ILC2 transfers to apolipoprotein E deficient mice reduce the lipid content of atherosclerotic lesions

The use of immunodeficient mice transplanted with human hematopoietic stem cells is an accepted approach to study human-specific infectious diseases such as HIV-1 and to investigate multiple aspects of human immune system development. However, mouse and human are different in sialylation patterns of proteins due to evolutionary mutations of the CMP-N-acetylneuraminic acid hydroxylase (CMAH) gene that prevent formation of N-glycolylneuraminic acid from N-acetylneuraminic acid. How changes in the mouse glycoproteins’ chemistry affect phenotype and function of transplanted human hematopoietic stem cells and mature human immune cells in the course of HIV-1 infection are not known. We mutated mouse CMAH in the NOD/scid-IL2Rγc−/− (NSG) mouse strain, which is widely used for the transplantation of human cells, using the CRISPR/Cas9 system. The new strain provides a better environment for human immune cells. Transplantation of human hematopoietic stem cells leads to broad B cells repertoire, higher sensitivity to HIV-1 infection, and enhanced proliferation of transplanted peripheral blood lymphocytes. The mice showed no effect on the clearance of human immunoglobulins and enhanced transduction efficiency of recombinant adeno-associated viral vector rAAV2/DJ8. NSG-cmah−/− mice expand the mouse models suitable for human cells transplantation, and this new model has advantages in generating a human B cell repertoire. This strain is suitable to study different aspects of the human immune system development, provide advantages in patient-derived tissue and cell transplantation, and could allow studies of viral vectors and infectious agents that are sensitive to human-like sialylation of mouse glycoproteins.     

A novel immunodeficient mouse model--RAG2 x common cytokine receptor gamma chain double mutants--requiring exogenous cytokine administration for human hematopoietic stem cell engraftment.

Gene transduction into immature human hematopoietic cells collected from umbilical cord blood, bone marrow, or mobilized peripheral blood cells could be useful for the treatment of genetic and acquired disorders of the hematopoietic system. Immunodeficient mouse models have been used frequently as recipients to assay the growth and differentiation of human hematopoietic stem/progenitor cells. Indeed, high levels of human cell engraftment were first reported in human/murine chimeras using NOD/SCID mice, which now are considered as the standard for these types of experiments. However, NOD/SCID mice have some clear disadvantages (including spontaneous tumor formation) that limit their general use. We have developed a new immunodeficient mouse model by combining recombinase activating gene-2 (RAG2) and common cytokine receptor gamma chain (gamma c) mutations. The RAG2-/-/gamma c- double mutant mice are completely alymphoid (T-, B-, NK-), show no spontaneous tumor formation, and exhibit normal hematopoietic parameters. Interestingly, human cord blood cell engraftment in RAG2-/-/gamma c- mice was greatly enhanced by the exogenous administration of human cytokines interleukin-(IL-3) granulocyte-macrophage colony-stimulating factor, (GM-CSF), and erythropoietin in contrast to the NOD/SCID model. This unique feature of the RAG2-/-/gamma c- mouse model should be particularly well suited for assessing the role of different cytokines in human lymphopoiesis and stem/progenitor cell function in vivo.     

Sonic hedgehog induces the proliferation of primitive human hematopoietic cells via BMP regulation

A pool of stem cells that arise from the mesoderm during embryogenesis initiates hematopoiesis. However, factors that regulate the expansion of blood stem cells are poorly understood. We show here that cytokine-induced proliferation of primitive human hematopoietic cells could be inhibited with antibodies to hedgehog (Hh). Conversely, Sonic hedgehog (Shh) treatment induced the expansion of pluripotent human hematopoietic repopulating cells detected in immunodeficient mice. Noggin, a specific inhibitor of bone morphogenetic protein 4 (BMP-4), was capable of inhibiting Shh-induced proliferation in a similar manner to anti-Hh; however, anti-Hh had no effect on BMP-4-induced proliferation. Our study shows that Shh functions as a regulator of primitive hematopoietic cells via mechanisms that are dependent on downstream BMP signals.     

CD26 inhibition on CD34+ or lineage- human umbilical cord blood donor hematopoietic stem cells/hematopoietic progenitor cells improves long-term engraftment into NOD/SCID/Beta2null immunodeficient mice

Given the tremendous need for and potential of umbilical cord blood (CB) to be utilized as a donor source for hematopoietic stem cell (HSC) transplantation in adults, there is a strong push to overcome the constraints created by the limited volumes and subsequent limited HSC and hematopoietic progenitor cell (HPC) numbers available for HSC transplantation from a single collection. We have previously described the use of CD26 inhibitor treatment of donor cells as a method to increase the transplant efficiency of mouse HSCs and HPCs into a mouse recipient. To study the use of CD26 inhibitors as a method of improving the transplantation of human CB HSCs and HPCs, we utilized the nonobese diabetic/severe combined immunodeficient/beta 2 microglobulin null (NOD/SCID/B2m(null)) immunodeficient mouse model of HSC transplantation. We report here significant improvements in the engraftment of long-term repopulating cells following the treatment of either CD34(+) or lineage negative (lin()) donor CB with the CD26 inhibitor, Diprotin A, prior to transplant. These results establish a basis on which to propose the use of CD26 inhibitor treatment of donor CB units prior to transplantation for the purpose of improving transplant efficiency and subsequently patient outcome.     

T lymphocytes determine the development of xeno GVHD and of human hemopoiesis in NOD/SCID mice following human umbilical cord blood transplantation

Over the past decade the human-immunodeficient mouse chimera has become a well-established in vivo model for studying the human immune system and/or hemopoiesis. Under certain experimental conditions and depending on the composition of the human cell graft, the recipient mice may develop a fatal disease, designated as discordant xenogenic graft-versus-host-disease (GVHD), which differs in target tissues and histopathology from allogenic GVHD. Experimental evidence is presented that immunodeficient mice are equally susceptible to allogenic GVHD as normal immunocompetent mice. Whole human cord blood and distinct cellular subpopulations from a single cord blood harvest were transplanted in NOD/severe combined immunodeficient mice and the repopulation of human cells was monitored over time. Depending on the ratio of lymphocytes to hemopoietic stem cells, proliferation of human T cells, hemopoiesis or a combination of the two is observed in widely varying proportions. When the graft contains a preponderance of lymphocytes, fatal protracted discordant xenogenic GVHD develops. Mice receiving purified CD34 cells survived up to 207 days in good health with more than 95% human cells in the bone marrow. In those mice all lineages (B and T lymphocytes, monocytes, granulocytes, erythrocytes and thrombocytes) were demonstrated in the bone marrow and peripheral blood.     

Instant conditional transgenesis in the mouse hematopoietic compartment

Adoptive transfer of retrovirally transduced stem cells has recently been described for instant transgenesis in the hematopoietic compartment of mice. This method circumvents the need to manipulate the germline. However, cell type specific gene expression in this ‘retrogenic’ mouse model has remained tedious. Here we report a single retroviral vector-based method to rapidly generate conditional retrogenic mice. For this purpose, mutated loxP-flanked DNA segments are transduced into hematopoietic stem cells isolated from Cre recombinase transgenic mice, which are subsequently transferred into immunodeficient mice. In this way gene expression can be restricted to hematopoietic cell lineages of choice in the acquired immune system.     

Interleukin-3 promotes proliferation and differentiation of human hematopoietic stem cells but reduces their repopulation potential in NOD/SCID mice

In the present study we explored systematically the influence of human interleukin-3 (IL-3) on the cord blood (CB) cell-derived production of human hematopoietic cells in the bone marrow, blood, and spleen of chimeric nonobese/severe combined immunodeficient mice ((NOD/SCID) mice. CB mononuclear cells and MACS-enriched CB CD34(+) cells were injected into irradiated NOD/SCID mice. The mice were additionally transplanted with a stably transfected rat fibroblast cell line expressing the human IL-3 gene (Rat-IL-3) constitutively, or with the nontransfected rat fibroblast cell line as a control (Rat-1). Rat-IL-3 mice displayed a higher engraftment of human hematopoietic cells in bone marrow, spleen, and peripheral blood compared with mice with Rat-1 cotransplantation. When we transplanted their total bone marrow cell population into secondary mice, surprisingly, mice transplanted with bone marrow cells from Rat-1 mice displayed a higher proportion of human hematopoietic cells compared with Rat-IL-3 mice. As expected, bone marrow cultures (BMCs) from Rat-IL-3 mice contained a higher proportion of human cells than Rat-1 bone marrow cells. However, when BMCs were passaged to new flasks, we observed a higher proportion of human cells in BMCs from Rat-1 mice compared with BMCs from Rat-IL-3 mice. IL-3 promotes the proliferation and differentiation of hematopoietic stem cells in chimeric bone marrow. In addition, IL-3 may play a role in the depletion of hematopoietic stem cells in chimeric bone marrow. In the absence of IL-3, the hematopoietic stem cells may remain in a quiescent state and proliferation can be induced by stimuli, including secondary transplantation or cell passage.     

Hematopoietic repopulating ability of cord blood CD34(+) cells in NOD/Shi-scid mice

Although umbilical cord blood (CB) is increasingly being used as an alternative to bone marrow (BM) as a source of transplantable hematopoietic stem cells (HSC), information on the hematopoietic repopulating ability of CB HSC is still limited. We recently established a xenotransplantation system in NOD/Shi-scid mice to evaluate human stem cell activity. In the present study, we transplanted 5 to 10 x 10(4) CB CD34(+) cells into six NOD/Shi-scid mice treated with anti-asialo GM1 antiserum to investigate the hematopoietic repopulating ability of CB. The BM of all recipients contained human CD45(+) cells 10 to 12 weeks after the transplantation (43.8 +/- 17.7%). Clonal culture of the recipient BM cells revealed the formation of various types of human hematopoietic colonies, including myelocytic, erythroid, megakaryocytic, and multilineage colonies, indicating that CB HSC can differentiate into hematopoietic progenitors of various lineages. However, the extent of the differentiation and maturation differed with each lineage. CD13(+)/CD14(+)/CD33(+) myelocytic cells were mainly repopulated in BM and peripheral blood (PB). While CD41(+) megakaryocytic cells and platelets were present, few glycophorin A(+)CD71(+) or hemoglobin alpha-containing erythroid cells were detected. CD19(+) B cells were the most abundantly repopulated in NOD/Shi-scid mice, but their maturational stage differed among the hematopoietic organs. Most of the BM CD19(+) cells were immature B cells expressing CD10 but not surface immunoglobulin (Ig) M, whereas more mature CD19(+)CD10(-) surface IgM(+) B cells were predominantly present in spleen and PB. CD3(+) T cells were not detected even in the recipient thymus. The transplantation to the NOD/Shi-scid mouse may provide a useful tool for evaluating the repopulating ability of transplantable human HSC.     

Current advances in humanized mouse models

Humanized mouse models that have received human cells or tissue transplants are extremely useful in basic and applied human disease research. Highly immunodeficient mice, which do not reject xenografts and support cell and tissue differentiation and growth, are indispensable for generating additional appropriate models. Since the early 2000s, a series of immunodeficient mice appropriate for generating humanized mice has been successively developed by introducing the IL-2Rγ^null gene (e.g., NOD/SCID/γc^null and Rag2^nullγc^null mice). These strains show not only a high rate of human cell engraftment, but also generate well-differentiated multilineage human hematopoietic cells after human hematopoietic stem cell (HSC) transplantation. These humanized mice facilitate the analysis of human hematology and immunology in vivo. However, human hematopoietic cells developed from HSCs are not always phenotypically and functionally identical to those in humans. More recently, a new series of immunodeficient mice compensates for these disadvantages. These mice were generated by genetically introducing human cytokine genes into NOD/SCID/γc^null and Rag2^nullγc^null mice. In this review, we describe the current knowledge of human hematopoietic cells developed in these mice. Various human disease mouse models using these humanized mice are summarized.     

Parameters for establishing humanized mouse models to study human immunity: Analysis of human hematopoietic stem cell engraftment in three immunodeficient strains of mice bearing the IL2rγ mutation

“Humanized” mouse models created by engraftment of immunodeficient mice with human hematolymphoid cells or tissues are an emerging technology with broad appeal across multiple biomedical disciplines. However, investigators wishing to utilize humanized mice with engrafted functional human immune systems are faced with a myriad of variables to consider. In this study, we analyze HSC engraftment methodologies using three immunodeficient mouse strains harboring the IL2rγ^null mutation; NOD-scid IL2rγ^null, NOD-Rag1^null IL2rγ^null, and BALB/c-Rag1^null IL2rγ^null mice. Strategies compared engraftment of human HSC derived from umbilical cord blood following intravenous injection into adult mice and intracardiac and intrahepatic injection into newborn mice. We observed that newborn recipients exhibited enhanced engraftment as compared to adult recipients. Irrespective of the protocol or age of recipient, both immunodeficient NOD strains support enhanced hematopoietic cell engraftment as compared to the BALB/c strain. Our data define key parameters for establishing humanized mouse models to study human immunity.     

Cotransplantation of ex vivo expanded and unexpanded cord blood units in immunodeficient mice using insulin growth factor binding protein-2-augmented mesenchymal cell cocultures

Ex vivo expansion of cord blood (CB) hematopoietic stem cells and cotransplantation of 2 CB units (CBUs) could enhance the applicability of CB transplantation in adult patients. We report an immunodeficient mouse model for cotransplantation of ex vivo expanded and unexpanded human CB, showing enhanced CB engraftment and provide proof of concept for this transplantation strategy as a means of overcoming the limiting cell numbers in each CBU. CBUs were expanded in serum-free medium supplemented with stem cell factor, Flt-3 ligand, thrombopoietin, and insulin growth factor binding protein-2 together with mesenchymal stromal cell coculture. Unexpanded and expanded CB cells were cotransplanted by tail vein injection into 45 sublethally irradiated nonobese diabetic SCID-IL2γ(-/-) (NSG) mice. Submandibular bleeding was performed monthly, and mice were sacrificed 4 months after transplantation to analyze for human hematopoietic engraftment. Expansion of non-CD34(+) selected CB cells yielded 40-fold expansion of CD34(+) cells and 3.1-fold expansion of hematopoietic stem cells based on limiting dilution analysis of NSG engraftment. Mice receiving expanded grafts exhibited 4.30% human cell repopulation, compared with 0.92% in mice receiving only unexpanded grafts at equivalent starting cell doses, even though the unexpanded graft predominated in long-term hematopoiesis (P = .07). Ex vivo expanded grafts with lower initiating cell doses also showed equivalent engraftment to unexpanded grafts with higher cell dose (8.0% versus 7.9%; P = .93). In conclusion, ex vivo expansion resulted in enhanced CB engraftment despite eventual rejection by the unexpanded CBU.     

Rejection of human islets and human HLA-A2.1 transgenic mouse islets by alloreactive human lymphocytes in immunodeficient NOD-scid and NOD-Rag1(null)Prf1(null) mice

Immunodeficient NOD mice engrafted with human peripheral blood mononuclear cells (PBMCs) were used in two models of human islet allograft rejection. Model one: human PBMCs were engrafted into chemically diabetic NOD-scid mice bearing established subrenal human islet allografts. Inflammation and often complete islet allograft rejection were observed. Model 2 incorporated three key advances. First, we developed a new immunodeficient recipient, NOD-RagI(null)Prf1(null) mice. Second, graft-lymphocyte interactions were optimized by intrasplenic co-transplantation of islets and human PBMC. Third, NOD-scid islets expressing human HLA-A2.1 were used as allograft targets. Diabetic NOD-RagI(null)Prf1(null) recipients of HLA-A2.1 transgenic mouse islets, alone or co-engrafted with HLA-A2-positive human PBMC, exhibited durable graft survival and euglycemia. Contrastingly, co-transplantation with HLA-A2-negative human PBMC led to islet graft rejection without evidence of graft-vs.-host disease (GVHD). We propose that diabetic NOD-RagI(null)Prf1(null) mice co-engrafted with HLA-A2 mouse transgenic islets and allogeneic human PBMC provide an effective in vivo model of human islet allograft rejection.     

Lentiviral vector transduction of hematopoietic stem cells that mediate long-term reconstitution of lethally irradiated mice

Lentiviral vectors efficiently transduce human CD34(+) cells that mediate long-term engraftment of nonobese diabetic/severe combined immunodeficient mice. However, hematopoiesis in these animals is abnormal. Typically, 95% of the human cells in peripheral blood are B lymphocytes. To determine whether lentiviral vectors efficiently transduce stem cells that maintain normal hematopoiesis in vivo, we isolated Sca-1(+)c-Kit(+)Lin(-) bone marrow cells from mice without 5-fluorouracil treatment, and transduced these cells in the absence of cytokine stimulation with a novel lentiviral vector containing a GFP (green flourescent protein) reporter gene. These cells were transplanted into lethally irradiated C57Bl/6 mice. In fully reconstituted animals, GFP expression was observed in 8.0% of peripheral blood mononuclear cells for 20 weeks posttransplantation. Lineage analysis demonstrated that a similar percentage (approximately 8.0%) of GFP-positive cells was detected in peripheral blood B cells, T cells, granulocytes and monocytes, bone marrow erythroid precursor cells, splenic B cells, and thymic T cells. In secondary transplant recipients, up to 20% of some lineages expressed GFP. Our results suggest that quiescent, hematopoietic stem cells are efficiently transduced by lentiviral vectors without impairing self-renewal and normal lineage specification in vivo. Efficient gene delivery into murine stem cells with lentiviral vectors will allow direct tests of genetic therapies in mouse models of hematopoietic diseases such as sickle cell anemia and thalassemia, in which corrected cells may have a selective survival advantage.     

Generation of hematopoietic humanized mice in the newborn BALB/c-Rag2null Il2rγnull mouse model: a multivariable optimization approach

Hematopoietic humanized mice generated via transplantation of human hematopoietic stem cells (hHSCs) into immunodeficient mice are a valuable tool for studying development and function of the human immune system. This study was performed to generate a protocol that improves development and quality of humanized mice in the BALB/c-Rag2(null)Il2rγ(null) strain, testing route of injection, in vitro culture and freezing of hHSCs, types of cytokines in the culture, and co-injection of lineage-depleted CD34(-) cells. Specific hHSC culturing conditions and the addition of support cells were found to increase the frequency, and human hematopoietic chimerism, of humanized mice. The optimized protocol resulted in BALB/c-Rag2(null)Il2rγ(null) humanized mice displaying more consistent human hematopoietic and lymphoid engraftment. Thus, hematopoietic humanized mice generated on a BALB/c immunodeficient background represent a useful model to study the human immune system.     

Engraftment kinetics of human cord blood and murine fetal liver stem cells following in utero transplantation into immunodeficient mice

This study was undertaken to evaluate the kinetics of engraftment after in utero transplantation of murine fetal liver and human cord blood stem cells in the nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse model. NOD/SCID fetuses were injected with murine fetal liver or human cord blood CD34+ cells at day 13.5 of gestation. Frequencies of donor cells were analyzed by flow cytometry up to 48 h post transplantation and 4-16 weeks postnatally. Hematopoietic multilineage reconstitution capacity was assessed. Both types of donor cells home rapidly. However, the frequency of human cord blood stem cells rapidly diminished while the murine fetal liver stem cells expanded over time, resulting in multilineage hematopoietic reconstitution. Differences in long-term reconstitution of allogeneic versus xenogeneic donor cells were ascribed to the inability of the human cells to self-renew and differentiate in the fetal mouse environment, demonstrating the limitations of this commonly used xenograph.     

Selective Purging of Human Multiple Myeloma Cells from Autologous Stem Cell Transplantation Grafts using Oncolytic Myxoma Virus

Autologous stem cell transplantation and novel therapies have improved overall survival of patients with multiple myeloma; however, most patients relapse and eventually succumb to their disease. Evidence indicates that residual cancer cells contaminate autologous grafts and may contribute to early relapses after autologous stem cell transplantation. Here, we demonstrate that ex?vivo treatment with an oncolytic poxvirus called myxoma virus results in specific elimination of human myeloma cells by inducing rapid cellular apoptosis while fully sparing normal hematopoietic stem and progenitor cells. The specificity of this elimination is based on strong binding of the virus to myeloma cells coupled with an inability of the virus to bind or infect CD34(+) hematopoietic stem and progenitor cells. These 2 features allow myxoma to readily identify and distinguish even low levels of myeloma cells in complex mixtures. This ex?vivo rabbit-specific oncolytic poxvirus called myxoma virus treatment also effectively inhibits systemic in?vivo engraftment of human myeloma cells into immunodeficient mice and results in efficient elimination of primary CD138(+) myeloma cells contaminating patient hematopoietic cell products. We conclude that ex?vivo myxoma treatment represents a safe and effective method to selectively eliminate myeloma cells from hematopoietic autografts before reinfusion.     

Humanized mice: novel model for studying mechanisms of human immune-based therapies

The lack of relevant animal models is the major bottleneck for understanding human immunology and immunopathology. In the last few years, a novel model of humanized mouse has been successfully employed to investigate some of the most critical questions in human immunology. We have set up and tested in our laboratory the latest technology for generating mice with a human immune system by reconstituting newborn immunodeficient NOD/SCID-γ ^−/−_c mice with human fetal liver-derived hematopoietic stem cells. These humanized mice have been deemed most competent as human models in a thorough comparative study with other humanized mouse technologies. Lymphocytes in these mice are of human origin while other hematopoietic cells are chimeric, partly of mouse and partly of human origin. We demonstrate that human CD8 T lymphocytes in humanized mice are fully responsive to our novel cell-based secreted heat shock protein gp96^HIV-Ig vaccine. We also show that the gp96^HIV-Ig vaccine induces powerful mucosal immune responses in the rectum and the vagina, which are thought to be required for protection from HIV infection. We posit the hypothesis that vaccine approaches tested in humanized mouse models can generate data rapidly, economically and with great flexibility (genetic manipulations are possible), to be subsequently tested in larger nonhuman primate models and humans.

     

Induction of human humoral immune responses in a novel HLA-DR-expressing transgenic NOD/Shi-scid/γcnull mouse

Mounting evidence has demonstrated that NOD-Shi/scid/γc(null) (NOG) mice are one of the most suitable mouse strains for humanized mouse technologies, in which various human cells or tissues can be engrafted without rejection and autonomously maintained. We have characterized and analyzed various features of the human immune system reconstituted in NOG mice by transplanting human hematopoietic stem cells (hu-HSC). One of the problems of the quasi-immune system in these hu-HSC NOG mice is that the quality of immune responses is not always sufficient, as demonstrated by the lack of IgG production in response to antigen challenge. In this study, we established a novel transgenic NOG sub-strain of mice bearing the HLA-DRA and HLA-DRB1:0405 genes, which specifically expresses HLA-DR4 molecules in MHC II-positive cells. This mouse strain enabled us to match the haplotype of HLA-DR between the recipient mice and human donor HSC. We demonstrated that T-cell homeostasis was differentially regulated in HLA-matched hu-HSC NOG mice compared with HLA-mismatched control mice, and antibody class switching was induced after immunization with exogenous antigens in HLA-matched mice. This novel mouse strain improves the reconstituted human immune systems that develop in humanized mice and will contribute to future studies of human humoral immune responses.     

Shortening the immunodeficient period after hematopoietic stem cell transplantation

The delayed reconstitution of the T-lymphoid compartment represents a major clinical challenge after HLA-mismatched hematopoietic stem cell transplantation. The generation of new T lymphocytes deriving from transplanted hematopoietic stem cells requires several months, a period associated with an increased risk of opportunistic infections and relapses. Recently, the early steps of human lymphopoiesis and the nature of the thymus-seeding progenitors were described. Moreover several scientific groups succeeded to generate T-cell precursors from murine and human hematopoietic stem cells in vitro by transitory exposition to Notch-ligands. Here we summarize and discuss these results and their possible usage in the development of new cell therapies to shorten the immunodeficient period following hematopoietic stem cell transplantation.     

APOA-1 is a Novel Marker of Erythroid Cell Maturation from Hematopoietic Stem Cells in Mice and Humans

The mechanism that regulates the terminal maturation of hematopoietic stem cells into erythroid cells is poorly understood. Therefore, identifying genes and surface markers that are restricted to specific stages of erythroid maturation will further our understanding of erythropoiesis. To identify genes expressed at discrete stages of erythroid development, we screened for genes that contributed to the proliferation and maturation of erythropoietin (EPO)-dependent UT-7/EPO cells. After transducing erythroid cells with a human fetal liver (FL)-derived lentiviral cDNA library and culturing the cells in the absence of EPO, we identified 17 candidate genes that supported erythroid colony formation. In addition, the mouse homologues of these candidate genes were identified and their expression was examined in E12.5 erythroid populations by qRT-PCR. The expression of candidate erythroid marker was also assessed at the protein level by immunohistochemistry and ELISA. Our study demonstrated that expression of the Apoa-1 gene, an apolipoprotein family member, significantly increased as hematopoietic stem cells differentiated into mature erythroid cells in the mouse FL. The Apoa-1 protein was more abundant in mature erythroid cells than hematopoietic stem and progenitor cells in the mouse FL by ELISA. Moreover, APOA-1 gene expression was detected in mature erythroid cells from human peripheral blood. We conclude that APOA-1 is a novel marker of the terminal erythroid maturation of hematopoietic stem cells in both mice and humans.