Brain Research Bulletin 69 (2006) 647–651 Peripheral and mesencephalic transfer of a synthetic gene for the thymic peptide thymulin Gustavo R. Morel a , Oscar A. Brown a , Paula C. Reggiani a , Claudia B. Here ˜ n´ u a , Enrique L. Portiansky b , Gustavo O. Zuccolilli c , Jean M. Pl´ eau d , Mireille Dardenne d , Rodolfo G. Goya a,∗ a Institute for Biochemical Research-Histology B (INIBIOLP), Faculty of Medicine, Argentina b Institute of Pathology, School of Veterinary Sciences, UNLP, La Plata, Argentina c Institute of Anatomy, School of Veterinary Sciences, UNLP, La Plata, Argentina d CNRS UMR 8147, Universit´ e Paris V, Hˆ opital Necker, Paris, France Received 28 January 2006; received in revised form 9 March 2006; accepted 17 March 2006 Available online 18 April 2006 Abstract Thymulin is a thymic peptide with antiinflammatory activity in the brain. We constructed a recombinant adenoviral vector, RAd-FTS, expressing a synthetic DNA sequence encoding met-FTS, a biologically active analog of thymulin and used it for peripheral and central gene transfer in rats. Thymulin concentration in serum and brain tissue was determined by bioassay. Reporter gene expression in the substantia nigra (SN) was quantitated by enzymohistochemistry or fluorescence microscopy using an appropriate image analysis software. A single intramuscular injection (10 8 plaque forming units (pfu)/animal) of RAd-FTS in thymectomized rats (nondetectable serum thymulin) induced supraphysiologic serum thymulin levels for at least 110 days (123 ± 22 fg/ml versus 598 ± 144 fg/ml in intact and vector-injected rats, respectively). Stereotaxic intranigral injection of RAd-FTS induced steady expression levels of met-FTS for at least 90 days, whereas expression of adenovirally transferred reporter genes coding for green fluorescent protein fused to HSV thymidine kinase (GFP-TK) fus or E. coli -galactosidase (-gal), declined drastically within a month (% transgene expression in the SN on post-injection day 30 relative to day 2 was: 18, <1 and 125%, for -gal, (GFP-TK) fus and met-FTS, respectively). We conclude that RAd-FTS constitutes a suitable biotechnological tool for the assessment of peripheral and central thymulin gene therapy in animal models of nigral dopaminergic neurodegeneration induced by pro-inflammatory agents. © 2006 Elsevier Inc. All rights reserved. Keywords: Thymulin; Synthetic gene; Adenoviral vector; Substantia nigra; Gene transfer; Long-term expression; Antiinflammatory action 1. Introduction Thymulin is a thymic hormone involved in several aspects of intra- and extrathymic T-cell differentiation [1,3]. Thymulin, which is exclusively produced by the thymic epithelial cells (TEC), consists of a nonapeptide component (termed facteur thymique serique or FTS), coupled in an equimolecular ratio to the ion zinc [8], which confers biological activity to this molecule [6]. ∗ Corresponding author at: INIBIOLP, Faculty of Medicine, UNLP, CC 455, Zip 1900, La Plata, Argentina. Tel.: +54 221 425 6735: fax: +54 221 425 0924. E-mail addresses: gustavmorel@Argentina.com (G.R. Morel), rgoya@netverk.com.ar (R.G. Goya). There is increasing evidence that thymulin and thymulin analogs possess antiinflammatory and analgesic activity in the brain, which makes this peptide a molecule of clinical interest [16,18]. Unfortunately, efforts to clone the gene for thymulin have been unrewarding so far. This has prevented the assess- ment of thymulin gene therapy strategies in relevant animal models [9]. This situation led us to undertake the design and con- struction of a synthetic DNA sequence encoding a biologically active analog of thymulin [14]. We report here the utilization of a recombinant adenoviral vector (RAd) to transfer a synthetic gene for thymulin peripherally and in the substantia nigra (SN) of rats. We also document that transgenic thymulin has a longer expression in the SN than the adenovirally transferred genes for a fusion variant of enhanced green fluorescent protein (GFP) or for E. coli -galactosidase (-gal), two reporter genes. 0361-9230/$ – see front matter © 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.brainresbull.2006.03.015