延龄草总皂苷对缺血卒中后认知障碍大鼠Tau蛋白磷酸化及p38MAPK/NF-κB通路的影响

Effects of total saponins from Trillium tschonoskii maxin on Tau phosphorylation and p38MAPK/NF-κB pathway in rats with cognitive impairment after ischemic stroke

目的基于Tau蛋白磷酸化及p38丝裂原活化蛋白激酶(p38MAPK)/核因子-κB(NF-κB)通路探讨延龄草总皂苷对缺血卒中后认知障碍大鼠的神经保护作用。方法将50只雄性SD大鼠随机分为假手术组、模型组和延龄草总皂苷低、中、高剂量组,每组10只。除假手术组外,其余组大鼠均采用大脑中动脉闭塞(MCAO)法建立缺血卒中后认知障碍模型。术后延龄草总皂苷低、中、高剂量分别给予延龄草总皂苷50 mg/kg、100 mg/kg、200 mg/kg灌胃(灌胃量5 mL/kg),假手术组、模型组给予等量双蒸水灌胃,均1次/d,连续灌胃8周。末次灌胃结束后,采用Zausinger六分法对大鼠进行神经功能缺损程度评分,Morris水迷宫试验评估大鼠的认知功能情况,尼氏染色观察海马神经元损伤情况,RT-qPCR技术检测海马组织中桥接整合因子1(Bin1)mRNA表达情况,Western blot法检测海马组织中核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、Tau、磷酸化Tau(p-Tau)、p38MAPK、NF-κB蛋白表达情况。结果与假手术组比较,模型组大鼠出现明显神经功能缺损和认知功能障碍,海马皮质区及CA1区神经元数量较少,尼氏小体数量明显减少(P<0.05),海马组织中Bin1 mRNA相对表达量和NLRP3、Tau、p-Tau、p38MAPK、NF-κB蛋白相对表达量均明显升高(P均<0.05);与模型组比较,延龄草总皂苷各组大鼠神经功能缺损评分均明显降低(P均<0.05),逃避潜伏期均明显缩短(P均<0.05),穿越平台次数均明显增加(P均<0.05),海马CA1区尼氏小体数量明显增多(P均<0.05),海马组织中Bin1 mRNA相对表达量和NLRP3、Tau、p-Tau、p38MAPK、NF-κB蛋白相对表达量均明显降低(P均<0.05)。结论延龄草总皂苷对缺血卒中后认知障碍大鼠具有神经保护作用,可改善大鼠的认知功能,机制可能与其抑制p38 MAPK/NF-κB信号通路,下调NLRP3、Tau、p-Tau、p38MAPK、NF-κB蛋白表达有关。

Objective It is to investigate the neuroprotective effects of total saponins from Trillium tschonoskii maxin(TST)in rats with cognitive impairment after ischemic stroke based on Tau phosphorylation and p38 mitogen activated protein kinase(p38MAPK)/nuclear factor kappa-B(NF-κB)pathway.Methods Fifty male SD rats were randomly divided into sham operation group,model group and low dose,medium dose and high dose groups of TST,with 10 rats in each group.The rats of all groups except for the sham operation group were used to establish models of cognitive impairment after ischemic stroke by middle cerebral artery occlusion(MCAO)method.After surgery,the rats in the low dose,medium dose and high dose groups of TST were gavaged with 50 mg/kg,100 mg/kg,and 200 mg/kg of TST,respectively(the volume of gavage was 5 mL/kg),and the sham operation group and model group were given the same amount of double-distilled water by gavage,all once daily,continuously treated for 8 weeks.At the end of the final gavage,the rats were scored for the degree of neurological deficit by Zausinger six-point scale,the cognitive function of the rats were evaluated by Morris water maze test,the damage of hippocampal neurons was observed by Nichols staining,the expression of recombinant bridging integrator 1(Bin1)mRNA in hippocampal tissues was detected by RT-qPCR,and the protein expressions of nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),Tau,phosphorylated Tau(p-Tau),p38MAPK,and NF-κB in hippocampal tissue were detected by Western blot method.Results Compared with the sham operation group,the rats in the model group had obvious neurological deficits and cognitive dysfunction,with less neurons and much less Nystagmus vesicles in hippocampal cortical area and CA1 area(all P<0.05),and the relative expression of Bin1 mRNA and the relative expressions of NLRP3,Tau,p-Tau,p38MAPK,and NF-κB proteins in hippocampal tissues were all significantly increased(all P<0.05);compared with the model group,the neurological deficit scores of rats in all groups of TST were significantly reduced(all P<0.05),their escape latencies were significantly shortened(all P<0.05),the numbers of crossing the plateau were significantly increased(all P<0.05),and the number of Nycthemeral vesicles in hippocampal CA1 area was significantly increased(all P<0.05),and the relative expression of Bin1 mRNA and relative expressions of NLRP3,Tau,p-Tau,p38MAPK,and NF-κB proteins in hippocampal tissues were significantly decreased(all P<0.05).Conclusion The saponins from Trillium tschonoskii maxin have neuroprotective effects on rats with cognitive impairment after ischemic stroke,and can improve their cognitive function,the mechanism may be related to inhibiting the p38 MAPK/NF-κB signaling pathway and down-regulating the protein expressions of NLRP3,Tau,p-Tau,p38MAPK,and NF-κB.