技术资料

Fibrodysplasia ossificans progressiva (FOP) is a rare and intractable disease characterized by extraskeletal bone formation through endochondral ossification. Patients with FOP harbor point mutations in ACVR1,a type I receptor for BMPs. Although mutated ACVR1 (FOP-ACVR1) has been shown to render hyperactivity in BMP signaling,we and others have uncovered a mechanism by which FOP-ACVR1 mistransduces BMP signaling in response to Activin-A,a molecule that normally transduces TGF-β signaling. Although Activin-A evokes enhanced chondrogenesis in vitro and heterotopic ossification (HO) in vivo,the underlying mechanisms have yet to be revealed. To this end,we developed a high-throughput screening (HTS) system using FOP patient-derived induced pluripotent stem cells (FOP-iPSCs) to identify pivotal pathways in enhanced chondrogenesis that are initiated by Activin-A. In a screen of 6,809 small-molecule compounds,we identified mTOR signaling as a critical pathway for the aberrant chondrogenesis of mesenchymal stromal cells derived from FOP-iPSCs (FOP-iMSCs). Two different HO mouse models,an FOP model mouse expressing FOP-ACVR1 and an FOP-iPSC-based HO model mouse,revealed critical roles for mTOR signaling in vivo. Moreover,we identified ENPP2,an enzyme that generates lysophosphatidic acid,as a linker of FOP-ACVR1 and mTOR signaling in chondrogenesis. These results uncovered the crucial role of the Activin-A/FOP-ACVR1/ENPP2/mTOR axis in FOP pathogenesis. View Publication

This report describes the establishment directly from normal preimplantation mouse embryos of a cell line that forms teratocarcinomas when injected into mice. The pluripotency of these embryonic stem cells was demonstrated conclusively by the observation that subclonal cultures,derived from isolated single cells,can differentiate into a wide variety of cell types. Such embryonic stem cells were isolated from inner cell masses of late blastocysts cultured in medium conditioned by an established teratocarcinoma stem cell line. This suggests that such conditioned medium might contain a growth factor that stimulates the proliferation or inhibits the differentiation of normal pluripotent embryonic cells,or both. This method of obtaining embryonic stem cells makes feasible the isolation of pluripotent cells lines from various types of noninbred embryo,including those carrying mutant genes. The availability of such cell lines should made possible new approaches to the study of early mammalian development. View Publication

Under appropriate conditions in culture,embryonic stem cells will differentiate and form embryoid bodies that have been shown to contain cells of the hematopoietic,endothelial,muscle and neuronal lineages. Many aspects of the lineage-specific differentiation programs observed within the embryoid bodies reflect those found in the embryo,indicating that this model system provides access to early cell populations that develop in a normal fashion. Recent studies involving the differentiation of genetically altered embryonic stem cells highlight the potential of this in vitro differentiation system for defining the function of genes in early development. View Publication

The available techniques for directed gene manipulation in the mouse are unprecedented in any multicellular organism and make the mouse an invaluable tool for unraveling all aspects of mammalian biology. To realize fully the potential of these genetic tools requires that phenotypic analysis be efficient,rapid,and complete. Genetic chimeras and mosaics,in which mutant cells are mixed with wild-type cells,can be used to augment standard analysis of intact mutant animals and alleviate the time required and the expense involved in generating and maintaining multiple strains of mutant mice. View Publication