Dear Prof. Pawitan,

I thank you for your attention to my work and your precious comments. Hereatfer, Iwould like to answer to your review.

Kind regards,

Ari Massoudi 

Discrepancy Zuk et al. and my work:

I have also tested the Differentiation medium used by Zuk et al. (50µM hydrocortisone, 10% FBS and 5% horse serum). I did not obtain mucle differentiation in this medium (myotube formation), and I did not detected the expression of MRF (MyoD or myf5) at the protein level (by immunocytochemistry). These data were not reported in my article (Massoudi A. WebmedCentral STEM CELL RESEARCH 2011;2(9):WMC002229.)

Zuk et al. reported experession of MRFs at the mRNA level (RT-PCR) under their differentation medium. But Zuk et al. did not report any data about expression of the MRFs at the protein level, neither myotube formation in their culture condition.

Zuk et al. reported experession of MRFs at the mRNA level (RT-PCR) under their differentation medium. But Zuk et al. did not report any data about expression of the MRFs at the protein level, neither myotube formation in their culture condition. Adult stem cells are undiferentied cells, therfore they could have a basal or stochastic mRNA expression of a wide range of genes. Therefore, to avoid misinterpretation, it is very important to investigate both at the protein level and at the phenotype level, the determination and differention of stem cells. That is what I tryed to do with my work.

I used human adipose tissue derived mesenchymal stem cells provided by Christian Dani. Dani previously reported an article (Rodriguez et al. J Exp Med. 2005 May 2;201(9):1397-405.) in which hMADS cells were able to express Myod (in vitro under differentiaton medium that I also tested) and massively fuse in vivo with regenerationg muscle of mdx mice.

http://www.ncbi.nlm.nih.gov/pubmed/15867092

Of course, I do not exclude the fact that in human adipose tissue, adult stem cell population could be heterogenous, and it could exist a sub-population having the capacity to produce myotubes. My goal was to expertise how hMADS cells were able to provide the spectacular data reported by Rodriguez et al. Therefore, I cultured hMADS cells exactly as these cells were cultured in the Lab and for the Rodriguez' paper.

Discrepancy Di Rocco G. et al. and my work:

Concerning, the fibronectin hypothesis: I did not test fibronecting as Di Rocco G et al., but I tested the Matrigel. I did not obtain any evidence of muscle determination or differention in mono-culture of hMADS cells. Matrigel is a complex extracellular material having the composition of the lamina basalis which is well-known to promote muscle differentation.

Sub-cloning of myogenic human mesenchymal stem cells from adipose tissue should be investigated; but this is another work.

The discrepency between Di Rocco G. and my results could also be due the higher per se plasticity of murine adult stem cells comapred to human ones. Indeed, the results reported in the Di Rocco G. article were obtained with murin cells, and not with human cells.

Dear Prof. Pawitan,

I thank you for your attention to my work and your precious comments. Hereatfer, Iwould like to answer to your review.

Kind regards,

Ari Massoudi 

Discrepancy Zuk et al. and my work:

I have also tested the Differentiation medium used by Zuk et al. (50µM hydrocortisone, 10% FBS and 5% horse serum). I did not obtain mucle differentiation in this medium (myotube formation), and I did not detected the expression of MRF (MyoD or myf5) at the protein level (by immunocytochemistry). These data were not reported in my article (Massoudi A. WebmedCentral STEM CELL RESEARCH 2011;2(9):WMC002229.)

Zuk et al. reported experession of MRFs at the mRNA level (RT-PCR) under their differentation medium. But Zuk et al. did not report any data about expression of the MRFs at the protein level, neither myotube formation in their culture condition.

Zuk et al. reported experession of MRFs at the mRNA level (RT-PCR) under their differentation medium. But Zuk et al. did not report any data about expression of the MRFs at the protein level, neither myotube formation in their culture condition. Adult stem cells are undiferentied cells, therfore they could have a basal or stochastic mRNA expression of a wide range of genes. Therefore, to avoid misinterpretation, it is very important to investigate both at the protein level and at the phenotype level, the determination and differention of stem cells. That is what I tryed to do with my work.

I used human adipose tissue derived mesenchymal stem cells provided by Christian Dani. Dani previously reported an article (Rodriguez et al. J Exp Med. 2005 May 2;201(9):1397-405.) in which hMADS cells were able to express Myod (in vitro under differentiaton medium that I also tested) and massively fuse in vivo with regenerationg muscle of mdx mice.

http://www.ncbi.nlm.nih.gov/pubmed/15867092

Of course, I do not exclude the fact that in human adipose tissue, adult stem cell population could be heterogenous, and it could exist a sub-population having the capacity to produce myotubes. My goal was to expertise how hMADS cells were able to provide the spectacular data reported by Rodriguez et al. Therefore, I cultured hMADS cells exactly as these cells were cultured in the Lab and for the Rodriguez' paper.

Discrepancy Di Rocco G. et al. and my work:

Concerning, the fibronectin hypothesis: I did not test fibronecting as Di Rocco G et al., but I tested the Matrigel. I did not obtain any evidence of muscle determination or differention in mono-culture of hMADS cells. Matrigel is a complex extracellular material having the composition of the lamina basalis which is well-known to promote muscle differentation.

Sub-cloning of myogenic human mesenchymal stem cells from adipose tissue should be investigated; but this is another work.

The discrepency between Di Rocco G. and my results could also be due the higher per se plasticity of murine adult stem cells comapred to human ones. Indeed, the results reported in the Di Rocco G. article were obtained with murin cells, and not with human cells.