RESUMEN
Heterogeneous populations of human bone marrow-derived stromal cells (BMSC) are among the most frequently tested cellular therapeutics for treating degenerative and immune disorders, which occur predominantly in the aging population. Currently, it is unclear whether advanced donor age and commonly associated comorbidities affect the properties of ex vivo-expanded BMSCs. Thus, we stratified cells from adult and elderly donors from our biobank (n = 10 and n = 13, mean age 38 and 72 years, respectively) and compared their phenotypic and functional performance, using multiple assays typically employed as minimal criteria for defining multipotent mesenchymal stromal cells (MSCs). We found that BMSCs from both cohorts meet the standard criteria for MSC, exhibiting similar morphology, growth kinetics, gene expression profiles, and pro-angiogenic and immunosuppressive potential and the capacity to differentiate toward adipogenic, chondrogenic, and osteogenic lineages. We found no substantial differences between cells from the adult and elderly cohorts. As positive controls, we studied the impact of in vitro aging and inflammatory cytokine stimulation. Both conditions clearly affected the cellular properties, independent of donor age. We conclude that in vitro aging rather than in vivo donor aging influences BMSC characteristics.
Asunto(s)
Células Madre Adultas/citología , Células Madre Adultas/fisiología , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/fisiología , Adipogénesis , Adulto , Células Madre Adultas/inmunología , Anciano , Envejecimiento/inmunología , Envejecimiento/patología , Envejecimiento/fisiología , Bancos de Muestras Biológicas , Diferenciación Celular , Linaje de la Célula , Proliferación Celular , Células Cultivadas , Senescencia Celular/inmunología , Senescencia Celular/fisiología , Condrogénesis , Comorbilidad , Humanos , Inmunofenotipificación , Células Madre Mesenquimatosas/inmunología , Osteogénesis , Fenotipo , Donantes de Tejidos , TranscriptomaRESUMEN
Reprogramming to induced pluripotent stem cells (iPSCs) and differentiation of pluripotent stem cells (PSCs) are regulated by epigenetic machinery. Tripartite motif protein 28 (TRIM28), a universal mediator of Krüppel-associated box domain zinc fingers (KRAB-ZNFs), is known to regulate both processes; however, the exact mechanism and identity of participating KRAB-ZNF genes remain unknown. Here, using a reporter system, we show that TRIM28/KRAB-ZNFs alter DNA methylation patterns in addition to H3K9me3 to cause stable gene repression during reprogramming. Using several expression datasets, we identified KRAB-ZNFs (ZNF114, ZNF483, ZNF589) in the human genome that maintain pluripotency. Moreover, we identified target genes repressed by these KRAB-ZNFs. Mechanistically, we demonstrated that these KRAB-ZNFs directly alter gene expression of important developmental genes by modulating H3K9me3 and DNA methylation of their promoters. In summary, TRIM28 employs KRAB-ZNFs to evoke epigenetic silencing of its target differentiation genes via H3K9me3 and DNA methylation.
Asunto(s)
Diferenciación Celular/genética , Células Madre Pluripotentes/metabolismo , Proteínas Represoras/genética , Proteína 28 que Contiene Motivos Tripartito/genética , Sitios de Unión , Autorrenovación de las Células/genética , Reprogramación Celular/genética , Metilación de ADN/genética , Represión Epigenética , Regulación del Desarrollo de la Expresión Génica/genética , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Células Madre Pluripotentes/citología , Regiones Promotoras GenéticasRESUMEN
Reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) requires profound alterations in the epigenetic landscape. During reprogramming, a change in chromatin structure resets the gene expression and stabilises self-renewal. Reprogramming is a highly inefficient process, in part due to multiple epigenetic barriers. Although many epigenetic factors have already been shown to affect self-renewal and pluripotency in embryonic stem cells (ESCs), only a few of them have been examined in the context of dedifferentiation. In order to improve current protocols of iPSCs generation, it is essential to identify epigenetic drivers and blockages of somatic cell reprogramming.
