人参皂苷Rb1和Rg1对海马神经元的影响

Effects of ginsenoside Rb1 and Rg1 on hippocampal neurons

目的探讨人参皂苷(ginsenoside)Rb1和Rg1(GRb1,GRg1)对海马神经元突起生长及神经保护作用及其机制。方法培养SD新生大鼠海马神经元,分为:正常组、Rb1组、Rg1组、Rb1+API-2(Akt抑制剂)、Rg1+API-2、Rb1+PD98059(MEK抑制剂)和Rg1+PD98059组,培养1 d,用免疫染色观察神经突起的生长。加入Aβ25-35制备海马神经元损伤模型,分为正常组、损伤组(Aβ3组)、Rb1组、Rg1组、Rb1+API-2、Rg1+API-2、Rb1+PD98059和Rg1+PD98059,培养48 h,用Hoechst33258染色观察活细胞与凋亡细胞。用Elisa观察培养上清液里神经营养因子的浓度(nerve growth factor,NGF;brain-derived neurotrophic factor,BDNF;neurotrophin-3,NT-3)。结果 Rb1和Rg1组神经突起生长高于对照组(P<0.05);API-2和PD98059显著抑制了Rb1和Rg1诱导的神经突起生长(P<0.01),API-2的抑制效果强于PD98059;Rg1+API-2组BDNF高于对照组和Rg1组(P<0.05),其余各组间差异无统计学意义(P>0.05);Rg1组NT-3的浓度高于对照组(P<0.05);Rb1+PD98059组的NGF高于Rb1组。Rb1和Rg1组海马神经元的凋亡率显著低于损伤组(P<0.01);PD98059和API-2阻断了Rb1和Rg1对神经元的保护作用(P<0.01);神经营养因子水平各组间差异无统计学意义。结论 Rb1和Rg1促进海马神经元突起生长及抵抗Aβ25-35损伤,促进神经元的存活。其机制与Akt和ERK1/2的信号通路激活有关,与增加神经营养因子的分泌无关。

Objective To investigate the effects of ginsenoside Rbl and Rg1 （Rbl, Rg1 ） on neurite outgrowth of hippocampal neurons and protection hippocampal neurons against injury caused by A[325 - 35. Methods Cultured hippocampal neurons from newborn SD rats were divided into groups： normal, Rbl, Rgl, Rb1 ＋ API-2 （Akt inhibitor）, Rgl ＋ API-2, Rbl ＋ PD98059 （MEK inhibitor）,and Rgl ＋ PD98059. After 24 h of culture the neurite outgrowth was evaluated by immunostaining of neuronal growth associated protein-43 （ GAP- 43 ）. Hippocampal neurons were treated with Aβ25-35 and divided into groups： injured Aβ group, Rbl, Rgl, Rbl ＋ API-2, Rg1 ＋ API-2, Rb1 ＋ PD98059 and Rgl ＋ PD98059. After 48 h of cultured the Hoechst33258 staining was performed to evaluate the rate of apoptosis. In addition, the nerve growth factor （NGF）, brain-derived neurotrophic factor （BDNF） and neurotrophin-3 （NT-3） were analyzed by ELISA method. Results The neurites were significantly longer and the number increased in Rbl and Rgl groups compared with normal group（ P 〈 0.05 ）. API-2 and PD98059 blocked neurite outgrowth significantly（ P 〈 0.01 ）, and API-2 was more efficient than PD98059. No significant difference of BDNF from supernatant was observed between groups except that BDNF in Rg1 ＋ API-2 group was higher compared with Rgl group; NT-3 in Rgl group was significantly higher than that in normal group; NGF in Rb1 ＋ PD98059 was significantly higher than that in Rbl group （ P 〈 0. 05 ）. Apoptotic rates of hippocampal neurons in Rbl and Rgl groups decreased significantly （P 〈0.01 ） compared with Aβ group. Moreover, API-2 and PD98059 significantly inhibited neuroprotection of Rbl and Rgl against Aβ25-35 injury （P 〈 0.05）. No differences of NGF, NT-3 and BDNF were observed among groups （P 〉 0.05）. Conclusion Ginsenosides Rb1 and Rgl can induce neurite outgrowth of hippocampal neurons and protect neurons against Aβ25 - 35 injury. Their mechanisms are related to Akt and ERK1/2 signal transduction pathways but not to the increase of neurotrophins secretion.