处理来评估PPAR-γ对PGA(1)神经保护作用的贡献。缺血后观察到磷酸化IκKα水平短暂升高，IκBα迅速降低。PGA(1)可阻断缺血诱导的磷酸化IκKα水平升高，并逆转IκBα水平的下降。PGA(1)可减弱缺血诱导的NF-κB p65核转位。PGA(1)还可抑制缺血诱导的NF-κB靶基因c-Myc mRNA表达增加。免疫组织化学显示，pMCAO后3小时，纹状体细胞核中PPAR-γ免疫反应性增加。蛋白质免疫印迹分析显示，PPAR-γ蛋白表达在12小时时显著增加，并在24小时时达到峰值。PGA(1)增强了缺血触发的PPAR-γ蛋白诱导。用不可逆的PPAR-γ拮抗剂GW9662进行预处理可减弱PGA(1)对缺血的神经保护作用。这些发现表明，PGA(1)介导的对缺血的神经保护作用似乎与阻断NF-κB激活有关，并且可能与上调PPAR-γ表达有关。
