Characterization of human mesenchymal stem cells from ewing sarcoma patients. Pathogenetic implications.
BACKGROUND
Ewing Sarcoma (EWS) is a mesenchymal-derived tumor that generally arises in bone and soft tissue. Intensive research regarding the pathogenesis of EWS has been insufficient to pinpoint the early events of Ewing sarcomagenesis. However, the Mesenchymal Stem Cell (MSC) is currently accepted as the most probable cell of origin.


METHODS
In an initial study regarding a deep characterization of MSC obtained specifically from EWS patients (MSC-P), we compared them with MSC derived from healthy donors (MSC-HD) and EWS cell lines. We evaluated the presence of the EWS-FLI1 gene fusion and EWSR1 gene rearrangements in MSC-P. The presence of the EWS transcript was confirmed by q-RT-PCR. In order to determine early events possibly involved in malignant transformation, we used a multiparameter quantitative strategy that included both MSC immunophenotypic negative/positive markers, and EWS intrinsic phenotypical features. Markers CD105, CD90, CD34 and CD45 were confirmed in EWS samples.


RESULTS
We determined that MSC-P lack the most prevalent gene fusion, EWSR1-FLI1 as well as EWSR1 gene rearrangements. Our study also revealed that MSC-P are more alike to MSC-HD than to EWS cells. Nonetheless, we also observed that EWS cells had a few overlapping features with MSC. As a relevant example, also MSC showed CD99 expression, hallmark of EWS diagnosis. However, we observed that, in contrast to EWS cells, MSC were not sensitive to the inhibition of CD99.


CONCLUSIONS
In conclusion, our results suggest that MSC from EWS patients behave like MSC-HD and are phenotypically different from EWS cells, thus raising important questions regarding MSC role in sarcomagenesis.
Molecular Pathology Program, Institute of Biomedical Research of Salamanca-Centro de Investigación del Cáncer, Centro de Investigación del Cáncer (IBSAL-CIC), Salamanca, Spain ; Instituto de Biomedicina de Sevilla (IBiS), CSIC-Universidad de Sevilla, Department of Pathology and Biobank, Hospital Universitario Virgen del Rocío, Seville, Spain.
Ana Teresa
AT
Amaral
CRS Sviluppo di Terapie Biomolecolari, Oncologia Sperimentale, Istituto Ortopedico Rizzoli (IOR), Bologna, Italy.
Maria Cristina
MC
Manara
Department of Pediatrics and Biobank, Leiden University Medical Center (LUMC), Leiden, The Netherlands.
Dagmar
D
Berghuis
Molecular Pathology Program, Institute of Biomedical Research of Salamanca-Centro de Investigación del Cáncer, Centro de Investigación del Cáncer (IBSAL-CIC), Salamanca, Spain.
José Luis
JL
Ordóñez
Instituto de Biomedicina de Sevilla (IBiS), CSIC-Universidad de Sevilla, Department of Pathology and Biobank, Hospital Universitario Virgen del Rocío, Seville, Spain.
Michele
M
Biscuola
Instituto de Biomedicina de Sevilla (IBiS), CSIC-Universidad de Sevilla, Department of Pathology and Biobank, Hospital Universitario Virgen del Rocío, Seville, Spain.
Maria Angeles
MA
Lopez-García
Molecular Pathology Program, Institute of Biomedical Research of Salamanca-Centro de Investigación del Cáncer, Centro de Investigación del Cáncer (IBSAL-CIC), Salamanca, Spain.
Daniel
D
Osuna
Osteoarticolar Regeneration Laboratory, Istituto Ortopedico Rizzoli (IOR), Bologna, Italy.
Enrico
E
Lucarelli
Dipartimento di Istologia, Embriologia e Biologia, Istituto Ortopedico Rizzoli (IOR), Bologna, Italy.
Francesco
F
Alviano
Department of Pediatrics and Biobank, Leiden University Medical Center (LUMC), Leiden, The Netherlands.
Arjan
A
Lankester
CRS Sviluppo di Terapie Biomolecolari, Oncologia Sperimentale, Istituto Ortopedico Rizzoli (IOR), Bologna, Italy.
Katia
K
Scotlandi
Molecular Pathology Program, Institute of Biomedical Research of Salamanca-Centro de Investigación del Cáncer, Centro de Investigación del Cáncer (IBSAL-CIC), Salamanca, Spain ; Instituto de Biomedicina de Sevilla (IBiS), CSIC-Universidad de Sevilla, Department of Pathology and Biobank, Hospital Universitario Virgen del Rocío, Seville, Spain.
Enrique
E
de Álava
D000071443:12E7 Antigen; D015703:Antigens, CD / Q000378:metabolism; D018952:Antigens, CD34 / Q000378:metabolism; D002148:Calmodulin-Binding Proteins / Q000235:genetics*; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D000071063:Endoglin; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D015321:Gene Rearrangement; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D017493:Leukocyte Common Antigens / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism* / Q000473:pathology; D015514:Oncogene Proteins, Fusion / Q000235:genetics*; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics*; D034802:RNA-Binding Protein EWS / Q000235:genetics*; D016601:RNA-Binding Proteins / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012512:Sarcoma, Ewing / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D018800:Thy-1 Antigens / Q000378:metabolism
D000071443:12E7 Antigen; D015703:Antigens, CD; D018952:Antigens, CD34; C061828:CD99 protein, human; D002148:Calmodulin-Binding Proteins; D015815:Cell Adhesion Molecules; C064206:ENG protein, human; C106031:EWS-FLI fusion protein; C500986:EWSR1 protein, human; D000071063:Endoglin; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins; D011956:Receptors, Cell Surface; D018800:Thy-1 Antigens; D017493:Leukocyte Common Antigens; C516769:PTPRC protein, human