人参皂苷Rb_(1)通过PI3K/Akt信号通路对LPS炎症小鼠的神经保护作用

Neuroprotective Effect of Ginsenoside Rb1 on Lipopolysaccharide-induced Neuroinflammation in Mice Based on PI3K/Akt Signaling Pathway

目的:观察人参皂苷Rb1对脂多糖(LPS)炎症小鼠的神经保护作用,初步探讨其作用机制。方法:ICR小鼠70只,随机分为空白组、模型组、地塞米松磷酸钠注射液组、人参皂苷Rb1低、高剂量组,每组14只。采用LPS剂量递增方法制备小鼠大脑神经炎症模型,以1 mg·kg^(-1)为起始剂量,每48 h(隔天早上)腹腔注射1次,每次以2 mg·kg^(-1)的增加量递增剂量,共注射7次,末次LPS剂量为13 mg·kg^(-1)。地塞米松磷酸钠注射液组5 mg·kg^(-1)·d^(-1)腹腔注射,人参皂苷Rb1低、高剂量组分别以10、20 mg·kg^(-1)·d^(-1)腹腔注射,空白组与模型组腹腔注射等体积生理盐水,干预14 d。观察LPS小鼠活动行为,Y迷宫和高架十字迷宫检测小鼠焦虑样行为;酶联免疫吸附测定法(ELISA)检测小鼠大脑单核细胞趋化蛋白-1(MCP-1)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)水平;免疫荧光染色法检测小鼠大脑小胶质细胞神经元损伤、小胶质细胞及星形胶质细胞活化状态;蛋白免疫印迹法(Western blot)检测小鼠大脑磷脂酰肌醇3-激酶(PI3K)、蛋白激酶B(Akt)及核转录因子-κB(NF-κB)蛋白表达。结果:与空白组比较,模型组小鼠Y迷宫和高架十字迷宫焦虑探索样行为明显提高(P<0.05,P<0.01),脑内MCP-1、TNF-α、IL-1β含量显著增加(P<0.01),小鼠大脑体感区皮层及海马CA1区神经元阳性细胞数显著减少(P<0.01),脑内小胶质细胞、星形胶质细胞显著被激活(P<0.01),大脑磷酸化PI3K、Akt及NF-κB蛋白表达显著上升(P<0.01);与模型组比较,人参皂苷Rb1低、高剂量组小鼠Y迷宫和高架十字迷宫焦虑样行为明显减少(P<0.05,P<0.01),脑内MCP-1、TNF-α、IL-1β含量显著降低(P<0.01),小鼠大脑体感区皮层及海马CA1区神经元阳性细胞数显著增多(P<0.01),脑内小胶质细胞、星形胶质细胞激活明显被抑制(P<0.05,P<0.01),大脑磷酸化PI3K、Akt及NF-κB蛋白表达明显下降(P<0.05,P<0.01)。结论:人参皂苷Rb1对LPS炎症小鼠具有神经保护作用,PI3K/Akt信号通路可能参与人参皂苷Rb1对LPS小鼠的神经保护作用。

Objective:To observe the neuroprotective effects of ginsenoside Rb1 on lipopolysaccharide(LPS)-induced neuroinflammation in mice and to preliminarily investigate its mechanism of action.Method:Seventy ICR mice were randomly divided into blank group,model group,dexamethasone sodium phosphate injection group,and low-dose,high-dose ginsenoside Rb1 groups,with 14 mice in each group.A mouse brain neuroinflammation model was prepared using the LPS dose escalation method,starting with a dose of 1 mg·kg^(-1)and administered via intraperitoneal injection every 48 h(every other morning).Each subsequent dose increased by 2 mg·kg^(-1),for a total of 7 injections,culminating in a final dose of 13 mg·kg^(-1).The dexamethasone sodium phosphate injection group received an intraperitoneal injection at 5 mg·kg^(-1)·d^(-1).The low-dose and high-dose ginsenoside Rb1 groups were given intraperitoneal injections at 10 mg·kg^(-1)·d^(-1)and 20 mg·kg^(-1)·d^(-1),respectively,while the blank and model groups received the same volume of normal saline for 14 days.The behavioral activity of LPS mice was observed,anxiety-like behavior was assessed using the Y-maze and elevated plus maze,and brain levels of monocyte chemoattractant protein-1(MCP-1),tumor necrosis factor-α(TNF-α),and interleukin-1β(IL-1β)were measured by enzyme-linked immunosorbent assay(ELISA).Neuronal damage of microglia,and the activation status of microglia and astrocytes in the brain were assessed using immunofluorescence staining.The protein expression of phosphatidylinositol-3-kinase(PI3K),protein kinase B(Akt),and nuclear factor-κB(NF-κB)in mouse brain were detected by Western blot.Result:Compared with the blank group,the model group showed significantly increased anxiety-like behavior in the Y-maze and elevated plus maze(P<0.05,P<0.01),significantly elevated levels of MCP-1,TNF-α,and IL-1βin the brain(P<0.01),a significant decrease in the number of neuronal positive cells in the somatosensory cortex and hippocampus CA1 region(P<0.01),significant activation of microglia and astrocytes(P<0.01),and a significant increase in the expression of phosphorylated PI3K,Akt,and NF-κB proteins(P<0.01).Compared with the model group,the ginsenoside Rb1 low-dose and high-dose groups showed significantly reduced anxietylike behavior in the Y-maze and elevated plus maze(P<0.05,P<0.01),significantly decreased levels of MCP-1,TNF-α,and IL-1βin the brain(P<0.01),a significant increase in the number of neuronal positive cells in the somatosensory cortex and hippocampus CA1 region(P<0.01),significant inhibition of microglia and astrocyte activation(P<0.05,P<0.01),and a significant decrease in the expression of phosphorylated PI3K,Akt,and NF-κB proteins(P<0.05,P<0.01).Conclusion:Ginsenoside Rb1 has neuroprotective effects on LPSinduced inflammation in mice,which may involve the regulation of the PI3K/Akt signaling pathway.