**7. Muscle progenitor cell transplantation: Causes of failure and new perspectives**

So far, in pre-clinical models, the exploration of cell-based therapy for muscle degeneration in MDs showed promising results. However, both technical and ethical issues are still the main determinants that hinder this therapeutic approach in clinics. The low abundance of satellite cells in human muscle, their heterogeneity and the reduced myogenic potential when expand‐ ed *in vitro* are the reasons why this source of stem cell is not ideal for the treatment of muscular dystrophies. Previously, satellite cells showed a general limited cell migration from the transplantation site and were not able to cross the endothelial barrier when injected systemi‐ cally [12]. To overcome this important issue, MABs were proposed as an alternative source of cells. This is due to their ability to cross the vessel walls after intra-arterial delivery in dystro‐ phic mice [11, 12]. Nevertheless, a large number of cell deaths can still occur if the immune response occurs within a few hours of the treatment. Thus, immune suppression is needed in the heterologous transplantation, as planned in the ongoing phase I/II clinical trial where Duchenne patients were transplanted intra-arterially with HLA-identical allogeneic MABs (EudraCTno. 2011-000176-33). This clinical trial faced three problems: the age of enrolled patients (frequently in advanced stage of diseases), the low dose of cells transplanted (from 1/5 to 1/10 of that administered to the GRMD dogs) and the intra femoral arteries delivery (limiting the treatment only for the muscle located downstream of femoral artery). For the treatment of muscular dystrophies, pluripotent stem cells have been recently explored. In such pre-clinical studies, the major problem encountered was controlling their myogenic differen‐ tiation and avoiding tumour formation. In addition, the therapeutic use of ES cells has been strongly contested within the scientific community and from public opinion. This is because of ethical concerns (due to the embryonic origin of ES cells) and it does not seem that the employment of iPS cells will help this.

derm, ectoderm and endoderm. It was demonstrated in the early 1990s that, if cultured *in vitro*, murine ES cells can develop aggregates of cells (embryoid bodies) and differentiate in skeletal muscle cells expressing myogenic markers in the same muscle-specific determina‐ tion genes order observed during embrional development: myf5, myogenin, myoD and myf6 [47]. Later on, both *in vitro* and *in vivo* studies confirmed their myogenic differentiation potential [13]. Nonetheless, the possibility of a therapeutic adoption of ES cells met the criticisms of both civil and scientific communities (see below). In 2006, Yamanaka publish‐ ed a revolutionary, paradigm-shift study. For the first time, a fate conversion of somatic cells (fibroblasts) into pluripotency was demonstrated [48]. As a result of this study, Yamanaka won the Nobel Prize in 2012 and began a new era for pluripotent stem cells-based therapeu‐ tic approach of chronic illness. So far, the myogenic potential of the induced pluripotent stem (iPS) cells, either from mouse or human origin, have been provided to counteract muscle degeneration in MDs [13] (Figure 2). In particular, *in vitro* and *in vivo* analyses showed that myogenic precursors generated from iPS cells could produce chimeric myotubes if cocultured with C2C12 myogenic cell line. Furthermore, if transplanted in dystrophic mus‐

cles, their contractile properties could also be improved [49].

**perspectives**

404 Muscle Cell and Tissue

**7. Muscle progenitor cell transplantation: Causes of failure and new**

So far, in pre-clinical models, the exploration of cell-based therapy for muscle degeneration in MDs showed promising results. However, both technical and ethical issues are still the main determinants that hinder this therapeutic approach in clinics. The low abundance of satellite cells in human muscle, their heterogeneity and the reduced myogenic potential when expand‐ ed *in vitro* are the reasons why this source of stem cell is not ideal for the treatment of muscular dystrophies. Previously, satellite cells showed a general limited cell migration from the transplantation site and were not able to cross the endothelial barrier when injected systemi‐ cally [12]. To overcome this important issue, MABs were proposed as an alternative source of cells. This is due to their ability to cross the vessel walls after intra-arterial delivery in dystro‐ phic mice [11, 12]. Nevertheless, a large number of cell deaths can still occur if the immune response occurs within a few hours of the treatment. Thus, immune suppression is needed in the heterologous transplantation, as planned in the ongoing phase I/II clinical trial where Duchenne patients were transplanted intra-arterially with HLA-identical allogeneic MABs (EudraCTno. 2011-000176-33). This clinical trial faced three problems: the age of enrolled patients (frequently in advanced stage of diseases), the low dose of cells transplanted (from 1/5 to 1/10 of that administered to the GRMD dogs) and the intra femoral arteries delivery (limiting the treatment only for the muscle located downstream of femoral artery). For the treatment of muscular dystrophies, pluripotent stem cells have been recently explored. In such pre-clinical studies, the major problem encountered was controlling their myogenic differen‐ tiation and avoiding tumour formation. In addition, the therapeutic use of ES cells has been strongly contested within the scientific community and from public opinion. This is because

In conclusion, many scientific issues can be solved by further investigations in the cell biology of myogenic stem cells. With this in mind, further studies on mechanisms regulating skeletal muscle regeneration in basic and applied research are needed in order to solve several practical problems, as described above.
