Project description:Kidney development depends on a balance of nephron progenitor self-renewal and commitment. Here we examine how cells shift from a motile progenitor state to a static committed state during nephrogenesis. Cells that express Wnt4, an early marker of commitment, give rise to both the nephron lineage and a migrating sub-population that may re-enter the progenitor pool. Single cell transcriptional profiling reveals Wnt4-lineage cells across a range of transcriptional states, including cells that return to an uninduced progenitor state.
Project description:MCMV has been reported to infect neural stem cells in the subventricular zone. These same cells have been shown to give rise to gliomas. We wanted to test expression levels of RNA in this cell population after MCMV infection in a mouse model of glioma.
Project description:Limited stem cell migration capacities and undirected homing hamper their therapeutic efficacy. We report here the novel technology to isolate stem cell subpopulations with high migration potential. Hereby, we identified podoplanin-dependent stem cell mobility as important mechanism for their in vivo homing capacities and therapeutic efficacy as validated in preclinical models of Alzheimer´s disease and mouse glioma. In vivo applications of highly migrating stem cells resulted in remarkable improvement of targeted homing and engraftment rates. Moreover, animals receiving highly migrating stem cells showed higher recovery rates compared to unselected stem cells. This novel technology provides useful tool to improve therapeutic efficacy of stem cells and give insights into stem cell subpopulations heterogeneity with the potential of general applicability for cellular therapies.
Project description:Neural stem cells can migrate towards tumors of both neural and non-neural origins, which is crucial for the success in treating disseminated tumors. Although the understanding of the molecular mechanisms underlying NSC tumor tropism is limited, it has been noted that several cytokines, growth factors and receptors direct the migration in vitro. A proper understanding of the basic molecular mechanisms of NSC migration towards tumors, especially identification of key cellular regulators of the migration, will have important implications in improving the effectiveness of engineering and employing NSCs as tumor therapy agents. We compared gene expression profiles between migratory and non-migratory hiPSC-NSCs towards cancer cells using cDNA microarray profiling. We collected human iPSCs derived NSCs migrating and not migrating towards human U87 glioma cells in an in vitro migration system for total RNA extraction and hybridization to Affymetrix microarrays
Project description:Glioblastoma multiforme is the most common and most aggressive type of primary brain tumor. The brain-infiltrative character of glioblastoma makes complete surgical removal of the tumor impossible and neither radiation nor current chemotherapy provide cure. Recent evidence shows that glioblastoma multiforme consists of heterogeneous cell populations which differ in tumor-forming potential. Enriched tumor-initiating capacity has been linked to poorly differentiated glioblastoma cells sharing features with neural stem cells. Thus, these cells are important targets for new therapeutic strategies. We aim to identify novel targets controlling maintenance and differentiation in glioblastoma-initiating cells through high throughput screening. To this end, we utilized libraries of small chemical compounds and small interference RNAs in combination with automated imaging and data analysis. Patient-derived glioblastoma cells were expanded and characterized using neural stem cell conditions. In culture, the cells showed low differentiation but expression of neural stem cell markers such as Nestin and Sox2. Upon intracranial injection into SCID mice these cells gave rise to tumors displaying the hallmarks of the human disease. Differentiation of glioblastoma-initiating cells (for example elicited through bone morphogenetic protein, BMP) was associated with strong morphological changes. Hence, cellular morphology, as well as markers specific for differentiation or death were used as screen readout. Lentiviral RNA interference-based screening yielded several gene knockdowns leading to ‘forced’ differentiation of glioblastoma-initiating cells. For example, knockdown of TRRAP (transformation/transcription domain associated protein) led to strongly increased differentiation and loss of proliferative and self-renewing capacity in these cells. TRRAP is an adapter protein implicated in oncogenic transformation through c-MYC transcription activation, also participating in chromatin remodeling and DNA repair. Glioblastoma-initiating cells with reduced TRRAP displayed increased apoptosis upon treatment with the genotoxic agent temozolomide. In vivo, Trapp knockdown cells were not able to give rise to glioblastoma upon transplantation into the brain of SCID mice. Together, these findings support a crucial role for TRRAP in maintenance and tumorigenicity of glioblastoma-initiating cells and might offer future therapeutic options. Two treatments compared to control: two different shRNA sequences for TRRAP were compared to a control shRNA sequence in their effects on global transcription in brain tumor initiating cells
Project description:Hematopoietic stem cells give rise to all blood lineages, can fully re-populate the bone marrow, and easily outlive the host organism. To better understand how stem cells remain fit during aging, we analyzed the proteome of hematopoietic stem and progenitor cells.
Project description:Skeletal muscle harbors quiescent stem cells termed satellite cells and proliferative progenitors termed myoblasts, which play pivotal roles during muscle regeneration. However, current technology does not allow permanent capture of these cell populations in vitro. Here, we show that ectopic expression of the myogenic transcription factor MyoD, combined with exposure to small molecules, reprograms mouse fibroblasts into expandable induced myogenic progenitor cells (iMPCs). iMPCs express key skeletal muscle stem and progenitor cell markers including Pax7 and Myf5 and give rise to Dystrophin-expressing myofibers upon transplantation, a subset of which maintain Pax7 expression in vivo and sustain serial regenerative responses. Similar to satellite cells, iMPCs originate from Pax7+ cells and require Pax7 itself for maintenance. Finally, we show that iMPCs can be established from muscle tissue following small molecule exposure alone. This study thus reports on a robust approach to derive expandable myogenic stem/progenitor-like cells from multiple differentiated cell types.
Project description:Skeletal muscle harbors quiescent stem cells termed satellite cells and proliferative progenitors termed myoblasts, which play pivotal roles during muscle regeneration. However, current technology does not allow permanent capture of these cell populations in vitro. Here, we show that ectopic expression of the myogenic transcription factor MyoD, combined with exposure to small molecules, reprograms mouse fibroblasts into expandable induced myogenic progenitor cells (iMPCs). iMPCs express key skeletal muscle stem and progenitor cell markers including Pax7 and Myf5 and give rise to Dystrophin-expressing myofibers upon transplantation, a subset of which maintain Pax7 expression in vivo and sustain serial regenerative responses. Similar to satellite cells, iMPCs originate from Pax7+ cells and require Pax7 itself for maintenance. Finally, we show that iMPCs can be established from muscle tissue following small molecule exposure alone. This study thus reports on a robust approach to derive expandable myogenic stem/progenitor-like cells from multiple differentiated cell types.
Project description:Germline-competent embryonic stem cells (ESCs) have been derived from mice and rats, but not from other species. Here, we report the development of conditions for the derivation of ESCs from chickens and several other avian species. Chicken ESCs express core pluripotency markers and can efficiently differentiate into cells of all three germ layers. Furthermore, chicken ESCs can form high rates of chimerism when injected into chicken embryos both ex ovo and in ovo. More importantly, chicken ESCs can give rise to germ cells both in vitro and in ovo, indicating that chicken ESCs are germline competent. Interspecies chimeras can also be generated by injecting other avian ESCs into chicken embryos. Establishment of germline competent avian ESCs opens up a new avenue for producing genetically engineered avian species.
Project description:Chimeric and germline-competent embryonic stem cells (ESCs) have been derived from mice and rats, but not from other species. Here, we report the development of conditions for the derivation of ESCs from chickens and several other avian species. Chicken ESCs express core pluripotency markers and can efficiently differentiate into cells of all three germ layers. Furthermore, chicken ESCs can form high rates of chimerism when injected into chicken embryos both ex ovo and in ovo. More importantly, chicken ESCs can give rise to germ cells both in vitro and in ovo, indicating that chicken ESCs are germline competent. Interspecies chimeras can also be generated by injecting other avian ESCs into chicken embryos. Establishment of germline competent avian ESCs opens up a new avenue for producing genetically engineered avian species.