Gene therapy of tyrosine hydroxylase, aromatic L-amino acid decarboxylase, and GTP cyclohydrolase genes in rat model of Parkinson's disease

Abstract

To detect the expression and function of enzyme genes involved in biosynthetic pathway for dopamine in vitro and assess their effect in rat model of Parkinson's disease. Cos7 cells were transfected with separate adeno-associated virus (AAV) expressing tyrosine hydroxylase (TH) gene, aromatic L-amino acid decarboxylase (AADC) gene and GTP cyclohydrolase I (GCH-I) gene. The expression and function of the three genes were detected by methods of immunohistochemistry, in situ hybridization and high performance liquid chromatograph and electrochemical detection (HPLC-ECD). Gene engineered cells were sequentially transplanted into the striatum of 6-hydroxy-dopamine-leisioned Parkinsonian rat by stereotaxic instrastriatal injection. The asymmetric rotations of these rats after apomorphine administration were detected every week after transplantation. 10 weeks after grafting, the animals were sacrificed and the dopamine produced in the striatum was detected by HPLC-ECD. In vitro experiments showed that the three genes were high expressed in Cos7 cells. When Cos7 cells expressing TH, AADC and GCH-I were cocultured, they produced large amount of dopamine in the condition of existance of L-tyrosine. Furthermore, triple genes therapy resulted in greater dopamine production in the striatum of Parkinsonian rats and improved the rotational behavior of the rats more efficiently than did single gene therapy. However, the production of dopamine in the rats with triple genes therapy is no more than double genes therapy. For gene therapy in Parkinson's disease, the amount of target genes to be used should be determined by the level of doperminergic neurons damaged. In the present study, the efficiency of multiple genes therapy is significantly better than that of single gene therapy.