白细胞介素-1β对大鼠视网膜Müller细胞磷酸化的信号转导及转录活化因子3表达的影响

摘要

目的 观察不同浓度白细胞介素-1β(IL-1β)对大鼠视网膜Müller细胞磷酸化的信号转导及转录活化因子3(pSTAT3)表达的影响.方法 将体外培养的Sprague-Dawley(SD)大鼠Müller细胞分别以0.0、0.1、1.0、5.0、10.0 ng/ml浓度的IL-1β刺激24 h后,免疫荧光法和蛋白质印迹(Western botting)检测细胞内pSTAT3的表达改变.SD大鼠32只,随机分为对照组、100、500、1000 ng/ml组,每组均为8只大鼠.分别将磷酸缓冲液(PBS)配制的浓度分别为100、500、1000 ng/ml的IL-1β注入大鼠的左眼玻璃体腔中,对照组大鼠左眼玻璃体腔注射相同容积的PBS,24 h后采用同样方法 检测pSTAT3在视网膜的表达情况.结果 免疫荧光法和Western botting检测结果 显示,IL-1β刺激24 h后,IL-1β浓度为0.0 ng/ml时,pSTAT3在细胞核内有极少量表达,当浓度达到1.0 ng/ml后,pSTAT3在细胞核内的表达随浓度增加而升高,差异有统计学意义(F=46.64,43.78;P<0.01).对照组大鼠视网膜内无明显pSTAT3的表达,当浓度达到100.0 ng/ml后,pSTAT3在视网膜内的表达随浓度增加而升高,差异有统计学意义(F=73.53,43.70;P<0.01);pSTAT3主要表达于内核层和神经节细胞层.双荧光标记显示,对照组大鼠视网膜内无pSTAT3表达,胶质纤维酸性蛋白(GFAP)主要表达于神经节细胞层,并向视网膜色素上皮层呈丝状放射;从100 ng/ml IL-1β刺激24 h后起,内核层就出现颗粒状GFAP和pSTAT3的双标染色.结论 IL-1β可以上调Müller细胞pSTAT3的表达,这可能是Müller细胞发生反应性胶质活化的通路之一.%Objective To observe the influence of interleukin-1β (IL-1β) on the expression of phosphorylated signal transducers and activators of transcription 3 (pSTAT 3) in rat retinal MOiler cells. Methods For in vitro study cultured Mailer cells were treated with IL-1β of different concentrations (0, 0.1, 1, 5 and 10 ng/ml) for 24 hours. For in vivo study, 32 Sprague-Dawley(SD)rats were divided into 4 groups randomly (control group, 100,500 and 1000 ng/ml group) with 8 rats in each group. After 24 hours of injection with phosphate buffered solution (PBS), or 100, 500, 1000 ng/ml IL-1β into the vitreous treated retinas were evaluated by indirect immunofluorescence and western blotting. Results After 24 hours of incubation without IL-1β, pSTAT3 has little expression in cultured Muller cells, but was up-regulated by 1 ng/ml or higher IL-1β in a dosage-dependent manner (F=46.64, 43.78; P<0.01). pSTAT3 was not expressed in adult rat retina, but was up-regulated by vitreous injection of 100 ng/ml or higher IL-1β in a dosage-dependent manner (F=73.53, 43.70; P<0.01). pSTAT3 expressed mainly in inner nuclear layer and ganglion cell layer. Double-labeling showed that there was no co-staining of pSTAT3 and glial fibrillary treated with IL-1β. Conclusions Expression of pSTAT3 in MUller cells could be activated by IL-1β which may represent one pathway link to reactive gliosis.