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Optimization of in vivo growth hormone gene transfer parameters for phenotypic correction of dwarf mice

Grant number: 14/07380-6
Support type:Scholarships in Brazil - Master
Effective date (Start): July 01, 2014
Effective date (End): January 31, 2016
Field of knowledge:Biological Sciences - Biochemistry
Cooperation agreement: Coordination of Improvement of Higher Education Personnel (CAPES)
Principal Investigator:Cibele Nunes Peroni
Grantee:Eliana Rosa Lima Filha
Home Institution: Instituto de Pesquisas Energéticas e Nucleares (IPEN). Secretaria de Desenvolvimento Econômico (São Paulo - Estado). São Paulo , SP, Brazil

Abstract

The electrotransfer of plasmid DNA containing the Genes for Human (hGH) or Mouse (mGH) growth hormone has been used by our group. For these studies of in vivo gene therapy, two animal models of GH deficiency, the dwarf (lit/lit) and immunodeficient dwarf (lit/scid) are used. With respect to the hGH gene, sustainable circulating levels of the hormone were obtained during 60 days, and a body weight gain of 33% for lit/scid mice after a single administration of the plasmid containing this gene, followed by electroporation into the quadriceps muscle. It was also found that this gene therapy strategy yielded effects comparable to those obtained after daily injections of recombinant hGH. When a long-term (~ 6 months) assay was conducted in mice, which received 3 administrations of plasmid, an increase of body weight of ~ 50% was obtained, which corresponds to an approximation of growth (catch-up growth) relative to a normal size mice (scid) of 27%. We also used a homologous electrotransfer model in which a plasmid containing the mGH gene was administered in immunocompetent mice lit/lit. In a 3 month assay, an increase in body weight of these animals of 34% was obtained, equivalent to a catch-up of 16 %. From these results, we concluded that the treatment was effective, but obviously the ideal would be to reach a catch-up of the order of 100 %, i.e., that the dwarf mice would present a complete phenotypic correction, reaching weight and size of the normal mouse, and also similar circulatory levels of the main effector of GH, the mouse insulin-like growth factor I (mIGF-I). In the present work, the aim is therefore to increase the catch-up and the main tools for this are: 1) use mice in their most significant growth phase (~ 40 days old); 2) increase the number of injection sites of the plasmid in the quadriceps muscle itself or another muscle, as the anterior tibialis; 3) optimize the amount of hyaluronidase enzyme used to facilitate the penetration of plasmid in muscle fibers; 4) employ a technique of gene transfer by attenuated hydrodynamic injection of plasmid, in order to reduce the aggressiveness of this methodology. In each condition studied and performed experimentally, should also be determined the mIGF-I increment compared with the previous situation. (AU)

Scientific publications
(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)
CECCHI, C. R.; HIGUTI, E.; LIMA, E. R.; VIEIRA, D. P.; SQUAIR, P. L.; PERONI, C. N.; BARTOLINI, P. Efficient Non-Invasive Plasmid-DNA Administration into Tibialis Cranialis Muscle of ``Little{''} Mice. CURRENT MOLECULAR MEDICINE, v. 17, n. 3, p. 230-235, 2017. Web of Science Citations: 0.
HIGUTI, ELIZA; CECCHI, CLAUDIA R.; OLIVEIRA, NELIO A. J.; LIMA, ELIANA R.; VIEIRA, DANIEL P.; AAGAARD, LARS; JENSEN, THOMAS G.; JORGE, ALEXANDER A. L.; BARTOLINI, PAOLO; PERONI, CIBELE N. Partial correction of the dwarf phenotype by non-viral transfer of the growth hormone gene in mice: Treatment age is critical. GROWTH HORMONE & IGF RESEARCH, v. 26, p. 1-7, FEB 2016. Web of Science Citations: 2.

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