JAK2/STAT3信号通路在蛛网膜下腔出血后早期脑损伤中的作用

The involvement of JAK2/STAT3 signaling pathway in early brain injury after subarachnoid hemorrhage

目的探讨酪氨酸激酶/信号传导和转录激活子(JAK2/STAT3)信号通路在小鼠蛛网膜下腔出血(SAH)后早期脑损伤(EBI)中的作用及可能机制。方法运用血管内穿刺法建立小鼠SAH模型。健康雄性C57BL/6J小鼠55只随机分为对照组(Control组,n=15)、手术+生理盐水组(SAH+Saline组,n=21)、手术+JAK2抑制剂组(SAH+AG490组,n=19)。SAH+AG490组小鼠伤后立即给予AG490腹腔注射,SAH+Saline组腹腔注射等量生理盐水。24h后运用疲劳转棒实验评价小鼠的神经功能;运用原位末端标记法检测神经细胞凋亡情况;运用蛋白质印迹法分析SAH后小鼠脑组织t-JAK2、p-JAK2、t-STAT3、p-STAT3的蛋白表达;运用酶联免疫吸附试验分析SAH后小鼠脑组织炎性细胞因子白细胞介素(IL)-1β、IL-6及肿瘤坏死因子α(TNF-α)的表达情况。结果与Control组比较,SAH+Saline组小鼠出现明显的神经细胞凋亡(P<0.05),平衡运动功能明显降低,转棒时间明显缩短(P<0.05);进一步研究发现,JAK2和STAT3蛋白磷酸化水平升高(P<0.05),下游炎性因子IL-1β、IL-6及TNF-α的表达水平明显升高(P<0.05)。与SAH+Saline组比较,SAH+AG490组小鼠脑组织JAK2和STAT3磷酸化水平明显降低(P<0.05),炎性因子IL-1β、TNF-α及IL-6的表达水平明显降低(P<0.05),小鼠神经细胞凋亡明显减少(P<0.05),神经功能障碍明显减轻(P<0.05)。结论小鼠SAH后早期JAK2/STAT3信号通路激活可能参与了EBI的发生、发展过程,其机制可能与调控SAH后早期的神经炎性反应有关。

Objective To investigate the involvement and possible mechanism of janus-activated kinase/signal transducer and activator of transcription 3(JAK2/STAT3)signaling pathway in early brain injury(EBI)after subarachnoid hemorrhage(SAH)in mice.Methods Experimental mice SAH model was established by using intravascular puncture method.A total of 55 healthy male C57BL/6J mice were randomly divided into the control group(n=15),the SAH+Saline group(n=21),and the SAH+JAK2 inhibitor AG490 group(SAH+AG490 group,n=19).The AG490 was administrated immediately to mice in the SAH+Saline group via intraperitoneal injection after the onset of SAH.Mice in the SAH+Saline group received equal amount of saline instead.Twenty-four hours after establishment of SAH model,rotarod test was used to evaluate mice neural motor function;TUNEL staining was used to detect neurons apoptosis.Western blot was used to analyse the protein expressions of t-JAK2,p-JAK2,t-STAT3 and p-STAT3.Enzyme-linked immunosorbent assay was used to detect the inflammation cytokines release,including IL-1β,IL-6 and TNF-αafter SAH.Results Compared with the control group,mice in the SAH+Saline group displayed significant neural apoptosis and neural motor dysfunctions(P<0.05),and the length of RR time was decreased(P<0.05).Further investigation found that JAK2 and STAT3 phosphorylation dramatically increased,while the release of IL-1β,IL-6 and TNF-αalso significantly increased(P<0.05).Compared with the SAH+Saline group,mice brain JAK2 and STAT3 phosphorylation levels were significantly decreased in the SAH+AG490 group(P<0.05),the inflammatory cytokines including IL-1β,IL-6 and TNF-αwere also significantly reduced(P<0.05).Simultaneously,the neural apoptosis was significantly reduced,while the mice neural dysfunctions were significantly alleviated(P<0.05).Conclusion The activation of JAK2/STAT3 signaling pathway may participate in the occurrence and development of EBI after SAH,the underlying mechanism may be associated with regulating the neuroinflammatory response after SAH.