腺病毒介导的胶质细胞源性神经营养因子基因脑内转移对帕金森病大鼠模型的保护作用

Protective effects of the intracerebral transfer of the adenoviral-mediated GDNF gene in a rat model of Parkinson's disease

目的 探讨腺病毒介导的胶质细胞源性神经营养因子 (GDNF)基因直接转移对帕金森病 (PD)的保护作用。 方法 实验SD大鼠分重组GDNF腺病毒 (Ad GDNF)实验组、LacZ腺病毒 (Ad LacZ)对照组和磷酸缓冲液 (PBS)对照组。将重组腺病毒及PBS定向注射至一侧黑质附近 ,1周后于同侧纹状体注射 6 羟基多巴胺 (6 OHDA)诱发多巴胺 (DA)能神经元进行性变性。通过旋转行为观察和中脑酪氨酸羟化酶 (TH)免疫组化染色及纹状体单胺类递质高压液相色谱 电化学仪 (HPLC ECD)检测评估其治疗效应 ;通过RT PCR、ELISA检测Ad GDNF在脑内的表达情况。 结果 Ad GDNF组阿扑吗啡诱发的旋转次数明显低于 2个对照组 ;Ad GDNF组约 70 %的黑质DA能神经元得以保存 ,而Ad LacZ及PBS对照组仅有 30 %左右 ;Ad GDNF组纹状体DA含量也显著高于Ad LacZ及PBS对照组 ;Ad GDNF在脑内可有效表达 ,注射后 5周黑质附近GDNF含量达 1ng/ 10mg脑组织湿重 ,是对照组的16～ 2 0倍。 结论 腺病毒介导的GDNF基因脑内直接转移可阻止 6 OHDA诱发的大鼠DA能神经元进行性变性 ,提示这一手段在PD保护性治疗方面具有一定的应用价值。

Objective To study the neuroprotective effects of adeno viral mediated glial cell line derived neurotrophic factor(GDNF) gene transfer in the treatment of Parkinson's disease. Methods Thirty five SD rats were divided into 3 groups which received perinigral injections of recombinant adenovirus encoding GDNF (Ad GDNF)/ LacZ(Ad LacZ) and PBS, respectively. One week later, intrastriatal injection of 6 hydroxydopamine (6 OHDA) was made to induce progressive degeneration of dopaminergic neurons. The neuroprotective effects of Ad GDNF were evaluated by apomorphine induced rotational behavior, immunohistochemical assay of the tyrosine hydroxylase(TH) positive neurons in the midbrain and measurement of monoamine level in the striatum. RT PCR and ELISA were performed to check the expression of the exogenous GDNF gene in the brain. Results Ad GDNF treated rats showed improved motor functions, better survival of TH positive cells in the lesioned substantia nigra (70% vs 30%) and higher DA levels in the lesioned striatum. The exogenous GDNF gene was efficiently expressed in the midbrain. GDNF protein level in the injection site reached 1 ng/10 mg wet tissue 5 weeks after the adenoviral vector delivery, being 16 20 times of that of the Ad LacZ delivery or PBS treated groups. Conclusions Adeno viral mediated GDNF gene intracerebral transfer significantly protected the dopaminergic neurons of nigrostriatal system from 6 OHDA induced injury and is valuable in the treatment of Parkinson's disease.