Molecular mechanism of cerebral ischemia through Jak-Stat signaling

脑缺血诱导了许多生长因子和细胞因子的表达，这些生长因子和细胞因子可以保护神经元免受缺血性脑损伤。例如，纤毛神经营养因子（CNTF）、表皮生长因子（EGF）和白细胞介素-6 （IL-6）已被证明可促进Stat3的激活。这些生长因子和细胞因子可以激活Janus蛋白酪氨酸激酶(Jaks)/信号转导和转录激活因子（Stat）通路。最近，我们报道了Stat3在小鼠[1]脑缺血后被激活。然而，对于激活的Stat3在缺血性脑中的功能知之甚少。本研究表明，激活Stat3表达的增加促进了培养星形胶质细胞的细胞死亡。方法与结果：培养新生大鼠前脑星形胶质细胞。表达Stat3野生型的重组腺病毒(St3。Wt)， Stat3显性阴性型（St3.DN）和LacZ按照前面描述的[2]构建。细胞暴露于…更多表达Stat3的腺病毒载体。Wt, Stat3。DN或LacZ（感染倍数10）孵育2h，在无病毒新鲜培养基中孵育至72h。3天后，用表达St3的病毒载体接种。wt和St3。与对照（LacZ）相比，DN显著诱导Stat3蛋白表达。免疫印迹也显示Ad.St3。wt显著诱导p-Stat3(Tyr-705)；DN和Ad。LacZ没有。结果表明，Ad.St3。Wt和Ad.St3。DN在培养的星形胶质细胞中诱导Stat3蛋白表达，仅Ad.St3表达。Wt可使Stat3在tyr705位点磷酸化并激活Stat3。LDH检测显示St3过表达。wt过表达St3可促进细胞死亡。DN和LacZ则没有。此外，St3的caspase-3样活性。Wt处理组较St3显著升高。病毒接种后2d DN组。结论：Stat3的过表达促进了培养的星形胶质细胞的死亡，caspase-3活性的激活可能参与了这种细胞死亡机制。这些数据表明Stat3的激活在缺血性细胞死亡机制中起着至关重要的作用。参考文献：[1]Wen等；神经科学通讯，303:153-156,(2001)[2]Nakajima等；[J]中国农业大学学报(自然科学版

Introduction :Cerebral ischemia induces the expression of a number of growth factors and cytokines that can protect neurons against ischemic brain injury. For example, ciliary neurotrophic factor (CNTF), epidermal growth factor (EGF) and interleukin-6 (IL-6) have been shown to promote Stat3 activation. These growth factors and cytokines can activate Janus protein tyrosine kinases (Jaks)/signal transducers and activators of transcription (Stat) pathways. Recently, we reported that Stat3 was activated following cerebral ischemia in mouse [1]. However, little is known about the function of activated Stat3 in the ischemic brain. Here we demonstrate the increased expression of activated Stat3 promotes cell death in cultured astrocyres.Methods & Results :Astrocytes from the forebrains of newborn rats were cultured. Recombinant adenovirus expressing Stat3 wild type (St3.Wt), Stat3 dominant negative type (St3.DN) and LacZ were constructed as described previously [2]. Astocytes were exposed to … More adenovirus vector expressing Stat3.Wt, Stat3.DN or LacZ (multiplicity of infection 10) for 2h and incubated with a virus free fresh medium up to 72h. Three days later, inoculation with the virus vectors expressing St3.wt and St3.DN remarkably induced Stat3 protein compared with the control (LacZ). Western blots also revealed that Ad.St3.wt remarkably induced p-Stat3(Tyr-705) while Ad.St3.DN and Ad.LacZ did not. These results demonstrated that both Ad.St3.Wt and Ad.St3.DN induced Stat3 protein in cultured astrocytes and that only Ad.St3.Wt could phosphorylate Stat3 at Tyr-705 site and activate Stat3. LDH assay revealed that overexpression of St3.wt promoted cell death while overexpression of St3.DN and LacZ did not. In addition, caspase-3 like activity of the St3.Wt treated group was significantly increased than that of the St3.DN group at 2d after virus inoculation.Conclusion :We have shown that over-expression of Stat3 promotes cell death in cultured astrocytes, and activation of caspase-3 activity may be involved in this cell death mechanism. These data have revealed that activation of Stat3 can play a crucial role in the mechanism of ischemic cell death.References :[1]Wen et al. ; Neurosci Lett, 303:153-156, (2001)[2]Nakajima et al. ; EMBO J, 15:3651-58 (1996) Less