携带A53T突变人α突触核蛋白转基因帕金森病大鼠模型的建立

目的大鼠在神经系统疾病模型方面比小鼠有更好的表现，建立人α-synA53T突变型转基因大鼠模型，为研究帕金森病发病提供更优良的疾病模型。方法构建人α—synA53T表达载体，显微注射法制备转基因大鼠。PCR法鉴定转基因首建鼠及其子代基因型。Western blot检测转基因大鼠脑组织中人α-syn蛋白的表达。免疫组化和免疫荧光观察中脑黑质酪氨酸羟化酶阳性神经元数目变化，观察人α-syn蛋白在转基因大鼠黑质神经元中的表达和分布。Rotatingrod实验和步态分析系统评价转基因大鼠的行为学改变。结果获得1个α-synA53T高表达且可以稳定传代的转基因大鼠品系。转基因大鼠中脑黑质酪氨酸羟化酶（TH）阳性神经元数目减少69％（P〈0．05），残存神经元胞浆中有大量的α—syn蛋白聚集。转基因大鼠在滚轴上保持平衡的时间显著减少40％～60％（P〈0．05），并表现出步态障碍如步宽加大、摆动期缩短和足迹最大接触面积减小等。结论建立了人α-synA53T突变型转基因的帕金森病大鼠模型。

Establishment of a transgenic rat model of Parkinson＇s disease with α-synuclein A53T mutation

Objective In view of better manifestation of rats than mice in nervous diseases studies, the aim of this study was to generate a transgenie rat model of Parkinson＇s disease （PD） expressing alpha-synuelein gene with A53T mutation and provide a better animal model for PD researches. Methods The transgenic plasmid was constructed by inserting the alpha-synuclein gene with A53T mutation into the downstream of human PDGF promoter. The transgenic rats were produced by microinjection and the genotype was detected by PCR. The protein expression levels in the brain tissues were determined by Western blotting. The expression of human alpha-synuclein in the brain tissues from transgenic rats was analyzed by immunohistochemical staining （IHC） and immunofluorescence （IF）. The changes of the number of tyrosine hydroxylase （TH）-positive neurons in the substantia nigra were observed by IHC and IF. The motor performances of transgenic rats were measured by rotating rod test and gait analysis. Results One transgenic rat line with high expression of human alpha-synuclein was established. Histopathological examination revealed the abnormal aggregation of dopamine （DA） neurons in the midbrain and significantly reduced number of TH-positive neurons （69% ,P 〈 0. 05 ） in the substantia nigra of transgenic rats. Rotating rod test and rat gait analysis indicated declined motor performances（40% -60% , P 〈 0.05）） and abnormal gait in the transgenic rats. Conclusions A human α-synuclein A53T transgenic rat model of Parkinson＇s disease has been established.