HMGB1 and HMGB2 proteins have been implicated in numerous cellular processes, including proliferation, differentiation, apoptosis, and tumor growth. It is unknown whether they are involved in regulating the typical functions of pluripotent human embryonic stem cells (hESCs) and/or those of the differentiated derivatives of hESCs. Using inducible, stably transfected hESCs capable of shRNA-mediated knockdown of HMGB1 and HMGB2, we provide evidence that downregulation of HMGB1 and/or HMGB2 in undifferentiated hESCs does not affect the stemness of cells and induces only minor changes to the proliferation rate, cell-cycle profile, and apoptosis. After differentiation is induced, however, the downregulation of those proteins has important effects on proliferation, apoptosis, telomerase activity, and the efficiency of differentiation toward the neuroectodermal lineage. Furthermore, those processes are affected only when one, but not both, of the two proteins is downregulated; the knockdown of both HMGB1 and HMGB2 results in a normal phenotype. Those results advance our knowledge of regulation of hESC and human neuroectodermal cell differentiation and illustrate the distinct roles of HMGB1 and HMGB2 during early human development.
- MeSH
- apoptóza genetika MeSH
- buněčná diferenciace * MeSH
- buněčná sebeobnova genetika MeSH
- buněčné linie MeSH
- buněčný cyklus genetika MeSH
- buněčný rodokmen genetika MeSH
- down regulace genetika MeSH
- histony metabolismus MeSH
- lidé MeSH
- lidské embryonální kmenové buňky cytologie metabolismus MeSH
- neurální ploténka cytologie MeSH
- proliferace buněk genetika MeSH
- protein HMGB1 metabolismus MeSH
- protein HMGB2 metabolismus MeSH
- telomerasa metabolismus MeSH
- transfekce MeSH
- tvar buňky genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Despite therapeutic advances, neurodegenerative diseases and disorders remain some of the leading causes of mortality and morbidity in the United States. Therefore, cell-based therapies to replace lost or damaged neurons and supporting cells of the central nervous system (CNS) are of great therapeutic interest. To that end, human pluripotent stem cell (hPSC) derived neural progenitor cells (hNPCs) and their neuronal derivatives could provide the cellular 'raw material' needed for regenerative medicine therapies for a variety of CNS disorders. In addition, hNPCs derived from patient-specific hPSCs could be used to elucidate the underlying mechanisms of neurodegenerative diseases and identify potential drug candidates. However, the scientific and clinical application of hNPCs requires the development of robust, defined, and scalable substrates for their long-term expansion and neuronal differentiation. In this study, we rationally designed a vitronectin-derived peptide (VDP) that served as an adhesive growth substrate for the long-term expansion of several hNPC lines. Moreover, VDP-coated surfaces allowed for the directed neuronal differentiation of hNPC at levels similar to cells differentiated on traditional extracellular matrix protein-based substrates. Overall, the ability of VDP to support the long-term expansion and directed neuronal differentiation of hNPCs will significantly advance the future translational application of these cells in treating injuries, disorders, and diseases of the CNS.
- MeSH
- biokompatibilní potahované materiály farmakologie MeSH
- buněčná adheze účinky léků MeSH
- buněčná diferenciace účinky léků MeSH
- extracelulární matrix - proteiny metabolismus MeSH
- lidé MeSH
- molekuly buněčné adheze metabolismus MeSH
- myši MeSH
- nervové kmenové buňky cytologie účinky léků metabolismus MeSH
- neurony cytologie účinky léků metabolismus MeSH
- peptidy farmakologie MeSH
- pluripotentní kmenové buňky cytologie účinky léků metabolismus MeSH
- proliferace buněk účinky léků MeSH
- vitronektin farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Cell cycle represents not only a tightly orchestrated mechanism of cell replication and cell division but it also plays an important role in regulation of cell fate decision. Particularly in the context of pluripotent stem cells or multipotent progenitor cells, regulation of cell fate decision is of paramount importance. It has been shown that human embryonic stem cells (hESCs) show unique cell cycle characteristics, such as short doubling time due to abbreviated G1 phase; these properties change with the onset of differentiation. This review summarizes the current understanding of cell cycle regulation in hESCs. We discuss cell cycle properties as well as regulatory machinery governing cell cycle progression of undifferentiated hESCs. Additionally, we provide evidence that long-term culture of hESCs is accompanied by changes in cell cycle properties as well as configuration of several cell cycle regulatory molecules.
- MeSH
- buněčná diferenciace MeSH
- buněčné kultury MeSH
- buněčný cyklus fyziologie MeSH
- embryonální kmenové buňky cytologie fyziologie MeSH
- kontrolní body buněčného cyklu fyziologie MeSH
- lidé MeSH
- pluripotentní kmenové buňky cytologie fyziologie MeSH
- proteiny buněčného cyklu metabolismus fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Embryonic stem cells progress very rapidly through the cell cycle, allowing limited time for cell cycle regulatory circuits that typically function in somatic cells. Mechanisms that inhibit cell cycle progression upon DNA damage are of particular importance, as their malfunction may contribute to the genetic instability observed in human embryonic stem cells (hESCs). In this study, we exposed undifferentiated hESCs to DNA-damaging ultraviolet radiation-C range (UVC) light and examined their progression through the G1/S transition. We show that hESCs irradiated in G1 phase undergo cell cycle arrest before DNA synthesis and exhibit decreased cyclin-dependent kinase two (CDK2) activity. We also show that the phosphatase Cdc25A, which directly activates CDK2, is downregulated in irradiated hESCs through the action of the checkpoint kinases Chk1 and/or Chk2. Importantly, the classical effector of the p53-mediated pathway, protein p21, is not a regulator of G1/S progression in hESCs. Taken together, our data demonstrate that cultured undifferentiated hESCs are capable of preventing entry into S-phase by activating the G1/S checkpoint upon damage to their genetic complement.
- MeSH
- buněčná diferenciace MeSH
- buněčné linie MeSH
- cyklin-dependentní kinasa 2 metabolismus MeSH
- fosfatasy cdc25 metabolismus MeSH
- G1 fáze účinky záření MeSH
- kmenové buňky cytologie metabolismus účinky záření MeSH
- lidé MeSH
- poškození DNA MeSH
- protein-serin-threoninkinasy metabolismus MeSH
- proteinkinasy metabolismus MeSH
- S fáze účinky záření MeSH
- signální transdukce MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
