Prasad K N, Clarkson E D, La Rosa F G, Edwards-Prasad J, Freed C R
Center for Vitamins and Cancer Research, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
Mol Genet Metab. 1998 Sep;65(1):1-9. doi: 10.1006/mgme.1998.2726.
Dopamine (DA) deficiency is one of the primary lesions in the pathogenesis of Parkinson disease (PD). Because of long-term toxicity of L-DOPA therapy, the grafting of fetal mesencephalic tissue containing dopamine neurons or homogeneous populations of DA neurons into striatum appears to be rational. Fetal tissue transplants have many problems which include legal (in some countries), ethical, paucity of tissue availability, heterogenicity of cell populations, and the presence of antigen-presenting cells that are responsible for rejection of allogeneic grafts. In order to resolve the above problems, we have established immortalized DA neurons from fetal rat mesencephalon by inserting the large T-antigen (LTa) gene of the SV40 virus into the cells. A clone of DA neurons (1RB3AN27) was isolated, characterized, and tested in 6-hydroxydopamine (6-OHDA)-lesioned rats (a model of PD). These cells divided with a doubling time of about 26 h, expressed the LTa gene, and contained the tyrosine hydroxylase and dopamine transporter proteins and their respective mRNAs, which became elevated upon differentiation. These cells were nontumorigenic and nonimmunogenic and improved the symptoms of neurological deficits (methamphetamine-induced rotation) in 6-OHDA-lesioned rats. The differentiated DA neurons were more effective than undifferentiated ones. These studies suggest that immortalized DA neurons generated in vitro by LTa gene insertion may be used in transplant therapy without fear of tumor formation or rejection.
多巴胺（DA）缺乏是帕金森病（PD）发病机制中的主要病变之一。由于左旋多巴治疗存在长期毒性，将含有多巴胺神经元的胎儿中脑组织或多巴胺神经元的同质群体移植到纹状体似乎是合理的。胎儿组织移植存在许多问题，包括法律问题（在一些国家）、伦理问题、组织供应不足、细胞群体的异质性以及负责排斥同种异体移植物的抗原呈递细胞的存在。为了解决上述问题，我们通过将SV40病毒的大T抗原（LTa）基因插入细胞，从胎鼠中脑建立了永生化多巴胺神经元。分离出一个多巴胺神经元克隆（1RB3AN27），对其进行了表征，并在6-羟基多巴胺（6-OHDA）损伤的大鼠（一种帕金森病模型）中进行了测试。这些细胞的倍增时间约为26小时，表达LTa基因，含有酪氨酸羟化酶和多巴胺转运蛋白及其各自的mRNA，分化后这些物质会升高。这些细胞无致瘤性和免疫原性，并改善了6-OHDA损伤大鼠的神经功能缺损症状（甲基苯丙胺诱导的旋转）。分化的多巴胺神经元比未分化的更有效。这些研究表明，通过插入LTa基因在体外产生的永生化多巴胺神经元可用于移植治疗，而无需担心肿瘤形成或排斥反应。
