Differential Gene Expression Profiling of Dystrophic Dog Muscle after MuStem Cell Transplantation

Abstract

BackgroundSeveral adult stem cell populations exhibit myogenic regenerative potential, thus representing attractive candidates for therapeutic approaches of neuromuscular diseases such as Duchenne Muscular Dystrophy (DMD). We have recently shown that systemic delivery of MuStem cells, skeletal muscle-resident stem cells isolated in healthy dog, generates the remodelling of muscle tissue and gives rise to striking clinical benefits in Golden Retriever Muscular Dystrophy (GRMD) dog. This global effect, which is observed in the clinically relevant DMD animal model, leads us to question here the molecular pathways that are impacted by MuStem cell transplantation. To address this issue, we compare the global gene expression profile between healthy, GRMD and MuStem cell treated GRMD dog muscle, four months after allogenic MuStem cell transplantation.ResultsIn the dystrophic context of the GRMD dog, disease-related deregulation is observed in the case of 282 genes related to various processes such as inflammatory response, regeneration, calcium ion binding, extracellular matrix organization, metabolism and apoptosis regulation. Importantly, we reveal the impact of MuStem cell transplantation on several molecular and cellular pathways based on a selection of 31 genes displaying signals specifically modulated by the treatment. Concomitant with a diffuse dystrophin expression, a histological remodelling and a stabilization of GRMD dog clinical status, we show that cell delivery is associated with an up-regulation of genes reflecting a sustained enhancement of muscle regeneration. We also identify a decreased mRNA expression of a set of genes having metabolic functions associated with lipid homeostasis and energy. Interestingly, ubiquitin-mediated protein degradation is highly enhanced in GRMD dog muscle after systemic delivery of MuStem cells.ConclusionsOverall, our results provide the first high-throughput characterization of GRMD dog muscle and throw new light on the complex molecular/cellular effects associated with muscle repair and the clinical efficacy of MuStem cell-based therapy.

Quantitative proteome profiling of dystrophic dog skeletal muscle reveals a stabilized muscular architecture and protection against oxidative stress after systemic delivery of MuStem cells

“…Multilevel analyses simultaneously investigate the transcriptome and the proteome of the tissue, and thus have greater potential for providing information on the biological pathways impacted by cell therapy. However, it is noteworthy that, among the 25 proteins overexpressed following cell transplantation, only ITIH4 was identified in our previous transcriptomic study, where it was also shown to be overexpressed [10]. Similarly, among the 46 proteins identified as underexpressed, only MYH2, MYH4, and MYH7 were identified at a transcriptomic level, and were conversely shown to be overexpressed.…”

“…MuStem cell delivery is associated with an upregulation of genes playing a role in the enhancement of skeletal muscle fiber regeneration. Simultaneously, we observed an overrepresentation of ubiquitin‐mediated protein reflecting a more active degradation process of the damaged tissue .…”

“…We have recently shown that systemic delivery of MuStem cells generates beneficial clinical and tissue impacts in the GRMD dog . Also, we implemented the remodeling of dystrophic muscle tissue by demonstrating the induction of differential gene expression profiling at the transcript level . In the present study, we use a large‐scale quantitative proteomic approach to complement these observations at the protein level.…”

Determination of qPCR Reference Genes Suitable for Normalizing Gene Expression in a Canine Model of Duchenne Muscular Dystrophy

“…Effective normalization requires appropriate reference genes, and indeed efforts to identify, validate and publicize such genes are becoming more common in a variety of disease states [20][21][22][23][24] and model organisms [25][26][27][28][29][30]. A review of the literature specifically within the DMD field however reveals a considerable number of candidates: selected examples in dystrophic dogs include GAPDH [31][32][33], RPS18 [34], HPRT1 [35,36], 18S [37]; in humans, TBP and GUSB [13]; and in mice GAPDH [33,38], ActB [39], 18S [40]. There appears to be minimal effort to apply reference genes consistently between studies (even varying from manuscript to manuscript within a research group), and data supporting the selection of the gene or genes used is rarely presented.…”

Vascular Delivery of Allogeneic MuStem Cells in Dystrophic Dogs Requires Only Short-Term Immunosuppression to Avoid Host Immunity and Generate Clinical/Tissue Benefits

“…Over the past years, we have isolated delayed adherent MDSCs from healthy dogs, which we have named MuStem cells, and demonstrated that their vascular delivery into GRMD dogs submitted to IS that started 1 week before cell administration and then maintained throughout the experiment (mentioned then as continuous IS) produce striking clinical stabilization and long-term muscle repair 58 , 59 . In the present study, we sought to determine the optimal immunosuppressive regimen required to obtain beneficial effects using the aforementioned allogeneic MuStem cells infusion protocol.…”