苁蓉精对MPP^+诱导多巴胺能神经细胞损伤后GSK-3β表达的影响

Effects of Congrongjing on the expression of GSK-3β protein after the injury of dopaminergic neurons induced by MPP^+

目的观察苁蓉精水提物及苁蓉精纳米微粉对1-甲基-4-苯基吡啶离子(MPP+)诱导损伤的多巴胺能神经细胞MES23.5细胞中帕金森病相关蛋白α-突触核蛋白(α-synuclein)、糖原合成酶激酶-3β(GSK-3β)、Tau蛋白表达的影响,探讨苁蓉精治疗帕金森病的作用机制。方法分别将苁蓉精水提物及纳米微粉经PBS溶解后加入培养基使终浓度为100、200、250μg/mL,孵育MPP+诱导的PD模型细胞24h后,并设模型对照(MPP+诱导的PD模型细胞,不加干预)及空白对照组(未经任何处理的MES23.5细胞),以MTT法检测细胞的存活率,Western Blot法测定各组细胞内α-synuclein、GSK-3β、Tau蛋白表达。结果同空白对照组比较,模型组及苁蓉精水提物及纳米微粉100、200、250μg/mL组α-synuclein、GSK-3β及Tau蛋白的磷酸化水平均升高(分别P<0.01、P<0.05);与模型组比较,苁蓉精水提物及纳米微粉100、200、250μg/mL组α-synuclein、GSK-3β及Tau蛋白的磷酸化水平均显著降低(分别P<0.01、P<0.05)。除苁蓉精纳米微粉100μg/mL组GSK-3β磷酸化水平与苁蓉精水提物100μg/mL组比较差异无统计学意义(P>0.05)外,其余各苁蓉精纳米微粉组α-synuclein、GSK-3β及Tau蛋白的磷酸化水平均低于相同浓度的苁蓉精水提物组(分别P<0.01、P<0.05)。结论苁蓉精水提物及苁蓉精纳米微粉均能够减轻MPP+诱导的PD模型细胞的损伤,且苁蓉精纳米微粉的效果相对较好。其机制可能与苁蓉精可以减少α-synuclein的聚集,降低GSK-3β的活性,抑制Tau蛋白的过度磷酸化有关。

Objective To observe the effect of Congrongjing aqueous extract and Congrongjingnanopowders on the expression of Parkinson＇s disease（PD）associated proteins,α-synuclein,GSK-3βand Tau in injured dopaminergic neurons induced by 1-methyl-4-phenyl-pyridinium（MPP＋）and investigate the mechanism of Congrongjing in the treatment of PD.Methods Congrongjing aqueous extract and Congrongjing nano-powders dissolved by PBS medium were added into the culture medium and different concentrations of solution（100,200,250μg/mL）were prepared.Then the PD model cells induced by MPP＋were cultured by the above culture medium for 24 h,model control group（PD model cells were induced by MPP＋,without intervention）and blank control group（MES23.5cells without any treatment）were set up,MTT method was used to detect the cell survival rate,changes in expression of a-synuclein,GSK-3βand Tau protein in cells were determined by Western blot.Results Compared with the blank control group,the expression levels ofα-synuclein protein,phosphorylation level of GSK-3βprotein and Tau protein increased in the model control group,Congrongjing aqueous extract and Congrongjing nano-powders 100,200,250μg/mL groups（P〈0.01,P〈0.05,respectively）.Compared with the model control group,the expression levels of α-synuclein protein,phosphorylation level of GSK-3βprotein and Tau protein decreased in the Congrongjing aqueous extract and Congrongjing nano-powders 100,200,250μg/mL groups（P〈0.01,P〈0.05,respectively）.Except for the phosphorylation level of GSK-3βprotein between the Congrongjing aqueous extract 100μg/mL groupand the Congrongjin gnano-powders 100 μg/mL group（P〉0.05）,the expression levels ofα-synuclein protein,phosphorylation level of GSK-3βprotein and Tau protein of Congrongjing nano-powders groups were significantly lower than those of the Congrongjing aqueous extract group（P〈0.01,P〈0.05,respectively）.Conclusions Congrongjing aqueous extract and Congrongjing nano-powders can reduce the injury of MES23.5cells induced by MPP＋and the effect of Congrongjing nano-powders is better.The mechanism may be related to reducing the accumulation of theα-synuclein and theactivity of GSK-3β,inhibiting the excessive phosphorylation of Tau protein.