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Gene therapy for long-term restoration of circulating thymulin in thymectomized mice and rats

Gene Therapy volume 13, pages 1214–1221 (2006)Cite this article

Abstract

Thymulin is a thymic peptide possessing hypophysiotropic activity and antiinflammatory effects in the brain. We constructed a synthetic DNA sequence encoding met-FTS, a biologically active analog of thymulin, and subsequently cloned it into different expression vectors. A sequence optimized for expression of met-FTS in rodents, 5′-ATGCAGGCCAAGTCGCAGGGGGGGTCGAACTAGTAG-3′, was cloned in the mammalian expression vectors pCDNA3.1(+) and phMGFP (which expresses the Monster Green Fluorescent Protein), thus obtaining pcDNA3.1-metFTS and p-metFTS-hMGFP, which express met-FTS and the fluorescent fusion protein metFTS-hMGFP, respectively. The synthetic sequence was also used to construct the adenoviral vector RAd-metFTS, which expresses met-FTS. Transfection of HEK293 and BHK cells with pcDNA3.1-metFTS (experimental groups) or pcDNA3.1 (control), led to high levels of thymulin bioactivity (>600 versus <0.1 pg/ml in experimental and control supernatants, respectively). Transfection of HEK293 and BHK cells with pmetFTS-hMGFP revealed a cytoplasmic and nuclear distribution of the fluorescent fusion protein. A single intramuscular (i.m.) injection (107 plaque forming units (PFU)/mouse or 108 PFU/rat) of RAd-metFTS in thymectomized animals (nondetectable serum thymulin) restored serum thymulin levels for at least 110 and 130 days post-injection in mice and rats, respectively. We conclude that RAd-metFTS constitutes a suitable biotechnological tool for the implementation of thymulin gene therapy in animal models of chronic brain inflammation.

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Acknowledgements

The authors are grateful to Ms Yolanda Sosa for technical assistance and Dr Maria José Bellini for assistance with digital microphotograpy. This work was supported in part by NIH Grant #R21TW6665, grants from the Argentine Research Council (CONICET) and Grant #PICT10663 from the National Agency for the Promotion of Science and Technology (ANPCyT) to RGG, and by the Institut National de la Santé et de la Recherche Médicale (INSERM), France, to MD and RGG. PCR and CBH are ANPCyT and CONICET doctoral fellows, respectively. OAB, OJR and RGG are career researchers of CONICET.

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  1. P C Reggiani and C B Hereñú: These authors contributed equally to this work.

Authors and Affiliations

  1. Institute for Biochemical Research-Histology B, Faculty of Medicine, National University of La Plata, La Plata, Argentina

    P C Reggiani, C B Hereñú, O J Rimoldi, O A Brown & R G Goya

  2. CNRS UMR 8147, Université Paris V, Hôpital Necker, Paris, France

    J-M Pléau, M Dardenne & R G Goya

Authors
  1. P C Reggiani
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  2. C B Hereñú
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  3. O J Rimoldi
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  4. O A Brown
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  5. J-M Pléau
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  6. M Dardenne
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  7. R G Goya
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Correspondence to R G Goya.

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Reggiani, P., Hereñú, C., Rimoldi, O. et al. Gene therapy for long-term restoration of circulating thymulin in thymectomized mice and rats. Gene Ther 13, 1214–1221 (2006). https://doi.org/10.1038/sj.gt.3302775

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