RESUMEN
Hematopoietic stem cells (HSCs) and downstream progenitors have long been studied based on phenotype, cell purification, proliferation, and transplantation into myeloablated recipients. These experiments, complemented by data on expression profiles, mouse mutants, and humans with hematopoietic defects, are the foundation for the current hematopoietic differentiation tree. However, there are fundamental gaps in our knowledge of the quantitative and qualitative operation of the HSC/progenitor system under physiological and pathological conditions in vivo. The hallmarks of HSCs, self-renewal and multipotency, are observed in in vitro assays and cell transplantation experiments; however, the extent to which these features occur naturally in HSCs and progenitors remains uncertain. We focus here on work that strives to address these unresolved questions, with emphasis on fate mapping and modeling of the hematopoietic flow from stem cells toward myeloid and lymphoid lineages during development and adult life.
Asunto(s)
Envejecimiento/inmunología , Diferenciación Celular , Hematopoyesis , Células Madre Hematopoyéticas/fisiología , Células Progenitoras Linfoides/fisiología , Animales , Linaje de la Célula , Autorrenovación de las Células , Humanos , Ratones , Modelos Teóricos , TranscriptomaRESUMEN
Developmental deconvolution of complex organs and tissues at the level of individual cells remains challenging. Non-invasive genetic fate mapping has been widely used, but the low number of distinct fluorescent marker proteins limits its resolution. Much higher numbers of cell markers have been generated using viral integration sites, viral barcodes, and strategies based on transposons and CRISPR-Cas9 genome editing; however, temporal and tissue-specific induction of barcodes in situ has not been achieved. Here we report the development of an artificial DNA recombination locus (termed Polylox) that enables broadly applicable endogenous barcoding based on the Cre-loxP recombination system. Polylox recombination in situ reaches a practical diversity of several hundred thousand barcodes, allowing tagging of single cells. We have used this experimental system, combined with fate mapping, to assess haematopoietic stem cell (HSC) fates in vivo. Classical models of haematopoietic lineage specification assume a tree with few major branches. More recently, driven in part by the development of more efficient single-cell assays and improved transplantation efficiencies, different models have been proposed, in which unilineage priming may occur in mice and humans at the level of HSCs. We have introduced barcodes into HSC progenitors in embryonic mice, and found that the adult HSC compartment is a mosaic of embryo-derived HSC clones, some of which are unexpectedly large. Most HSC clones gave rise to multilineage or oligolineage fates, arguing against unilineage priming, and suggesting coherent usage of the potential of cells in a clone. The spreading of barcodes, both after induction in embryos and in adult mice, revealed a basic split between common myeloid-erythroid development and common lymphocyte development, supporting the long-held but contested view of a tree-like haematopoietic structure.
Asunto(s)
Sitios de Ligazón Microbiológica/genética , Linaje de la Célula/genética , Rastreo Celular/métodos , Código de Barras del ADN Taxonómico/métodos , Células Madre Hematopoyéticas/citología , Recombinación Genética/genética , Análisis de la Célula Individual/métodos , Animales , Células Clonales/citología , Células Clonales/metabolismo , Embrión de Mamíferos/citología , Células Eritroides/citología , Células Eritroides/metabolismo , Femenino , Células Madre Hematopoyéticas/metabolismo , Integrasas/metabolismo , Linfocitos/citología , Linfocitos/metabolismo , Masculino , Ratones , Mosaicismo , Células Mieloides/citología , Células Mieloides/metabolismoRESUMEN
Haematopoietic stem cells (HSCs) are widely studied by HSC transplantation into immune- and blood-cell-depleted recipients. Single HSCs can rebuild the system after transplantation. Chromosomal marking, viral integration and barcoding of transplanted HSCs suggest that very low numbers of HSCs perpetuate a continuous stream of differentiating cells. However, the numbers of productive HSCs during normal haematopoiesis, and the flux of differentiating progeny remain unknown. Here we devise a mouse model allowing inducible genetic labelling of the most primitive Tie2(+) HSCs in bone marrow, and quantify label progression along haematopoietic development by limiting dilution analysis and data-driven modelling. During maintenance of the haematopoietic system, at least 30% or â¼5,000 HSCs are productive in the adult mouse after label induction. However, the time to approach equilibrium between labelled HSCs and their progeny is surprisingly long, a time scale that would exceed the mouse's life. Indeed, we find that adult haematopoiesis is largely sustained by previously designated 'short-term' stem cells downstream of HSCs that nearly fully self-renew, and receive rare but polyclonal HSC input. By contrast, in fetal and early postnatal life, HSCs are rapidly used to establish the immune and blood system. In the adult mouse, 5-fluoruracil-induced leukopenia enhances the output of HSCs and of downstream compartments, thus accelerating haematopoietic flux. Label tracing also identifies a strong lineage bias in adult mice, with several-hundred-fold larger myeloid than lymphoid output, which is only marginally accentuated with age. Finally, we show that transplantation imposes severe constraints on HSC engraftment, consistent with the previously observed oligoclonal HSC activity under these conditions. Thus, we uncover fundamental differences between the normal maintenance of the haematopoietic system, its regulation by challenge, and its re-establishment after transplantation. HSC fate mapping and its linked modelling provide a quantitative framework for studying in situ the regulation of haematopoiesis in health and disease.
Asunto(s)
Linaje de la Célula/fisiología , Hematopoyesis , Células Madre Hematopoyéticas/citología , Células Madre/citología , Envejecimiento , Animales , Animales Recién Nacidos , Trasplante de Médula Ósea , Proliferación Celular , Rastreo Celular , Femenino , Feto/citología , Feto/embriología , Fluorouracilo , Células Madre Hematopoyéticas/metabolismo , Masculino , Ratones , Receptor TIE-2/metabolismo , Células Madre/metabolismoRESUMEN
Most haematopoietic cells renew from adult haematopoietic stem cells (HSCs), however, macrophages in adult tissues can self-maintain independently of HSCs. Progenitors with macrophage potential in vitro have been described in the yolk sac before emergence of HSCs, and fetal macrophages can develop independently of Myb, a transcription factor required for HSC, and can persist in adult tissues. Nevertheless, the origin of adult macrophages and the qualitative and quantitative contributions of HSC and putative non-HSC-derived progenitors are still unclear. Here we show in mice that the vast majority of adult tissue-resident macrophages in liver (Kupffer cells), brain (microglia), epidermis (Langerhans cells) and lung (alveolar macrophages) originate from a Tie2(+) (also known as Tek) cellular pathway generating Csf1r(+) erythro-myeloid progenitors (EMPs) distinct from HSCs. EMPs develop in the yolk sac at embryonic day (E) 8.5, migrate and colonize the nascent fetal liver before E10.5, and give rise to fetal erythrocytes, macrophages, granulocytes and monocytes until at least E16.5. Subsequently, HSC-derived cells replace erythrocytes, granulocytes and monocytes. Kupffer cells, microglia and Langerhans cells are only marginally replaced in one-year-old mice, whereas alveolar macrophages may be progressively replaced in ageing mice. Our fate-mapping experiments identify, in the fetal liver, a sequence of yolk sac EMP-derived and HSC-derived haematopoiesis, and identify yolk sac EMPs as a common origin for tissue macrophages.
Asunto(s)
Linaje de la Célula , Eritrocitos/citología , Hematopoyesis , Macrófagos/citología , Células Madre/citología , Saco Vitelino/citología , Animales , Proliferación Celular , Rastreo Celular , Femenino , Feto/citología , Granulocitos/citología , Macrófagos del Hígado/citología , Células de Langerhans/citología , Hígado/citología , Hígado/embriología , Macrófagos Alveolares/citología , Masculino , Ratones , Microglía/citología , Monocitos/citología , Receptor de Factor Estimulante de Colonias de Macrófagos/metabolismo , Receptor TIE-2/metabolismo , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
Cell competition is an emerging principle underlying selection for cellular fitness during development and disease. Competition may be relevant for cancer, but an experimental link between defects in competition and tumorigenesis is elusive. In the thymus, T lymphocytes develop from precursors that are constantly replaced by bone-marrow-derived progenitors. Here we show that in mice this turnover is regulated by natural cell competition between 'young' bone-marrow-derived and 'old' thymus-resident progenitors that, although genetically identical, execute differential gene expression programs. Disruption of cell competition leads to progenitor self-renewal, upregulation of Hmga1, transformation, and T-cell acute lymphoblastic leukaemia (T-ALL) resembling the human disease in pathology, genomic lesions, leukaemia-associated transcripts, and activating mutations in Notch1. Hence, cell competition is a tumour suppressor mechanism in the thymus. Failure to select fit progenitors through cell competition may explain leukaemia in X-linked severe combined immune deficiency patients who showed thymus-autonomous T-cell development after therapy with gene-corrected autologous progenitors.
Asunto(s)
Transformación Celular Neoplásica , Células Madre Hematopoyéticas/citología , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Timo/citología , Animales , División Celular , Movimiento Celular , Transformación Celular Neoplásica/genética , Progresión de la Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Proteínas HMGA/genética , Células Madre Hematopoyéticas/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Receptor Notch1/genética , Linfocitos T/citología , Linfocitos T/metabolismo , Linfocitos T/patología , Timo/patología , Transcriptoma/genéticaRESUMEN
The cellular differentiation pathway originating from the bone marrow leading to early T lymphocytes remains poorly understood. The view that T cells branch off from a lymphoid-restricted pathway has recently been challenged by a model proposing a common progenitor for T cell and myeloid lineages. We generated interleukin-7 receptor alpha (Il7r) Cre recombinase knockin mice and traced lymphocyte development by visualizing the history of Il7r expression. Il7r fate mapping labeled all T cells but few myeloid cells. More than 85% of T cell progenitors were Il7r reporter(+) and, hence, had arisen from an Il7r-expressing pathway. In contrast, the overwhelming majority of myeloid cells in the thymus were derived from Il7r reporter(-) cells. Thus, lymphoid-restricted progenitors are the major route to T cells, and distinct origins of lymphoid and myeloid lineages represent a fundamental hallmark of hematopoiesis.
Asunto(s)
Linaje de la Célula , Células Mieloides/citología , Células Mieloides/inmunología , Linfocitos T/citología , Linfocitos T/inmunología , Timo/citología , Timo/inmunología , Alelos , Animales , Diferenciación Celular , Ratones , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/inmunologíaRESUMEN
PURPOSE OF REVIEW: Hematopoietic stem cell (HSC) transplantation has yielded tremendous information on experimental properties of HSCs. Yet, it remains unclear whether transplantation reflects the physiology of hematopoiesis. A limitation is the difficulty in accessing HSC functions without isolation, in-vitro manipulation and readout for potential. New genetic fate mapping and clonal marking techniques now shed light on hematopoiesis under physiological conditions. RECENT FINDINGS: Transposon-based genetic marks were introduced across the entire hematopoietic system to follow the clonal dynamics of these tags over time. A polyclonal source downstream from stem cells was found responsible for the production of at least granulocytes. In independent experiments, HSCs were genetically marked in adult mice, and the kinetics of label emergence throughout the system was followed over time. These experiments uncovered that during physiological steady-state hematopoiesis large numbers of HSCs yield differentiated progeny. Individual HSCs were active only rarely, indicating their very slow periodicity of differentiation rather than quiescence. SUMMARY: Noninvasive genetic experiments in mice have identified a major role of stem and progenitor cells downstream from HSCs as drivers of adult hematopoiesis, and revealed that post-transplantation hematopoiesis differs quantitatively from normal steady-state hematopoiesis.
Asunto(s)
Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Animales , Biomarcadores , Diferenciación Celular , Autorrenovación de las Células , Trasplante de Células Madre Hematopoyéticas , Humanos , Inmunofenotipificación , Fenotipo , Análisis de la Célula IndividualRESUMEN
Hepatitis B virus infection represents a major global health problem. Currently, there are more than 240 million chronically infected people worldwide. The development of chronic hepatitis B virus-mediated liver disease may lead to liver fibrosis, cirrhosis and eventually hepatocellular carcinoma. Recently, the discovery of the viral entry receptor sodium taurocholate cotransporting polypeptide has facilitated new approaches for a better understanding of viral physiopathology. Hopefully, these novel insights may give rise to the development of more effective antiviral therapy concepts during the next years. In this review, we will discuss the natural history of hepatitis B virus infection including the viral biology, the clinical course of infection and the role of the immune response.
Asunto(s)
Carcinoma Hepatocelular/patología , Virus de la Hepatitis B/fisiología , Hepatitis B Crónica/complicaciones , Hepatitis B Crónica/patología , Cirrosis Hepática/complicaciones , Cirrosis Hepática/patología , Neoplasias Hepáticas/patología , Carcinoma Hepatocelular/epidemiología , Carcinoma Hepatocelular/etiología , Hepatitis B Crónica/epidemiología , Hepatitis B Crónica/prevención & control , Humanos , Cirrosis Hepática/epidemiología , Cirrosis Hepática/etiología , Neoplasias Hepáticas/epidemiología , Neoplasias Hepáticas/etiologíaRESUMEN
Hematopoietic stem cells (HSCs) and multipotent progenitors (MPPs) generate the immune system in development, and contribute to its maintenance under steady-state conditions. How stem and progenitor cells respond to increased demand for mature cells upon injury is a fundamental question of stem cell biology. Several studies of murine hematopoiesis have reported increased proliferation of HSCs in situ when exposed to inflammatory stimuli, which has been taken as a proxy for increased HSC differentiation. Such surplus generation of HSC may fuel enhanced HSC differentiation or, alternatively, maintain HSC cellularity in the face of increased cell death without enhanced HSC differentiation. This key question calls for direct measurements of HSC differentiation in their natural niches in vivo. Here, we review work that quantifies native HSC differentiation by fate mapping and mathematical inference. Recent differentiation tracing studies show that HSC do not increase their differentiation rate upon a wide range of challenges, including systemic bacterial infection (sepsis), blood loss, and transient or persistent ablation of specific mature immune cells. By contrast, MPPs differentiate more rapidly in response to systemic infection to accelerate the production of myeloid cells. These new in vivo data identify MPPs as a major source of hematopoietic regeneration; HSCs might not contribute to regeneration while remaining protected.
Asunto(s)
Hematopoyesis , Células Madre Hematopoyéticas , Animales , Ratones , Células Madre Hematopoyéticas/metabolismo , Diferenciación Celular , Sistema InmunológicoRESUMEN
In response to infections and stress, hematopoiesis rapidly enhances blood and immune cell production. The stage within the hematopoietic hierarchy that accounts for this regeneration is unclear under natural conditions in vivo. We analyzed by differentiation tracing, using inducible Tie2- or Flt3-driven Cre recombinase, the roles of mouse hematopoietic stem cells (HSCs) and multipotent progenitors (MPPs). During polymicrobial sepsis, HSCs responded transcriptionally and increased their proliferation and cell death, yet HSC differentiation rates remained at steady-state levels. HSC differentiation was also independent from the ablation of various cellular compartments-bleeding, the antibody-mediated ablation of granulocytes or B lymphocytes, and genetic lymphocyte deficiency. By marked contrast, the fate mapping of MPPs in polymicrobial sepsis identified these cells as a major source for accelerated myeloid cell production. The regulation of blood and immune cell homeostasis by progenitors rather than stem cells may ensure a rapid response while preserving the integrity of the HSC population.
Asunto(s)
Células Madre Hematopoyéticas , Sepsis , Animales , Ratones , Diferenciación Celular/genética , Linaje de la Célula , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/metabolismo , Integrasas/metabolismo , Células Madre Multipotentes , Sepsis/metabolismo , Tirosina Quinasa 3 Similar a fms/metabolismo , Receptor TIE-2/metabolismoRESUMEN
Hematopoiesis serves as a paradigm for how homeostasis is maintained within hierarchically organized cell populations. However, important questions remain as to the contribution of hematopoietic stem cells (HSCs) toward maintaining steady state hematopoiesis. A number of in vivo lineage labeling and propagation studies have given rise to contradictory interpretations, leaving key properties of stem cell function unresolved. Using processed flow cytometry data coupled with a biology-driven modeling approach, we show that in vivo flux experiments that come from different laboratories can all be reconciled into a single unifying model, even though they had previously been interpreted as being contradictory. We infer from comparative analysis that different transgenic models display distinct labeling efficiencies across a heterogeneous HSC pool, which we validate by marker gene expression associated with HSC function. Finally, we show how the unified model of HSC differentiation can be used to simulate clonal expansion in the early stages of leukemogenesis.
Asunto(s)
Células Madre Hematopoyéticas/metabolismo , Leucemia/patología , Modelos Biológicos , Animales , Biomarcadores/metabolismo , Carcinogénesis/patología , Autorrenovación de las Células , Factores de Intercambio de Guanina Nucleótido/metabolismo , Integrasas/metabolismo , Cinética , Ratones Transgénicos , Receptor TIE-2/metabolismo , Coloración y EtiquetadoRESUMEN
Within the bone marrow microenvironment, endothelial cells (EC) exert important functions. Arterial EC support hematopoiesis while H-type capillaries induce bone formation. Here, we show that BM sinusoidal EC (BM-SEC) actively control erythropoiesis. Mice with stabilized ß-catenin in BM-SEC (Ctnnb1OE-SEC) generated by using a BM-SEC-restricted Cre mouse line (Stab2-iCreF3) develop fatal anemia. While activation of Wnt-signaling in BM-SEC causes an increase in erythroblast subsets (PII-PIV), mature erythroid cells (PV) are reduced indicating impairment of terminal erythroid differentiation/reticulocyte maturation. Transplantation of Ctnnb1OE-SEC hematopoietic stem cells into wildtype recipients confirms lethal anemia to be caused by cell-extrinsic, endothelial-mediated effects. Ctnnb1OE-SEC BM-SEC reveal aberrant sinusoidal differentiation with altered EC gene expression and perisinusoidal ECM deposition and angiocrine dysregulation with de novo endothelial expression of FGF23 and DKK2, elevated in anemia and involved in vascular stabilization, respectively. Our study demonstrates that BM-SEC play an important role in the bone marrow microenvironment in health and disease.
Asunto(s)
Anemia/genética , Médula Ósea/metabolismo , Moléculas de Adhesión Celular Neuronal/genética , Endotelio Vascular/metabolismo , Eritroblastos/metabolismo , Eritropoyesis/genética , beta Catenina/genética , Anemia/metabolismo , Anemia/mortalidad , Anemia/patología , Animales , Médula Ósea/irrigación sanguínea , Capilares/citología , Capilares/metabolismo , Moléculas de Adhesión Celular Neuronal/metabolismo , Diferenciación Celular , Células Endoteliales/clasificación , Células Endoteliales/citología , Células Endoteliales/metabolismo , Endotelio Vascular/citología , Eritroblastos/clasificación , Eritroblastos/citología , Femenino , Factor-23 de Crecimiento de Fibroblastos/genética , Factor-23 de Crecimiento de Fibroblastos/metabolismo , Regulación de la Expresión Génica , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Integrasas/genética , Integrasas/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Masculino , Ratones , Ratones Transgénicos , Osteogénesis , Reticulocitos/citología , Reticulocitos/metabolismo , Análisis de Supervivencia , Vía de Señalización Wnt , beta Catenina/metabolismoRESUMEN
Lineage tracing reveals hematopoietic stem cell (HSC) fates, while single-cell RNA sequencing identifies snapshots of HSC transcriptomes. To obtain information on fate plus transcriptome in the same cell, we developed the PolyloxExpress allele, enabling Cre-recombinase-dependent RNA barcoding in situ. Linking fates to single HSC transcriptomes provided the information required to identify transcriptional signatures of HSC fates, which were not apparent in single-HSC transcriptomes alone. We find that differentiation-inactive, multilineage, and lineage-restricted HSC clones reside in distinct regions of the transcriptional landscape of hematopoiesis. Differentiation-inactive HSC clones are closer to the origin of the transcriptional trajectory, yet they are not characterized by a quiescent gene signature. Fate-specific gene signatures imply coherence of clonal HSC fates, and HSC output toward short-lived lineage progenitors indicates stability of HSC fates over time. These combined analyses of fate and transcriptome under physiological conditions may pave the way toward identifying molecular determinants of HSC fates.
Asunto(s)
Células Madre Hematopoyéticas , Transcriptoma , Diferenciación Celular/genética , Linaje de la Célula/genética , Células Clonales , Hematopoyesis/genética , Transcriptoma/genéticaRESUMEN
Arterial macrophages have different developmental origins, but the association of macrophage ontogeny with their phenotypes and functions in adulthood is still unclear. Here, we combine macrophage fate-mapping analysis with single-cell RNA sequencing to establish their cellular identity during homeostasis, and in response to angiotensin-II (AngII)-induced arterial inflammation. Yolk sac erythro-myeloid progenitors (EMP) contribute substantially to adventitial macrophages and give rise to a defined cluster of resident immune cells with homeostatic functions that is stable in adult mice, but declines in numbers during ageing and is not replenished by bone marrow (BM)-derived macrophages. In response to AngII inflammation, increase in adventitial macrophages is driven by recruitment of BM monocytes, while EMP-derived macrophages proliferate locally and provide a distinct transcriptional response that is linked to tissue regeneration. Our findings thus contribute to the understanding of macrophage heterogeneity, and associate macrophage ontogeny with distinct functions in health and disease.
Asunto(s)
Arterias/citología , Arteritis/inmunología , Diferenciación Celular/fisiología , Homeostasis/fisiología , Macrófagos/fisiología , Envejecimiento/fisiología , Angiotensina II/administración & dosificación , Angiotensina II/inmunología , Animales , Arterias/fisiología , Médula Ósea/fisiología , Trasplante de Médula Ósea , Linaje de la Célula , Modelos Animales de Enfermedad , Femenino , Células Madre Hematopoyéticas/fisiología , Humanos , Masculino , Ratones , Ratones Transgénicos , RNA-Seq , Regeneración/fisiología , Análisis de la Célula Individual , Quimera por TrasplanteRESUMEN
Microvascular endothelial cells (ECs) are increasingly recognized as organ-specific gatekeepers of their microenvironment. Microvascular ECs instruct neighboring cells in their organ-specific vascular niches through angiocrine factors, which include secreted growth factors (angiokines), extracellular matrix molecules, and transmembrane proteins. However, the molecular regulators that drive organ-specific microvascular transcriptional programs and thereby regulate angiodiversity are largely elusive. In contrast to other ECs, which form a continuous cell layer, liver sinusoidal ECs (LSECs) constitute discontinuous, permeable microvessels. Here, we have shown that the transcription factor GATA4 controls murine LSEC specification and function. LSEC-restricted deletion of Gata4 caused transformation of discontinuous liver sinusoids into continuous capillaries. Capillarization was characterized by ectopic basement membrane deposition, formation of a continuous EC layer, and increased expression of VE-cadherin. Correspondingly, ectopic expression of GATA4 in cultured continuous ECs mediated the downregulation of continuous EC-associated transcripts and upregulation of LSEC-associated genes. The switch from discontinuous LSECs to continuous ECs during embryogenesis caused liver hypoplasia, fibrosis, and impaired colonization by hematopoietic progenitor cells, resulting in anemia and embryonic lethality. Thus, GATA4 acts as master regulator of hepatic microvascular specification and acquisition of organ-specific vascular competence, which are indispensable for liver development. The data also establish an essential role of the hepatic microvasculature in embryonic hematopoiesis.
Asunto(s)
Diferenciación Celular/fisiología , Embrión de Mamíferos/enzimología , Células Endoteliales/metabolismo , Endotelio/embriología , Factor de Transcripción GATA4/metabolismo , Hematopoyesis/fisiología , Hígado/embriología , Animales , Capilares/embriología , Factor de Transcripción GATA4/genética , Hígado/irrigación sanguínea , Ratones , Ratones Transgénicos , Especificidad de Órganos/fisiologíaRESUMEN
Tie1 is a mechanistically poorly characterized endothelial cell (EC)-specific orphan receptor. Yet, Tie1 deletion is embryonic lethal and Tie1 has been implicated in critical vascular pathologies, including atherosclerosis and tumor angiogenesis. Here, we show that Tie1 does not function independently but exerts context-dependent effects on the related receptor Tie2. Tie1 was identified as an EC activation marker that is expressed during angiogenesis by a subset of angiogenic tip and remodeling stalk cells and downregulated in the adult quiescent vasculature. Functionally, Tie1 expression by angiogenic EC contributes to shaping the tip cell phenotype by negatively regulating Tie2 surface presentation. In contrast, Tie1 acts in remodeling stalk cells cooperatively to sustain Tie2 signaling. Collectively, our data support an interactive model of Tie1 and Tie2 function, in which dynamically regulated Tie1 versus Tie2 expression determines the net positive or negative effect of Tie1 on Tie2 signaling.
Asunto(s)
Receptor TIE-1/fisiología , Receptor TIE-2/fisiología , Remodelación Vascular/fisiología , Animales , Diferenciación Celular/fisiología , Células Cultivadas , Células Madre Embrionarias/citología , Células Endoteliales/citología , Células Endoteliales/enzimología , Ratones , Ratones Endogámicos C57BL , Neovascularización Fisiológica/fisiología , Receptor TIE-1/genética , Receptor TIE-1/metabolismo , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Vasos Retinianos/fisiología , Transducción de SeñalRESUMEN
OBJECTIVE: In children with attention-deficit/hyperactivity disorder (ADHD), an increased theta/beta ratio in the resting EEG typically serves as a rationale to conduct theta/beta neurofeedback (NF) training. However, this finding is increasingly challenged. As NF may rather target an active than a passive state, we studied the EEG in a condition that requires attention. METHODS: In children with ADHD of the DSM-IV combined type (ADHD-C; N = 15) and of the predominantly inattentive type (ADHD-I; N = 9) and in typically developing children (N = 19), EEG spectral analysis was conducted for segments during the attention network test (ANT) without processing of stimuli and overt behavior. Frontal (F3, Fz, F4), central (C3, Cz, C4) and parietal (P3, Pz, P4) electrodes were included in the statistical analysis. To investigate if EEG spectral parameters are related to performance measures, correlation coefficients were calculated. RESULTS: Particularly in the ADHD-C group, higher theta and alpha activity was found with the most prominent effect in the upper-theta/lower-alpha (5.5-10.5 Hz) range. In the ADHD-I group, a significantly higher theta/beta ratio was observed at single electrodes (F3, Fz) and a tendency for a higher theta/beta ratio when considering all electrodes (large effect size). Higher 5.5-10.5 Hz activity was associated with higher reaction time variability with the effect most prominent in the ADHD-C group. A higher theta/beta ratio was associated with higher reaction times, particularly in the ADHD-I group. CONCLUSIONS: (1) In an attention demanding period, children with ADHD are characterized by an underactivated state in the EEG with subtype-specific differences. (2) The functional relevance of related EEG parameters is indicated by associations with performance (reaction time) measures. (3) Findings provide a rationale for applying NF protocols targeting theta (and alpha) activity and the theta/beta ratio in subgroups of children with ADHD.
RESUMEN
In attention-deficit/hyperactivity disorder (ADHD), a reduced phasic alerting response (event-related potential component P3 to cue stimuli) has been reported for different subtypes and task types in a series of studies. In order to get a refined picture of this attentional deficit, which is based on the analysis of averaged event-related potentials, we studied the distribution of single-trial cue-P3 amplitudes and the relation between the cue-P3 and the neural state (EEG spectral analysis) when expecting the stimulus. Brain electrical activity was recorded in children of different ADHD subtypes (combined type, predominantly inattentive) and typically developing children while conducting the attention network test. In children with ADHD of the combined type, smaller cue-P3 amplitudes in the averaged signal were due to a larger portion of single trials with reduced cue-P3 amplitudes whereas maximum amplitudes did not differ from typically developing children. In this ADHD subtype, larger activity in the upper theta/lower alpha range (5.5-10.5Hz) was strongly associated with the range (difference between 0.9 quantile and 0.1 quantile) of the cue-P3 amplitude in single trials (correlation coefficient r=0.77) indicating a suboptimal neural state before stimulus presentation. In children with ADHD of the predominantly inattentive subtype, single-trial P3 amplitudes were comparable at lower quantiles but maximum amplitudes were reduced. This result pattern indicates an intact triggering of the cue-P3 but a reduced capacity of resource allocation for the predominantly inattentive subtype. Though findings are limited by a relative small sample size, the cue-P3 may be considered as a neurophysiological marker of alerting deficits in ADHD reflecting different underlying mechanisms in ADHD subtypes.
Asunto(s)
Trastorno por Déficit de Atención con Hiperactividad/fisiopatología , Potenciales Evocados , Atención , Trastorno por Déficit de Atención con Hiperactividad/psicología , Encéfalo/fisiopatología , Estudios de Casos y Controles , Niño , Señales (Psicología) , Electroencefalografía , Femenino , Humanos , Masculino , Tiempo de ReacciónRESUMEN
Recently we have identified the novel mitochondrial peptidase responsible for degrading presequences and other short unstructured peptides in mitochondria, the presequence peptidase, which we named PreP peptidasome. In the present study we have identified and characterized the human PreP homologue, hPreP, in brain mitochondria, and we show its capacity to degrade the amyloid beta-protein (Abeta). PreP belongs to the pitrilysin oligopeptidase family M16C containing an inverted zinc-binding motif. We show that hPreP is localized to the mitochondrial matrix. In situ immuno-inactivation studies in human brain mitochondria using anti-hPreP antibodies showed complete inhibition of proteolytic activity against Abeta. We have cloned, overexpressed, and purified recombinant hPreP and its mutant with catalytic base Glu(78) in the inverted zinc-binding motif replaced by Gln. In vitro studies using recombinant hPreP and liquid chromatography nanospray tandem mass spectrometry revealed novel cleavage specificities against Abeta-(1-42), Abeta-(1-40), and Abeta Arctic, a protein that causes increased protofibril formation an early onset familial variant of Alzheimer disease. In contrast to insulin degrading enzyme, which is a functional analogue of hPreP, hPreP does not degrade insulin but does degrade insulin B-chain. Molecular modeling of hPreP based on the crystal structure at 2.1 A resolution of AtPreP allowed us to identify Cys(90) and Cys(527) that form disulfide bridges under oxidized conditions and might be involved in redox regulation of the enzyme. Degradation of the mitochondrial Abeta by hPreP may potentially be of importance in the pathology of Alzheimer disease.