Eletrotransferência do gene do hormônio de crescimento de camundongo associada à administração de células-tronco mesenquimais em modelo murino de osteogênese imperfeita

Abstract

GOMES, A. M. Electrotransference of the mouse growth hormone gene associated with the administration of mesenchymal stem cells, in a murine model of osteogenesis imperfecta, 2021. 131 p. Dissertação (Mestrado em Ciências) -Instituto de Pesquisas Energéticas e Nucleares -IPEN-CNEN/SP, São Paulo.Pre-clinical and clinical trials of cell and gene therapy have been developed and applied in a very promising manner, through the latest decades, facing the search for the cure or the effective improvement of several genetic disorders. Our Gene Therapy group of the Biotechnology Center, at IPEN, holds vast experience on in vivo delivery of plasmid DNA containing human growth hormone (hGH) and murine (mGH) genes for the treatment of GH-deficient mice, developing protocols that showed very relevant results in the last 10 years. Thus, in this work we propose a novel approach based on the association between in vivo gene therapy and allogeneic transplant of mesenchymal stem cells for treating osteogenesis imperfecta (OI) in a murine model, the oim mice. The main feature of OI is the bone fragility caused, mainly, by mutations in the genes that synthesize or process type I collagen, and in those responsible for osteoblast maturation. Initially, we evaluated two lineages of mesenchymal stem cells (MSCs) extracted from two different tissues, bone marrow (BM-MSC) and adipose tissue (AT-MSC), to comprehend which is the better bone healer on the oim mice femurs. These cells were applied locally through the femoral condyles of previously irradiated oim mice. The bioassay comparing these two types of cells showed positive results regarding the action of AT-MSC for the improvement of the femoral bone quality in these animals, through the dosage of the αII (I) chain of the type I collagen, and furthermore providing significant results (P < 0.05) acquired from biomechanics evaluation. Therefore, this lineage was chosen for the bioassay of pre-clinical evaluation, associating the allogeneic transplant of these cells to the electroporation of the mGH gene. The techniques applied were based on pre-clinical protocols performed by our gene therapy group for the phenotypical improvement of both GH-deficient models and heterozygous oim mice. The results revealed that the association between in vivo gene therapy and AT-MSC transplantation was statistically significant (P < 0.05) versus negative control for practically all the evaluated parameters, demonstrating that these cells possess their reparative activity, possibly, potentialized by over expression of mGH. Collectively, the data reported in this work unveil new horizons for the understanding of the action of AT-MSC in tissue reparation, as well as for the anabolic role of GH over type I OI phenotype improvement.