摘要
目的探讨白藜芦醇抑制新生大鼠生发基质出血(germinal matrix hemorrhage, GMH)后神经炎症和改善受损神经功能的作用。方法采用7日龄SD大鼠脑实质注射细菌性胶原酶方法制备GMH模型。108只大鼠随机分为18组, 每组6只, 包括4个假手术组、GMH(12 h、24 h、72 h、7 d)组、3个GMH+药物(二甲基亚砜)对照组、GMH+白藜芦醇(10、100、1 000 mg/kg)组(其中最佳剂量共3组)、2个GMH+白藜芦醇+白藜芦醇抑制剂(EX527)组。采用负向趋地试验和翻正反射试验评估大鼠短期神经行为;采用水迷宫、失足试验、旋转试验评估大鼠长期神经行为;采用免疫荧光方法评估大鼠脑血肿周围炎性指标分泌情况;采用蛋白印迹试验评估大鼠脑内目标蛋白表达情况。结果 GMH发生后24 h内源性去乙酰化酶1(sirtuin-1, SIRT1)下降至最低, 然后逐渐上升;磷酸化核转录因子κB(nuclear factor kappa B, NF-κB)于12 h开始升高, 7 d降至正常;与GMH+药物对照组及其他剂量组比较, GMH+白藜芦醇100 mg/kg组48 h及72 h短期行为学得分更高;与GMH+药物对照组比较, GMH+白藜芦醇100 mg/kg组22 d后水迷宫、失足试验、旋转试验得分更高;免疫荧光染色显示, GMH+白藜芦醇100 mg/kg组大鼠脑血肿周围白细胞介素1β、髓过氧化物酶阳性细胞数低于GMH+药物对照组和GMH+白藜芦醇+EX527组;蛋白印迹提示GMH+白藜芦醇100 mg/kg组大鼠脑内炎性因子白细胞介素1β、肿瘤坏死因子α和白细胞介素6下降, 使用抑制剂EX527后可抵消白藜芦醇的作用。结论白藜芦醇100 mg/kg可改善GMH大鼠短期和长期神经行为学功能, 减少GMH大鼠脑血肿周围炎性指标阳性细胞数, 其通过SIRT1/NF-κB途径抑制GMH大鼠脑内神经炎性指标的表达。
Objective To study the roles of resveratrol in reducing neuroinflammation and improving neurobehavioral functions after germinal matrix hemorrhage(GMH)in neonatal rat model.Methods GMH model was established intraparenchymally injecting bacterial collagenase in 7-day-old SD rats.108 rats were randomly assigned into 18 groups(6 in each group),including 4 sham groups,GMH(12 h,24 h,72 h,7 d)groups,3 GMH+vehicle(dimethylsulfoxide,DMSO)groups,5 GMH+resveratrol(10 mg/kg,100 mg/kg,1000 mg/kg)groups and 2 GMH+resveratrol+EX527(SIRT1 inhibitor)groups.Negative geotaxis and righting reflex tests were used to evaluate the short-term neurobehavior.Water maze,foot fault and Rotor-Rod tests were used to assess the long-term neurobehavior.Immunofluorescence was used to quantify the IL-1βand MPO positive cells(inflammatory markers)in peri-hematoma area.Western blot was used to evaluate the expression of relevant proteins in the brain.Results Endogenous sirtuin-1(SIRT1)decreased to the lowest level at 24 h and then increased gradually.Phosphorylated NF-κB increased at 12 h,peaked at 72 h and returned to normal level at 7 d after GMH.Compared with the control group and other doses groups,GMH treated with resveratrol(100 mg/kg)had higher short-term behavioral scores at 48 h and 72 h.Compared with the control group,the resveratrol(100 mg/kg)group also had higher scores in water maze,foot fault and Rotor-Rod tests 22 days later.Immunofluorescence showed less positive IL-1βand MPO cells around hematoma in GMH+resveratrol group than both GMH+vehicle group and GMH+resveratrol+EX527 group.Western blot indicated that IL-1,TNF-αand IL-6 expressions were decreased in GMH+resveratrol group and Ex527 could offset the effects of resveratrol.Conclusions Resveratrol(optimal dose:100 mg/kg)can improve the short-term and long-term neurobehavioral functions of neonatal GMH rats.It can reduce GMH cells with positive inflammatory markers around the hematoma,possibly via inhibition of the SIRT1/NF-κB pathway.Resveratrol may be promising for the tre
作者
刘胜鹏
张小丽
彭俊
高灵
张浩然
逯军
Liu Shengpeng;Zhang Xiaoli;Peng Jun;Gao Ling;Zhang Haoran;Lu Jun(Department of Pediatrics,Shenzhen People's Hospital(The Second Clinical Medical College,Jinan University The First Affiliated Hospital,Southern University of Science and Technology),Shenzhen 518020,China;Department of Neurosurgery,Central South University Xiangya School of Medicine Affiliated Haikou Hospital,Haikou 570208,China;Department of Pediatrics,Central South University Xiangya School of Medicine Affiliated Haikou Hospital,Haikou 570208,China)
