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1.
Development ; 138(23): 5067-78, 2011 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-22031545

RESUMEN

Successful brain development requires tight regulation of sequential symmetric and asymmetric cell division. Although Pax6 is known to exert multiple roles in the developing nervous system, its role in the regulation of cell division is unknown. Here, we demonstrate profound alterations in the orientation and mode of cell division in the cerebral cortex of mice deficient in Pax6 function (Pax6(Sey/Sey)) or after acute induced deletion of Pax6. Live imaging revealed an increase in non-vertical cellular cleavage planes, resulting in an increased number of progenitors with unequal inheritance of the apical membrane domain and adherens junctions in the absence of Pax6 function. This phenotype appears to be mediated by the direct Pax6 target Spag5, a microtubule-associated protein, reduced levels of which result in the replication of the Pax6 phenotype of altered cell division orientation. In addition, lack of Pax6 also results in premature delamination of progenitor cells from the apical surface due to an overall decrease in proteins mediating anchoring at the ventricular surface. Moreover, continuous long-term imaging in vitro revealed that Pax6-deficient progenitors generate daughter cells with asymmetric fates at higher frequencies. These data demonstrate a cell-autonomous role for Pax6 in regulating the mode of cell division independently of apicobasal polarity and cell-cell interactions. Taken together, our work reveals several direct effects that the transcription factor Pax6 has on the machinery that mediates the orientation and mode of cell division.


Asunto(s)
División Celular/fisiología , Polaridad Celular/fisiología , Corteza Cerebral/citología , Corteza Cerebral/embriología , Proteínas del Ojo/metabolismo , Proteínas de Homeodominio/metabolismo , Factores de Transcripción Paired Box/metabolismo , Proteínas Represoras/metabolismo , Células Madre/fisiología , Análisis de Varianza , Animales , Proteínas de Ciclo Celular , Inmunoprecipitación de Cromatina , Cartilla de ADN/genética , Electroporación , Femenino , Inmunohistoquímica , Luciferasas , Ratones , Factor de Transcripción PAX6 , Reacción en Cadena de la Polimerasa/métodos , Embarazo
2.
Cytometry A ; 79(8): 635-45, 2011 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-21735544

RESUMEN

Mesenchymal stromal cells (MSCs) do not express a unique definite epitope or marker gene. As such, minimal criteria were recently established for defining multipotent MSC. These criteria include expression of CD73, CD90, CD105, and a lack of hematopoietic marker expression. However, we detected binding of a CD14 antibody on bone marrow- and placenta-derived MSC and investigated the staining of CD14 antibodies on these MSC in more detail. The MSC were isolated from human bone marrow and placenta tissue, expanded, characterized by quantitative RT-PCR, flow cytometry, and immunocytochemistry and differentiated to generate osteoblasts, chondrocytes, and adipocytes. The CD14-cross-reactive MSCs were enriched by cell sorting. Human peripheral blood mononuclear cells, fibroblasts, and hematopoietic cell lines served as controls. Utilizing four different clones of CD14 monoclonal antibodies, we found that three CD14 reagents stained the MSC. Two CD14 antibodies (HCD14 and M5E2) clearly marked the CD90(+) MSC population with distinct intensities, clone 134 620 generated a shift in flow cytometry histograms, but clone MΦP9 did not stain MSC. Transcripts encoding CD14 or the CD14 protein were not detected in MSC. We confirm that bone marrow- and placenta-derived MSC do not express CD14 and that the CD14 antibody MΦP9 discriminates between monocytes and MSC more efficiently than the other antibodies employed here. This investigation does not contradict previous work but provides a more accurate characterization of MSC.


Asunto(s)
Anticuerpos/inmunología , Epítopos/inmunología , Receptores de Lipopolisacáridos/inmunología , Mesodermo/inmunología , Adipocitos/citología , Adipocitos/inmunología , Células de la Médula Ósea/citología , Células de la Médula Ósea/inmunología , Diferenciación Celular , Condrocitos/citología , Condrocitos/inmunología , Reacciones Cruzadas/inmunología , Epítopos/genética , Femenino , Citometría de Flujo , Humanos , Receptores de Lipopolisacáridos/genética , Receptores de Lipopolisacáridos/metabolismo , Mesodermo/citología , Monocitos/inmunología , Osteoblastos/citología , Osteoblastos/inmunología , Placenta/citología , Placenta/inmunología , Embarazo , Células del Estroma/citología , Células del Estroma/inmunología
3.
Stem Cells Dev ; 20(4): 635-46, 2011 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-21047215

RESUMEN

Mesenchymal stromal cells (MSC) can be isolated from different tissues. They are capable of differentiating in vitro, for example, to osteoblasts, chondrocytes, or adipocytes. In contrast to CD34 for hematopoietic stem cells, a distinct MSC-defining antibody is not available. Further, for hematopoietic cells lineage-defining antigens such as CD3 or CD20 are known. In contrast, for MSC-derived cells lineage-associated cell surface markers are far from being established. We therefore investigated expression of cell surface antigens on human term placenta-derived MSC (pMSC) in more detail and correlated expression pattern to the osteogenic differentiation capacity of the MSC. We report that pMSC expressed the typical cell surface antigens at levels comparable to bone marrow-derived MSC (bmMSC), including CD73, CD90, and CD105, but did not express CD11b, CD34, and CD45. Further, CD164, TNAP, and the W5C5 antigens were detected on pMSC, whereas CD349 was not observed. Some pMSC expressed CD146 at low or moderate levels, and their osteogenic differentiation potential was weak. In contrast, bmMSC expressed CD146 at high levels, expression of alkaline phosphatase was significantly higher, and they presented a pronounced osteogenic differentiation potential. We conclude that MSC from different sources differ in their expression of distinct markers, and that this may correlate in part with their lineage determination. Thus, a higher percentage of bmMSC expressed CD146 at prominent levels and such cells may be better suited for bone repair. In contrast, many pMSC expressed CD146 at low or moderate levels. They, therefore, may be suitable for applications in which osteogenic differentiation is undesirable.


Asunto(s)
Células de la Médula Ósea/citología , Células Madre Mesenquimatosas/citología , Osteoblastos/citología , Placenta/citología , Anciano , Fosfatasa Alcalina/metabolismo , Antígenos de Diferenciación/metabolismo , Células de la Médula Ósea/metabolismo , Antígeno CD146/metabolismo , Diferenciación Celular , Células Cultivadas , Pruebas de Enzimas , Femenino , Fibroblastos/metabolismo , Citometría de Flujo , Humanos , Masculino , Células Madre Mesenquimatosas/metabolismo , Persona de Mediana Edad , Osteogénesis , Fenotipo , Placenta/metabolismo , Embarazo , Células del Estroma/citología , Células del Estroma/metabolismo
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