RESUMEN
Although generating new neurons in the ischemic injured brain would be an ideal approach to replenish the lost neurons for repairing the damage, the adult mammalian brain retains only limited neurogenic capability. Here, we show that direct conversion of microglia/macrophages into neurons in the brain has great potential as a therapeutic strategy for ischemic brain injury. After transient middle cerebral artery occlusion in adult mice, microglia/macrophages converge at the lesion core of the striatum, where neuronal loss is prominent. Targeted expression of a neurogenic transcription factor, NeuroD1, in microglia/macrophages in the injured striatum enables their conversion into induced neuronal cells that functionally integrate into the existing neuronal circuits. Furthermore, NeuroD1-mediated induced neuronal cell generation significantly improves neurological function in the mouse stroke model, and ablation of these cells abolishes the gained functional recovery. Our findings thus demonstrate that neuronal conversion contributes directly to functional recovery after stroke.
Asunto(s)
Isquemia Encefálica , Accidente Cerebrovascular , Ratones , Animales , Microglía/metabolismo , Accidente Cerebrovascular/metabolismo , Macrófagos/metabolismo , Encéfalo/metabolismo , Neuronas/metabolismo , Isquemia Encefálica/metabolismo , Infarto de la Arteria Cerebral Media/metabolismo , MamíferosRESUMEN
Neuronal regeneration to replenish lost neurons after injury is critical for brain repair. Microglia, brain-resident macrophages that have the propensity to accumulate at the site of injury, can be a potential source for replenishing lost neurons through fate conversion into neurons, induced by forced expression of neuronal lineage-specific transcription factors. However, it has not been strictly demonstrated that microglia, rather than central nervous system-associated macrophages, such as meningeal macrophages, convert into neurons. Here, we show that NeuroD1-transduced microglia can be successfully converted into neurons in vitro using lineage-mapping strategies. We also found that a chemical cocktail treatment further promoted NeuroD1-induced microglia-to-neuron conversion. NeuroD1 with loss-of-function mutation, on the other hand, failed to induce the neuronal conversion. Our results indicate that microglia are indeed reprogrammed into neurons by NeuroD1 with neurogenic transcriptional activity.
Asunto(s)
Microglía , Neuronas , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Encéfalo/metabolismo , Microglía/metabolismo , Neurogénesis , Neuronas/metabolismo , Factores de Transcripción/metabolismo , Animales , RatonesRESUMEN
Several transcription factors, including NeuroD1, have been shown to act as neuronal reprogramming factors (RFs) that induce neuronal conversion from somatic cells. However, it remains unexplored whether expression levels of RFs in the original cells affect reprogramming efficiency. Here, we show that the neuronal reprogramming efficiency from two distinct glial cell types, microglia and astrocytes, is substantially dependent on the expression level of NeuroD1: low expression failed to induce neuronal reprogramming, whereas elevated NeuroD1 expression dramatically improved reprogramming efficiency in both cell types. Moreover, even under conditions where NeuroD1 expression was too low to induce effective conversion by itself, combined expression of three RFs (Ascl1, Brn2, and NeuroD1) facilitated the breaking down of cellular barriers, inducing neuronal reprogramming. Thus, our results suggest that a sufficiently high expression level of RFs, or alternatively their combinatorial expression, is the key to achieving efficient neuronal reprogramming from different cells.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Reprogramación Celular , Reprogramación Celular/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neuronas/metabolismo , Astrocitos/metabolismo , Factores de Transcripción/metabolismo , Neuroglía/metabolismoRESUMEN
Recent advances have enabled the direct induction of human tissue-specific stem and progenitor cells from differentiated somatic cells. However, it is not known whether human hepatic progenitor cells (hHepPCs) can be generated from other cell types by direct lineage reprogramming with defined transcription factors. Here, we show that a set of three transcription factors, FOXA3, HNF1A, and HNF6, can induce human umbilical vein endothelial cells to directly acquire the properties of hHepPCs. These induced hHepPCs (hiHepPCs) propagate in long-term monolayer culture and differentiate into functional hepatocytes and cholangiocytes by forming cell aggregates and cystic epithelial spheroids, respectively, under three-dimensional culture conditions. After transplantation, hiHepPC-derived hepatocytes and cholangiocytes reconstitute damaged liver tissues and support hepatic function. The defined transcription factors also induce hiHepPCs from endothelial cells circulating in adult human peripheral blood. These expandable and bipotential hiHepPCs may be useful in the study and treatment of human liver diseases.
Asunto(s)
Técnicas de Reprogramación Celular/métodos , Células Endoteliales/citología , Hepatocitos/citología , Células Madre/citología , Animales , Conductos Biliares/citología , Conductos Biliares/fisiología , Agregación Celular , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Células Cultivadas , Reprogramación Celular/genética , Reprogramación Celular/fisiología , Células Endoteliales/fisiología , Femenino , Factor Nuclear 1-alfa del Hepatocito/genética , Factor Nuclear 1-alfa del Hepatocito/fisiología , Factor Nuclear 3-gamma del Hepatocito/genética , Factor Nuclear 3-gamma del Hepatocito/fisiología , Factor Nuclear 6 del Hepatocito/genética , Factor Nuclear 6 del Hepatocito/fisiología , Hepatocitos/fisiología , Hepatocitos/trasplante , Xenoinjertos , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Esferoides Celulares/citología , Esferoides Celulares/fisiología , Células Madre/fisiologíaRESUMEN
Hepatic progenitor cells (HPCs) appear in response to several types of chronic injury in the human and rodent liver that often develop into liver fibrosis, cirrhosis, and primary liver cancers. However, the contribution of HPCs to the pathogenesis and progression of such liver diseases remains controversial. HPCs are generally defined as cells that can differentiate into hepatocytes and cholangiocytes. In this study, however, we found that HPCs isolated from the chronically injured liver can also give rise to myofibroblasts as a third type of descendant. While myofibroblast differentiation from HPCs is not significant in culture, during tumor development, HPCs can contribute to the formation of the tumor microenvironment by producing abundant myofibroblasts that might form a niche for tumor growth and survival. Thus, HPCs can be redefined as cells with a potential for differentiation into myofibroblasts that is specifically activated during tumor formation.
Asunto(s)
Hígado/citología , Miofibroblastos/citología , Células Madre/citología , Microambiente Tumoral , Animales , Recuento de Células , Diferenciación Celular , Separación Celular , Células Cultivadas , Células Clonales , Hígado/embriología , Ratones Endogámicos C57BL , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Intrahepatic cholangiocarcinoma (ICC) is a malignant epithelial neoplasm composed of cells resembling cholangiocytes that line the intrahepatic bile ducts in portal areas of the hepatic lobule. Although ICC has been defined as a tumor arising from cholangiocyte transformation, recent evidence from genetic lineage-tracing experiments has indicated that hepatocytes can be a cellular origin of ICC by directly changing their fate to that of biliary lineage cells. Notch signaling has been identified as an essential factor for hepatocyte conversion into biliary lineage cells at the onset of ICC. However, the mechanisms underlying Notch signal activation in hepatocytes remain unclear. Here, using a mouse model of ICC, we found that hepatic macrophages called Kupffer cells transiently congregate around the central veins in the liver and express the Notch ligand Jagged-1 coincident with Notch activation in pericentral hepatocytes. Depletion of Kupffer cells prevents the Notch-mediated cell-fate conversion of hepatocytes to biliary lineage cells, inducing hepatocyte apoptosis and increasing mortality in mice. These findings will be useful for uncovering the pathogenic mechanism of ICC and developing prevenient and therapeutic strategies for this refractory disease.