Lactacystin对酪氨酸羟化酶基因和蛋白表达的影响

Effects of Lactacystin on the Expressions of Gene and Protein of Tyrosine Hydroxylase

目的:探讨lactacystin对酪氨酸羟化酶(TH)基因及蛋白表达的影响。方法:在大鼠单侧黑质致密部(SNc)立体定向注射蛋白酶体抑制剂lactacystin 0 .2、2、8μg抑制α- synuclein的降解,对照组注射等量生理盐水,观察大鼠自主行为和阿扑吗啡诱导的旋转行为变化;应用免疫组化观察黑质细胞-αsynuclein和TH蛋白表达;采用RT- PCR方法测定黑质细胞THmRNA的表达。结果:注射lactacystin 0 .2 μg组未见异常;2和8μg组大鼠出现进行性的运动迟缓、少动、头向对侧倾斜、震颤,但后者更加明显并出现阿扑吗啡诱导的向对侧旋转运动;给药后3周注射lactacystin 2和8μg组大鼠黑质细胞α-synuclein表达分别较对照组高出5 6.71% (P <0 .0 1)和12 2 .16% (P <0 .0 0 1) ;部分细胞胞质内出现αsynuclein免疫反应呈强阳性的Lewy小体;THmRNA和蛋白的表达分别较对照组减少了63 .3 4% (P <0 .0 0 1)、60 .5 5 % (P <0 .0 5 )和81.95 % (P <0 .0 0 1)、89.5 3 % (P <0 .0 1)。结论:lactacystin对中脑黑质细胞TH基因及蛋白表达均有显著的抑制作用,参与了lactacystin所致的帕金森病发病机制。

Aim:To study the effects of lactacystin on the expressions of gene and protein of tyrosine hydroxylase. Methods: By stereotaxic unilateral injection of different doses of lactacystin, a selective proteasome inhibitor, into the substantia nigral pars compacta, the spontaneous and apomorphine-induced contralateral behaviors of rats were observed; the expressions of α-synuclein and tyrosine hydroxylase were investigated by immunohistochemistry; the levels of tyrosine hydroxylase mRNA were determined by RT-PCR.Results: There was no difference between 0.2 μg group and control treated with sodium saline. Animals treated with 2 μg and 8 μg lactacystin developed progressively bradykinetic and displayed contralateral head tilting and tremor; apomorphine-induced contralateral behavior was notably observed only in rats of 8 μg group; 3 weeks later, the expressions of α-synuclein in 2 μg and 8 μg groups were increased to 56.71%( P <0.01)and 122.16%( P <0.001)of the control respectively; some of nigral cells contained acidophilic Lewy body with intense immune response to α-synuclein; the levels of protein and mRNA of tyrosine hydroxylase in 2 μg and 8 μg groups were decreased to 63.34%( P <0.001),60.55%( P <0.05)and 81.95%( P <0.001),89.53%( P <0.01),respectively,compared to the control. Conclusions:The lactacystin may reduce the levels of tyrosine hydroxylase mRNA and protein,which might be involved in the pathogenesis of Parkinson′s disease.