Effect of cytokines on the proliferation/differentiation of stroma-initiating cells

A culture system that identifies the precursor of murine bone marrow fibroblastic stromal cells (stroma-initiating cells, SIC) has been developed. In this system, mature fibroblasts are depleted by adherence to plastic dishes and the nonadherent cells are seeded at a low density, which results in the formation of colonies composed of fibroblastic cells. Macrophage colony-stimulating factor (M-CSF) has been shown to accelerate the colony formation in the system. In this study, we examined the stroma-inducing activity of a number of cytokines. Neither granulocyte-CSF, stem cell factor, interleukin (IL)-1, IL-6, transforming growth factor, epidermal growth factor, insulin-like growth factor, platelet-derived growth factor, nor fibroblast growth factor showed the activity. Similarly, tumor necrosis factor (TNF) did not show any stroma-inducing activity, but the factor inhibited the stromal colony formation induced by M-CSF. In this study, we found that granulocyte/macrophage-CSF (GM-CSF) and IL-3, as well as M-CSF had the stroma-inducing activity. Neither an additive nor synergistic effect was observed when the three factors were assayed in various combinations. The stroma-inducing activity of M-CSF, GM-CSF and IL-3 was observed even if lineage-negative bone marrow cells were used as target cells, suggesting that mature hematopoietic cells such as macrophages and granulocytes were not involved in the induction of stromal colony formation by these factors. Our results raise the possibility that GM-CSF and IL-3 as well as M-CSF stimulate the proliferation or differentiation of the precursor of bone marrow fibroblastic stromal cells.     

Mononuclear macrophage (M-CFC) colony formation in semisolid media in the absence of exogenous GM-CSF

Human fetal bone marrow (FBM) cells were examined for the ability to form colonies in the absence of exogenous colony-stimulating factor (CSF) in double layer agar, methylcellulose (MC), and in agar-MC (agar underlayer, MC overlayer) culture systems. Without exogenous CSF, macrophage colonies (M-CFC) were formed in a combined culture of agar and MC. Aggregates of 5-40 cells were observed on day 7. Gradually, large compact colonies which survived for 10-12 weeks of cultivation, were formed. They were composed of mononuclear monocytes and multinucleated cells. M-CFC progenitors were nonadherent, but their progeny became adherent during differentiation within the colony. Colony formation was cell-dose-dependent. Depletion of monocytes increased the number of colonies in agar-MC cultures and stimulated the development of some macrophage colonies in MC. Survival of monocyte progenitors was not dependent on CSF. Neither was their proliferation nor partial differentiation in agar-MC cultures. CSF increased M-CFC colony efficiency, however, if it was present when cultures were initiated. Addition of CSF to M-CFC growing for 2-5 weeks in CSF-deprived medium stimulated monocytes proliferation and transformation into macrophages. Epithelioid cells, an increase in the number of giant multinucleated cells, and granulocyte multiplication were also observed. The absolute dependence of macrophage colony formation on CSF described by others might be a result of inadequate culture conditions due to agar rather than an intrinsic physiological requirement.     

Hypothesis: the target cell of GM-CSF is a macrophage precursor capable to produce cells with the property to secrete a G-CSF like activity.

The induction of granulocyte and macrophage colony formation by the granulocyte-macrophage colony stimulating factor (GM-CSF) on bone marrow cells (BMC) was evaluated as a function of time in agar cultures. We found that while macrophage cell clusters were very abundant on the first two days of culture, granulocytic cell clusters did not appear until the third day. We also found that macrophage colonies were present from the fourth day of culture, while granulocyte colonies did not appear until the fifth day. When two day cell clusters were transferred to cultures with GM-CSF we observed that only macrophage-colonies developed. On the other hand, when four day clusters were transferred, both granulocyte and macrophage colony formation was obtained in a similar way as the one obtained when using GM-CSF with fresh BMC. Two day clusters did not respond to granulocyte colony stimulating factor (G-CSF) while fourth day clusters generated granulocytic colonies in a similar way as when G-CSF was used with fresh BMC. In order to test the hypothesis that granulocyte colony formation in these assays could be a result of the secretion of G-CSF by the macrophages previously induced by GM-CSF, lysates from macrophage colonies were used to induce colony formation on BMC. We observed that colonies, mainly granulocytic, were induced in a similar way as when G-CSF was used. Finally, the possibility that GM-CSF is just a macrophage inducer with the property to produce cells that secrete G-CSF is discussed.     

Role of granulocyte/macrophage colony-stimulating factor in the regulation of murine alveolar macrophage proliferation and differentiation

Granulocyte/macrophage (GM)-CSF is one of the hemopoietic growth factors that stimulates neutrophilic granulocyte and macrophage production by bone marrow progenitor cells. In this study, the effect of GM-CSF on the growth and differentiation of murine pulmonary alveolar macrophages (PAM) was investigated. In the presence of GM-CSF, normal murine PAM were induced to proliferate and develop into macrophage colonies with a dose-response curve similar to that of bone marrow GM colony-forming cells. PAM also responded to CSF-1, a lineage-restricted growth factor, but required much higher doses of CSF-1 and a longer incubation time for optimal colony formation. The proliferative response of PAM to CSF-1, however, was greatly enhanced by the concurrent addition of low doses of GM-CSF. In contrast, low doses of CSF-1 failed to potentiate the proliferative response of PAM to GM-CSF. Macrophages derived from GM-CSF cultures were rounder and less stretched and possessed less FcR-mediated phagocytic activity than cells produced in CSF-1 cultures. A study with hydrocortisone-induced monocytopenia showed that nearly one half of lung macrophages may be sustained by local proliferation of PAM without the continuous migration of blood monocytes. This study suggests that GM-CSF may play a major role in the production of PAM by two modes of action, 1) direct stimulation of cell proliferation and 2) enhancement of their responsiveness to CSF-1, thereby producing more mature and functionally competent macrophages.     

A recombinant human G-CSF/GM-CSF fusion protein from E. coli showing colony stimulating activity on human bone marrow cells

Granulocyte-Macrophage colony stimulating factor (GM-CSF) and Granulocyte colony stimulating factor (G-CSF) are cytokines involved in the differentiation of bone marrow progenitor cells into myeloid cells. They also activate mature myeloid cells to mediate a variety of antimicrobial activities and inflammatory responses. Recombinant GM-CSF and G-CSF proteins have been used to treat various diseases including cancer and hematopoietic diseases and to isolate peripheral blood progenitor cells for bone marrow transplantation. A plasmid construct expressing recombinant human G-CSF/GM-CSF fusion protein has now been prepared by linking the human G-CSF and GM-CSF coding regions and the recombinant fusion protein has been successfully expressed in E. coli. The recombinant human G-CSF/GM-CSF fusion protein was extracted and purified from the cellular inclusion and refolded into the biologically active form to show colony stimulating activity. The recombinant fusion protein exhibited colony stimulating activity on human bone marrow cell cultures, indicating that the linkage of GM-CSF and G-CSF by a linker peptide may not interrupt activities of the cytokines in the fusion protein. The colony forming unit of the fusion protein was also higher than those of the cultures treated with the same molar numbers of the recombinant human GM-CSF and G-CSF separately, which suggests that the fusion protein presumably retains both G-CSF and GM-CSF activities.     

The dependence of the formation of stromal CFU-f colonies on the stimulating action of hematopoietic cells

CFU-f-derived stromal colony formation was accomplished in adherent marrow cell cultures (AMCC) with serum-rich medium. It turned out to require additional stimulation by hemopoietic feeder cells: by irradiated marrow cells and spleen cells if they possess megakaryocytes and platelets or by platelets from the blood. PDGF, EGF and IL-3 did not substitute the colony stimulating activity of feeder cells. Thymus, lymph node cells and blood leucocytes had no colony stimulating activity. At low oxygen concentrations which improve colony formation the stimulating activity of hemopoietic feeder cells was expressed, as well. Thus, CFU-f colony formation depends on stimulation by hemopoietic cells in addition to serum growth factors. In full populations of marrow cells the CFU-f colony formation is stimulated by marrow cells which accompany the CFU-f.     

The effect of 1,25-dihydroxyvitamin D3 on hematopoiesis in long-term human bone marrow cultures

The modulatory effect of 1,25-dihydroxyvitamin D3 (vit D) on the growth of myeloid progenitors and on the composition of the stromal layer in human bone marrow long-term cultures was studied. Vit D (2 X 10(-8) M) caused an enhancement in myeloid progenitor cell (CFU-C) growth in the nonadherent and adherent layers during the entire 5-week incubation period. The vitamin did not alter the differentiation pattern of CFU-C (monocyte-macrophage progenitors CFU-M, granulocytic progenitors CFU-G, or monocyte-granulocyte progenitors CFU-GM). Vit D caused a marked increase in the percentage of lipid-containing cells in the adherent layer and an increase in the number of cells that specifically bound My4 monoclonal antibody (McAb), that reacted positively to fluoride-sensitive alpha-naphthyl acetate esterase, and that phagocytosed Candida albicans (CA). Concentrated supernatants harvested from control cultures showed significant levels of myeloid colony stimulating factor (CSF) activity. The addition of vit D to cultures for 5 weeks did not alter CSF levels. These results suggest that vit D may play a role in hematopoiesis by acting directly on the progenitor cells or via the stromal cell production of stimulatory factor(s).     

In vitro actions on hemopoietic cells of recombinant murine GM-CSF purified after production in Escherichia coli: comparison with purified native GM-CSF

Recombinant murine GM-CSF produced in Escherichia coli was purified to homogeneity and tested in parallel with purified native GM-CSF. Both recombinant and native GM-CSF stimulated granulocyte and/or macrophage colony formation by adult and fetal mouse progenitor cells, and with adult marrow cells the specific activity of the recombinant GM-CSF (25 X 10(8) U/mg) was similar to that of the native form (15 X 10(8) U/mg). At high concentrations (greater than 200 U/ml), both forms of GM-CSF also stimulated eosinophil colony formation by adult marrow cells and, at very high concentrations (greater than 800 U/ml), megakaryocyte and some erythroid and mixed-erythroid colony formation. Recombinant GM-CSF was as effective in stimulating the proliferation of the GM-CSF-dependent cell line FD as the native molecule. Both recombinant and native GM-CSF were able to induce partial differentiation in colonies of WEHI-3B myeloid leukemic cells. Recombinant GM-CSF competed effectively for the binding of 125I-labeled native GM-CSF to hemopoietic cells, and antiserum to recombinant GM-CSF also neutralized the biological activity of native GM-CSF. The bacterially synthesized GM-CSF was a slightly more effective stimulus for megakaryocyte colony formation than the native molecule. The demonstration that purified bacterially synthesized GM-CSF is biologically active in vitro now permits studies to be undertaken on the in vivo effects of this material.     

The suppressive effects of recombinant human tumor necrosis factor-alpha on normal and malignant myelopoiesis: synergism with interferon-gamma

The modulation of growth of normal and leukemic myeloid progenitor cells in soft agar cultures by recombinant human tumor necrosis factor-alpha (TNF alpha) and recombinant human interferon-gamma (IFN gamma) was investigated. TNF alpha inhibited colony formation of all colony types representing different maturational stages of normal progenitor cells committed to the myeloid lineage with different orders of sensitivity. Blast-type colonies derived from patients with acute myelogenous leukemia were more sensitive to TNF alpha inhibition than progenitor cells purified from normal bone marrow or bone marrow from patients with stable-phase chronic myelogenous leukemia. The response of most colony types to IFN gamma was poor. However, when IFN gamma was administered together with TNF alpha, synergistically enhanced antiproliferative effects were detected in all colony types tested. The antiproliferative action of IFN gamma on myelopoiesis was enhanced in culture by the presence of autologous monocytes, presumedly by inducing endogenous production of TNF alpha. However, TNF alpha seemed to act directly on the progenitor cells themselves to suppress their clonal growth, rather than involving accessory marrow elements such as monocytes and/or T lymphocytes.     

The effect of vitamin D3 metabolites on normal and leukemic bone marrow cells in vitro

We studied the effects of 1,25-dihydroxyvitamin D3 and other metabolites of vitamin D3 on the maturation in liquid culture and on colony formation in semisolid media of marrow and buffy coat cells from patients with myeloid leukemias and from normal individuals. After incubation with 1,25-dihydroxy-vitamin D3, a proportion of both normal and leukemic myeloid cells resembled cells of the monocyte-macrophage lineage; these cells expressed alpha-naphthylacetate esterase and were able to phagocytize and kill candida organisms. When granulocyte-macrophage progenitor cells (CFU-GM) were incubated with 1,25-dihydroxyvitamin D3, the number of monocyte-macrophage colonies was increased and the number of granulocyte colonies was reduced; megakaryocyte colony formation (CFU-Mk) was inhibited substantially; and there was no effect on erythroid (BFU-E) or multilineage (CFU-GEMM) progenitor cell colony formation. We propose that 1,25-dihydroxyvitamin D3 may induce cells that are normally committed to differentiate along the granulocytic pathways to differentiate instead along the monocyte-macrophage pathway. If these in vitro observations reflect the in vivo activity of 1,25-dihydroxyvitamin D3, it may be involved in the modulation of collagen deposits in the bone marrow.     

The effect of colony stimulating factor on the synthesis of ribonucleic acid by mouse bone marrow cells in vitro.

The effect of granulocyte-macrophage colony stimulating factor (GM-CSF) on the synthesis of RNA in liquid cultures of mouse bone marrow, spleen, thymus, peritoneal, peripheral blood leukocytes and lymph node cells was investigated. GM-CSF appeared to stimulate RNA-synthesis in syngeneic bone marrow cells within ten minutes of adding it to the culture. In the presence of GM-CSF bone marrow cultures maintained their initial rate of RNA synthesis for approximately ten hours. GM-CSF had no apparent effect on the uptake of 3H-uridine into bone marrow cells. This stimulation was still observed in the presence of puromycin and cycloheximide, but was abrogated by actinomycin D. The magnitude of the stimulation was not affected by the density of cells between 1 and 20 x 10(6) cells/ml but was slightly smaller at 0.1 and 40 x 10(6) cells/ml. Increasing concentration of GM-CSF (up to 2 X 105 units per ml) led to increased stimulation of RNA synthesis in bone marrow cells, but a significant stimulation could be detected at concentrations as low as 800 units/ml. GM-CSF did not significantly stimulate RNA synthesis in spleen, thymus, mesenteric or subcutaneous lymph node cells. However a small stimulation was observed in peripheral blood leukocytes and peritoneal cells. Autoradiographic studies showed that GM-CSF stimulated RNA synthesis in blast cells, myelocytes, metamyelocytes and polymorphs. Nucleated erythroid cells showed no increased labeling with GM-CFS. Labeling in lymphoid-like cells was highly variable but the level of labeling did not appear to be influenced by GM-CSF.     

Osteocalcin promotes differentiation of osteoclast progenitors from murine long-term bone marrow cultures

Murine long-term bone marrow cultures (LTBMCs) were used to generate hematopoietic cells free from marrow stromal cells. These progenitor cells were treated with GM-CSF (5 U/ml) with or without rat bone osteocalcin or rat serum albumin in either α-MEM with 2% heat-inactivated horse serum alone (α) or supplemented with 10% L-cell-conditioned medium (as a source of M-CSF) (L10). Few substrate-attached cells survived in basal α medium, but when treated with L10 medium or GM-CSF, they survived and proliferated. Osteocalcin did not significantly affect survival or proliferation. Subcultures of cells treated with GM-CSF had large numbers of multinucleated cells, more than half of which were tartrate-resistant acid phosphatase–positive (TRAP). Osteocalcin further promoted the development of TRAP-positive multinucleated cells; a dose of 0.7 μg/ml osteocalcin promoted osteoclastic differentiation by 60%. Using a novel microphotometric assay, we detected significantly more tartrate-resistant acid phosphatase activity in the osteocalcin plus GM-CSF group (75.6 ± 14.2) than in GM-CSF alone (53.3 ± 7.3). In the absence of M-CSF, GM-CSF stimulated tartrate-resistant acid phosphatase activity, but osteocalcin did not have an additional effect. These studies indicate that osteocalcin promotes osteoclastic differentiation of a stromal-free subpopulation of hematopoietic progenitors in the presence of GM-CSF and L-cell-conditioned medium. These results are consistent with the hypothesis that this bone-matrix constituent plays a role in bone resorption. © 1994 Wiley-Liss, Inc.     

Effects of GM-CSF deprivation on precursors of granulocytes and macrophages

Culture of C57BL bone marrow cells in the absence of GM-CSF led to a loss of recoverable granulocyte-macrophage colony-forming cells of 2% per hour. The rate of loss of progenitor cells in cultures of CBA fetal liver cells was 5–6% per hour. Surviving colony-forming cells exhibited a normal responsiveness to GM-CSF but generated smaller colonies than normal when subsequently stimulated by GM-CSF. Transfer of washed individual day-3 granulocyte-macrophage colony cells to cultures lacking GM-CSF indicated that most cells were unable to survive or proliferate in the absence of GM-CSF. Death of transferred cells was rapid and invariable when the cells were from macrophage-forming colonies. However some cells from 40–70% of granulocyte-forming colonies were able to undergo one or two divisions in the absence of GM-CSF. This phenomenon was seen most often with cells from colonies where matching colony cells exhibited a higher-than-average proliferative capacity in parallel stimulated cultures. The results indicate the difficulty that will be encoutered in obtaining valid metabolic dta from unstimulated populations of granulocyte-macrophage precursor cells. The ability of some granulocyte precursor cells to exhibit limited proliferation following GM-CSF deprivation suggests that significant amounts of GM-CSF may be bound to or be internalized in some precursor cells and result in cell division in the absence of GM-CSF from culture medium.     

Maintenance of granulopoiesis in long-term bone marrow cultures from W/Wv mice and effects of lipopolysaccharide on granulopoiesis in culture

An attempt was made to establish long-term cultures of marrow cells from genetically anaemic W/Wv mice. Two batches of horse sera were used. One batch of horse serum (HS-lot A) supported long-term maintenance (up to 20 weeks) of granulopoiesis in vitro. The number of suspension cells in W/Wv marrow culture was maintained at the same level as that in the control +/+ culture, but the number of granulocyte-macrophage progenitor cells (GM-CFC) and the ratio of immature to mature granulocytes were at a lower level than those in +/+ culture. These data suggest that haemopoietic progenitors in W/Wv cultures maintain a higher level of differentiation, and hence an increased self-renewal than those in +/+ cultures. Another batch of horse serum (HS-lot B) was less effective in the maintenance of the cultures, and the cultures deteriorated within 10 weeks. Addition of bacterial lipopolysaccharide (LPS) induced increased granulopoiesis in +/+ cultures, whereas such treatment resulted in the depletion of suspension cells in W/Wv cultures. The results suggest that haemopoietic cells of W/Wv mouse cannot cope with the strong stimulus for differentiation that occurs after the administration of LPS, although the cells can continue a moderately increased self-renewal and differentiation, as indicated by the results in the culture with HS-lot A.     

Fibroblast colonies in monolayer cultures of human bone marrow

In a liquid culture of human bone marrow, the development of fibroblast colonies takes place on days 6 to 9. Twenty percent fetal calf serum is used as the stimulus for fibroblast colony growth. Human bone marrow cells are plated as 2 × 10₅ cells in the culture. Normal human bone marrow yields 47 ± 4 fibroblasts colonies per 2 × 10₅ cells plated. Bone marrow fibroblast cultures using agar or methylcellulose restrict colony formation. Marked colony suppression was observed in acute leukemia, and a discrete colony number was observed in hypoplastic anemia. This fibroblast culture method should be applied to a larger number of patients to determine whether it has a pathognomonic value and clinical significance.     

Erythroid lineage-specific activity in conditioned medium derived from cloned human marrow stromal cells (CFU-RF)

Using long-term culture techniques, it has been shown that stromal cells in the marrow microenvironment are essential for the continued production and self-renewal of hematopoietic stem cells. We previously reported the development of a methylcellulose colony assay for a population of marrow stromal progenitors called CFU-RF. In this paper, a method is described for subculturing cells from individual CFU-RF-derived colonies to allow conditioned medium production (StCM). StCM, prepared in this way, was found to possess an erythroid lineage-specific activity that stimulated the formation of macroscopic erythroid colonies in cultures containing erythropoietin (epo). Using dose-response curves, the KG1 colony assay, and antibody neutralization, it was shown that the activity could not be attributed to interleukin 3 (IL3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). However, it was further shown that a monolayer of stromal cells, which had earlier been producing the erythroid activity, could be stimulated by IL1 to produce granulocytic colony-stimulating activity, but only as long as IL1 was present in the culture medium. These findings indicate a mechanism whereby the same stromal population could be modulated to promote growth and differentiation of different hematopoietic lineages.     

Functional activities of acidic isoferritins and lactoferrin in vitro and in vivo

The functional activities of acidic isoferritins (AIF) and lactoferin (LF) were evaluated. The inhibitory activity of AIF (AIFIA) was inactivated by preincubation with a monoclonal antibody (2A4) against AIF, but AIFIA was not inactivated by another monoclonal antibody against AIF (1C5), by a monoclonal antibody (3A5) against basic isoferritins, or by a heteroantiserum (LFT) against basic isoferritins. Monoclonal 2A4 also inactivated the inhibitory activity against colony formation by granulocyte-macrophage (CFU-GM) progenitor cells that was constitutively released by human monocytes or induced by human monocytes in the presence of OKT4+ lymphocytes. In addition to OKT4+ lymphocytes, the release of AIFIA from human monocytes was modulated by iron-saturated human LF and OKT8+ lymphocytes, both of which suppressed the release of AIFIA. Evidence for the physiologic relevance of AIF as a regulator of myelopoiesis was presented, in that human AIF suppressed the numbers of CFU-GM, BFU-E, and CFU-GEMM per femur and the cycling status of these cells in mice recovering from a sublethal dosage of Cytoxan. Abnormalities in LF and AIF interactions were found with cells from a pediatric patient with neutrophilia of unknown etiology that were consistent with the disease manifestations of neutrophilia. Polymorphonuclear neutrophils (PMN) from the patient contained low levels (1%-10% of control) of immunologically reactive LF and the LF found was ineffective as a suppressor molecule for the release of GM-CSF from normal mononuclear blood cells. In addition, the patient's GM-CSF releasing mononuclear blood cells were insensitive to the suppressive effects of purified LF, and colony formation by the patient's CFU-GM, but not BFU-E or CFU-GEMM, were insensitive to the suppressive effects of purified AIF. When the activity of purified AIF was assessed against mouse bone marrow cells under serum-free conditions, it was apparent that serum was not needed for the suppressive activity of AIF and that in some cases, serum actually masked the effects of AIF. Human monoblast cell line U937 was found to be a good model in vitro for the actions of LF and AIF; U937 cells induced for Ia-antigens by human gamma interferon were separated into populations of Ia-antigen+ and Ia-antigen- cells by fluorescence activated cell sorting (FACS), and LF and AIF suppressed colony formation only by the Ia-antigen+ U937 cells. A comparative analysis of bovine and human LF against release of GM-CSF from human mononuclear cells demonstrated that both were active in their iron-saturated form.(ABSTRACT TRUNCATED AT 400 WORDS)     

1,25-Dihydroxyvitamin D3 modulates bone marrow macrophage precursor proliferation and differentiation. Up-regulation of the mannose receptor

1,25-Dihydroxyvitamin D3 (1,25-(OH)2D3), the biologically active form of vitamin D3, has been shown to inhibit proliferation and promote monocytic differentiation of leukemic cell lines. In the present communication, we extend these observations to normal bone marrow macrophage precursors, and 1) identify the stage of monocytic maturation wherein the steroid exerts its antiproliferative effect, and 2) demonstrate that 1,25-(OH)2D3 promotes bone marrow macrophage differentiation as manifest by specific up-regulation of the lineage-specific membrane protein, the mannose-fucose receptor. In these experiments, the 1,25-(OH)2D3-mediated inhibitory effect on colony formation was shown to be independent of attendant levels of colony stimulating factor-1 and targeted through the adherent bone marrow macrophage precursor. Examination of this steroid-sensitive adherent precursor population demonstrates that its specific binding of 125I-mannose bovine serum albumin spontaneously and progressively increases with time in culture. Whereas adherent bone marrow macrophages cultured for 2 days express 3 X 10(4) mannose receptors/cell, the number of binding sites increases to 7 X 10(4)/cell by day 4. When bone marrow macrophage precursors are exposed to 1,25-(OH)2D3, an additional stepwise enhancement of 125I-mannose bovine serum albumin obtains with time. Four days of culture with the steroid results in 1.6 X 10(5) mannose receptors/cell, a 100% increase as compared to control cells. Neither duration of culture nor exposure to 1,25-(OH)2D3 alters the KD of 125I-mannose bovine serum albumin which approximates 3-5 X 10(-9) ml-1. Finally, the "specificity" of vitamin D-mediated up-regulation of the mannose receptor was established by demonstrating that the steroid does not alter binding of 125I-alpha-thrombin by bone marrow-derived macrophage precursors.     

Lactoferrin, transferrin and acidic isoferritins: regulatory molecules with potential therapeutic value in leukemia

In the process of evaluating roles for purified preparations of lactoferrin, transferrin and acidic isoferritins in the regulation of myelopoiesis, it was found that: (1) values reported for lactoferrin in the serum and plasma of normal donors are in most cases an over-estimation, (2) lactoferrin suppresses the production/release of granulocyte-macrophage colony stimulatory factors (GM-CSF) from monocytes in the absence of T-lymphocytes and also suppresses the production/release of acidic isoferritin-inhibitory activity from monocytes, (3) lactoferrin, transferrin and acidic isoferritins act on their specific target cells which express Ia-like antigens, (4) lactoferrin and transferrin act in vivo to suppress rebound myelopoiesis in mice recovering from sublethal dosages of Cytoxan, with preliminary observations suggesting that lactoferrin has a greater apparent effect on the bone marrow and transferrin has a greater apparent effect on the spleen, (5) active lactoferrin derives from Fc receptor positive subpopulations of PMN from patients with CML as well as from normal donors, but the percentage of Fc receptor containing PMN is lower in CML, as is the amount of active lactoferrin found in their PMN, and (6) lactoferrin, transferrin and acidic isoferritins suppress the colony formation of U937 clonogenic cells, with lactoferrin and transferrin decreasing the release of growth factors from U937 cells which are needed to stimulate U937 colony formation, and lactoferrin and acidic isoferritins suppress the colony formation of WEHI-3 cells, with lactoferrin decreasing the release of growth factors from WEHI-3 cells which are needed to stimulate WEHI-3 colony formation. Speculation on the potential usefulness of these iron binding glycoproteins to control of disease progression is given in the discussion.