Expression of CD4 by human hematopoietic progenitors [see comments]

It has been recently reported that murine hematopoietic stem cells and progenitors express low levels of CD4. In this study, we have investigated by phenotypic and functional analysis whether the CD4 molecule was also present on human hematopoietic progenitors. Unfractionated marrow cells or immunomagnetic bead-purified CD34+ cells were analyzed by two-color fluorescence with an anti-CD4 and an anti- CD34 monoclonal antibody (MoAb). A large fraction (25% to 50%) of the CD34+ cells was weakly stained by anti-CD4 antibodies. Moreover, in further experiments analyzing the expression of CD4 in different subpopulations of CD34+ cells, we found that CD4 was predominantly expressed in phenotypically primitive cells (CD34+ CD38-/low CD71low Thy-1high, HLA-DR+/low). However, the presence of CD4 was not restricted to these primitive CD34+ cell subsets and was also detected in a smaller fraction of more mature CD34+ cells exhibiting differentiation markers. Among those, subsets with myelo-monocytic markers (CD13, CD33, CD14, and CD11b) have a higher CD4 expression than the erythroid or megakaryocytic subsets. In vitro functional analysis of the sorted CD34+ subsets in colony assays and long-term culture- initiating cell (LTC-IC) assays confirmed that clonogenic progenitors (colony-forming unit-granulocyte-macrophage, burst-forming unit- erythroid, and colony-forming unit-megakaryocyte) and LTC-IC were present in the CD4low population. However, most clonogenic progenitors were recovered in the CD4- subset, whereas the CD4low fraction was greatly enriched in LTC-IC. In addition, CD4low LTC-IC generated larger numbers of primitive clonogenic progenitors than did CD4- LTC-IC. These observations suggest that, in the progenitor compartment, the CD4 molecule is predominantly expressed on very early cells. The CD4 molecule present on CD34+ cells appeared identical to the T-cell molecule because it was recognized by three MoAbs recognizing different epitopes of the molecule. Furthermore, this CD4 molecule is also functional because the CD34+ CD4low cells are able to bind the human immunodeficiency virus (HIV) gp120. This observation might be relevant to the understanding of the mechanisms of HIV-induced cytopenias.     

Circulating erythroid progenitors in polycythemia vera are hypersensitive to insulin-like growth factor-1 in vitro: studies in an improved serum-free medium [see comments]

We have investigated the question of erythropoietin (Epo) hypersensitivity versus Epo independence as the basis for the endogenous erythroid bursts (EEBs) that develop in cultures without added Epo from hematopoietic cells of polycythemia vera (PV) patients. Using an improved serum-free (SF) medium containing interleukin (IL)-3, but no insulin-like growth factor-1 (IGF-1), and devoid of contaminants that influence erythropoiesis, we compared circulating normal and PV early erythroid progenitors (BFU-E) with respect to their responses in vitro to recombinant human (rHu) Epo. Cultures were seeded with Ficoll- Hypaque density-separated peripheral blood (PB) mononuclear cells (MNCs), and erythroid bursts, together with their component colonies of > or = 50 cells, were scored in situ at 13 to 16 days of culture. The Epo dose-response curve of BFU-E from PV patients was found to be statistically indistinguishable from that of normal subjects. This observation provides compelling evidence against the Epo- hypersensitivity hypothesis. In the complete SF medium minus Epo, the sensitivity of BFU-E to IGF-1 was much greater in PV than in normals, the dose-response curve being shifted to the left by at least 2 orders of magnitude. These data show that the erythroid progenitor cell response in PV is hypersensitive to IGF-1, and independent of Epo. The data also emphasize the importance of truly SF medium conditions for assessment of progenitor cell sensitivities to recombinant growth factors. Depletion of adherent cells totally prevented erythroid burst formation by normal circulating progenitors, but did not prevent the hypersensitive response to IGF-1 of such cells from PV patients. Hence, again unlike its normal counterpart, the progenitor cell response in PV appears to be independent of adherent cell control.     

Thrombopoietin alone stimulates the early proliferation and survival of human erythroid, myeloid and multipotential progenitors in serum-free culture

We examined the effects of recombinant human thrombopoietin (TPO, c-Mpl ligand) on the proliferation and differentiation of human haemopoietic progenitors other than megakaryocytic progenitors using serum-free cultures. TPO alone supported the generation of not only megakaryocytic (MK) but also blast cell (blast) colonies from cord blood CD34⁺ cells. Delayed addition of a cytokine cocktail (cytokines; interleukin (IL)-3, IL-6, stem cell factor, erythropoietin, granulocyte-macrophage colony-stimulating factor, and TPO) to cultures with TPO alone on day 7 induced various colonies including granulocyte-macrophage (GM) colonies, erythroid bursts (E), granulocyte-erythrocyte-macrophage-megakaryocyte (GEMM) colonies. Replating experiments of blast colonies supported by TPO alone for culture with cytokines revealed that approximately 60% of the blast colonies contained various haemopoietic progenitors. Single cell cultures of clone-sorted CD34⁺ cells indicated that TPO supported the early proliferation and/or survival of both primitive and committed haemopoietic progenitors. In serum-free suspension cultures, TPO alone significantly stimulated the production of progenitors for MK, GM, E and GEMM colonies as well as long-term culture-initiating cells. These effects were completely abrogated by anti-TPO antibody. These results suggest that TPO is an important cytokine in the early proliferation of human primitive as well as committed haemopoietic progenitors, and in the ex vivo manipulation of human haemopoietic progenitors.     

Gene transfer into human bone marrow hematopoietic cells mediated by adenovirus vectors [see comments]

Human bone marrow mononuclear cells (BMMNCs) and enriched CD34 positive (CD34+) cells were transduced with adenovirus vectors encoding Escherichia coli beta-galactosidase gene. Tranductions were carried out by 24-hour coincubation with adenovirus vectors at different multiplicities of infections (moi). Efficacy of gene transfer into BM cells and expression of the gene product (ie, beta-galactosidase) were studied using X-Gal histochemical staining and flow cytometric analysis. X-Gal staining demonstrated that the percentage of positive cells at mois of 5 to 500 was 3.4% to 34.5% for BMMNCs and 6.0% to 20.0% for enriched CD34+ cells. Similar results (1.5% to 35.7% for BMMNCs and 5.4% to 24.2% for enriched CD34+ cells) were obtained with flow cytometric analysis using fluorescein di-beta-D-galactopyranoside (FDG). Multicolor flow cytometry analysis, which included FDG, demonstrated that BM progenitors (CD34+ or CD34+CD38-), T cells (CD2+), B cells (CD19+), natural killer cells (CD56+), granulocytes, and monocytes all expressed the adenovirus transgene. To ascertain the effects of adenovirus vectors on normal BM progenitors, the numbers of colony forming unit-granulocyte/macrophage (CFU-GM), burst-forming unit- erythrocyte (BFU-E), and high-proliferative potential-colony-forming cells (HPP-CFC) after 24-hour coincubation with adenovirus vectors were determined. When BMMNCs or enriched CD34+ cells were incubated with adenovirus vectors at mois of 5 and 50, no significant differences in the numbers of CFU-GM, BFU-E, and HPP-CFC were observed compared with the uninfected control cells. However, the numbers of CFU-GM were significantly (P < .01) decreased when BMMNCs or enriched CD34+ cells were incubated with adenovirus vectors at a moi of 500, compared with the uninfected control cells. The adenovirus infected cells, purified by cell sorting for FDG expression, were capable of growing in culture and gave rise to various colonies (ie, CFU-GM, BFU-E, and HPP-CFC). These data indicate that recombinant adenovirus vectors can be used to transfer genes to human BM hematopoietic cells with expression of the exogenous gene at a high transduction efficiency.     

Hemoglobin switching in culture: evidence for a humoral factor that induces switching in adult and neonatal but not fetal erythroid cells.

An erythropoietic activity that exerts a profound effect on fetal Hb synthesis is present in fetal sheep sera and it attains a peak concentration at the end of the second to the middle of the third trimester of fetal life. The activity consistently inhibits the increased synthesis of fetal Hb in cultures of burst-forming units (BFUes) from normal adults. In cultures of BFUes from homozygous beta+-thalassemias the activity produces a striking decline in gamma chain synthesis, a decline in G gamma/A gamma chain synthesis ratio, and an increase in delta/gamma and alpha/non-alpha ratios--i.e., findings suggesting a genuine gamma-to-beta switch. The activity accelerates Hb F-to-Hb A switching in neonatal BFUe cultures but it has no effect on fetal Hb synthesis in cultures of BFUe obtained from human fetuses. These findings provide direct evidence that (a) humoral factors play a role in the regulation of the switch from fetal to adult Hb formation, and (b) progenitor cells from various stages of ontogeny respond differently to these factors. The results are compatible with the hypothesis that Hb switching during development is mediated through a change in a developmental program which controls the responsiveness of progenitor cells to "switching" activities in their environment.     

BCL-2 but not its Epstein-Barr virus-encoded homologue, BHRF1, is commonly expressed in posttransplantation lymphoproliferative disorders [see comments]

Posttransplantation lymphoproliferative disease (PTLD) is virtually always associated with Epstein-Barr virus (EBV) infection. BCL-2 and other proteins that confer resistance to apoptosis have been implicated in the pathogenesis of a variety of malignancies including lymphomas. One EBV protein, BHRF1, is a homologue of BCL-2, whereas another, the latency membrane protein 1 (LMP-1), upregulates BCL-2 expression in vitro. In the present study, we used immunohistochemistry to study the expression of these viral and cellular proteins as well as a variety of other EBV-encoded proteins in PTLD. BHRF1 was not detected in any PTLD specimen, whereas BCL-2 was shown in 12 of 17 lesions examined. With one exception, all LMP1-positive cases also expressed BCL-2 and the absence of LMP1 was always associated with a lack of BCL-2 expression. The results do not support a role for the EBV homologue of BCL-2 in PTLD, but they do support a role for viral induction of BCL-2 expression.     

Splenectomy is safe and effective in human immunodeficiency virus- related immune thrombocytopenia [see comments]

Sixty-eight patients, followed in a prospective cohort study of 185 human immunodeficiency virus (HIV)-infected patients with severe immune thrombocytopenia (platelets < 50 x 10(9)/L), underwent splenectomy, 2 to 41 months (median: 10 months) after immune thrombocytopenic purpura (ITP) was diagnosed. The mean platelet count increased from 18 x 10(9)/L to 223 x 10(9)/L with a persistent increase in 56 (82%). It also led to a significant increase of the mean CD4 cell count from 475 x 10(6)/L to 725 x 10(6)/L within a mean delay of 10 months. In the whole cohort, with a mean follow-up of 63 months (range, 6 to 126), the 5-year estimated rate for progression to acquired immunodeficiency syndrome (AIDS) was 23% (95% confidence interval [CI], 15% to 31%) and the AIDS-free survival was 69% (95% CI, 61% to 77%). To investigate the potential impact of splenectomy, a Cox's multiple regression model was used; as splenectomy was not randomly assigned, it was incorporated as a time-dependent covariate. After adjustment on the CD4 cell count, no statistically significant differences were observed between the splenectomized and the nonsplenectomized patients: AIDS progression rate (P = 0.23), survival (P = 0.64) and AIDS-free survival (P = 0.72) were not influenced by splenectomy. Splenectomy is both effective and safe in the treatment of severe, refractory ITP associated with HIV infection.     

Mitochondrial dysfunctions in circulating T lymphocytes from human immunodeficiency virus-1 carriers [see comments]

In several models of lymphocyte apoptosis, two alterations of mitochondrial function precede advanced DNA fragmentation: (1) a reduction of mitochondrial transmembrane potential (delta psi m) and (2) an increase in mitochondrial generation of superoxide anion. Here we show that two fluorochromes allow for the identification of analogous mitochondrial perturbations in circulating T lymphocytes from human immunodeficiency virus (HIV)-1+ donors. The first among these fluorochromes, the cationic lipophilic dye DiOC6(3), measures delta psi m; the second marker, hydroethidine (HE), is nonfluorescent, unless it is oxidized by superoxide anions to the product ethidium (Eth). CD4+ or CD8+ cells from clinically asymptomatic HIV-1 carriers contain a significantly elevated percentage of cells endowed with enhanced HE --> Eth conversion and/or reduced DiOC6(3) uptake as compared with normal controls. Phenotypic characterization of (HE --> Eth)high cells from HIV+ donors shows that these cells possess a low delta psi m, thus demonstrating a functional alteration of mitochondria. In addition, (HE --> Eth)high cells display a reduced incorporation of the cardiolipin- specific dye nonyl-acridine orange (NAO), showing a structural defect of the cardiolipin-containing inner mitochondrial membrane. Control experiments involving rotenone, an inhibitor of the respiratory chain complex I, indicate that the reactive oxygen species responsible for HE --> Eth conversion is generated during mitochondrial electron transport. In synthesis, it appears that mitochondrial alterations occur in a significant percentage of circulating T lymphocytes from HIV- 1 carriers. The extent of delta psi m reduction, as determined ex vivo, correlates with the frequency of cells undergoing DNA fragmentation after overnight in vitro culture. These observations may be important for the understanding and for the direct ex vivo quantitation of HIV- triggered lymphocyte destruction.     

Transplantation of allogeneic peripheral blood stem cells mobilized by recombinant human granulocyte colony-stimulating factor [see comments]

Peripheral blood stem cells (PBSCs) are widely used in autologous transplantation because of ease of collection and rapid hematopoietic reconstitution. However, PBSCs have rarely been used for allogeneic transplantation because of concerns about donor toxicities from cytokine administration and the theoretical increased risk of graft- versus-host-disease (GVHD) from the large number of T cells infused. Eight patients with advanced malignancies received allogeneic PBSC transplants from genotypically HLA-identical sibling donors. All donors received 5 days of recombinant human granulocyte colony-stimulating factor (rhG-CSF; 16 micrograms/kg/day) subcutaneously and were leukapheresed for 2 days. After treatment of the patient with total body irradiation and cyclophosphamide (n = 7) or etoposide, thiotepa, and cyclophosphamide (n = 1), PBSCs were infused immediately after collection and without modification. All patients received cyclosporine and either methotrexate (n = 6) or prednisone (n = 2) for GVHD prophylaxis, rhG-CSF was well tolerated with mild bone pain requiring acetaminophen occurring in two donors. All patients engrafted and in seven hematopoietic recovery was rapid, with 500 neutrophils/microL achieved by day 18 and 20,000 platelets/microL by day 12. Complete donor engraftment was documented by Y chromosome analysis in all four sex-mismatched donor-recipient pairs tested and by DNA analysis in two sex-matched pairs. One patient died on day 18 of veno-occlusive disease of the liver with engraftment but before chromosome analysis could be performed (results are pending in 1 patient). A second patient died of fungal infection 78 days after transplant. Grade 2 acute GVHD occurred in two patients and grade 3 GVHD occurred in one patient. One patient is 301 days from transplant in remission with chronic GVHD; the remaining five patients are alive and disease free 67 to 112 days after transplantation. Preliminary results indicate that allogeneic PBSCs mobilized by rhG-CSF can provide rapid hematologic recovery without an appreciably greater incidence of acute GVHD than would be expected with marrow. Further follow-up is required to determine the incidence of chronic GVHD and any potential beneficial effects on relapse after transplant.     

Accelerated programmed cell death (apoptosis) in erythroid precursors of patients with severe beta-thalassemia (Cooley's anemia) [see comments]

The profound and life-threatening anemia in patients with Cooley's anemia is ascribed primarily to intramedullary hemolysis (ineffective erythropoiesis), the cause of which is obscure. Based on prior morphologic data showing nuclear abnormalities, we hypothesized that accelerated apoptosis could occur in these erythroid precursors. The highly successful bone marrow (BM) transplantation program for patients with Cooley's anemia provided us with a unique opportunity to test this hypothesis. We obtained pretransplantation BM aspiration samples from patients undergoing BM transplantation in Pesaro, Italy and from their allogeneic donors. The erythroid precursors were isolated using ficoll sedimentation and then panning selecting fro CD45- cells. Cytospin and Giemsa staining showed that the separation provided greater than 90% erythroblasts. Five million of these erythroblasts were lysed and their DNA was isolated. There were obvious ladder patterns of DNA breakdown products in beta-thalassemia major samples, with less occurring in beta- thalassemia trait. Normal individuals showed only a slight smear of breakdown of DNA. These results indicate there is enhanced apoptosis in the erythroblasts in the BMs of Cooley's anemia patients. This finding might partially explain why most of these erythroblasts never survive to become mature erythrocytes.     

BCR-ABL antisense oligodeoxyribonucleotides suppress the growth of leukemic and normal hematopoietic cells by a sequence-specific but nonantisense mechanism [see comments]

We have examined the effect of BCR/ABL junctional antisense phosphodiester oligodeoxyribonucleotides (ODNs) on BV173 and other chronic myeloid leukemia (CML) cell lines. Various control ODNs were used to understand the mechanism of the observed antiproliferative effect. Not only the antisense ODNs but also several control ODNs inhibit the proliferation of the leukemic cell lines. All the ODNs that inhibit the cell proliferation share a TAT consensus sequence at their 3' end. A 1-base mismatch within this consensus sequence abolishes the antiproliferative effect. Mismatches of several bases at any other position within the sequence of the active ODNs do not suppress the observed effect. Similar experiments on normal or CML CD34+ cell fraction led to the same observations. We conclude that the antiproliferative effect of the phosphodiester BCR/ABL antisense ODNs cannot be attributed to an antisense mechanism but rather to a nonelucidated effect of a 3' terminal TAT sequence. This effect is not CML specific.     

Clinical importance of extraordinary integration patterns of human T- cell lymphotropic virus type I proviral DNA in adult T-cell leukemia/lymphoma [see comments]

The proviral DNA of human T-cell lymphotrophic virus type I (HTLV-I) is known to be integrated monoclonally in the malignant cells of adult T- cell leukemia/lymphoma (ATL), which is a peripheral T-cell malignancy caused by this virus. We studied the relationship between the integration patterns of HTLV-I and clinical characteristics in 89 patients with ATL. The proviral DNA of HTLV-I was examined by the standard Southern blot analysis using the endonucleases EcoRI and Pst I. One clear band of greater than 9 kb was detected in most of the patients (83 case) when cellular DNA was digested with EcoRI. On the other hand, extraordinary integration patterns of HTLV-I proviral DNA were detected in 6 patients; 3 of them showed two bands, while the other 3 showed one band smaller than 9 kb. When cellular DNA was digested with PstI, the band patterns of these patients were quite different from those of typical patients. The patients with the extraordinary integration patterns had clinical characteristics dissimilar to those of the other 83 patients with the ordinary integration pattern. The patients with two bands by EcoRI digestion always had severe hypoxemia with extremely high levels of serum lactate dehydrogenase at first presentation and showed peculiar organ infiltrations, such as retina and muscle, which were less frequent in the other ordinary 83 patients. They all died within 8 months after the onset. In contrast, the patients with one smaller band by EcoRI digestion always had small and mature T lymphocytes with bilobulated nuclei without lymphadenopathy and showed a favorable clinical course, which was uncommon in the ordinary cases. They were alive 20 to 38 months after diagnosis. Rearranged bands of the T-cell receptor gene were detected in all patients with unusual integration. These findings indicate that the integration patterns of HTLV-I proviral DNA have a clinical implication and may be one of the explanations for heterogeneity in the behavior of this disease.     

Production of granulocyte colony-stimulating factor in vitro by monocytes from preterm and term neonates [see comments]

We postulated that defective generation of granulocyte colony- stimulating factor (G-CSF) by cells of newborn infants might underlie their deficiencies in upregulating neutrophil production and function during bacterial infection. To test this, we isolated monocytes from the blood of preterm neonates, term neonates, and adults and, after stimulation with various concentrations of interleukin-1 alpha (IL-1 alpha) or lipopolysaccharide (LPS), quantified G-CSF concentrations in cell supernatants and G-CSF mRNA in cell lysates. When stimulated with plateau concentrations of IL-1 alpha for 24 hours, G-CSF concentrations were higher in supernatants of adult cells (8,699 +/- 5,529 pg/10(6) monocytes) than in those from term infants (2,557 +/- 442 pg, P < .05) or from preterm infants (879 +/- 348 pg, P < .05 v adults). When stimulated with plateau concentrations of LPS, supernatants of monocytes from preterm neonates had less G-CSF than did those from term neonates or adults. G-CSF mRNA content was low in cells from preterm infants, higher in those from term infants, and highest in those from adults. On the basis of the in vitro studies, we speculated that serum G-CSF concentrations might be less elevated in neutropenic neonates than in neutropenic adults. Indeed, serum concentrations were relatively low in all nonneutropenic subjects; 92 +/- 34 pg/mL (mean +/- SEM) in 10 preterm neonates, 114 +/- 21 pg/mL in 16 term neonates, and 45 +/- 13 pg/mL in 11 healthy adults. Serum concentrations were not elevated in 7 neutropenic neonates (39 +/- 17 pg/mL) but were in 8 neutropenic adults (2101 +/- 942 pg/mL, P < .05 v healthy adults). Other studies suggested that the lower G-CSF production in neonates is not counterbalanced by a heightened sensitivity of G-CSF--responsive progenitors to G-CSF. Therefore, we speculate that newborn infants, particularly those delivered prematurely, generate comparatively low quantities of G-CSF after inflammatory stimulation, and that this might constitute part of the explanation for their defective upregulation of neutrophil production and function during infection.     

Failure of recombinant human interleukin-3 therapy to induce erythropoiesis in patients with refractory Diamond-Blackfan anemia [see comments]

In two previous studies, we observed that recombinant human interleukin- 3 (IL-3) induced an increase in marrow burst-forming unit-erythroid- derived colonies in vitro in some patients with Diamond-Blackfan anemia (DBA). To determine whether a similar erythropoietic response could be induced in vivo, we treated 13 patients with DBA (aged 4 to 19 years) with two preparations of IL-3. All patients had absent absolute reticulocyte counts and markedly reduced to absent recognizable bone marrow erythroid elements; patients with circulating reticulocytes in the previous 12 months were excluded from study. All patients except 1 had failed steroid therapy and had been transfusion-dependent since infancy; 1 patient was maintained on high-dose prednisone at the time of enrollment. On the first arm of the study, IL-3 (Immunex Corp, Seattle, WA) was administered subcutaneously using a dose escalation regimen of 125 to 500 micrograms/m2/day in divided dosage at 12-hour intervals, coadministered with 1.5 mg/kg/d of oral ferrous sulphate. Of the 13 patients that entered the trial, 4 stopped prematurely because of adverse side effects. In the other 9 evaluable cases, reticulocytes increased transiently in 1 patient from 0 to 65 x 10(9)/L after 35 days of IL-3 therapy at 250 micrograms/m2, but transfusion dependency persisted. One transient peak in absolute reticulocyte count was noted in 6 other patients, but no erythroid response was observed after completion of a full course of IL-3. Oral prednisone at 0.5 mg/kg/d was then coadministered with IL-3 at 500 micrograms/m2 to 5 of the patients without effect, and treatment was stopped. In 2 patients, a second preparation of IL-3 (Sandoz Canada Inc, Dorval, Quebec, Canada) was initiated in a dose escalation regimen of 2.5 to 10 micrograms/kg and was coadministered with ferrous sulphate. No erythroid response was observed in either patient, and in one of the two, alternate-day subcutaneous recombinant erythropoietin at 300 U/kg was administered for 3 weeks in combination with daily IL-3 at 10 micrograms/kg, but no increased erythropoiesis was seen. Significant increases in white blood cell and eosinophil counts during administration of both preparations of IL-3 were observed in all patients. These data show that the response of DBA patients to IL-3 in vivo is heterogeneous and cannot be predicted from in vitro studies. The absence of a corrective effect of IL-3 in these patients with DBA indicates that a deficiency of the cytokine is not central in the pathogenesis of the disorder.     

Erythropoietin induces the expansion of c-kit+ progenitors for myeloid and erythroid cells, but not for lymphoid cells, in the bone marrow and liver

In humans, the numbers of erythrocytes and granulocytes, but not that of lymphocytes, tend to increase in parallel. To determine the mechanism, we investigated how the administration of erythropoietin induces the expansion of erythroid cells and other lineage cells in the bone marrow, liver, and other organs of mice. When mice were injected twice (days 1 and 2) with erythropoietin at a dose of 20 or 200 IU/day/ mouse, a prominent expansion of TER 19+ (erythroid cells) and Gr-1high cells (granulocytes) occurred in the liver, spleen, and bone marrow day 3 after the initial injection. On the other hand, lymphoid cells, including NK cells, extrathymic T cells, and conventional T cells, did not expand. In parallel with the expansion of erythroid cells and granulocytes, the levels of c-kit(+)Lin- cells increased in the liver and bone marrow. Despite the increase in the proportion of c-kit(+) Lin(-) cells, the generation of lymphocytes (e.g., T cells) decreased when such bone marrow cells were injected to scid mice. These results suggest that erythropoietin has the ability to induce the expansion of not only erythroid cells but also granulocytes in the liver, spleen, and bone marrow. Furthermore, c-kit+ progenitors which may commit themselves to erythroid and myeloid cells, but not to lymphoid cells, were also activated in the liver and bone marrow of mice treated with erythropoietin.     

Dengue virus, a flavivirus, propagates in human bone marrow progenitors and hematopoietic cell lines [see comments]

Dengue and other arbovirus diseases are frequently associated with bone marrow failure. We show that dengue type 4 (DEN4) propagates in colonies derived from immature human bone marrow progenitors. DEN4 was propagated in BFU-E-derived colonies and replication was dependent on erythropoietin. DEN4 was not cytotoxic. In inoculated cultures, diffuse bursts with many clusters contained large amounts of DEN4 RNA. In contrast to dengue infection of macrophages, virus propagation in semisolid culture was sustained and not enhanced by subneutralizing amounts of antibody. DEN4 also was efficiently propagated in human hematopoietic cell lines, especially those with erythroid properties. In K562 cells, DEN4 infection persisted for months; greatly slowed cell growth, again without cytotoxicity; and resulted in cytopathic changes in cell appearance. Flaviviruses can infect human hematopoietic cells and alter their proliferative capacity.