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In VivoL-DOPA Production by Genetically Modified Primary Rat Fibroblast or 9L Gliosarcoma Cell Grafts via Coexpression of GTPcyclohydrolase I with Tyrosine Hydroxylase

Authors
Journal
Experimental Neurology
0014-4886
Publisher
Elsevier
Publication Date
Volume
151
Issue
2
Identifiers
DOI: 10.1006/exnr.1998.6803
Keywords
  • Gene Therapy
  • Parkinson'S Disease
  • Rats
  • Transgenes
  • Dopamine
  • L-Dihydroxyphenylalanine
  • Tetrahydrobiopterin.
Disciplines
  • Biology
  • Chemistry
  • Medicine

Abstract

Abstract To investigate the biochemical requirements for in vivoL-DOPA production by cells genetically modified ex vivoin a rat model of Parkinson's disease (PD), rat syngeneic 9L gliosarcoma and primary Fischer dermal fibroblasts (FDFs) were transduced with retroviral vectors encoding the human tyrosine hydroxylase 2 (hTH2) and human GTP cyclohydrolase I (hGTPCHI) cDNAs. As GTPCHI is a rate-limiting enzyme in the pathway for synthesis of the essential TH cofactor, tetrahydrobiopterin (BH 4), only hTH2 and GTPCHI cotransduced cultured cells produced L-DOPA in the absence of added BH 4. As striatal BH 4levels in 6-hydroxydopamine (6-OHDA)-lesioned rats are minimal, the effects of cotransduction with hTH2 and hGTPCHI on L-DOPA synthesis by striatal grafts of either 9L cells or FDFs in unilateral 6-OHDA-lesioned rats were tested. Microdialysis experiments showed that those subjects that received cells cotransduced with hTH2 and hGTPCHI produced significantly higher levels of L-DOPA than animals that received either hTH2 or untransduced cells. However, animals that received transduced FDF grafts showed a progressive loss of transgene expression until expression was undetectable 5 weeks after engraftment. In FDF-engrafted animals, no differential effect of hTH2 vs hTH2 + hGTPCHI transgene expression on apomorphine-induced rotation was observed. The differences in L-DOPA production found with cells transduced with hTH2 alone and those cotransduced with hTH2 and hGTPCHI show that BH 4is critical to the restoration of the capacity for L-DOPA production and that GTPCHI expression is an effective means of supplying BH 4in this rat model of PD.

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