Comparison of various bone marrow fractions in the ability to participate in vascular remodeling after mechanical injury

In contrast to conventional assumption, recent reports propose the possibility that hematopoietic stem cells (HSCs) may have broader potential to differentiate into various cell types. Here, we tested the pluripotency of HSCs by comparing vascular lesions induced by mechanical injury after bone marrow reconstitution with total bone marrow (TBM) cells, c-Kit+ Sca-1+ Lin- (KSL) cells, or a single HSC cell (Tip-SP CD34-KSL cell, CD34- c-Kit+ Sca-1+ Lin- cell with the strongest dye-efflux activity) harboring green fluorescent protein (GFP). The lesions contained a significant number of GFP-positive cells in the TBM and KSL groups, whereas GFP-positive cells were rarely detected in the HSC group. These results suggest that transdifferentiation of a highly purified HSC seems to be a rare event, if it occurs at all, whereas bone marrow cells including the KSL fraction can give rise to vascular cells that substantially contribute to repair or lesion formation after mechanical injury.     

Characterization of a lineage-negative stem-progenitor cell population optimized for ex vivo expansion and enriched for LTC-IC

Current hematopoietic stem cell transplantation protocols rely heavily upon CD34+ cells to estimate hematopoietic stem and progenitor cell (HSPC) yield. We and others previously reported CD133+ cells to represent a more primitive cell population than their CD34+ counterparts. However, both CD34+ and CD133+ cells still encompass cells at various stages of maturation, possibly impairing long-term marrow engraftment. Recent studies demonstrated that cells lacking CD34 and hematopoietic lineage markers have the potential of reconstituting long-term in vivo hematopoiesis. We report here an optimized, rapid negative-isolation method that depletes umbilical cord blood (UCB) mononucleated cells (MNC) from cells expressing hematopoietic markers (CD45, glycophorin-A, CD38, CD7, CD33, CD56, CD16, CD3, and CD2) and isolates a discrete lineage-negative (Lin-) cell population (0.10% +/- 0.02% MNC, n=12). This primitive Lin- cell population encompassed CD34+/- and CD133+/- HSPC and was also enriched for surface markers involved in HSPC migration, adhesion, and homing to the bone marrow (CD164, CD162, and CXCR4). Moreover, our depletion method resulted in Lin- cells being highly enriched for long-term culture-initiating cells when compared with both CD133+ cells and MNC. Furthermore, over 8 weeks in liquid culture stimulated by a cytokine cocktail optimized for HSPC expansion, TPOFLK (thrombopoietin 10 ng/ml, Flt3 ligand 50 ng/ml, c-Kit ligand 20 ng/ml) Lin- cells underwent slow proliferation but maintained/expanded more primitive HSPC than CD133+ cells. Therefore, our Lin- stem cell offers a promising alternative to current HSPC selection methods. Additionally, this work provides an optimized and well-characterized cell population for expansion of UCB for a wider therapeutic potential, including adult stem cell transplantation.     

Hierarchical structure of human megakaryocyte progenitor cells

Megakaryocytopoiesis is a complex biological process involving a series of cellular events that begins with the pluripotent hematopoietic stem cell and ultimately results in the biogenesis of platelets. A hierarchy of megakaryocyte (MK) progenitor cells has been previously defined based upon studies of in vitro megakaryocytopoiesis. Ontogeny-related changes in MK progenitor cells were analyzed in order to further define this cellular hierarchy. Unifocal colony-forming unit-megakaryocyte (CFU-MK)-derived colonies cloned from fetal bone marrow (FBM) formed after fewer days of in vitro culture and were 2.6-fold larger than those colonies cloned from adult bone marrow (ABM). The frequency of CFU-MK-derived colonies cloned from ABM was significantly greater. MK colonies, however, cloned from FBM morphologically consisted of both pure MK colonies and mixed colonies containing MKs, in which a core of CD41- cells were surrounded by CD41+ MKs. Large colonies resembling the primitive BFU-MK also were assayed from both FBM and ABM. These BFU-MK-derived colonies appeared after fewer days of incubation when FBM was assayed, compared to ABM, but at a significantly lower frequency. In addition, large unifocal MK colonies consisting of >300 cells (300-1000) appeared from cells cloned from fetal, but not adult, marrow. This type of colony represents a unique type of MK progenitor cell, termed the high-proliferative-potential cell-MK. Such colonies represent the progeny of the most primitive human MK progenitor cell identified to date. We also attempted to investigate the process of commitment of stem cells to the MK lineage. We explored the actions of thrombopoietin (TPO) on primitive hematopoietic cells in order to gain an understanding of stem cell commitment. CD34+ Thy-1+ Lin- marrow cells, which are enriched for pluripotent hematopoietic stem cells, were shown to express c-Mpl by the polymerase chain reaction. In addition, TPO alone was capable of inducing CD34+ Thy-1+ Lin- cells after two to three weeks to produce progeny composed entirely of MKs. These studies indicate that TPO has a profound effect on hematopoietic stem cells, and that the hierarchy of MK progenitor cells begins with the pluripotent hematopoietic stem cell.     

Unexpectedly efficient homing capacity of purified murine hematopoietic stem cells

Single-cell transplantation analysis revealed that the cells that had the strongest dye efflux activity ("Tip"-SP cells) and had the phenotype CD34- c-Kit+ Sca-1+ Lin- (CD34- KSL cells) exhibited very strong proliferation and multilineage differentiation capacity. Ninety-six percent of the lethally irradiated mice that received a single "Tip"-SP CD34- KSL cell showed significant donor cell engraftment for long term. These findings support the hypothesis that "Tip"-SP CD34- KSL cells represent the most primitive hematopoietic stem cells that are capable of migrating into the primary site and surviving and/or proliferating with nearly absolute efficiency. This led us to propose high marrow-seeding efficiency as a specific characteristic of primitive HSCs, in addition to their self-renewal and multipotent capacity.     

Catecholaminergic neurotransmitters regulate migration and repopulation of immature human CD34+ cells through Wnt signaling

Catecholamines are important regulators of homeostasis, yet their functions in hematopoiesis are poorly understood. Here we report that immature human CD34+ cells dynamically expressed dopamine and beta2-adrenergic receptors, with higher expression in the primitive CD34+CD38(lo) population. The myeloid cytokines G-CSF and GM-CSF upregulated neuronal receptor expression on immature CD34+ cells. Treatment with neurotransmitters increased the motility, proliferation and colony formation of human progenitor cells, correlating with increased polarity, expression of the metalloproteinase MT1-MMP and activity of the metalloproteinase MMP-2. Treatment with catecholamines enhanced human CD34+ cell engraftment of NOD-SCID mice through Wnt signaling activation and increased cell mobilization and bone marrow Sca-1+c-Kit+Lin- cell numbers. Our results identify new functions for neurotransmitters and myeloid cytokines in the direct regulation of human and mouse progenitor cell migration and development.     

含人AGM区、胎肝及骨髓基质细胞培养体系程序化诱导小鼠胚胎干细胞向造血干细胞的分化

背景：前期已分别制备人主动脉-性腺-中肾区基质细胞系及胎肝基质细胞系,发现前者可促进小鼠胚胎干细胞定向分化为造血干细胞。 目的：模拟胚胎发育过程中永久造血发育的时空顺序,探讨人主动脉-性腺-中肾(AGM)区、胎肝(FL)及骨髓(BM)基质细胞对小鼠胚胎干细胞体外诱导分化为造血干细胞的支持作用,以寻求更佳的诱导条件。 方法：将小鼠E14胚胎干细胞诱导为拟胚体(EB),并利用Transwell非接触共培养体系依次在人主动脉-性腺-中肾区、胎肝及骨髓基质细胞饲养层上进一步诱导分化,按不同诱导阶段分为拟胚体对照、EB/AGM、EB/AGM+FL和EB/AGM+FL+BM共4组。共培养6 d后分别收获各组拟胚体来源细胞,以流式细胞仪检测Sca-1+c-Kit+细胞含量,进行各系造血细胞集落形成单位分析并观察细胞形态。 结果与结论：①EB/AGM+FL组和EB/AGM+FL+BM组收获细胞涂片均发现原始造血细胞。②拟胚体来源细胞经AGM区基质细胞诱导后Sca-1+c-Kit+ 细胞明显升高(P < 0.05)。③拟胚体对照组造血细胞集落形成单位低于其他各组(P < 0.05), 而EB/AGM+FL、EB/AGM+FL+BM组造血细胞集落形成单位计数亦较EB/AGM组明显增高。提示AGM+FL和AGM+FL+骨髓基质细胞微环境对原始造血干细胞的扩增效应均明显高于单纯主动脉-性腺-中肾饲养层。     

Transplantation of bone marrow-derived very small embryonic-like stem cells attenuates left ventricular dysfunction and remodeling after myocardial infarction

Adult bone marrow (BM) contains Sca-1+/Lin-/CD45- very small embryonic-like stem cells (VSELs) that express markers of several lineages, including cardiac markers, and differentiate into cardiomyocytes in vitro. We examined whether BM-derived VSELs promote myocardial repair after a reperfused myocardial infarction (MI). Mice underwent a 30-minute coronary occlusion followed by reperfusion and received intramyocardial injection of vehicle (n= 11), 1 x 10(5) Sca-1+/Lin-/CD45+ enhanced green fluorescent protein (EGFP)-labeled hematopoietic stem cells (n= 13 [cell control group]), or 1 x 10(4) Sca-1+/Lin-/CD45- EGFP-labeled cells (n= 14 [VSEL-treated group]) at 48 hours after MI. At 35 days after MI, VSEL-treated mice exhibited improved global and regional left ventricular (LV) systolic function (echocardiography) and attenuated myocyte hypertrophy in surviving tissue (histology and echocardiography) compared with vehicle-treated controls. In contrast, transplantation of Sca-1+/Lin-/CD45+ cells failed to confer any functional or structural benefits. Scattered EGFP+ myocytes and capillaries were present in the infarct region in VSEL-treated mice, but their numbers were very small. These results indicate that transplantation of a relatively small number of CD45- VSELs is sufficient to improve LV function and alleviate myocyte hypertrophy after MI, supporting the potential therapeutic utility of these cells for cardiac repair. Disclosure of potential conflicts of interest is found at the end of this article.     

Differential expression of alpha2 integrin separates long-term and short-term reconstituting Lin-/loThy1.1(lo)c-kit+ Sca-1+ hematopoietic stem cells

Self-renewing, multipotent hematopoietic stem cells are highly enriched within the Lin- Thy1.1(lo)c-kit+ Sca-1+ subset of mouse bone marrow. However, heterogeneous expression within this population of certain cell surface markers raises the possibility that it may be further fractionated phenotypically and perhaps functionally. We previously identified alpha2-integrin (CD49b) as a surface marker with heterogeneous expression on Lin(-/lo)Thy1.1(lo)c-kit+ Sca-1+ stem cells. To determine whether differences in alpha2 expression were indicative of differences in stem cell function, we purified alpha2- and alpha2hi stem cells by fluorescence-activated cell sorting and analyzed their function in long- and short-term hematopoietic reconstitution assays. Both alpha2- and alpha2hi cells could give rise to mature lymphoid and myeloid cells after transplantation into lethally irradiated congenic recipients. However, alpha2hi cells supported hematopoiesis for only a short time (<4 weeks), whereas alpha2- cells reproducibly yielded robust, long-term (>20 weeks) reconstitution, suggesting that alpha2- cells represent a more primitive population than do alpha2hi cells. Consistent with this idea, alpha2- Lin(-/lo)Thy1.1(lo)c-kit+ Sca-1+ cells exhibited an approximately sixfold decreased frequency of spleen colony-forming units (day 12) versus alpha2hi cells. Furthermore, bone marrow cells isolated from animals transplanted >20 weeks previously with 20 alpha2- Lin(-/lo)Thy1.1(lo)c-kit+ Sca-1+ cells included both alpha2- and alpha2hi stem cells of donor origin, indicating that alpha2hi cells are likely lineal descendents of alpha2- cells. Interestingly, alpha2 integrin expression is significantly reduced on lineage-restricted oligopotent progenitors in the marrow, suggesting that high level expression of alpha2 selectively marks a subset of primitive hematopoietic cells which retains multilineage reconstitution potential but exhibits reduced self-renewal capacity.     

Expression of CXCR4, the receptor for stromal cell-derived factor-1 on fetal and adult human lympho-hematopoietic progenitors.

Stromal cell-derived factor-1 (SDF-1) is a CXC chemokine produced by stromal cells that acts as a chemoattractant for human CD34+ progenitor cells. We investigated the expression of CXCR4, the receptor for SDF-1, on CD34+ cells from different hematopoietic sites and developmental stages. CXCR4 was detected by flow cytometry on 37 % of fetal bone marrow (BM) [gestation weeks (gw) 14-23] and 40% of adult BM CD34+ cells. Interestingly, in fetal liver CD34+ cells, CXCR4 was expressed at lower levels at later stages (9%, gw 20-23) compared to early stages of development (39%, gw 7.5-18), suggesting a development-related change in the migratory capacity of progenitors. CXCR4 was detected at similar levels on both phenotypically primitive and committed progenitors from fetal and adult sites. However, B cell lineage progenitor and precursor cells expressed CXCR4 at the highest density (80% of BM CD34+/CD10+ pro-B cells are CXCR4+). CXCR4 was also expressed in the fetal thymus in early T cell precursors and found to be down-regulated during T cell maturation. Finally, we found that stem cell factor, alone or in combination with other cytokines, can up-modulate CXCR4 expression on CD34+ cells by three- to fourfold. In conclusion, our results suggest that CXCR4 may play an important role in the local and systemic trafficking of human CD34+ cells as well as in human B lymphopoiesis and that its expression can be modulated by cytokines.     

Murine side population cells contain cobblestone area-forming cell activity in mobilized blood

Primitive hematopoietic stem cells (HSCs) can be purified from murine bone marrow by sorting Hoechst 33342-effluxing side population (SP) cells. The aim of this study was to establish whether SP cells from peripheral blood contain primitive HSCs and whether this is altered in mice following mobilization. SP cells were analyzed and isolated from bone marrow and blood of mice after mobilization; the HSC content of isolated SP cells was determined through surrogate cobblestone area-forming cell (CAFC) assays. SP cells in normal blood were not found in the high Hoechst dye effluxing portion of the SP tail, did not express the stem cell markers c-Kit and CD34, and did not have measurable CAFC activity. In contrast, SP cells in mobilized blood expressed both stem cell markers, contained cells in the high dye efflux portion of the SP tail, and displayed significant day- 28 to day-35 CAFC activity with 165- to 334-fold enrichment. In comparison to mobilized blood SP cells, normal marrow SP cells contained a higher proportion of cells expressing c-Kit and CD34 and had a greater percentage of cells in the high Hoechst dye-effluxing portion of the SP tail. Analysis of SP cells in the bone marrow after mobilization revealed a decrease in the frequency of SP cells, in expression of c-Kit and Sca+ CD34(+)/CD34(-), and in day-7 to day-35 CAFC activity, consistent with mobilization into blood. We conclude that murine SP cells mobilized into blood contain primitive hematopoietic stem cell activity (day-28 to day-35 CAFC activity). This model offers a means to study the mechanisms of mobilization of primitive stem cells directly in a murine model.     

Purified canine CD34+Lin- marrow cells transduced with retroviral vectors give rise to long-term multi-lineage hematopoiesis.

Human CD34+ cells have been shown to retain long-term hematopoietic engrafting potential in preclinical and clinical studies. However, recent studies of human and murine CD34- stem cells suggest that these are functionally important early progenitors. Using autologous transplantation, we investigated whether canine CD34 and CD34- marrow cells could be transduced and give rise to long-term hematopoiesis. CD34+Lin- and CD34-Lin- cell populations purified by fluorescence-activated cell sorting were separately cocultivated with retroviral vectors LN (CD34+Lin-) and LNY (CD34-Lin-), which carry the neomycin (neo) gene. After myeloablative total body irradiation (920 cGy), 3 dogs received transplants of both CD34+Lin- cells and CD34-Lin- cells and 2 dogs received only CD34-Lin- cells. Untransduced autologous marrow cells were given to ensure hematopoietic recovery. Using CFU-C assays, transduction efficiencies of CD34+Lin- cells ranged from 6% to 18% with no CFU-C formation from CD34-Lin- cells. PCR-based detection of the neo gene from WBCs was used to detect transduced cells weekly after transplantation. Additional PCR studies in 3 dogs given both CD34+Lin- and CD34-Lin- cells were performed on monocytes, granulocytes, and T cells (2 dogs, one at 7.5 months and the other at 9 months) and granulocytes (1 dog at 12 months). LN was detected up to 12 months posttransplantation in WBCs and mono-myeloid and lymphoid populations from 3 dogs receiving transplants of transduced CD34+Lin- cells. LNY was not detected at any time after transplantation in 5 dogs that received transduced CD34-Lin- cells. Whereas canine CD34+Lin- marrow cells contributed to long-term multilineage hematopoiesis, progeny of CD34-Lin- progenitor cells were not detected after transplantation in these experiments.     

Viable c-Kit(W/W) mutants reveal pivotal role for c-kit in the maintenance of lymphopoiesis

Mice lacking the receptor tyrosine kinase c-Kit (c-Kit(W/W)) have hematopoietic defects causing perinatal death. We have identified a viable c-Kit(W/W) mouse, termed the "Vickid" mouse. Around birth, c-Kit plays a redundant role in T and no role in B cell development. Here, we show an age-dependent, progressive decline of pro-T and pro-B cells accompanied by loss of common lymphoid progenitors in the bone marrow in adult mice lacking c-Kit. Adult c-Kit(W/W) hematopoietic stem cells can engraft in host bone marrow but fail to radioprotect, form spleen colonies, or establish sustained lymphopoiesis. These defects in adult T and B cell development are also evident in a second viable c-Kit(W/W) strain, rescued by overexpression of erythropoietin.     

Comparison of hematopoietic activities of human bone marrow and umbilical cord blood CD34 positive and negative cells.

Although the hematopoietic activities of human CD34+ bone marrow (BM) and cord blood (CB) cells have been well characterized, the phenotype of nonobese-diabetic severe combined immunodeficient (NOD/SCID) mice repopulating cells (SRCs) in CB and BM has not yet been fully examined. To address this issue, various hematopoietic activities were compared in terms of total and CD34+ CB and BM cells. Clonal culture of fluorescence-activated cell sorter (FACS) CD34+ CB and BM cells revealed a higher incidence of colony-forming cells with greater proliferation capacity in CB over BM CD34+ cells. CB CD34+ cells also demonstrated higher secondary plating efficiency over BM cells. In addition, we demonstrated that mice transplanted with CB mononuclear cells (MNCs) showed significantly higher levels of chimerism than those transplanted with BM MNCs. However, recipients of FACS-sorted CD34+ CB cells showed significantly lower levels of chimerism than those that received total CB MNCs, suggesting a role of facilitating cells in the CD34- cell population. To further analyze the role of CD34- cells, the NOD/SCID repopulating ability of FACS-sorted CB CD34-c-kit+Lin- and CD34-c-kit-Lin- cells were examined. However, SRCs were not detected in those cells. Taken together, these data suggest that CB is a better source of hematopoietic stem cells and that there are cells in the CD34- fraction that facilitate repopulation of hematopoiesis in the NOD/SCID environment.     

The lineage-c-Kit+Sca-1+ cell response to Escherichia coli bacteremia in Balb/c mice

During bacterial infection, the bone marrow hematopoietic activity shifts toward granulocyte production, which is critical for host defenses. Along with this enhancement of granulopoiesis, the bone marrow also increases its release of hematopoietic precursors. At the present time, little is known about the commitment of hematopoietic precursor cells, including hematopoietic stem cells and progenitors, in this response. To investigate the hematopoietic precursor cell response to bacterial infection, bacteremia was established in Balb/c mice by i.v. injection of Escherichia coli. Bacteremia caused a 10-fold increase in the number of lineage (lin)-c-kit+Sca-1+ cells in the bone marrow. This dramatic expansion of the lin-c-kit+Sca-1+ cell pool resulted from both increased mitosis of these cells and inversion from lin-c-kit+Sca-1- cell phenotype. Lipopolysaccharide, tumor necrosis factor-alpha, and interleukin-6 were potent factors capable of mediating phenotypic inversion of lin-c-kit+Sca-1- cells. Cells in the expanded lin-c-kit+Sca-1+ cell pool contained more colony-forming unit-granulocyte/macrophage. Mobilization of lin-c-kit+Sca-1+ cells into the circulation was significantly enhanced following bacteremia. These results demonstrate that the lin-c-kit+Sca-1+ cell population in the bone marrow constitutes a key component of the host defense response to bacteremia. Functional modifications of these primitive hematopoietic precursors are critical for enhancing granulocyte production following bacterial infection.     

Hematopoietic stem cells from fancc(-/-) mice have lower growth and differentiation potential in response to growth factors

Fanconi anemia (FA) is a complex recessive genetic disease characterized by progressive bone marrow (BM) failure. We have previously shown that stem cells from the FA group C mouse model have lower long-term primary and secondary reconstitution ability, and that bone marrow of Fancc(-/-) mice contained fewer lineage-negative (Lin(-))Thy1.2(low)Sca-1(+)c-kit(+) CD34(+) cells but normal levels of Lin(-)Thy1.2(low)Sca-1(+)c-kit(+)CD34(-) primitive cells. These data suggest that CD34(+) primitive cells have either a lower growth or differentiation potential, or that these cells have greater apoptosis levels. To investigate the role Fancc might have on the growth and differentiation potentials of primitive hematopoietic stem cells, we used a single-cell culture system and monitored cell viability, doubling potential, and apoptosis levels of Fancc(-/-) primitive Lin(-)Thy1.2(-)Sca-1(+) (LTS)-CD34(+) and LTS-CD34(-) stem cells. Results showed that Fancc(-/-) LTS-CD34(-) and LTS-CD34(+) cells had altered growth and apoptosis responses to combinations of stimulatory cytokines, most dramatically in response to a combination of factors that included interleukin-3 (IL-3) and IL-6. In addition, Fancc(-/-) LTS-CD34(-) and LTS-CD34(+) cells showed a lower differentiation potential than Fancc(+/+) cells. These results support a role for Fancc in the growth and differentiation of primitive hematopoietic cells and suggest that an altered response to stimulatory cytokines may contribute to BM aplasia in FA patients.     

Fibroblast growth factor-1 and -2 preserve long-term repopulating ability of hematopoietic stem cells in serum-free cultures

In this study, we demonstrate that extended culture of unfractionated mouse bone marrow (BM) cells, in serum-free medium, supplemented only with fibroblast growth factor (FGF)-1, FGF-2, or FGF-1 +2 preserves long-term repopulating hematopoietic stem cells (HSCs). Using competitive repopulation assays, high levels of stem cell activity were detectable at 1, 3, and 5 weeks after initiation of culture. FGFs as single growth factors failed to support cultures of highly purified Lin(-)Sca-1(+)c-Kit(+)(LSK) cells. However, cocultures of purified CD45.1 LSK cells with whole BM CD45.2 cells provided high levels of CD45.1 chimerism after transplant, showing that HSC activity originated from LSK cells. Subsequently, we tested the reconstituting potential of cells cultured in FGF-1 + 2 with the addition of early acting stimulatory molecules, stem cell factor +interleukin-11 + Flt3 ligand. The addition of these growth factors resulted in a strong mitogenic response, inducing rapid differentiation and thereby completely overriding FGF-dependent stem cell conservation. Importantly, although HSC activity is typically rapidly lost after short-term culture in vitro, our current protocol allows us to sustain stem cell repopulation potential for periods up to 5 weeks.     

Vaccinia virus infection modulates the hematopoietic cell compartments in the bone marrow

Successful proliferation and differentiation of hematopoietic progenitor cells in bone marrow (BM) is essential to generate all mature blood cell types, including those involved in the immune response. Although vaccinia virus (VV) is known to induce a strong immune response, the effect of VV infection on hematopoiesis remains largely unknown. Here, we showed that in vivo VV infection results in the expansion of c-Kit(hi)Sca-1+Lin- (KSL) hematopoietic stem cells. The in vivo expansion of the KSL population requires MyD88 that is a critical adaptor for Toll-like receptor-mediated signaling. Moreover, in BM of VV-infected mice, common myeloid progenitors (CMP) was decreased because of the rapid differentiation of CMP to more mature cells. However, the CMP compartment was not affected by VV infection in the absence of MyD88. The common lymphoid progenitor (CLP) cell population was increased regardless of MyD88 status, suggesting the independent regulation of CMP and CLP compartments by VV infection. VV infection also enhanced the potential of progenitors that preferentially induce the programming of dendritic cell (DC) development toward plasmacytoid DC. Therefore, the host immune response is gearing toward antiviral responses as early as at the precursor level upon VV infection.     

AGM区来源的基质细胞促进造血干细胞扩增的体外实验研究

目的： 探讨主动脉-性腺-中肾（aorta-gonad-mesonephros，AGM）来源的基质细胞对造血干细胞（HSC）增殖的促进作用，为探寻HSC的体外扩增方法奠定实验基础。 方法： 分别从孕11 d BALB/c小鼠胚胎AGM区及6周龄小鼠骨髓分离、培养基质细胞，流式细胞仪等对基质细胞进行鉴定；利用小鼠胚胎干细胞（ESC）向造血细胞定向分化的模型，结合高增殖潜能集落（HPP-CFC）、原始细胞集落（BL-CFC）形成实验及流式细胞仪分析CD34+、CD34+Sca-1+细胞比例，对比研究AGM及骨髓基质细胞对ESC来源的HSC的扩增作用。 结果： 小鼠AGM和骨髓基质细胞在形态及表型上基本相似，均符合基质细胞的特征。AGM和骨髓基质细胞均可促进ESC来源的HPP-CFC的形成，但AGM基质细胞还可促进ESC来源的 BL-CFC的形成；流式细胞仪检测发现：在骨髓基质细胞支持下，CD34+细胞增加了3-4倍，但CD34+/Sca-1+却无明显增加；而在AGM基质细胞支持下CD34+、CD34+Sca-1+细胞均明显增加了4-5倍。 结论： AGM基质细胞在有效扩增小鼠HSC同时，能很好地维持HSC自我更新及多向分化的潜能。