THERAPEUTIC EFFECTS OF ASTROCYTES EXPRESSING BOTH TYROSINE HYDROXYLASE AND BRAIN-DERIVED NEUROTROPHIC FACTOR ON A RAT MODEL OF PARKINSON’S DISEASE Z. H. WANG, a Y. JI, a;b W. SHAN, c B. ZENG, a N. RAKSADAWAN, a G. M. PASTORES, a T. WISNIEWSKI a;b and E. H. KOLODNY a a Department of Neurology, New York University School of Medicine, 550 First Avenue, New York, NY 10016, USA b Department of Pathology, New York University School of Medicine, 550 First Avenue, New York, NY 10016, USA c Department of Biochemistry and Molecular Biology, Mount Sinai School of Medicine, New York, NY 10029, USA AbstractöTyrosine hydroxylase (TH) and brain-derived neurotrophic factor (BDNF), expressed in normal astrocytes, were used in combination for the treatment of Parkinson’s disease (PD) symptoms in a rat model. Normal neonatal rat astrocytes were co-transfected with a vector expressing BDNF (AAVBDNF) and a retroviral vector expressing TH (termed TH-BDNF-DA þ cells), and then implanted into the striatum of PD rats induced by 6-hydroxydopamine. TH-BDNF-DA þ cells compensated for a severe insu⁄ciency of endogenous dopaminergic neurons in the PD rats, resulting in a signi¢cant improvement of PD symptoms. The decrease in the rotational rate of PD rats implanted with TH-BDNF-DA þ cells was more marked than that in PD rats implanted with normal astrocytes expressing either TH or BDNF alone (termed TH þ and BDNF þ cells, P 6 0.01 and 0.001, respectively), and suggested a synergistic e¡ect between TH and BDNF. In contrast, the rotational rate was not altered from the baseline in PD rats without treatment or implanted with parental rat astrocytes alone (P s 0.05). BDNF protected the dopaminergic neurons from apoptosis induced by 6-hydroxydopamine, and signi¢cantly increased the long-term survival of TH-positive cells in the striatum. Our data indicate that the combined use of TH and BDNF has a synergistic therapeutic e¡ect, and is more e⁄cient for the treatment of PD than a single gene therapy using either TH or BDNF alone. ß 2002 IBRO. Published by Elsevier Science Ltd. All rights reserved. Key words: apoptosis, astrocytes, brain-derived neurotrophic factor, gene therapy, Parkinson’s disease, tyrosine hydroxylase. Parkinson’s disease (PD), a common neurodegenerative disorder, is characterized by a progressive loss of dopa- minergic (DA) cells in the substantia nigra and a severe decrease in dopamine in the striatum. Although treat- ment of PD with L-DOPA is initially e¡ective, its e⁄cacy decreases with progression of the disease, and is often associated with increasing side e¡ects such as dyskinesia. Local production of dopamine in the striatum can be obtained by intracerebral grafting of fetal mesencephalic cells, but serious ethical issues and poor clinical out- comes have limited this approach (During et al., 1994; Freed et al., 2001; Horellou and Mallet, 1997; Levivier et al., 1995). Gene therapy of PD, an alternative therapy, is actively under investigation. Both neurotrophic factors inhibiting neurodegenerative processes and neurotransmitter-syn- thesizing enzymes provide the basis for current gene ther- apy of PD (Horellou and Mallet, 1997; Ridet et al., 1999). Tyrosine hydroxylase (TH) and the neurotrophic factor have individually been utilized for the treatment of animal models with PD (Cao et al., 1995; Choi- Lundberg et al., 1997; During et al., 1994; Fan et al., 1998; Frim et al., 1994; Kaplitt et al., 1994; Kordower et al., 2000; Levivier et al., 1995; Spina et al., 1992; Yoshimoto et al., 1995). TH converts tyrosine to L-DOPA, and is the key rate-limiting enzyme for the production of dopamine. TH activity progressively decreases following the loss of DA neurons in the sub- stantia nigra of PD patients. The use of the TH gene improves symptoms in animal models with PD (Cao et al., 1995; During et al., 1994; Kaplitt et al., 1994). The pathogenesis of PD is associated with a cascade of events that include oxidative stress, abnormalities of mitochondrial function, excitotoxicity, in£ammatory fac- tors and, ¢nally, death of DA neurons, probably through apoptosis (Marsden and Olanow, 1998). Neurotrophic 629 *Corresponding author. Tel.: +1-212-263-6549; fax: +1-212-263- 8228. E-mail address: edwin.kolodny@med.nyu.edu (E. H. Kolodny). Abbreviations: 6-OHDA, 6-hydroxydopamine; AADC, L-amino acid decarboxylase; AAV, adeno-associated viral; BDNF, brain-derived neurotrophic factor; BDNF þ cells, normal neona- tal rat astrocytes expressing BDNF alone; BH 4 , tetrahydrobiop- terin; DA, dopaminergic; DMEM, Dulbecco’s modi¢ed Eagle’s medium; GCH, GTP-cyclohydrolase; HPLC, high-performance liquid chromatography; PBS, phosphate-bu¡ered saline; PD, Parkinson’s disease; TH, tyrosine hydroxylase; TH-BDNF- DA þ cells, normal neonatal rat astrocytes expressing both TH and BDNF; TH þ cells, normal neonatal rat astrocytes expressing TH alone; TUNEL, terminal deoxynucleotidyl transferase (TdT)- mediated dUTP-£uorescein nick end labeling. NSC 5649 26-7-02 Cyaan Magenta Geel Zwart www.neuroscience-ibro.com Neuroscience Vol. 113, No. 3, pp. 629^640, 2002 ß 2002 IBRO. Published by Elsevier Science Ltd All rights reserved. Printed in Great Britain PII:S0306-4522(02)00204-X 0306-4522 / 02 $22.00+0.00