RESUMEN
Increasing evidence suggests that mesenchymal stem/stromal cells (MSCs) carrying specific mutations are at the origin of some sarcomas. We have reported that the deficiency of p53 alone or in combination with Rb (Rb(-/-) p53(-/-)) in adipose-derived MSCs (ASCs) promotes leiomyosarcoma-like tumors in vivo. Here, we hypothesized that the source of MSCs and/or the cell differentiation stage could determine the phenotype of sarcoma development. To investigate whether there is a link between the source of MSCs and sarcoma phenotype, we generated p53(-/-) and Rb(-/-)p53(-/-) MSCs from bone marrow (BM-MSCs). Both genotypes of BM-MSCs initiated leiomyosarcoma formation similar to p53(-/-) and Rb(-/-)p53(-/-) ASCs. In addition, gene expression profiling revealed transcriptome similarities between p53- or Rb-p53-deficient BM-MSCs/ASCs and muscle-associated sarcomagenesis. These data suggest that the tissue source of MSC does not seem to determine the development of a particular sarcoma phenotype. To analyze whether the differentiation stage defines the sarcoma phenotype, BM-MSCs and ASCs were induced to differentiate toward the osteogenic lineage, and both p53 and Rb were excised using Cre-expressing adenovectors at different stages along osteogenic differentiation. Regardless the level of osteogenic commitment, the inactivation of Rb and p53 in BM-MSC-derived, but not in ASC-derived, osteogenic progenitors gave rise to osteosarcoma-like tumors, which could be serially transplanted. This indicates that the osteogenic differentiation stage of BM-MSCs imposes the phenotype of in vivo sarcoma development, and that BM-MSC-derived osteogenic progenitors rather than undifferentiated BM-MSCs, undifferentiated ASCs or ASC-derived osteogenic progenitors, represent the cell of origin for osteosarcoma development.
Asunto(s)
Carcinogénesis , Diferenciación Celular , Células Madre Mesenquimatosas/patología , Fenotipo , Proteína de Retinoblastoma/deficiencia , Sarcoma/patología , Proteína p53 Supresora de Tumor/deficiencia , Tejido Adiposo/patología , Animales , Células de la Médula Ósea/patología , Ciclo Celular , Eliminación de Gen , Regulación Neoplásica de la Expresión Génica , Leiomiosarcoma/genética , Leiomiosarcoma/metabolismo , Leiomiosarcoma/patología , Ratones , Osteogénesis , Osteosarcoma/genética , Osteosarcoma/metabolismo , Osteosarcoma/patología , Proteína de Retinoblastoma/genética , Sarcoma/genética , Sarcoma/metabolismo , Proteína p53 Supresora de Tumor/genéticaRESUMEN
BACKGROUND: Human embryonic stem cells (hESCs) have opened up a new area of research in biomedicine. The efficiency of hESC derivation from frozen poor-quality embryos is low and normally achieved by plating embryos on mouse or human foreskin feeders (HFFs). We attempted to optimize embryo survival and hESC derivation. METHODS: Three conditions were tested on frozen poor-quality embryos: (i) embryo treatment with the Rho-associated kinase (ROCK) inhibitor, Y-27632; (ii) use of human mesenchymal stem cells (hMSCs) as feeders; and (iii) laser drilling (LD) for inner cell mass (ICM) isolation. Two hundred and nineteen thawed embryos were randomly treated with (n = 110) or without (n = 109) 10 microM Y-27632. Surviving embryos that developed to blastocyst stage (n = 50) were randomly co-cultured on HFFs (n = 21) or hMSCs (n = 29). ICM isolation was either by whole-blastocyst culture (WBC) or WBC plus LD. RESULTS: Embryo survival was 52% higher with Y-27632. hMSCs appeared to facilitate ICM outgrowth and hESC derivation: three hESC lines were derived on hMSCs (10.3% efficiency) whereas no hESC line was derived on HFFs. ROCK inhibition and ICM isolation method did not affect hESC efficiency. The lines derived on hMSCs (AND-1, -2, -3) were characterized and showed typical hESC morphology, euploidy, surface marker and transcription factor expression and multilineage developmental potential. The hESC lines have been stable for over 38 passages on hMSCs. CONCLUSION: Our data suggest that Y-27632 increases post-thaw embryo survival and that hMSCs may facilitate the efficiency of hESC derivation from frozen poor-quality embryos.