黑质LAMP2A过表达改善帕金森病恒河猴模型的运动功能

Overexpression of LAMP2A in substantia nigra improves motor function in rhesus monkey models of Parkinson’s disease

目的在帕金森病(Parkinson’s disease,PD)恒河猴模型上观察黑质过表达溶酶体相关膜蛋白2A(lysosome associated membrane protein 2A,LAMP2A)上调分子伴侣介导的自噬(chaperone-mediated autophagy,CMA)对实验猴运动行为的影响。方法应用立体定位注射仪向7只恒河猴的右脑纹状体区注射PD患者死亡后脑组织(自愿捐献)的路易小体(Lewy bodies,LBs)提取物,同时在左脑纹状体区注射非PD病人死亡患者的脑组织(自愿捐献)提取物来构建PD恒河猴模型。模型动物随机分为两组,LAMP2A过表达组(4只)在其黑质区双侧注射过表达LAMP2A的腺相关病毒载体;对照组(3只)在黑质区双侧注射对照病毒载体。通过阶梯实验、平板取食实验评价PD恒河猴模型运动能力和LAMP2A对该PD模型运动功能的影响。利用ELISA法对实验猴脑脊液、血清的总α-突触核蛋白(α-synuclein,α-syn)及磷酸化α-syn水平进行测定。结果阶梯实验和平板取食实验中对照组恒河猴的左侧上肢的精细动作水平相较于其右侧出现下降;LAMP2A过表达组恒河猴的左右两侧上肢的精细运动水平没有出现明显差异;ELISA测定对照组术后12月脑脊液、血清的总α-syn水平及磷酸化α-syn水平显著增高;LAMP2A过表达组脑脊液磷酸化α-syn水平和血清磷酸化α-syn水平在8月之后均低于同期的对照组。结论过表达LAMP2A可能通过增加α-syn的CMA清除改善PD恒河猴模型精细运动能力。

Objective To observe the effect of overexpression of lysosome associated membrane protein 2A(LAMP2A)up-regulating chaperone-mediated autophagy(CMA)in the substantia nigra on the locomotor behavior of a Parkinson’s disease(PD)rhesus monkey model.Methods We established a rhesus monkey model of PD.A stereotactic injection device was used to inject Lewy bodies(LBs)extracts from the brain tissues donated by PD patients into the right striatum of seven rhesus monkeys.At the same time,extracts of brain tissue donated by control(non-PD)patients were injected into the striatum of the left brain.The animals were randomly divided into two groups.The LAMP2A overexpression group(n=4)was bilaterally injected with an adeno-associated virus vector overexpressing LAMP2A;the control group(n=3)was injected with a control virus vector in both sides of the substantia nigra.The motor function of monkeys was evaluated by the hill and valley staircase experiment and the pick-up test.The levels of totalα-synuclein(α-syn)and phosphorylatedα-syn in the cerebrospinal fluid and serum of experimental monkeys were determined by ELISA.Results In the valley staircase task and the outside board pick up test,the fine motor level of the left upper limb of the control group of rhesus monkeys was lower than that of the right side.There was no significant difference in the fine motor level of the upper limbs on the left and right sides of the rhesus monkeys in the LAMP2A overexpression group.Cerebrospinal fluid and serum totalα-syn levels and phosphorylatedα-syn levels of the control group increased significantly at 12 months after operation.The levels of phosphorylatedα-syn in cerebrospinal fluid and serum phosphorylatedα-syn in the LAMP2A overexpression group were lower than those in the control group at the same time after 8 months.Conclusions Overexpression of LAMP2A improved the fine motor ability of experimental monkeys in the rhesus monkey model of PD through up-regulating CMA-mediatedα-syn elimination.