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C M Verfaillie

Publications and source records attributed to C M Verfaillie.

At least 55 records · Page 3Linked to original sources
PubMed1998

Human CD34(+) bone marrow cells regulate stromal production of interleukin-6 and granulocyte colony-stimulating factor and increase the colony-stimulating activity of stroma.

Cytokines produced by stromal cells induce the proliferation and differentiation of hematopoietic cells in the marrow microenvironment. We hypothesized that cross-talk between hematopoietic cells at different stages of differentiation and stromal cells influences stromal cytokine production and is responsible for maintaining steady-state hematopoiesis and responding to stress situations. We show that coculture of primitive CD34(+) cells in contact with or separated by a transwell membrane from irradiated human bone marrow stromal layers induces a fourfold to fivefold increase in interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF) levels in the stromal supernatant (SN) during the first week. Levels of both cytokines decreased to baseline after coculture of CD34(+) cells for 3 to 5 weeks. Coculture of more mature CD15(+)/CD14(-) myeloid precursors induced only a transient 1.5- to 2-fold increase in IL-6 and G-CSF at 48 hours. Neither CD34(+) nor CD15(+)/CD14(-) cells produced IL-6, G-CSF, IL-1beta, or tumor necrosis factor alpha. When CD34(+) cells were cultured in methylcellulose medium supplemented with cytokines at concentrations found in stromal SN or supplemented with stromal SN, a fourfold to fivefold increase in colony formation was seen over cultures supplemented with erythropoietin (EPO) only. When cultures were supplemented with the increased concentrations of IL-6 and G-CSF detected in cocultures of stroma and CD34(+) cells or when CD34(+) cells were cocultured in methylcellulose medium in a transwell above a stromal layer, a further increase in the number and size of colonies was seen. The colony-forming unit-granulocyte-macrophage-stimulating activity of stromal SN was neutralized by antibodies against G-CSF or IL-6. These studies indicate that primitive CD34(+) progenitors provide a soluble positive feedback signal to induce cytokine production by stromal cells and that the observed increase in cytokine levels is biologically relevant.

Antigens, CD34↗
PubMed1998

Inhibition of BCR-ABL expression with antisense oligodeoxynucleotides restores beta1 integrin-mediated adhesion and proliferation inhibition in chronic myelogenous leukemia hematopoietic progenitors.

Chronic myelogenous leukemia (CML) is characterized by the continuous proliferation and abnormal circulation of malignant hematopoietic progenitors. This may be related to the unresponsiveness of CML progenitors to beta1 integrin adhesion receptor-mediated inhibition of progenitor proliferation by the marrow microenvironment. In hematopoietic cell lines, the BCR-ABL oncogene product, p210(BCR-ABL), interacts with a variety of cytoskeletal elements important for normal integrin signaling. We studied the role of p210(BCR-ABL) in abnormal integrin function in CML by evaluating the effect of inhibition of BCR-ABL expression with antisense oligodeoxynucleotides (AS-ODNs) on integrin-mediated adhesion and proliferation inhibition of malignant primary progenitors from CML marrow. Preincubation of CML CD34(+)HLA-DR+ (DR+) cells with breakpoint-specific AS-ODNs significantly increased adhesion of CML progenitors to stroma and fibronectin (FN). Pretreatment with breakpoint-specific ODNs also resulted in significant inhibition of CML progenitor proliferation after ligand or antibody-mediated beta1 integrin engagement. Breakpoint-specific ODNs were significantly more effective in restoring CML progenitor adhesion and proliferation inhibition than control ODNs. BCR-ABL mRNA and p210(BCR-ABL) levels in CML CD34(+) cells were significantly reduced after incubation with breakpoint-specific AS-ODN. These studies indicate a role for BCR-ABL in abnormal circulation and defective integrin-dependent microenvironmental regulation of proliferation of CML hematopoietic progenitors.

Cell Adhesion↗
PubMed1998

Interferon-alpha restores beta1-integrin-dependent, collagen-mediated platelet aggregation in a patient with chronic myelogenous leukemia.

Although interferon-alpha (IFN-alpha) induces hematologic remissions in 70% to 80% of patients with chronic myelogenous leukemia (CML) and complete or near-complete cytogenetic remissions in 10% to 20% of patients, the exact mechanisms underlying these clinical results remain unclear. We have hypothesized that IFN-alpha acts at least in part through restoration of beta1-integrin function on malignant hematopoietic progenitors that can promote adhesion of malignant progenitors to the marrow microenvironment. This may then restore microenvironmental inhibition of progenitor proliferation and induce tumor dormancy. We demonstrate that IFN-alpha administration to a patient suffering from a clinically severe bleeding diathesis reversed the defective collagen-mediated aggregation of platelets expressing normal numbers of functionally inactive collagen receptors. This is the first in vivo demonstration that IFN-alpha can up-regulate the function of adhesion receptors in CML and supports the premise that IFN-alpha induces remissions by restoring normal integrin-mediated interactions between progenitors and microenvironmental components.

Collagen↗
PubMed1998

Biology of chronic myelogenous leukemia.

This article reviews the biology of chronic myelogenous leukemia (CML) and its effect on the process of hematopoiesis. The relevance of the BCR-ABL fusion protein as well as murine models are also discussed. CML has been studied more extensively than any other malignancy, yet the correlation between the clinical symptoms of chronic phase CML and the BCR-ABL oncoprotein is poorly understood. Insights from recent efforts both to develop a good in vivo animal model and to characterize the effect of the BCR-ABL oncoprotein on relevant signal molecules may lead to a better understanding of the pathophysiology of chronic phase CML and, thereby, to the development of targeted therapeutic approaches.

Animals↗
PubMed1998

Chronic myelogenous leukemia: too much or too little growth, or both?

CML, characterized by the BCR/ABL gene rearrangement has been more extensively studied than any other malignancy. Over the last decade, significant progress has been made in our understanding of BCR-ABL-induced alterations in intracellular signaling. Unfortunately, we still only poorly understand the correlation between the clinical symptoms of chronic phase CML and the BCR-ABL oncoprotein. This is in part due to lack of a good in vivo animal model of chronic phase CML. In vivo and in vitro studies from the Clarkson group, recently reviewed in this journal (Leukemia 1997; 11: 1404-1428), have significantly enhanced our understanding of the pathophysiology of CML. However, further characterization of the effect of the BCR-ABL oncoprotein on signal molecules involved with cell differentiation, cell proliferation, cell survival and cell adhesion in primary Ph+ CML progenitors or in vivo models of CML will be needed to provide a full understanding of the pathophysiology of chronic phase CML.

Animals↗
PubMed1998

Tyrphostin AG957, a tyrosine kinase inhibitor with anti-BCR/ABL tyrosine kinase activity restores beta1 integrin-mediated adhesion and inhibitory signaling in chronic myelogenous leukemia hematopoietic progenitors.

Abnormal beta1 integrin receptor function may contribute to the continuous proliferation and abnormal circulation of malignant hematopoietic progenitors in chronic myelogenous leukemia (CML). Previous studies suggest that abnormal integrin function in CML progenitors is related to the presence of the BCR/ABL oncogene. BCR/ABL may alter integrin function in CML by phosphorylating cytoskeletal and/or signaling proteins important for normal integrin function. We evaluated the effect of Tyrphostin AG957, a protein tyrosine kinase (PTK) inhibitor which has activity against the p210BCR/ABL kinase, on beta1 integrin function in CML progenitors. Incubation of CML marrow CD34+HLA-DR+ cells with Tyrphostin AG957 at concentrations that did not affect colony-forming cells (CFC) viability, but which partly inhibited p210BCR/ABL kinase activity, significantly increased CML CFC adhesion to stroma and alpha4beta1 and alpha5beta1 integrin binding fragments of fibronectin (FN). CML CFC proliferation, unlike that of normal CFC, is not inhibited following integrin receptor engagement with FN or anti-integrin antibodies. AG957 did not alter CML CFC proliferation by itself, but resulted in significant inhibition of CML CFC proliferation following integrin engagement. Another PTK inhibitor, Tyrphostin AG555, which does not have anti-p210BCR/ABL kinase activity, did not affect CML CFC adhesion or proliferation. Neither AG957 nor AG555 affected normal CFC adhesion or proliferation. In BCR/ABL expressing cells, AG957 partially inhibited phosphorylation of several proteins that are BCR/ABL PTK substrates and are involved in normal integrin signaling. These studies suggest that abnormal tyrosine phosphorylation may play an important role in defective integrin function in CML progenitors.

Cell Adhesion↗
PubMed1998

The effect of interferon-alpha on beta-1 integrin mediated adhesion and growth regulation in chronic myelogenous leukemia.

There is considerable interest in understanding the mechanisms by which interferon-alpha can induce hematological and cytogenetic remissions and prolong survival in patients with chronic myelogenous leukemia (CML). There is evidence that the selective expansion and growth advantage of malignant hematopoietic progenitors in CML may be related, at least in part, to their deficient responsiveness to normal negative regulation by the marrow stromal microenvironment and that interferon-alpha may restore normal hematopoiesis in CML by restoring normal microenvironmental regulation of malignant CML progenitor proliferation. In normal hematopoiesis, beta1 integrin receptors mediate progenitor adhesion to stroma. Stimulation of beta1 integrin receptors on normal progenitors results in transmission of proliferation inhibitory signals. CML progenitors demonstrate defective beta1 integrin receptor-mediated adhesion and regulation of proliferation. We have shown that interferon-alpha can restore both beta1 integrin mediated adhesion as well as integrin-mediated proliferation inhibition of CML progenitors. Interferon-alpha enhances CML integrin function through direct effects on CML progenitors as well as indirect effects on stroma. Restored beta1 integrin-mediated regulation of malignant CML progenitor proliferation may result in restoration of normal hematopoiesis in CML patients treated with interferon-alpha. There is evidence that abnormal integrin mediated signaling in CML progenitors may result from abnormalities in integrin-cytoskeletal interactions induced by the p210BCR/ABL tyrosine kinase. Although the exact mechanisms by which interferon-alpha restores normal integrin signaling in CML are not known, preliminary studies indicate that this may at least partly be related to the restoration of normal integrin-cytoskeletal interactions. The above studies offer some insights into the mechanisms of abnormal hematopoietic regulation in CML and the mechanisms by which interferon-alpha may restore normal hematopoiesis, in this disease.

Antineoplastic Agents↗
PubMed1997

Gene therapy for chronic myelogenous leukemia (CML): a retroviral vector that renders hematopoietic progenitors methotrexate-resistant and CML progenitors functionally normal and nontumorigenic in vivo.

Chronic myelogenous leukemia (CML) is a malignant disease of the human hematopoietic stem cell caused by the BCR/ABL gene rearrangement. The only curative therapy is allogeneic transplantation. Although autologous transplants may prolong survival, most patients relapse because of disease persisting in the host and in the graft. Continued administration of chemotherapy after transplant could reduce the incidence of relapse provided that the autograft can be protected by transfer of a drug-resistance gene. However, CML autografts will almost certainly contain malignant stem cells that will also be rendered drug-resistant. The presence of the BCR/ABL oncoprotein is necessary and sufficient for malignant transformation seen in CML. We thus hypothesized that transfer of a vector that combines a drug-resistance gene with anti-BCR/ABL antisense (AS) sequences may allow for posttransplant chemotherapy to decrease persistent disease while rendering inadvertently transduced CML stem and progenitor cells functionally normal. We constructed a retroviral vector, LasBD, that combines the methotrexate (MTX)-resistant tyrosine-22 dihydrofolate-reductase (tyr22-DHFR) gene and AS sequences directed at the b3a2 BCR/ABL breakpoint. b3a2 BCR/ABL containing 32D and MO7e cells were transduced with LasBD and selected in MTX for 14 days. Expression of the AS sequences reduced BCR/ABL mRNA and p210(BCR/ABL) protein levels by 6- to 10-fold in most cells. This subsequently led to the restoration of normal function of BCR/ABL cDNA+ cells: they grew significantly slower in the presence of interleukin-3 (IL-3); they underwent apoptotic cell death when cultured without IL-3; and they had restored expression and function of adhesion receptors. These effects were specific, because LasBD-containing AS sequences directed at the b3a2 BCR/ABL breakpoint did not affect p190(BCR/ABL)-containing cells. LasBD also rendered 20% to 30% of primary Ph- and Ph+ CD34(+) cells MTX-resistant and decreased BCR/ABL mRNA levels in MTX resistant Ph+ CD34(+) cells by 10-fold. Expression of the MTX-resistant DHFR gene and the AS sequences has been stable for at least 1 year in vitro and for more than 70 days in vivo. Finally, LasBD decreased tumorigenicity of 32DBCR/ABL cells in vivo by 3 to 4 logs. In conclusion, the tyr22-DHFR gene in the LasBD vector can protect normal hematopoietic cells from MTX-mediated toxicity, whereas the AS sequences in LasBD can suppress expression of the BCR/ABL gene and restore normal function of BCR/ABL cDNA-containing cells. The LasBD vector may therefore prove to be an extremely useful adjunct in autologous transplantation for CML.

Animals↗
PubMed1997

Soluble factor(s) produced by adult bone marrow stroma inhibit in vitro proliferation and differentiation of fetal liver BFU-E by inducing apoptosis.

Hematopoiesis occurs in different organs during fetal development. Several studies suggest that the growth of hematopoietic progenitors at one stage of ontogenic maturation may not be supported by a microenvironment from a different ontogenic stage. To determine if human fetal liver (FL) clonogenic progenitors can develop in an adult bone marrow (ABM) microenvironment, we compared growth of BFU-E and CFU-GM from 7-14-wk-old FL, 11-20-wk-old fetal bone marrow (FBM), umbilical cord blood (UCB), or ABM in clonogenic medium with or without ABM stroma. In contrast to BFU-E from FBM, UCB, or ABM, soluble factor(s) produced by ABM stroma severely suppressed growth of 98% of FL BFU-E by inducing apoptosis of cells beyond early erythroblast stage. The nature of the soluble factor remains unknown, although we have evidence that it is heat labile with molecular mass < 10 kD. Antibody neutralization studies indicate that TGF-beta1, IL-1, TNF-alpha, macrophage inflammatory protein (MIP)-1alpha, or IFN-gamma are not responsible. The observation that FL progenitors may not be capable of differentiating when transferred to an ABM microenvironment may have important implications for FL transplantation into postnatal recipients. Further, this demonstrates that ontogenic stage-specific interactions between hematopoietic progenitors and their microenvironment are important for the normal development of hematopoiesis.

Age Factors↗
PubMed1997

Primitive long-term culture initiating cells (LTC-ICs) in granulocyte colony-stimulating factor mobilized peripheral blood progenitor cells have similar potential for ex vivo expansion as primitive LTC-ICs in steady state bone marrow.

We have recently shown that more than 90% of long-term culture initiating cells (LTC-IC) mobilized in the peripheral blood (PB) of normal individuals express HLA-DR and CD38 antigens and can sustain hematopoiesis for only 5 weeks. However, 10% of LTC-IC in mobilized PB are CD34+ HLA-DR- and CD34+ CD38- and can sustain hematopoiesis for at least 8 weeks. We now examine the ex vivo expansion potential of CD34+ HLA-DR+ cells (rich in mature LTC-IC) and CD34+ HLA-DR- cells (rich in primitive LTC-IC) in granulocyte colony-stimulating factor (G-CSF) mobilized PB progenitor cells (PBPC). Cells were cultured in contact with M2-10B4 cells (contact) or in transwells above M2-10B4 (noncontact) without and with interleukin-3 (IL-3) and macrophage inflammatory protein (MIP-1alpha) for 2 and 5 weeks. Progeny were evaluated for the presence of colony-forming cells (CFC) and LTC-IC. When CD34+ HLA-DR+ PB cells were cultured in contact cultures without cytokines, a threefold expansion of CFC was seen at 2 weeks, but an 80% decrease in CFC was seen at week 5. Further, the recovery of LTC-IC at week 2 was only 17% and 1% at week 5. This confirms our previous observation that although CD34+ HLA-DR+ mobilized PB cells can initiate long-term cultures, they are relatively mature and cannot sustain long-term hematopoiesis. In contrast, when CD34+ HLA-DR- mobilized PB cells were cultured in contact cultures without cytokines, CFC expansion persisted until week 5 and 49% and 11% of LTC-IC were recovered at week 2 and 5, respectively. As we have shown for steady state bone marrow (BM) progenitors, recovery of LTC-IC was threefold higher when CD34+ HLA-DR- PBPC were cultured in noncontact rather than contact cultures, and improved further when IL-3 and MIP-1alpha were added to noncontact cultures (96 +/- 2% maintained at week 5). We conclude that although G-CSF mobilizes a large population of "mature" CD34+ HLA-DR+ LTC-IC with a limited proliferative capacity, primitive CD34+ HLA-DR- LTC-IC present in mobilized PB have similar characteristics as LTC-IC from steady state BM: (1) they can be maintained in noncontact cultures containing IL-3 and MIP-1alpha for at least 5 weeks; (2) they are subject to the same proliferation inhibitory influences of contact with stroma. Since the absolute number of primitive LTC-IC (week 8 LTC-IC) per mL of G-CSF mobilized PB is similar to that per mL of steady state BM, these studies further confirm that G-CSF mobilized PBPC may have similar long-term repopulating abilities as steady state BM.

Antigens, CD34↗
PubMed1997

Pathophysiology of CML: do defects in integrin function contribute to the premature circulation and massive expansion of the BCR/ABL positive clone?

Hematopoiesis takes place in close contact with the marrow microenvironment. Normal progenitors adhere through a variety of receptors to stroma and extracellular matrix components, including fibronectin. Adhesion through beta1-integrin receptors to fibronectin not only anchor progenitors to the stroma but also result in direct adhesion-mediated signaling that inhibits progenitor proliferation. In contrast to normal hematopoiesis, chronic myelogenous leukemia (CML) is characterized not only by abnormal, premature circulation of primitive progenitors in the blood but also by continuous progenitor proliferation. Although CML progenitors express the same integrin receptors as normal progenitors, they fail to adhere to stroma and fibronectin, suggesting structural or functional abnormalities of these receptors. Furthermore, CML cells present in contact with stroma or fibronectin continue to proliferate, suggesting that failure to adhere through integrin receptors may also underlie the abnormal proliferation of CML progenitors. The observation that integrin-mediated adhesion and proliferation-inhibitory signaling can be restored through treatment with interferon-alpha or an activating anti-beta1-integrin antibody suggests a functional rather than structural defect that may be related to the presence of the BCR/ABL gene rearrangement in these cells. Insights into the role of integrins as adhesion molecules but also receptors that instruct hematopoietic progenitors to survive, proliferate, and possibly differentiate will not only further our understanding of the normal hematopoietic process but also provide insights into diseases characterized by deranged adhesion and proliferation that may lead to novel therapeutic approaches.

Animals↗
PubMed1997

Propagation and titration of murine cytomegalovirus in a continuous bone marrow-derived stromal cell line (M2-10B4).

Murine cytomegalovirus (MCMV) can only be propagated effectively in mouse embryo fibroblast (MEF) cells. We demonstrate that MCMV replicates significantly better in M2-10B4 cells, a continuous line of murine bone marrow stromal cells. M2-10B4 cells were also comparable to MEF cells for detection of small amounts of MCMV reactivating from latently infected spleen explants. M2-10B4 cells will be very useful for studies of MCMV pathogenesis.

Animals↗
PubMed1997

Stem cells in chronic myelogenous leukemia.

This article discusses briefly the molecular consequences of the BCR-ABL fusion gene. It then reviews the current evidence supporting the notion that chronic myelogenous leukemia in its chronic phase is a clonal, hematopoietic, stem cell disease in which malignant hematopoietic stem and progenitor cells respond to "normal" external proliferation and differentiation stimuli, but in which such responses are altered owing to defects in the stem and progenitor cells as a result of the BCR-ABL oncogene.

Cell Adhesion↗
PubMed1997

Activation of beta1 integrins on CML progenitors reveals cooperation between beta1 integrins and CD44 in the regulation of adhesion and proliferation.

Adhesion of normal colony-forming cells (CFC) to bone marrow (BM) stroma and the extracellular matrix (ECM) component fibronectin (FN) depends at least in part on the alpha4beta1 and alpha5beta1 integrins and the CD44 receptor. Aside from anchoring progenitors in the marrow microenvironment, beta1 integrin-dependent adhesion of normal CFC is associated with inhibition of their proliferation. In contrast to normal CFC, chronic myelogenous leukemia (CML) Ph+ CFC adhere significantly less to either stroma or FN. CML Ph+ CFC proliferation is also not inhibited by coculture with stroma or FN. However, equal numbers of alpha4, alpha5, and beta1 integrins and CD44 are present on CML and normal CD34+ cells. We have previously demonstrated that beta1-dependent adhesion to and subsequent proliferation inhibition by FN can be restored when CML Ph+ CFC are incubated with the beta1 integrin activating antibody, 8A2, and demonstrated a role for the alpha5beta1 integrin in this phenomenon. Since the integrin alpha4beta1 and the proteoglycan form of CD44 may cooperate in establishing normal CFC adhesion to FN, we examined if treatment of CML Ph+ CFC with 8A2 also restores the cooperativity between beta1 integrins and CD44. We demonstrate that 8A2 induces adhesion of CML Ph+ CFC not only to intact FN but also to alpha4beta1, alpha5beta1, and proteoglycan binding fragments of FN. 8A2-induced adhesion to these fragments and peptides also results in a significant inhibition of the proliferation of CML Ph+ CFC. Addition of antibodies to either the alpha5, alpha4, or beta1 integrins, antibodies against the CD44 receptor, or removal of chondroitin sulfate glycosaminoglycans from the surface of CML CD34+ HLA-DR+ cells significantly reduced the 8A2-induced adhesion to and adhesion-mediated inhibition of proliferation by FN. These studies demonstrate that activation of beta1 integrins on CML Ph+ CFC not only results in upregulation of beta1 integrin-dependent adhesion and adhesion-mediated inhibition of proliferation, but also in the restoration of cooperation between beta1 integrins and CD44. These studies suggest that decreased beta1 integrin avidity may also affect the function of the proteoglycan adhesion receptor CD44, both of which may contribute to the abnormal circulation and expansion of malignant progenitors in CML.

Antigens, CD↗
PubMed1997

Integrin-mediated regulation of hematopoiesis: do BCR/ABL-induced defects in integrin function underlie the abnormal circulation and proliferation of CML progenitors?

Hematopoiesis takes place in close contact with the marrow microenvironment. Normal progenitors adhere through a variety of receptors to stroma and extracellular matrix components, including fibronectin. Adhesion through integrins to fibronectin may not only serve to anchor progenitors to the microenvironment but also to directly alter the proliferative behavior of normal hematopoietic progenitors. Chronic myelogenous leukemia (CML) is a malignant disease of the hematopoietic stem cell. At the molecular level, CML is characterized by the BCR/ABL gene rearrangement which encodes for the oncoprotein, p210bcr-abl. Presence of the p210bcr-abl tyrosine kinase is necessary and sufficient for the malignant transformation of hematopoietic cells. Clinically, CML is characterized by an abnormal, premature release of primitive progenitors and precursors in the blood and by the continuous proliferation of the malignant progenitor population. In vitro, CML progenitors fail to adhere to or be regulated by marrow stroma. Since CML progenitors express similar numbers of integrin adhesion receptors as normal progenitors, functional rather than quantitative differences of these receptors on CML progenitors may be responsible for the abnormal circulation and proliferation of the malignant clone. In this manuscript we will review the role of integrin adhesion receptors present on normal hematopoietic progenitors in the regulation of their proliferation and discuss signal transduction mechanisms that may be responsible for these effects. We will also discuss the integrin defect in CML which may be caused by the presence of the oncoprotein, P210bcr-abl, and may explain the abnormal trafficking and proliferation observed in CML.

Bone Marrow↗
PubMed1997

Monoclonal antibody crosslinking of the alpha 4 or beta 1 integrin inhibits committed clonogenic hematopoietic progenitor proliferation.

Adhesion receptors can serve as primary signal transduction molecules that convey information into cells that can affect cell proliferation and differentiation. Since hematopoietic progenitors adhere to marrow stroma and fibronectin via the alpha 4 beta 1 integrin and CD44, we examined the role of these receptors in the transfer of proliferation-regulatory signals to progenitors. Actively proliferating colony-forming cells (CFCs) present in cultured CD34+ cells were incubated with mouse monoclonal antibodies against the alpha 4, beta 1, or CD44 receptors and crosslinking was performed with a secondary goat-anti-mouse antibody. The effect on CFC proliferation was examined with a 3H thymidine suicide assay. Compared with controls (39 to 51% kill), crosslinking the alpha 4 or beta 1 integrins significantly reduced CFC proliferation (12 to 26% kill, p = 0.01), indicating that proliferation-inhibitory signals are transmitted through the VLA-4 integrin. Cytochalasin D, a compound that prevents actin polymerization, prevented not only alpha 4 receptor capping, but also the inhibition of CFC proliferation observed following alpha 4 crosslinking. However, crosslinking of the CD44 receptor with the antibodies Hermes-3 and 50B4, which inhibit adhesion of CFC to fibronectin, failed to cap the CD44 receptor in the majority of CD34+ cells. Furthermore, crosslinking of the CD44 receptor with these antibodies also failed to inhibit proliferation of CFCs. These studies demonstrate that adhesion receptor crosslinking of the alpha 4 beta 1 integrin, together with subsequent changes in F-actin polymerization, negatively regulates hematopoietic progenitor proliferation in a manner independent of the shape change associated with adhesion.

Animals↗
PubMed1997

Phenotypic and functional characterization of committed and primitive myeloid and lymphoid hematopoietic precursors in human fetal liver.

We studied the phenotypic and functional properties of colony-forming cells (CFCs), primitive long-term culture initiating cells (LTC-ICs) and lymphoid precursors present in human fetal liver (FL) and compared these with their adult bone marrow (BM) counterparts. FL (7-14-week) cells were selected by fluorescence-activated cell sorting based on increasing CD34 antigen expression (34+, 34++, or 34 ), CD38 antigen expression (CD34++/ CD38+, or CD38-), and HLA-DR antigen expression (CD34++/ HLA-DR+ or HLA-DR-). 13 +/- 0.6% of FL CD34-positive cells were 34 . Significantly more FL CD34++/ cells were CD38- (49 +/- 2.4%) and HLA-DR-(72 +/- 6.7%) than BM CD34++ cells (6.8 +/- 0.7% CD38- and 13.3 +/- 3.2% HLA-DR-). FL and BM CFCs were CD34+/++, CD38+, and HLA-DR+. However, significantly more FL CFCs were erythroid (40%) than adult BM CFCs (15%), and FL colonies were larger (8111 +/- 738 cells/CFC) than BM colonies (3466 +/- 272 cells/CFC, p < 0.001). As is seen in adult BM, FL LTC-ICs were CD34++/ CD38-. In contrast to BM LTC-ICs, FL LTC-ICs were almost exclusively CD34++/ HLA-DR+. In addition, a single FL LTC-IC gave rise to >30 CFCs at 5 weeks compared with only 5 +/- 0.9 CFCs per LTC-IC from BM. Finally, we demonstrate that the FL CD34++/ /CD38-/HLA-DR+ population, which contains 3.7% LTC-ICs, also contains primitive lymphoid progenitors capable of differentiating into natural killer (NK) cells. In conclusion, the phenotype of primitive human FL progenitors such as LTC-IC and primitive NK progenitors is CD34++/ /CD38-/HLA-DR+, suggesting that this population may contain FL hematopoietic stem cells. The phenotypic characterization of FL primitive LTC-ICs and NK progenitors will facilitate further studies of the functional properties of these progenitors.

ADP-ribosyl Cyclase↗