Primary astrocytes retrovirally transduced with a tyrosine hydroxylase transgene driven by a glial-specific promoter elicit behavioral recovery in experimental Parkinsonism

Abstract

We used a retroviral-mediated gene transfer system to transduce primary rat astrocytes with a transgene in which the activity of a tyrosine hydroxylase (TH) cDNA is under the transcriptional control of a human promoter of the glial fibrillary acidic protein (GFAP). The engineered cells were tested for their therapeutic efficacy in a rodent model of Parkinson's disease (PD). The method is based both on the properties of astrocytes, as well as on those of the promoter. Astrocytes are an integral part of the neural tissue, have a long life span, are more resistant to oxidative stress than neurons, and possess an efficient secretory system. The GFAP promoter is active throughout postnatal life, and its activity is up-regulated by many insults to the brain, including PD. Transduced astrocytes were implanted into the striata of rats lesioned with 6-hydroxydopamine (6-OHDA), and the efficacy of grafted cells tested. Implanted astrocytes induced a significant reduction in the turning behavior that occurs in response to apomorphine for at least 4 weeks after grafting, and transgenic mRNA and protein could be detected in implanted brains. These results indicate that the gFa2-TH construct can be readily adapted to be used with a retroviral gene transfer system to obtain nontumorigenic cells that sustain a sufficient level of transgene activity to enable therapeutic effectiveness for prolonged periods. These results further endorse the use of astrocytes for gene therapy in the central nervous system.

Gene Therapy for Parkinson??s Disease

“…To overcome this severe limitation, we turned to primary astrocytes. In agreement with the vector expression results, behavioral recovery, as determined by the decrease in turning behavior, was maintained longer and at lower levels [73] than in previous reports where engineered astrocytes have been used. [73] Transduced astrocytes survived, expressed the TH transgene, and induced behavioral recovery for up to 4 weeks, without evidence of tumor formation.…”

“…C6 cells, derived from a rat glioblastoma, were stably transfected with the gfa2-TH transgene, and we observed expression of both transgenic mRNA and protein. [73] Transduced astrocytes survived, expressed the TH transgene, and induced behavioral recovery for up to 4 weeks, without evidence of tumor formation. [70] As a proofof-concept experiment, C6 cells carrying the gfa2-TH transgene were implanted into the striata of 6-OHDA-lesioned rats.…”

“…[70] Furthermore, gene activity and protein expression were up-regulated by forskolin, an adenylate cyclase activator, indicating that the transcriptional properties of the gfa2 promoter were preserved. [73] Both the endogenous GFAP promoter, and the gfa2 promoter transmitted in transgenic mice remain active for the lifetime of the organism. Engineered cells survived in the lesioned brain, expressed transgenic messenger RNA (mRNA) and protein, and were capable of reducing apomorphine-induced turning behavior.…”

Brain engraftment of autologous macrophages transduced with a lentiviral flap vector: an approach to complement brain dysfunctions

“…Numerous cell types such as neural progenitors, astrocytes, fibroblasts and immortalized cells have been genetically modified for their further transplantation into the rat brain. [1][2][3][4][5][6] For example, in a rat model of Parkinson's disease, the intrastriatal graft of dopaminergic embryonic neurons expressing the human Cu/Zn superoxide dismutase gene, was found to improve cell survival and to correct motor symptoms partially. 7,8 In the same model, the graft of astrocytes expressing tyrosine hydroxylase significantly improved motor deficits.…”

“…7,8 In the same model, the graft of astrocytes expressing tyrosine hydroxylase significantly improved motor deficits. 1 Other CNS diseases like peroxisomal or abnormal lysosomal storage are known to involve macrophage defects. 9,10 In the murine Gaucher disease, it has been reported that glucocerebrosidase-transduced mononuclear phagocytes injected intravenously migrated into the brain.…”