抗帕颗粒对帕金森病模型小鼠酪氨酸羟化酶神经元的影响

目的 探讨抗帕颗粒对1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP)帕金森病(PD)模型小鼠黑质纹状体酪氨酸羟化酶(TH)阳性神经元的影响。方法 90只健康雄性C57BL/6小鼠,鼠龄8～12周,并随机分为3组,即正常对照组30只、PD模型对照组30只、PD模型干预组30只; MPTP腹腔注射(40 mg·kg^-1·d^-1×7 d)制备小鼠PD模型; 正常对照组及PD模型对照组予生理盐水1 mL·d^-1灌胃,PD模型干预组给予抗帕颗粒40 mg·kg^-1·d^-1灌胃,连续喂养4个月; 黑质纹状体切片、HE染色、免疫组织化学染色TH神经元及Western blotting检测TH蛋白的表达量。结果 ①正常对照组30只(30/30只)最终均存活,PD模型对照组4个月存活27只(27/30只),PD模型干预组4个月存活28只(28/30只); ②PD模型对照组、PD模型干预组小鼠每次注射MPTP后先有短暂兴奋持续(7.61±2.17)min],表现为四处窜跳,随即出现全身中重度震颤,皮毛及尾巴时有竖立,活动减少,持续(24.23±3.89)min后震颤消失,随后出现活动减少; ③HE染色显示正常对照组大量褐色TH阳性细胞,PD模型对照组TH阳性细胞数明显减少,PD模型干预组TH阳性细胞数有所增加; ④免疫组织化学染色后经Imagepro-Plus 5.1系统分析,正常对照组TH阳性细胞面积为64 145 μm²,高倍镜下可见大量胞质为褐色颗粒的TH阳性细胞; PD模型对照组TH阳性细胞染色面积为40 012 μm²,高倍镜下见TH细胞数明显减少; PD模型干预组TH阳性细胞染色面积为60 952 μm²,高倍镜下见TH阳性细胞数较PD模型对照组增加; ⑤Western blotting检测显示正常对照组TH蛋白表达量与PD模型对照组和PD模型干预组比较均有明显差异(P<0.001),PD模型对照组TH蛋白表达量与PD模型干预组比较也有明显差异(P<0.05)。结论 抗帕颗粒可使PD小鼠黑质纹状体中多巴胺能神经元一定程度地减少丢失,对多巴胺能神经元的数量、形态及功能具有一定的保护作用。

Effect of anti-Parkinson granule on TH positive-neurons in model mice with Parkinson disease

ObjectiveTo explore the pharmaceutical effect of anti-Parkinson granule on substantia nigra and striatum Tyrosine hydroxylase(TH)-positive neurons in model mice with Parkinson disease(PD)induced by 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP).Methods 90 healthy male C57BL/ 6 mice,aged from 8 to 12 weeks,were randomly divided into 3 groups(n=30): normal control group,PD model control group and PD model intervention group.The PD model group was made through intraperitoneal injection with MPTP(40 mg·kg^-1·d^-1×7 d).The normal control group and the PD model control group were treated with normal saline(1 mL·d^-1)intragastrically.The PD model intervention group was treated with anti-Parkinson granule solution(40 mg·kg^-1·d^-1)intragastrically for 4 months.TH protein expression was detected by HE staining,and immunohistochemistry staining of Western blotting in substantia nigra and striatum slices.At the end of the forth month,TH-positive neurons in substantia nigra and striatum among groups was contrasted and analyzed.Results ①30 mice(30/30 mice)were living in the normal control group,27 mice(27/30 mice)in the PD model control group,and 28 mice(28/30 mice)in the PD model intervention group; ②After injection with MPTP,mice in the PD model control group and the PD model intervention group had short excitementscontinued(7.61±2.17)min] with jumping around,followed with the whole body's moderate or severe tremors,fur and tails erected timely and less movement.Tremors ceased after(24.23±3.89)min,and then the movement became less; ③HE staining showed a large number of brown TH positive cells in the control group,the number of PD model control group was significantly reduced,and the number of TH positive cells in the PD model group increased significantly; ④After analysis by Imagepro-Plus 5.1 system,TH positive cells in the normal control group ranged 64 145 μm² ,and the mass amount of TH positive cells with brown cytoplasmic granular could be observed under high power microscope.TH positive cells in the PD model control group ranged 40 012 μm²,and the number of TH positive cells decreased significantly.TH positive cells in the PD model intervention group ranged 60 952 μm²,and the number of TH positive cells under high power microscope was significantly increased than that in the PD model control group; ⑤The results of TH protein expression detected by Western Blotting were as follows: the comparison between the normal control group and the PD model group was P<0.001,between the normal control group and the PD model intervention group was P<0.001,and between the PD model control group and the PD model intervention group was P<0.05.Conclusion Anti-Parkinson granule can weaken the lost of TH-positive neurons in PD mice substantia nigra and striatum to some degree.Anti-Parkinson granule can partly protect dopaminergic neurons in aspects of quantity,morphology and function,and may have a hope to improve PD treatment status and a wide application in clinic.