Publications (3)23.34 Total impact
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Article: An important role for CDK2 in G1 to S checkpoint activation and DNA damage response in human embryonic stem cells.
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ABSTRACT: A precise understanding of mechanisms used by human embryonic stem cells (hESCs) to maintain genomic integrity is very important for their potential clinical applications. The G1 checkpoint serves to protect genomic integrity and prevents cells with damaged DNA from entering S-phase. Previously, we have shown that downregulation of cyclin-dependent kinase 2 (CDK2) in hESC causes G1 arrest, loss of pluripotency, upregulation of cell cycle inhibitors p21 and p27 and differentiation toward extraembryonic lineages. In this study, we investigate in detail the role of CDK2 in cellular processes, which are crucial to the maintenance of genomic stability in hESC such as G1 checkpoint activation, DNA repair, and apoptosis. Our results suggest that downregulation of CDK2 triggers the G1 checkpoint through the activation of the ATM-CHK2-p53-p21 pathway. Downregulation of CDK2 is able to induce sustained DNA damage and to elicit the DNA damage response (DDR) as evidenced by the formation of distinct γ-H2.AX and RAD52-BRCA1 foci in hESC nuclei. CDK2 downregulation causes high apoptosis at the early time points; however, this is gradually decreased overtime as the DDR is initiated. Our mass spectrometry analysis suggest that CDK2 does interact with a large number of proteins that are involved in key cellular processes such as DNA replication, cell cycle progression, DNA repair, chromatin modeling, thus, suggesting a crucial role for CDK2 in orchestrating a fine balance between cellular proliferation, cell death, and DNA repair in hESC.Stem Cells 02/2011; 29(4):651-9. · 7.78 Impact Factor -
Article: Opposing putative roles for canonical and noncanonical NFκB signaling on the survival, proliferation, and differentiation potential of human embryonic stem cells.
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ABSTRACT: The canonical and noncanonical NFκB signaling pathways regulate a variety of cellular activities; however, their functions in human embryonic stem cells (hESCs) have not been fully investigated. Expression studies during hESC differentiation indicated a significant increase in the expression of two key components of the canonical NFκB pathway (p50 and Ser529 phosphorylated form of p65) as well as a significant reduction in expression of key components of the noncanonical NFκB pathway [v-rel reticuloendotheliosis viral oncogene homolog B (RELB), p52, NIK]. Inhibition of canonical NFκB resulted in hESC apoptosis, changes in cell cycle distribution, and reduced hESC proliferation. In addition, inhibition of canonical NFκB was associated with significant changes in NANOG and OCT4 expression, suppression of differentiation toward all primitive extraembryonic and embryonic lineages with the exception of primitive ectoderm and ectodermal lineages. Inhibition of noncanonical NFκB via small interfering RNA-mediated downregulation of RELB resulted in reduced hESC proliferation and opposite changes to expression of key differentiation lineage markers genes when compared with downregulation of canonical NF-κB. Chromatin immunoprecipitation assays indicated binding of p65 and RELB to regulatory regions of key differentiation marker genes suggesting a direct transcriptional role for both branches of this pathway in hESC. These findings coupled with opposing trends in expression of key components during hESC differentiation, suggests a fine and opposing balance between the two branches of NFκB signaling pathways and their involvement in two distinct processes: the canonical pathway regulating hESC differentiation and the noncanonical pathway maintaining hESC pluripotency.Stem Cells 09/2010; 28(11):1970-80. · 7.78 Impact Factor -
Article: Activated spinal cord ependymal stem cells rescue neurological function.
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ABSTRACT: Spinal cord injury (SCI) is a major cause of paralysis. Currently, there are no effective therapies to reverse this disabling condition. The presence of ependymal stem/progenitor cells (epSPCs) in the adult spinal cord suggests that endogenous stem cell-associated mechanisms might be exploited to repair spinal cord lesions. epSPC cells that proliferate after SCI are recruited by the injured zone, and can be modulated by innate and adaptive immune responses. Here we demonstrate that when epSPCs are cultured from rats with a SCI (ependymal stem/progenitor cells injury [epSPCi]), these cells proliferate 10 times faster in vitro than epSPC derived from control animals and display enhanced self renewal. Genetic profile analysis revealed an important influence of inflammation on signaling pathways in epSPCi after injury, including the upregulation of Jak/Stat and mitogen activated protein kinase pathways. Although neurospheres derived from either epSPCs or epSPCi differentiated efficiently to oligodendrocites and functional spinal motoneurons, a better yield of differentiated cells was consistently obtained from epSPCi cultures. Acute transplantation of undifferentiated epSPCi or the resulting oligodendrocyte precursor cells into a rat model of severe spinal cord contusion produced a significant recovery of motor activity 1 week after injury. These transplanted cells migrated long distances from the rostral and caudal regions of the transplant to the neurofilament-labeled axons in and around the lesion zone. Our findings demonstrate that modulation of endogenous epSPCs represents a viable cell-based strategy for restoring neuronal dysfunction in patients with spinal cord damage.Stem Cells 02/2009; 27(3):733-43. · 7.78 Impact Factor
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- Stem Cells (3)
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2011
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Centro de Investigación Príncipe Felipe
Valencia, Valencia, Spain
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