A New Immunodeficient Mouse Model for Human Myoblast Transplantation

Abstract

Design of efficient transplantation strategies for myoblast-based gene therapies in humans requires animal models in which xenografts are tolerated for long periods of time. In addition, such recipients should be able to withstand pretransplantation manipulations for enhancement of graft growth. Here we report that a newly developed immunodeficient mouse carrying two known mutations (the recombinase activating gene 2, RAG2, and the common cytokine receptor gamma, gammac) is a candidate fulfilling these requirements. Skeletal muscles from RAG2(-/-)/gammac(-/-) double mutant mice recover normally after myotoxin application or cryolesion, procedures commonly used to induce regeneration and improve transplantation efficiency. Well-differentiated donor-derived muscle tissue could be detected up to 9 weeks after transplantation of human myoblasts into RAG2(-/-)/gammac(-/-) muscles. These results suggest that the RAG2(-/-)/gammac(-/-) mouse model will provide new opportunities for human muscle research.

Normal growth and regenerating ability of myoblasts from unaffected muscles of facioscapulohumeral muscular dystrophy patients

“…[55][56][57][58] In the present study, human FSHD myoblasts were injected into immunodeficient Rag mice, a validated mouse model for the analysis of muscle regeneration in the context of xenogenic MT. The implantation results we obtained were close to that reported previously [47][48][49] and confirmed that myoblasts prepared from unaffected FSHD muscles, even used at the 6th and 7th passages, take part into in vivo muscle regeneration. The overall number of dystrophin-positive fibers was inferior to that reported previously in immunocompetent models of muscular dystrophies in the context of syngenic or allogenic MT.…”

“…In the present study, the evaluation of transplantation outcome has been performed 1 month after transplantation, a time course that is of general use in human and animal MT assays. 9,16,17,20,31,47,49 This model was not regarded helpful to evaluate the longterm persistence of muscle fibers harboring the human mutated genes. Indeed, a negative selection of muscle fibers expressing the mutated gene could occur with time in the healthy environment of mouse muscle fibers, and this selection would not be representative of the situation presented in FSHD.…”

“…At 1 month after MT, the expression of human proteins validated the implantation of human cells into mouse muscle fibers. 16,[47][48][49] Human dystrophin was observed in all cell-injected muscles, and was absent from muscles injected with a saline solution (Table 3, Figure 5). No significant differences were observed between muscles receiving FSHD or control myoblasts (P40.8).…”

“…46 The cell implantation model had been validated previously. 47 Aliquots of cells were thawed and grown in one single plate (600 cm 2 , Nunc) in proliferation medium. The myoblasts were then used at P6 and P7.…”

“…The mice were killed 4 weeks after the last series of cell injections, which is a delay compatible with complete mouse muscle regeneration and the expression of donor markers. 16,[47][48][49] The muscles were snap frozen in nitrogen-cooled isopentan and serially sectioned (8 mm) using a cryostat. The participation of human cells in muscle formation was detected by the specific expression of human dystrophin beneath the basement membrane of muscle fibers.…”