补肾通腑方对脂多糖诱导的认知障碍模型大鼠学习记忆能力的影响及作用机制研究

doi:10.16431/j.cnki.1671-7236.2024.05.032

摘要/Abstract

摘要： 【目的】探讨补肾通腑方对脂多糖(lipopolysaccharide,LPS)诱导的认知障碍模型大鼠学习记忆能力的改善作用及抗炎保护机制。【方法】选取6周龄健康SPF级雄性SD大鼠,采用LPS侧脑室注射法制备认知障碍大鼠模型。选取造模成功的大鼠,随机分为模型组,甘露特钠胶囊组,补肾通腑方高、中、低剂量组,并设假手术组。分别给予对应的药物灌胃,1 次/d,连续8周,其中补肾通腑方高、中、低剂量组的灌胃剂量分别为15.12、7.56、3.78 g/kg,甘露特钠胶囊组灌胃剂量为0.094 g/kg。模型组和假手术组给予等体积的纯水灌胃。于第8周末,采用Morris水迷宫法观察各组大鼠空间学习记忆能力的变化,采用苏木素伊红(HE)染色观察大鼠海马区病理形态学的变化,采用免疫荧光检测大鼠海马区小胶质细胞活化情况,采用Western blotting方法检测大鼠海马区肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、IL-1β、N-甲基-D-天门冬胺酸受体(N-methyl-D-aspartate receptor,NMDA)、γ-氨基丁酸(γ-aminobutyric acid,GABA)、乙酰胆碱(acetylcholine,Ach)等的表达。【结果】与假手术组比较,模型组大鼠空间逃避潜伏期、游泳路程明显延长,撤除平台后跨越原平台次数显著减少(P＜0.05);HE染色结果显示,细胞排列紊乱、稀疏,有细胞空洞等损伤表现;免疫荧光结果显示,诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)/离子钙结合适配器分子1(ionized calcium binding adapter molecule 1,Iba1)共表达升高、精氨酸酶1(arginase-1,Arg1)/Iba1共表达降低(P＜0.05);海马区TNF-α、IL-6、IL-1β等的表达显著升高,NMDA、GABA、Ach等的表达显著降低(P＜0.05)。与模型组比较,补肾通腑方高、中、低剂量组甘露特钠胶囊组逃避潜伏期、游泳路程显著缩短,撤除平台后跨越原平台次数显著增多(P＜0.05),HE染色结果显示细胞排列较紧密、整齐,细胞形态趋于正常,iNOS/Iba1共表达显著降低,仅补肾通腑方高剂量组Arg1/Iba1共表达显著升高(P＜0.05);海马区TNF-α、IL-6、IL-1β等的表达显著降低,NMDA、GABA、Ach等的表达显著升高(P＜0.05)。【结论】补肾通腑方能够改善LPS诱导的认知障碍模型大鼠的学习记忆能力,其作用机制可能与抑制促炎型小胶质细胞的激活,降低海马区TNF-α、IL-6、IL-1β等炎症因子水平,提高海马区NMDA、GABA、Ach等记忆相关因子水平有关。

关键词: 认知障碍; 补肾通腑方; 中枢神经炎症; 脂多糖

Abstract: 【Objective】 This study was aimed to investigate the improvement effect and anti-inflammatory protective mechanism of Bushen Tongfu decoction(BTD) on the learning and memory abilities of LPS-induced cognitive impairment model rats.【Method】 SPF grade male SD rats of 6-week-old were selected.The rat models of cognitive impairment were established by LPS lateral ventricle injection method.All the successful modeled rats were divided into model group,sodium oligomannate capsules (SOC) group,BTD high-dose,middle-dose and low-dose groups randomly.At the same time,the sham operation control group was set.Drugs were administered to rats by gastric perfusion once a day for 8 weeks.BTD high-dose,middle-dose and low-dose groups were give the drug at 15.12,7.56,3.78 g/kg.SOC group was given the drug at 0.094 g/kg.The model and sham operation control groups were administered with equal volumes of pure water.On the 8th weekend,all the rats were detected the spatial learning and memory ability by Morris water maze.The pathological changes in the hippocampus of rats were detected by hematoxylin eosin (HE) staining.The activation of microglia in the hippocampus was detected by immunofluorescence.The expression levels of tumor necrosis factor-α (TNF-α),interleukin-6 (IL-6),IL-1β,N-methyl-D-aspartate receptor(NMDA),γ-aminobutyric acid (GABA) and acetylcholine (Ach) in hippocampus neurons were detected by Western blotting analysis.【Result】 Compared with sham operation group,the escape latency and swimming distance of rats in model group were significantly prolonged,the number of across the platform after remove platform were decreased significantly (P＜0.05).The cells were arranged irregularly,sparse,and there were cell vacuoles by the HE staining,the expression of inducible nitric oxide synthase (iNOS)/ ionized calcium binding adapter molecule 1 (Iba1) were increased and the expression of arginase-1(Arg1)/Iba1 were decreased significantly (P＜0.05),the expression levels of TNF-α,IL-6 and IL-1β were increased significantly and the expression levels of NMDA,GABA and Ach were decreased significantly in hippocampus neurons (P＜0.05).Compared with model group,the escape latency and swimming distance in BDT high-dose,middle-dose,low-dose groups and SOC group were significantly shortened,and the number of times of crossing the original platform after removing the platform was increased significantly (P＜0.05),the cells were arranged tightly and neatly,and the cell morphology tended to be normal by the HE staining,the expression of iNOS/ Iba1 were decreased significantly (P＜0.05),the expression of Arg1/Iba1 were decreased significantly only in the BTD high-dose (P＜0.05),the expression levels of TNF-α,IL-6,IL-1β were decreased significantly and the expression levels of NMDA,GABA,Ach were increased significantly (P＜0.05).【Conclusion】 BTD could improve the learning and memory ability of rats with LPS-induced cognitive impairment model.The possible mechanism was inhibiting the activation of proinflammatory microglia and decreasing the levels of TNF-α,IL-6,IL-1β and increasing the levels of NMDA,GABA,and Ach in the hippocampus.

Key words: cognitive impairment; Bushen Tongfu decoction; neuroinflammation; LPS

中图分类号:

R285.5

张杰, 李思敏, 宋晓雨, 王旭, 许玉珉, 段建平, 马云枝, 赵敏. 补肾通腑方对脂多糖诱导的认知障碍模型大鼠学习记忆能力的影响及作用机制研究[J]. 中国畜牧兽医, 2024, 51(5): 2110-2121.

ZHANG Jie, LI Simin, SONG Xiaoyu, WANG Xu, XU Yumin, DUAN Jianping, MA Yunzhi, ZHAO Min. Study on the Effect and Mechanism of Bushen Tongfu Decoction on the Learning and Memory Ability of LPS-induced Cognitive Impairment Model Rats[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(5): 2110-2121.

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