麦冬多糖对急性脑梗死大鼠认知功能及Th17/Treg细胞平衡的影响

Effects of Ophiopogon Polysaccharide on Cognitive Function and Th17/Treg Cell Balance in Rats with Acute Cerebral Infarction

SD大鼠通过中动脉闭塞/再灌注手术建立ACI模型,随机分为模型组、麦冬多糖低剂量(150 mg/kg)组、麦冬多糖高剂量(300 mg/kg)组、麦冬多糖高剂量(300 mg/kg)+MHY1485(m TOR激活剂,10 mg/kg)组,每组18只,另取18只大鼠不插入线栓组赛血流的情况下进行相同手术操作,作为假手术组.以麦冬多糖和MHY1485分组干预后通过Morris水迷宫、新物体识别实验评测大鼠认知功能;TTC、TUNEL染色分别评测大鼠脑梗死与海马神经元凋亡情况;通过流式细胞实验检测各组大鼠外周血Th17/Treg细胞平衡;用ELASA试剂盒测量大鼠血清Treg、Th17细胞因子水平;通过免疫印迹实验检测各组大鼠脑组织m TOR/P70S6K/4EBP1通路蛋白表达.结果显示与假手术组相比,模型组大鼠逃避潜伏期、脑梗死面积占比、海马神经元凋亡指数、Th17比例、Th17/Treg、TNF-α与IL-17A水平、p-m TOR/m TOR、p-P70S6K/P70S6K、p-4EBP1/4EBP1升高(P<0.05),穿台次数、识别指数、Treg比例、IL-4与IL-10水平降低(P<0.05);麦冬多糖可逆转ACI大鼠以上病理指标的变化,且高剂量麦冬多糖作用更强(P<0.05);与麦冬多糖高剂量组相比,麦冬多糖高剂量+MHY1485组大鼠逃避潜伏期、脑梗死面积占比、海马神经元凋亡指数、Th17比例、Th17/Treg、TNF-α与IL-17A水平、p-m TOR/m TOR、p-P70S6K/P70S6K、p-4EBP1/4EBP1升高(P<0.05),穿台次数、识别指数、Treg比例、IL-4与IL-10水平降低(P<0.05).表明麦冬多糖可通过阻断m TOR/P70S6K/4EBP1信号通路激活传导而调节Th17/Treg细胞平衡,促使其偏移向Treg,从而抑制ACI大鼠炎症反应,减轻脑组织梗死及海马神经元凋亡,最终改善ACI大鼠认知功能.

This study aims to investigate the effects of ophiopogon polysaccharide on cognitive function and Th17/Treg cell balance in acute cerebral infarction(ACI)rats by regulating the mammalian rapamycin target protein(mTOR)/p70 ribosomal protein S6 kinase(P70S6K)/4E binding protein 1(4EBP1)pathway.SD rats were selected to establish an ACI model through middle artery occlusion/reperfusion surgery.They were randomly separated into a model group,a low-dose(150 mg/kg)ophiopogon polysaccharide group,a high-dose(300 mg/kg)Ophiopogon polysaccharide group,and a high-dose(300 mg/kg)ophiopogon polysaccharide+MHY1485 group(mTOR activator,10 mg/kg),with 18 rats in each group.An additional 18 rats were selected as the sham surgery group without inserting thread plugs to compete for blood flow.After intervention with Ophiopogon polysaccharide and MHY1485,cognitive function of rats was evaluated through Morris water maze and new object recognition experiments.TTC and TUNEL staining were applied to evaluate the apoptosis of hippocampal neurons in rats with cerebral infarction,respectively.Flow cytometry was applied to detect the balance of Th17/Treg cells in peripheral blood of rats in each group.ELASA assay kit was applied to measure serum Treg and Th17 cytokine levels in rats.Immunoblotting experiment was applied to detect the expression of mTOR/P70S6K/4EBP1 pathway proteins in the brain tissues of rats in each group.The results show that compared with the sham surgery group,the escape latency,proportion of cerebral infarction area,hippocampal neuronal apoptosis index,Th17 ratio,Th17/Treg,TNF-αand IL-17A levels,p-mTOR/mTOR,p-P70S6K/P70S6K,and p-4EBP1/4EBP1 in the model group increased(P<0.05),the times of plate crossings,recognition index,Treg ratio,and IL-4 and IL-10 levels decreased(P<0.05).ophiopogon polysaccharides was able to reverse the changes in the above pathological indicators in ACI rats,and high-dose ophiopogon polysaccharides had a stronger effect(P<0.05).Compared with the high-dose Ophiopogon polysaccharide group,the escape latency,proportion of cerebral infarction area,hippocampal neuronal apoptosis index,Th17 ratio,Th17/Treg,TNF-αand IL-17A levels,p-mTOR/mTOR,p-P70S6K/P70S6K,and p-4EBP1/4EBP1 in the high-dose ophiopogon polysaccharide+MHY1485 group increased(P<0.05),the times of plate crossings,recognition index,Treg ratio,and IL-4 and IL-10 levels decreased(P<0.05).The results indicate that ophiopogon polysaccharide can regulate the balance of Th17/Treg cells by blocking the mTOR/P70S6K/4EBP1 signaling pathway,promoting their migration towards Treg,thereby inhibiting the inflammatory response in ACI rats,reducing brain tissue infarction and hippocampal neuronal apoptosis,and ultimately improving cognitive function in ACI rats.