Tissue resident mesenchymal stem cells (MSC) are important regulators of tissue repair or regeneration, fibrosis, inflammation, angiogenesis and tumor formation. Here we define a population of resident lung mesenchymal stem cells (luMSC) that function to regulate the severity of bleomycin injury via modulation of the T-cell response. Bleomycin induced loss of these endogenous luMSC and elicited fibrosis (PF), inflammation and pulmonary arterial hypertension (PAH). Replacement of resident stem cells by administration of isolated luMSC attenuated the bleomycin-associated pathology and mitigated the development of PAH. In addition, luMSC modulated a decrease in numbers of lymphocytes and granulocytes in bronchoalveolar fluid and demonstrated an inhibition of effector T cell proliferation in vitro. Global gene expression analysis indicated that the luMSC are a unique stromal population differing from lung fibroblasts in terms of proinflammatory mediators and pro-fibrotic pathways. Our results demonstrate that luMSCs function to protect lung integrity following injury however when endogenous MSC are lost this function is compromised illustrating the importance of this novel population during lung injury. The definition of this population in vivo in both murine and human pulmonary tissue facilitates the development of a therapeutic strategy directed at the rescue of endogenous cells to facilitate lung repair during injury.

Reprogramming somatic cells into an embryonic stem (ES) cell-like state, or induced pluripotent stem (iPS) cells, has emerged as a promising new venue for customized cell therapies. In this study, we performed directed differentiation to assess the ability of murine iPS cells to differentiate into bone, cartilage and fat in vitro and to maintain an osteoblast phenotype on a scaffold in vitro and in vivo. Embryoid bodies derived from murine iPS cells were cultured in differentiation medium for eight to twelve weeks. Differentiation was assessed by lineage specific morphology, gene expression, histological stain and immunostaining to detect matrix deposition. After 12 weeks of expansion, iPS derived osteoblasts were seeded in a gelfoam matrix followed by subcutaneous implantation in syngenic ICR mice. Implants were harvested at 12 weeks, and histological analyses of cell, mineral and matrix content were performed. Differentiation of iPS cells into mesenchymal lineages of bone, cartilage and fat was confirmed by morphology, and expression of lineage specific genes. Isolated implants of iPS cell derived osteoblasts expressed matrices characteristic of bone, including osteocalcin and bone sialoprotein. Implants were also stained with alizarin red and von Kossa, demonstrating mineralization and persistence of an osteoblast phenotype. Recruitment of vasculature and microvascularization of the implant was also detected. Taken together, these data demonstrate functional osteoblast differentiation from iPS cells both in vitro and in vivo and reveal a source of cells which merit evaluation for their potential uses in orthopaedic medicine and understanding of molecular mechanisms of orthopaedic disease.

EWS/FLI-1 is a chimeric oncogene generated by chromosomal translocation in Ewing tumors, a family of poorly differentiated pediatric tumors arising predominantly in bone but also in soft tissue. The fusion gene combines sequences encoding a strong transactivating domain from the EWS protein with the DNA binding domain of FLI-1, an ETS transcription factor. A related fusion, TLS/ERG, has been found in myeloid leukemia. To determine EWS/FLI-1 function in vivo, we engineered mice with Cre-inducible expression of EWS/FLI-1 from the ubiquitous Rosa26 locus. When crossed with Mx1-cre mice, Cre-mediated activation of EWS/FLI-1 resulted in the rapid development of myeloid/erythroid leukemia characterized by expansion of primitive mononuclear cells causing hepatomegaly, splenomegaly, severe anemia, and death. The disease could be transplanted serially into naïve recipients. Gene expression profiles of primary and transplanted animals were highly similar, suggesting that activation of EWS/FLI-1 was the primary event leading to disease in this model. The Cre-inducible EWS/FLI-1 mouse provides a novel model system to study the contribution of this oncogene to malignant disease in vivo.Specific chromosomal translocations are a common genetic mechanism in certain types of sarcomas and leukemias that create novel oncogenes presumed to play an important role in the initiation and/or progression of tumors that harbor them. Ewing tumors are defined at the molecular level by the presence of chromosomal translocations that produce chimeric oncogenes encoding the N-terminal transactivation domain of EWS fused to the C-terminal DNA binding domain from one of several ETS family genes (4). EWS belongs to a small family of RNA binding molecules termed TET proteins that includes TLS/FUS, EWS, and TAFII68 and appears to function in RNA transcription and/or mRNA processing (10,11,34,77). The ETS transcription factor family is a diverse group of proteins that cooperate with other factors to regulate a wide range of cellular processes, such as proliferation, differentiation, apoptosis, and senescence (52). EWS/FLI-1 is the most common TET/ETS fusion found in Ewing tumors, occurring in 85% of reported cases, while other fusions, such as EWS/ERG, EWS/FEV, EWS/ETV1, EWS/E1AF, and TLS/ERG, occur at much lower frequencies (31).Although Ewing tumors are extremely aggressive clinically, EWS/FLI-1 possesses only weak transforming activity in a traditional NIH 3T3 transformation assay (45). Moreover, its expression appears to be toxic to many primary cultured cells, suggesting that cellular context may be critical for the tumorigenic effects of EWS/FLI-1 (17). The cellular origin of Ewing tumors remains unknown, but recent studies suggest that this cell may be a progenitor associated with bone (13, 60, 71). In a previous study, we showed that expression of EWS/FLI-1 inhibits osteogenic and adipogenic differentiation of mesenchymal progenitors isolated from mouse bone marrow (71), implying that inappropriate expression of EWS/FLI-1 during embryogenesi...