亚甲基蓝改善脑炎性衰老大鼠认知功能及机制探索

Improved effect and underlying mechanism of methylene blue on cognitive function in brain-inflammatory-aging rats

目的 探讨亚甲基蓝(MB)改善脑炎性衰老大鼠认知功能及机制。方法 选用健康12月龄SD大鼠38只,随机分为健康对照组8只,脂多糖组、MB溶剂组和MB组各10只。建立脂多糖诱导的脑慢性炎性衰老模型,MB经皮下缓释泵注射1周[0.5 mg/(kg·d)]。采用T型迷宫实验和新物体识别实验测试大鼠的空间学习记忆;使用免疫荧光染色检测大鼠海马CA1区小胶质细胞和星形胶质细胞的激活;应用酶联免疫吸附测定试剂盒检测炎性因子白细胞介素(IL)1β和IL-6的释放;检测海马CA1区神经元死亡数量。结果 脂多糖组和MB溶剂组探索新、旧物体时间比较无显著差异(P>0.05),MB组探索新物体时间显著多于旧物体[(22.50±4.32)s vs(11.60±3.01)s,P=0.000]。与健康对照组比较,脂多糖组和MB溶剂组新臂交替选择率、神经元核抗原(NeuN)表达显著降低,离子钙接头蛋白1(Iba-1)、胶质纤维酸性蛋白(GFAP)表达和IL-1β、IL-6表达显著升高(P<0.05)。与MB溶剂组比较,MB组新臂交替选择率、NeuN表达显著升高,Iba-1、GFAP表达和IL-1β、IL-6表达显著降低(P<0.05)。结论MB皮下给药能显著抑制脂多糖诱导的脑炎性衰老对空间学习记忆的损害,其机制可能与MB对炎性小胶质细胞的抑制以及海马体神经元保护紧密相关。

Objective To determine the improved effect of methylene blue(MB)on cognitive func-tion in brain-inflammatory-aging rats and investigate the underlying mechanism.Methods A total of 38healthy 12-month-old SD rats were randomly divided into healthy control group,lipopo-lysaccharide(LPS)group,MB vehicle group and MB group,with 8rats in the control and 10rats in the other three groups.LPS was injected into the fourth ventricle with aid of a subcutaneous sustained release pump to establish a rat model of brain chronic inflammatory aging.MB of 0.5 mg/(kg·d)was added into the pump in the rats from the MB group.T-maze test and new object recognition test were employed to evaluate the learning and memory abilities of the rats.The acti-vation of microglia and astrocytes in the hippocampal CA1region of the rats was detected by im-munofluorescence assay.The release of inflammatory factors IL-1βand IL-6 was measured by ELISA,and neuronal death in the CA1region was assessed by neuronal nuclei(NeuN)fluores-cence staining.Results There was no significant difference in the exploration time for new and old objects between the LPS group and the MB solvent group(P>0.05).The MB group spent significantly longer time in exploring the new objects than the old object(22.50±4.32svs 11.60±3.01s,P=0.000).The alternating selection rate of new arm and expression level of NeuN antigen were significantly decreased,and the expression levels of ionized calcium binding adaptor mole-cule-1(Iba-1)and glial fibrillary acidic protein(GFAP)and the contents of IL-1βand IL-6were obviously increased(P<0.05)in the LPS group and the MB vehicle group than the healthy con-trol group.Compared with the MB vehicle group,the MB group had notably increased alternating selection rate of new arms and higher NeuN expression level,and decreased Iba-1and GFAP ex-pression and IL-1βand IL-6contents(P<0.05).Conclusion Subcutaneous administration of MB could significantly inhibit the damages of spatial learning and memory abilities in the LPS-induced brain chronic inflammatory aging rats.The mechanism may be closely associated with MB inhibi-ting inflammatory glial cells and protecting hippocampal pyramidal neurons.