摘要
目的探究黑逍遥散调控丝裂原活化蛋白激酶磷酸酶-1(MKP-1)/c-Jun氨基末端激酶(JNK)信号通路干预阿尔茨海默病(AD)大鼠海马Tau蛋白表达、抑制神经炎症的作用,及其作用机制。方法将SPF级Wistar雄性大鼠随机分为空白组、假手术组、模型组、对照组和低、中、高剂量实验组,每组10只。除空白组和假手术组外,其余5组大鼠双侧海马区注射β-淀粉样蛋白1-42(Aβ1-42)溶液复制AD大鼠模型,假手术组相同方法注射等量0.9%NaCl。将造模成功的大鼠随机分为模型组、对照组(0.5 mg·kg^(-1)盐酸多奈哌齐)和低、中、高剂量实验组(3.82、7.65、15.30 g·kg^(-1)黑逍遥散汤剂)。空白组、假手术组和模型组均灌胃给予等体积的0.9%NaCl。7组大鼠每天灌胃1次,连续灌胃给药42 d。用Morris水迷宫检测大鼠学习记忆能力,用酶联免疫吸附试验法检测海马组织中肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)水平,用蛋白质印迹法检测MKP-1、磷酸化JNK(p-JNK)、磷酸化Tau(p-Tau)蛋白的表达水平。结果中、高剂量实验组和对照组、模型组、假手术组、空白组第5天逃避潜伏期分别为(8.28±7.67)、(7.89±4.18)、(7.86±2.68)、(16.55±4.16)、(6.46±3.30)和(3.60±1.53)s,TNF-α水平分别为(406.56±28.44)、(404.17±22.84)、(402.28±28.36)、(665.89±61.15)、(226.44±34.84)和(218.50±30.16)pg·mL^(-1),IL-6水平分别为(136.54±7.04)、(121.67±5.19)、(119.15±5.87)、(166.27±8.91)、(88.75±5.28)和(79.58±7.53)ng·L^(-1),MKP-1蛋白相对表达水平分别为2.31±0.34、2.59±0.38、2.58±0.37、1.23±0.25、2.64±0.19和2.84±0.18,p-JNK蛋白相对表达水平分别为3.46±0.35、3.45±0.31、3.20±0.23、4.48±0.30、2.87±0.51和2.30±0.26,p-Tau蛋白相对表达水平分别为3.46±0.33、3.24±0.48、3.09±0.31、4.85±1.06、2.69±0.34和2.40±0.55。中、高剂量实验组的上述指标与模型组比较,在统计学上差异均有统计学意义(P<0.05,P<0.01)。结论黑逍遥散可显著改善AD大鼠的学习记忆能力,其作用机制可能与调控MKP-1与JNK蛋白,从而抑制Tau蛋白磷酸化水平、减轻神经炎症有关。
Objective To investigate the effect and mechanism of Heixiaoyao powder in regulating mitogen-activated protein kinase phosphatase-1(MKP-1)/c-Jun N-terminal kinase(JNK)signaling pathway on the level of Tau protein and neuroinflammation in the hippocampus of Alzheimer’s disease(AD)rats.Methods Male Wi sta r rats with SPF grade were randomly divided into blank,sham-operation,model,control and experimental-L,-M,-H groups with 10 rats per group.In addition to the blank and sham-operation groups,the other 5 groups of rats were injected withβ-amyloid 1-42(Aβ1-42)solution in bilateral hippocampus to replicate AD rat model,and the sham-operation group was injected with the same amount of 0.9%NaCl in the same way.Animals successfully replicated in the model were randomly divided into model group,control group(0.5 mg·kg^(-1)donepezil hydrochloride)and experimental-L,-M,-H groups(3.82,7.65,15.30 g·kg^(-1)Heixiaoyao powder decoction).The blank,sham-operation and model groups were given equal volume of 0.9%NaCl by gavage.The drug was given by gavage once a day for 42 days.Morris water maze was used to detect the learning and memory ability of rats.The levels of tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)in hippocampus were detected by enzyme-linked immunosorbent assay.Western blot was used to detect the expression of MKP-1,phospho JNK(p-JNK)and phospho Tau(p-Tau)proteins.Results The escape latency on day 5 of the experimental-M,-H groups,control group,model group,sham-operation group and blank group were(8.28±7.67),(7.89±4.18),(7.86±2.68),(16.55±4.16),(6.46±3.30)and(3.60±1.53)s;the levels of TNF-αin the above groups were(406.56±28.44),(404.17±22.84),(402.28±28.36),(665.89±61.15),(226.44±34.84)and(218.50±30.16)pg·mL^(-1);IL-6 levels were(136.54±7.04),(121.67±5.19),(119.15±5.87),(166.27±8.91),(88.75±5.28)and(79.58±7.53)ng·L^(-1);the relative expression levels of MKP-1 protein were 2.31±0.34,2.59±0.38,2.58±0.37,1.23±0.25,2.64±0.19 and 2.84±0.18;the relative expression levels of p-JNK protein were 3.46±0.35,3.45±0.31,3.20±0.23,4.48±0.30,2.87±0.51 and 2.30±0.26;the relative expression levels of p-Tau protein were 3.46±0.33,3.24±0.48,3.09±0.31,4.85±1.06,2.69±0.34 and 2.40±0.55,respectively.Compared with the model group and the normal group,compared with the experimental group and the model group,the differences of above indexes were statistically significant(P<0.05,P<0.01).Conclusion Heixiaoyao powder can improve the learning and memory ability of AD rats,and its mechanism may be related to the regulation of MKP-1 and JNK proteins,thus inhibiting the phosphorylation level of Tau protein and alleviating neuroinflammation.
作者
王虎平
孟志鹏
胡韵韵
吕育洁
杨娇
陈怡琴
WANG Hu-ping;MENG Zhi-peng;HU Yun-yun;LÜYu-jie;YANG Jiao;CHEN Yi-qin(School of Basic Medicine,Gansu University of Chinese Medicine,Lanzhou 730000,Gansu Province,China;Gansu Key Laboratory of Traditional Chinese Medicines Excavation and Innovative Transformation,Gansu Engineering Laboratory for New Products of Traditional Chinese Medicines,Lanzhou 730000,Gansu Province,China)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2024年第17期2518-2522,共5页
The Chinese Journal of Clinical Pharmacology
基金
国家自然科学基金资助项目(82160862,81960828)
兰州市科技计划基金资助项目(2020-ZD-53)
第五批全国中医临床优秀人才研修基金资助项目(国中医药人教函〔2022〕239号)
首批陇原青年英才基金资助项目(〔2022〕25号)。
