RESUMEN
Recent developments indicate that the regeneration of beta cell function and mass in patients with diabetes is possible. A regenerative approach may represent an alternative treatment option relative to current diabetes therapies that fail to provide optimal glycemic control. Here we report that the inactivation of GSK3 by small molecule inhibitors or RNA interference stimulates replication of INS-1E rat insulinoma cells. Specific and potent GSK3 inhibitors also alleviate the toxic effects of high concentrations of glucose and the saturated fatty acid palmitate on INS-1E cells. Furthermore, treatment of isolated rat islets with structurally diverse small molecule GSK3 inhibitors increases the rate beta cell replication by 2-3-fold relative to controls. We propose that GSK3 is a regulator of beta cell replication and survival. Moreover, our results suggest that specific inhibitors of GSK3 may have practical applications in beta cell regenerative therapies.
Asunto(s)
Glucógeno Sintasa Quinasa 3/fisiología , Animales , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Fragmentación del ADN , Inhibidores Enzimáticos/farmacología , Ácidos Grasos/química , Regulación de la Expresión Génica , Silenciador del Gen , Glucosa/metabolismo , Glucógeno Sintasa Quinasa 3/antagonistas & inhibidores , Células Secretoras de Insulina/metabolismo , Palmitatos/química , Interferencia de ARN , RatasRESUMEN
Modifications of DNA and chromatin are fundamental for the establishment and maintenance of cell type-specific gene expression patterns that constitute cellular identities. To test whether the developmental potential of fetal brain-derived cells that form floating sphere colonies (neurospheres) can be modified by destabilizing their epigenotype, neurosphere cells were treated with chemical compounds that alter the acetylation and methylation patterns of chromatin and DNA. Intravenous infusion of bulk or clonally derived neurosphere cells treated with a combination of trichostatin A (TSA) plus 5-aza-2'-deoxycytidine (AzaC) (TSA/AzaC neurosphere cells) yielded long-term, multilineage and transplantable neurosphere-derived haematopoietic repopulation. Untreated neurosphere cells exhibited no haematopoietic repopulation activity. The neurosphere-derived haematopoietic cells showed a diploid karyotype, indicating that they are unlikely to be products of cell fusion events, a conclusion strengthened by multicolour fluorescence in situ hybridization. Our results indicate that altering the epigenotype of neurosphere cells followed by transplantation enables the generation of neurosphere-derived haematopoietic cells.
Asunto(s)
Azacitidina/análogos & derivados , Azacitidina/farmacología , Cromatina/efectos de los fármacos , Hematopoyesis/efectos de los fármacos , Ácidos Hidroxámicos/farmacología , Animales , Decitabina , Diploidia , Genes bcl-2 , Proteínas Fluorescentes Verdes/genética , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , Técnicas In Vitro , Cariotipificación , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Células Madre Multipotentes/citología , Células Madre Multipotentes/efectos de los fármacos , Neuronas/citología , Neuronas/efectos de los fármacos , Esferoides CelularesRESUMEN
OBJECTIVE: In contrast to embryonic stem (ES) cells, which are able to give rise to all cell types of the body, somatic stem cells have been thought to be more limited in their differentiation potential in that they are committed to generate only cells of their tissue of origin. Unexpectedly, some recent data suggest that somatic stem cells isolated from one tissue can also generate cells of heterologous tissues and organs, implying that somatic stem cells have a greater potential for differentiation. METHODS: To explore further the developmental potential of murine neural stem cells (NSCs) we injected cultured NSCs as neurospheres into preimplantation blastocysts and determined the seeding by donor cells in tissues of developing chimeric fetal and adult animals. RESULTS: We frequently detected progeny of injected NSCs both in embryos and in adult animals. In embryos we observed transient seeding of donor cells to hematopoietic tissues and generation of NSC-derived cells that express globin genes and an erythroid-specific cell-surface marker. In adults progeny of NSCs were mostly detected in neural tissues. The observed low level of chimerism of wild-type NSCs was increased if we injected stem cells expressing a bcl-2 transgene, without changing the seeding pattern. CONCLUSION: These results suggest that cultured NSCs, following their injection into blastocysts, generate at mid-gestation erythroid-like cells but later, in adult chimeric mice, engraftment mainly persisted in neural tissues.
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Blastocisto/citología , Corteza Cerebral/inmunología , Células Madre Hematopoyéticas/citología , Células Madre/citología , Animales , Blastocisto/fisiología , Desarrollo Embrionario y Fetal , Regulación del Desarrollo de la Expresión Génica , Hígado/citología , Hígado/embriología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Reacción en Cadena de la Polimerasa , Proteínas Proto-Oncogénicas c-bcl-2/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Trasplante de Células Madre , Quimera por TrasplanteRESUMEN
It has been suggested that the embryonic microenvironment can control the survival and the transformed phenotype of tumour cells. Here, we addressed the hypothesis that the murine embryonic microenvironment can induce the differentiation of human tumour cells. To examine such interactions, we injected human leukaemic cells into preimplantation murine blastocysts at embryonic day 3.5 of gestation (E3.5). Microinjection of human KG-1 myeloid leukaemia cells and primary human acute myeloid leukaemia (AML) cells led to the generation of chimaeric embryos and adults. We observed that in E12.5 murine embryos, KG-1 cells were preferentially detected in yolk sac and peripheral blood, while primary AML cells mainly seeded the aorta gonad mesonephros region of chimaeric embryos. Analysis of the donor contribution in 15 different adult tissues showed that progeny of primary AML cells seeded to various haematopoietic and nonhaematopoietic tissues. Chimaeric embryos and adults showed no apparent tumour formation. Furthermore, analysis of chimaeric E12.5 embryos revealed that the progeny of human KG-1 cells activated erythroid-specific human globin and glycophorin A expression. In summary, our data indicate that human AML cells activate markers of erythroid differentiation after injection into early murine embryos.
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Blastocisto/fisiología , Leucemia Mieloide Aguda/patología , Quimera por Trasplante/fisiología , Trasplante Heterólogo/fisiología , Animales , Biomarcadores/análisis , Diferenciación Celular , Edad Gestacional , Glicoforinas/análisis , Hematopoyesis , Humanos , Riñón/embriología , Leucemia Mieloide Aguda/fisiopatología , Ratones , Microinyecciones , Células Tumorales CultivadasRESUMEN
Stem cell systems represent an effective and powerful approach for tissue development and regeneration of diverse tissue types. Common and defining features of these exceptional cells are the capacity for self-renewal and the potential for differentiation into multiple mature cell types. Recently, surprising new observations have indicated that stem cells isolated from one adult tissue can also give rise to mature cells of other cell lineages, irrespective of classical germ layer designations. This discovery has resulted in quantum leaps in both scientific knowledge and the potential applications of stem cells. The new findings contradict central dogmas of commitment and differentiation of stem and progenitor cells. However, the true potential of somatic stem cells is just emerging and the new findings have to be defined more fully and integrated into a unifying model of stem cell potential and behavior. Here we analyze the developmental potential of hematopoietic stem cells of mouse and man following their injection into the murine preimplantation blastocyst, an environment that allows the development of all cell lineages. In addition, we discuss the emerging lines of evidence of the developmental plasticity of hematopoietic and other somatic stem cells and consider how cellular memory of transcriptional states is established and may be potentially involved in this phenomenon.
Asunto(s)
Diferenciación Celular , Células Madre Hematopoyéticas/citología , Animales , Linaje de la Célula , Embrión de Mamíferos/citología , Regulación del Desarrollo de la Expresión Génica , Trasplante de Células Madre Hematopoyéticas , Humanos , Transducción de SeñalRESUMEN
Like many other animals, mammals develop from fertilized oocytes - the ultimate stem cells. As embryogenesis proceeds, most cells lose developmental potential and eventually become restricted to a specific cell lineage. The result is the formation of a complete and structured mature organism with complex organs composed of a great variety of mature, mostly mitotically quiescent effector cells. However, along the way, some exceptional cells, known as somatic stem cells (SSCs) are set aside and maintain a high proliferation and tissue-specific differentiation potential. SSCs, in contrast to embryonic stem (ES) cells, which are able to give rise to all cell types of the body, have been regarded as being more limited in their differentiation potential in the sense that they were thought to be committed exclusively to their tissue of origin. However, recent studies have demonstrated that somatic stem cells from a given tissue can also contribute to heterologous tissues and thus show a broad nontissue restricted differentiation potential. The question arises: how plastic are somatic stem cells? To provide a tentative answer, we describe and review here recent investigations into the developmental potentials of two somatic stem cell types, namely hematopoietic and neural stem cells.
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Diferenciación Celular/fisiología , Células Madre Hematopoyéticas/fisiología , Neuronas/fisiología , Animales , Biomarcadores , Linaje de la Célula , Separación Celular , Embrión de Mamíferos/anatomía & histología , Embrión de Mamíferos/fisiología , Ratones , Ratones TransgénicosRESUMEN
At different developmental stages, candidate human hematopoietic stem cells (HSCs) are present within the CD34+ CD38- population. By means of xenotransplantation, such CD34+CD38- cells were recently shown to engraft the hematopoietic system of fetal sheep and nonobese diabetic severe combined immunodeficient adult mice. Here it is demonstrated that, after their injection into murine blastocysts, human cord blood (CB)-derived CD34+ and CD34+ CD38- cells repopulate the hematopoietic tissues of nonimmunocompromised murine embryos and that human donor contribution can persist to adulthood. It is further observed that human hematopoietic progenitor cells are present in murine hematopoietic tissues of midgestational chimeric embryos and that progeny of the injected human HSCs activate erythroid-specific gene expression. Thus, the early murine embryo provides a suitable environment for the survival and differentiation of human CB CD34+ CD38- cells.