RESUMO
CD47 is a "don't eat me" signal to phagocytes that is overexpressed on many tumor cells as a potential mechanism for immune surveillance evasion. CD47 and its interaction with signal-regulating protein alpha (SIRPα) on phagocytes is therefore a promising cancer target. Therapeutic antibodies and fusion proteins that block CD47 or SIRPα have been developed and have shown activity in preclinical models of hematologic and solid tumors. Anemia is a common adverse event associated with anti-CD47 treatment, but mitigation strategies-including use of a low 'priming' dose-have substantially reduced this risk in clinical studies. While efficacy in single-agent clinical studies is lacking, findings from studies of CD47-SIRPα blockade in combination with agents that increase 'eat me' signals or with antitumor antibodies are promising. Magrolimab, an anti-CD47 antibody, is the furthest along in clinical development among agents in this class. Magrolimab combination therapy in phase Ib/II studies has been well tolerated with encouraging response rates in hematologic and solid malignancies. Similar combination therapy studies with other anti-CD47-SIRPα agents are beginning to report. Based on these early clinical successes, many trials have been initiated in hematologic and solid tumors testing combinations of CD47-SIRPα blockade with standard therapies. The results of these studies will help determine the role of this novel approach in clinical practice and are eagerly awaited.
RESUMO
We proposed and demonstrated that myelogenous leukemia has a preleukemic phase. In the premalignant phase, normal hematopoietic stem cells (HSCs) gradually accumulate mutations leading to HSC clonal expansion, resulting in the emergence of leukemic stem cells (LSCs). Here, we show that preleukemic HSCs are the basis of clonal hematopoiesis, as well as late-onset blood diseases (chronic-phase chronic myeloid leukemia, myeloproliferative neoplasms, and myelodysplastic disease). The clones at some point each trigger surface expression of "eat me" signals for macrophages, and in the clones and their LSC progeny, this is countered by upregulation of "don't eat me" signals for macrophages such as CD47,opening the possibility of CD47-based therapies. We include evidence that similar processes result in fibroblast expansion in a variety of fibrotic diseases, and arterial smooth muscle clonal expansion is a basis of atherosclerosis, including upregulation of both "eat me" and "don't eat me" molecules on the pathogenic cells.
Assuntos
Aterosclerose , Lesões Pré-Cancerosas , Antígeno CD47 , Células-Tronco Hematopoéticas , Humanos , MutaçãoRESUMO
Multiple myeloma is a hematological neoplasm characterized by the accumulation of clonal plasma cells in the bone marrow. Its frequent relapse following achievement of clinical remissions implicates the existence of therapy-resistant myeloma-initiating cells. To date, results on the identity of myeloma-initiating cells have differed. Here, we prospectively identified a myeloma-initiating population by fractionating and transplanting patient bone marrow cells into human bone-bearing immunocompromised mice. Xenotransplantation of fractionated CD138(+)/CD38(high) cells from 40% of patients (8/20) led to a repopulation of CD19(+)CD38(low) or CD138(+)CD38(+) B-lineage cells in human bone grafts; and these grafts were clonally derived from patient myeloma cells. Meanwhile, CD19(+)CD38(low) xenografts were detected in human bone-bearing mice transplanted with CD19(+)CD38(low/-) B cells from 8 of 22 samples but were not clonally related to patient myeloma cells. Further fractionation and xenotransplantation of CD138(+)CD38(high) cells demonstrated that (CD45(low/-) or CD19(-)) CD38(high)/CD138(+) plasma cells, but not (CD45(high) or CD19(+)) CD38(high)/CD138(+) plasmablasts enrich for myeloma-initiating cells. Quantitative reverse transcription-PCR of two serially transplantable xenografts, which were CD19(-)CD138(+), revealed that they were Pax5 (a B-cell-specific transactivator)-negative. These results suggest that CD19(-)CD45(low/-) fully differentiated plasma cells enrich for long-lived and tumor-initiating cells whereas B cells or plasmablasts do not.
Assuntos
ADP-Ribosil Ciclase 1/análise , Antígenos CD19/análise , Transplante de Medula Óssea , Antígenos Comuns de Leucócito/análise , Mieloma Múltiplo/patologia , Plasmócitos/patologia , Sindecana-1/análise , Animais , Western Blotting , Diferenciação Celular , Proteínas de Ligação a DNA/fisiologia , Feminino , Feto/patologia , Citometria de Fluxo , Humanos , Técnicas Imunoenzimáticas , Imunofenotipagem , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Mieloma Múltiplo/imunologia , Plasmócitos/imunologia , Estudos Prospectivos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
Multiple myeloma is a plasma cell neoplasm residing in bone marrow. Despite advances in myeloma therapies, novel therapies are required to improve patient outcomes. CD47 is highly expressed on myeloma cells and a potential therapeutic candidate for myeloma therapies. Flow cytometric analysis of patient bone marrow cells revealed that myeloma cells overexpress CD47 when compared with non-myeloma cells in 73% of patients (27/37). CD47 expression protects cells from phagocytosis by transmitting an inhibitory signal to macrophages. Here we show that blocking CD47 with an anti-CD47 monoclonal antibody increased phagocytosis of myeloma cells in vitro. In xenotransplantation models, anti-CD47 antibodies inhibited the growth of RPMI 8226 myeloma cells and led to tumor regression (42/57 mice), implicating the eradication of myeloma-initiating cells. Moreover, anti-CD47 antibodies retarded the growth of patient myeloma cells and alleviated bone resorption in human bone-bearing mice. Irradiation of mice before myeloma cell xenotransplantation abolished the therapeutic efficacy of anti-CD47 antibodies delivered 2 weeks after radiation, and coincided with a reduction of myelomonocytic cells in spleen, bone marrow and liver. These results are consistent with the hypothesis that anti-CD47 blocking antibodies inhibit myeloma growth, in part, by increasing phagocytosis of myeloma cells.
Assuntos
Anticorpos Anti-Idiotípicos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Osso e Ossos/imunologia , Antígeno CD47/imunologia , Antígeno CD47/farmacologia , Mieloma Múltiplo/prevenção & controle , Fagocitose/efeitos dos fármacos , Animais , Citotoxicidade Celular Dependente de Anticorpos , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/patologia , Proteínas de Ligação a DNA/fisiologia , Feminino , Feto/patologia , Citometria de Fluxo , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/patologia , Camundongos , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/patologia , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
Somatic cell nuclear transfer and transcription-factor-based reprogramming revert adult cells to an embryonic state, and yield pluripotent stem cells that can generate all tissues. Through different mechanisms and kinetics, these two reprogramming methods reset genomic methylation, an epigenetic modification of DNA that influences gene expression, leading us to hypothesize that the resulting pluripotent stem cells might have different properties. Here we observe that low-passage induced pluripotent stem cells (iPSCs) derived by factor-based reprogramming of adult murine tissues harbour residual DNA methylation signatures characteristic of their somatic tissue of origin, which favours their differentiation along lineages related to the donor cell, while restricting alternative cell fates. Such an 'epigenetic memory' of the donor tissue could be reset by differentiation and serial reprogramming, or by treatment of iPSCs with chromatin-modifying drugs. In contrast, the differentiation and methylation of nuclear-transfer-derived pluripotent stem cells were more similar to classical embryonic stem cells than were iPSCs. Our data indicate that nuclear transfer is more effective at establishing the ground state of pluripotency than factor-based reprogramming, which can leave an epigenetic memory of the tissue of origin that may influence efforts at directed differentiation for applications in disease modelling or treatment.
Assuntos
Epigênese Genética , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Reprogramação Celular/genética , Metilação de DNA/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Genoma/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Técnicas de Transferência Nuclear , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Many types of adult tissues, especially for high turnover tissues such as the blood and intestinal system, stand on a hierarchical tissue-specific stem cell system. Tissue-specific stem cells concurrently have self-renewal capacity and potential to give rise to all types of mature cells in their tissue. The differentiation process of the tissue-specific stem cell is successive restriction of these capacities. The first progeny of tissue-specific stem cells are multipotent progenitors (MPPs) that lose long-term self-renewal capacity yet have full lineage potential. MPPs in turn give rise to oligopotent progenitors, which then commit into lineage-restricted progenitors. This hierarchical system enables a lifelong supply of matured functional cells that generally have a short life span and a relatively high turnover rate. In this chapter, we review our findings and other key experiments that have led to the establishment of the current cellular stem and progenitor hierarchy in the blood-forming systems of mice and humans for both normal and leukemic hematopoiesis. We also review select signaling pathways intrinsic to normal hematopoietic and leukemic stem cell populations as well our recent findings elucidating the possible origin of the leukemia stem cell.
Assuntos
Hematopoese/fisiologia , Células-Tronco Hematopoéticas/citologia , Leucemia Mieloide/patologia , Células-Tronco Neoplásicas/patologia , Adulto , Células-Tronco Adultas/citologia , Animais , Apoptose , Diferenciação Celular , Movimento Celular , Proliferação de Células , Humanos , Leucemia Mieloide/imunologia , Camundongos , Modelos Biológicos , Células-Tronco Multipotentes/citologia , Células-Tronco Neoplásicas/imunologia , Especificidade da EspécieRESUMO
The Wilms' tumor gene WT1 is overexpressed in most of human leukemias regardless of disease subtypes. To characterize the expression pattern of WT1 during normal and neoplastic hematopoiesis, we generated a knock-in reporter green fluorescent protein (GFP) mouse (WT1(GFP/+)) and assayed for WT1 expression in normal and leukemic hematopoietic cells. In normal hematopoietic cells, WT1 was expressed in none of the long-term (LT) hematopoietic stem cells (HSC) and very few (<1%) of the multipotent progenitor cells. In contrast, in murine leukemias induced by acute myeloid leukemia 1 (AML1)/ETO+TEL/PDGFbetaR or BCR/ABL, WT1 was expressed in 40.5 or 38.9% of immature c-kit(+)lin(-)Sca-1(+) (KLS) cells, which contained a subset, but not all, of transplantable leukemic stem cells (LSCs). WT1 expression was minimal in normal fetal liver HSCs and mobilized HSCs, both of which are stimulated for proliferation. In addition, overexpression of WT1 in HSCs did not result in proliferation or expansion of HSCs and their progeny in vivo. Thus, the mechanism by which expansion of WT1-expressing cells occurs in leukemia remains unclear. Nevertheless, our results demonstrate that the WT1(GFP/+) mouse is a powerful tool for analyzing WT1-expressing cells, and they highlight the potential of WT1, as a specific therapeutic target that is expressed in LSCs but not in normal HSCs.
Assuntos
Regulação Neoplásica da Expressão Gênica , Proteínas de Fluorescência Verde/metabolismo , Hematopoese , Células-Tronco Hematopoéticas/metabolismo , Leucemia Experimental/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Proteínas WT1/fisiologia , Animais , Medula Óssea , Proliferação de Células , Ensaio de Unidades Formadoras de Colônias , Modelos Animais de Doenças , Feminino , Genes do Tumor de Wilms , Proteínas de Fluorescência Verde/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/patologia , Humanos , Imunofenotipagem , Lentivirus , Leucemia Experimental/genética , Leucemia Experimental/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Neoplásicas/patologia , Transfecção , Proteínas WT1/genéticaRESUMO
Like many epithelial tumors, head and neck squamous cell carcinoma (HNSCC) contains a heterogeneous population of cancer cells. We developed an immunodeficient mouse model to test the tumorigenic potential of different populations of cancer cells derived from primary, unmanipulated human HNSCC samples. We show that a minority population of CD44(+) cancer cells, which typically comprise <10% of the cells in a HNSCC tumor, but not the CD44(-) cancer cells, gave rise to new tumors in vivo. Immunohistochemistry revealed that the CD44(+) cancer cells have a primitive cellular morphology and costain with the basal cell marker Cytokeratin 5/14, whereas the CD44(-) cancer cells resemble differentiated squamous epithelium and express the differentiation marker Involucrin. The tumors that arose from purified CD44(+) cells reproduced the original tumor heterogeneity and could be serially passaged, thus demonstrating the two defining properties of stem cells: ability to self-renew and to differentiate. Furthermore, the tumorigenic CD44(+) cells differentially express the BMI1 gene, at both the RNA and protein levels. By immunohistochemical analysis, the CD44(+) cells in the tumor express high levels of nuclear BMI1, and are arrayed in characteristic tumor microdomains. BMI1 has been demonstrated to play a role in self-renewal in other stem cell types and to be involved in tumorigenesis. Taken together, these data demonstrate that cells within the CD44(+) population of human HNSCC possess the unique properties of cancer stem cells in functional assays for cancer stem cell self-renewal and differentiation and form unique histological microdomains that may aid in cancer diagnosis.
Assuntos
Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Separação Celular/métodos , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/patologia , Células-Tronco/metabolismo , Animais , Carcinoma de Células Escamosas/genética , Neoplasias de Cabeça e Pescoço/genética , Humanos , Receptores de Hialuronatos/metabolismo , Imuno-Histoquímica , Camundongos , Fenótipo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Bone marrow (BM) cells have recently been shown to give rise to skeletal, hepatic, cardiac, neural, and vascular endothelial tissues. However, it has been shown that this is the result of cell fusion rather than transdifferentiation of hematopoietic stem cells (HSC). For this study, we established a mouse model of brain tumor growth to investigate the differentiation potential of HSC into endothelial cells during brain tumor-induced angiogenesis. Nontransgenic (GFP(neg)) recipient mice were lethally irradiated, and their hematopoietic cells were subsequently repopulated by transplantation of a single green fluorescent protein (GFP)-expressing HSC. Rat glioma (RT-2/RAG) cells were then injected into the striatum of the chimeric mice 6-8 weeks post-transplantation. The animals were sacrificed 3-9 days after tumor implantation, and the mobilization, temporal-spatial distribution, and lineage-specific marker expression profile of the GFP(+) cells within the growing tumor were analyzed. We saw that GFP(+) cells gave rise to elongated, CD34(+)/Flk-1(+) cells that incorporated into the endothelium of tumor blood vessels. However, all GFP(+) cells were also CD45(+), and the presence of CD45 on the HSC-derived endothelial-like cells supports the hypothesis that the hematopoietic cells were recruited into the tumor milieu. The fact that we failed to demonstrate the expression of von Willebrand factor in these cells argues against a true endothelial identity. Nevertheless, the recruitment of HSC-derived endothelial-like cells was an extremely rare event in normal brain parenchyma, and, thus, the permissive influence afforded by the growing tumor appeared to enhance the perivascular tropism and acquisition of an endothelial phenotypes by a population of HSC-derived cells.
Assuntos
Neoplasias Encefálicas/patologia , Células Endoteliais/citologia , Células-Tronco Hematopoéticas/citologia , Neovascularização Patológica , Animais , Antígenos/metabolismo , Antígenos CD34/metabolismo , Biomarcadores/metabolismo , Neoplasias Encefálicas/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Linhagem da Célula , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Proteínas de Fluorescência Verde/metabolismo , Células-Tronco Hematopoéticas/fisiologia , Humanos , Antígenos Comuns de Leucócito/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Transplante de Neoplasias , Ratos , Quimeras de Transplante , Fator de von Willebrand/imunologiaRESUMO
We characterize the survival, migration, and differentiation of human neurospheres derived from CNS stem cells transplanted into the ischemic cortex of rats 7 days after distal middle cerebral artery occlusion. Transplanted neurospheres survived robustly in naive and ischemic brains 4 wk posttransplant. Survival was influenced by proximity of the graft to the stroke lesion and was negatively correlated with the number of IB4-positive inflammatory cells. Targeted migration of the human cells was seen in ischemic animals, with many human cells migrating long distances ( approximately 1.2 mm) predominantly toward the lesion; in naive rats, cells migrated radially from the injection site in smaller number and over shorter distances (0.2 mm). The majority of migrating cells in ischemic rats had a neuronal phenotype. Migrating cells between the graft and the lesion expressed the neuroblast marker doublecortin, whereas human cells at the lesion border expressed the immature neuronal marker beta-tubulin, although a small percentage of cells at the lesion border also expressed glial fibrillary acid protein (GFAP). Thus, transplanted human CNS (hCNS)-derived neurospheres survived robustly in naive and ischemic brains, and the microenvironment influenced their migration and fate.
Assuntos
Isquemia Encefálica/terapia , Movimento Celular , Neurônios/citologia , Transplante de Células-Tronco , Animais , Arteriopatias Oclusivas/terapia , Biomarcadores/análise , Isquemia Encefálica/patologia , Diferenciação Celular , Sobrevivência Celular , Córtex Cerebral/patologia , Proteína Duplacortina , Feto/citologia , Humanos , Ratos , Transplante HeterólogoRESUMO
In the adult, tissue-specific stem cells are thought to be responsible for the replacement of differentiated cells within continuously regenerating tissues, such as the liver, skin, and blood system. In this review, we will consider the factors that influence stem cell fate, taking as a primary example the cell fate determination of hematopoietic stem cells.
Assuntos
Linhagem da Célula/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Regeneração/fisiologia , Adulto , Animais , Apoptose/fisiologia , Adesão Celular/fisiologia , Diferenciação Celular/fisiologia , Divisão Celular/fisiologia , Movimento Celular/fisiologia , Humanos , Modelos BiológicosRESUMO
Hematopoietic stem cells (HSCs) reside predominantly in bone marrow, but low numbers of HSCs are also found in peripheral blood. We examined the fate of blood-borne HSCs using genetically marked parabiotic mice, which are surgically conjoined and share a common circulation. Parabionts rapidly established stable, functional cross engraftment of partner-derived HSCs and maintained partner-derived hematopoiesis after surgical separation. Determination of the residence time of injected blood-borne progenitor cells suggests that circulating HSCs/progenitors are cleared quickly from the blood. These data demonstrate that HSCs rapidly and constitutively migrate through the blood and play a physiological role in, at least, the functional reengraftment of unconditioned bone marrow.
Assuntos
Movimento Celular , Quimera/sangue , Hematopoese/fisiologia , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Animais , Circulação Sanguínea/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Homeostase , Antígenos Comuns de Leucócito/análise , Camundongos , Fatores de TempoRESUMO
Clonogenic multipotent mouse hematopoietic stem cells (HSCs) and progenitor cells are contained within the c-kit(+) (K) lineage(-/lo) (L) Sca-1(+) (S) population of hematopoietic cells; long-term (LT) and short-term (ST) HSCs are Thy-1.1(lo). c-kit is a member of the receptor tyrosine kinase family, a class of receptors that are important in the proliferation and differentiation of hematopoietic cells. To establish whether the Flk-2/Flt3 receptor tyrosine kinase was expressed on the most primitive LT-HSCs, we sorted highly purified multipotent stem and progenitor cells on the basis of Flk-2 surface expression and used them in competitive reconstitution assays. Low numbers of Flk-2(-) HSCs gave rise to long-term multilineage reconstitution in the majority of recipients, whereas the transfer of Flk-2(+) multipotent cells resulted in mostly short-term multilineage reconstitution. The KLS subset of adult mouse bone marrow was analyzed for Flk-2 and Thy-1.1 expression. Three phenotypically and functionally distinct populations were isolated: Thy(lo) Flk-2(-) (LT-HSCs), Thy(lo) Flk-2(+) (ST-HSCs), and Thy(-) Flk-2(+) multipotent progenitors. The loss of Thy-1.1 and gain of Flk-2 expression marks the loss of self-renewal in HSC maturation. The addition of Flk-2 antibody to the lineage mix allows direct isolation of LT-HSC from adult bone marrow as c-kit(+) lin(-) Sca-1(+) Flk-2(-) from many strains of mice. Fetal liver HSCs are contained within Flk-2(-) and Flk-2(+) KTLS cells.
Assuntos
Separação Celular/métodos , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Animais , Antígenos Ly/metabolismo , Biomarcadores , Diferenciação Celular , Feto/citologia , Feto/imunologia , Feto/metabolismo , Hematopoese , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/imunologia , Fígado/citologia , Fígado/imunologia , Fígado/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Fenótipo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Receptores Proteína Tirosina Quinases/genética , Antígenos Thy-1/metabolismo , Tirosina Quinase 3 Semelhante a fmsRESUMO
Multipotent stem cells are clonal cells that self-renew as well as differentiate to regenerate adult tissues. Whereas stem cells and their fates are known by unique genetic marker studies, the fate and function of these cells are best studied by their prospective isolation. This review is about the properties of various highly purified tissue-specific multipotent stem cells and purified oligolineage progenitors. We contend that unless the stem or progenitor cells in question have been purified to near homogeneity, one cannot know whether their generation of expected (or unexpected) progeny is a property of a known cell type. It is interesting that in the hematopoietic system the only long-term self-renewing cells in the stem and progenitors pool are the hematopoietic stem cells. This fact is discussed in the context of normal and leukemic hematopoiesis.
Assuntos
Células-Tronco Hematopoéticas/citologia , Células-Tronco/fisiologia , Animais , Diferenciação Celular , Linhagem da Célula , Separação Celular , Transformação Celular Neoplásica/patologia , Células Clonais , Hematopoese , Humanos , Sistema Nervoso/citologia , Fenótipo , RegeneraçãoRESUMO
Stem cell biology has come of age. Unequivocal proof that stem cells exist in the haematopoietic system has given way to the prospective isolation of several tissue-specific stem and progenitor cells, the initial delineation of their properties and expressed genetic programmes, and the beginnings of their utility in regenerative medicine. Perhaps the most important and useful property of stem cells is that of self-renewal. Through this property, striking parallels can be found between stem cells and cancer cells: tumours may often originate from the transformation of normal stem cells, similar signalling pathways may regulate self-renewal in stem cells and cancer cells, and cancer cells may include 'cancer stem cells' - rare cells with indefinite potential for self-renewal that drive tumorigenesis.
Assuntos
Neoplasias/patologia , Células-Tronco , Animais , Divisão Celular , Transformação Celular Neoplásica , Células-Tronco Hematopoéticas , Humanos , Leucemia/patologia , Mutação , Regeneração , Transdução de SinaisRESUMO
The recent application of new techniques, such as multi-color cell sorting and the production of transgenic and gene-knockout mice, has contributed to a better understanding of lymphocyte development from hematopoietic stem cells. Now that we can purify progenitors at different maturational stages during lymphocyte development, the challenge is to understand the processes that govern each developmental stage transition.