Bone Marrow Stromal Macrophages Research Articles

CD4+ T Cells Alter the Stromal Microenvironment and Repress Medullary Erythropoiesis in Murine Visceral Leishmaniasis.

Human visceral leishmaniasis, a parasitic disease of major public health importance in developing countries, is characterized by variable degrees of severity of anemia, but the mechanisms underlying this change in peripheral blood have not been thoroughly explored. Here, we used an experimental model of visceral leishmaniasis in C57BL/6 mice to explore the basis of anemia following infection with Leishmania donovani. 28 days post-infection, mice showed bone marrow dyserythropoiesis by myelogram, with a reduction of TER119+ CD71−/+ erythroblasts. Reduction of medullary erythropoiesis coincided with loss of CD169high bone marrow stromal macrophages and a reduction of CXCL12-expressing stromal cells. Although the spleen is a site of extramedullary erythropoiesis and erythrophagocytosis, splenectomy did not impact the extent of anemia or affect the repression of medullary hematopoiesis that was observed in infected mice. In contrast, these changes in bone marrow erythropoiesis were not evident in B6.Rag2−/− mice, but could be fully reconstituted by adoptive transfer of IFNγ-producing but not IFNγ-deficient CD4+ T cells, mimicking the expansion of IFNγ-producing CD4+ T cells that occurs during infection in wild type mice. Collectively, these data indicate that anemia during experimental murine visceral leishmaniasis can be driven by defects associated with the bone marrow erythropoietic niche, and that this represents a further example of CD4+ T cell-mediated immunopathology affecting hematopoietic competence.

Regulation of steady-state neutrophil homeostasis by macrophages

AbstractThe timely clearance of apoptotic neutrophils from inflammation sites is an important function of macrophages; however, the role of macrophages in maintaining neutrophil homeostasis under steady-state conditions is less well understood. By conditionally deleting the antiapoptotic gene cellular FLICE-like inhibitory protein (C-FLIP) in myeloid cells, we have generated a novel mouse model deficient in marginal zone and bone marrow stromal macrophages. These mice develop severe neutrophilia, splenomegaly, extramedullary hematopoiesis, decreased body weight, and increased production of granulocyte colony-stimulating factor (G-CSF) and IL-1β, but not IL-17. c-FLIPf/f LysM-Cre mice exhibit delayed clearance of circulating neutrophils, suggesting that failure of macrophages to efficiently clear apoptotic neutrophils causes production of cytokines that drive excess granulopoiesis. Further, blocking G-CSF but not IL-1R signaling in vivo rescues this neutrophilia, suggesting that a G-CSF–dependent, IL-1β–independent pathway plays a role in promoting neutrophil production in mice with defective clearance of apoptotic cells.

The role of the bone marrow in neutrophil clearance under homeostatic conditions in the mouse.

In humans, 1011 neutrophils are released from the bone marrow per day, and these cells have a half-life in the blood of only ∼6.5 h. Although it is generally believed that neutrophils are cleared from the circulation via the liver and spleen, in this study using 111In-labeled senescent neutrophils, we show that in mice, 32% of neutrophils are cleared from the circulation via the bone marrow. We have previously shown that senescent neutrophils home to the bone marrow in a CXCR4-dependent manner, and we show here that pretreatment of neutrophils with pertussis toxin significantly inhibits neutrophil clearance via the bone marrow (75%), consistent with a role for chemokines in this process. By labeling senescent neutrophils with inert fluorescent microspheres, we have tracked their fate and shown that in vivo, they are ultimately phagocytosed by bone marrow stromal macrophages. Finally, we show that under noninflammatory conditions, circulating levels of neutrophils are regulated by granulocyte-colony stimulating factor (G-CSF), but not interleukin-17. Interestingly, we report that the uptake of apoptotic neutrophils by bone marrow macrophages stimulates their production of G-CSF in vitro. Taken together, these data provide evidence that the bone marrow represents a major site of neutrophil clearance in mice.—Furze, R. C., Rankin, S. M. The role of the bone marrow in neutrophil clearance under homeostatic conditions in the mouse.

Patients with non-immune chronic idiopathic neutropenia syndrome have increased splenic volume on ultrasonography.

Clinically detectable splenomegaly is rarely seen in patients with non-immune chronic idiopathic neutropenia syndrome (NI-CINS). Using ultrasound, we estimated splenic volume in 52 NI-CINS patients and 14 age- and sex-matched normal controls by determining the "corrected splenic index" (CSI) from the product of length, width and thickness of the organ expressed in cm3/m2 body surface area. We found that CSI was significantly higher in the group of patients compared to controls (202.8 +/- 82.0 vs. 133.8 +/- 28.1 cm3/m2, P=0.003), and that individual CSI values was inversely correlated with the number of circulating neutrophils (r=-0.5097, P < 0.0001). About 48.1% of the patients had CSI above 190 cm3/m2 body surface, representing the upper 95% confidence limit of values found in the controls. Patients also had increased serum concentrations of pro-inflammatory cytokines and chemokines mainly produced by activated macrophages (IL-1beta, TNF-alpha, RANTES and IL-8), as well as increased serum levels of soluble cell adhesion molecules derived from activated endothelium (sE-Selectin, sICAM and sVCAM). We hypothesize that the increased splenic volume in NI-CINS patients may be due to the accumulation of activated macrophages inside the spleen, possibly as the result of an unrecognized low-grade chronic inflammatory process. The nature of such an inflammation is unknown. A study was designed to search for viral or bacterial genomic material in patients' bone marrow stromal macrophages in which the unknown causal agent might be located.

Leishmania donovani infection of bone marrow stromal macrophages selectively enhances myelopoiesis, by a mechanism involving GM-CSF and TNF-α

Alterations in hematopoiesis are common in experimental infectious disease. However, few studies have addressed the mechanisms underlying changes in hematopoietic function or assessed the direct impact of infectious agents on the cells that regulate these processes. In experimental visceral leishmaniasis, caused by infection with the protozoan parasite Leishmania donovani, parasites persist in the spleen and bone marrow, and their expansion in these sites is associated with increases in local hematopoietic activity. The results of this study show that L donovani targets bone marrow stromal macrophages in vivo and can infect and multiply in stromal cell lines of macrophage, but not other lineages in vitro. Infection of stromal macrophages increases their capacity to support myelopoiesis in vitro, an effect mediated mainly through the induction of granulocyte macrophage-colony stimulating factor and tumor necrosis factor-alpha. These data are the first to directly demonstrate that intracellular parasitism of a stromal cell population may modify its capacity to regulate hematopoiesis during infectious disease. (Blood. 2000;95:1642-1651)

Sialoadhesin expression by bone marrow macrophages derived from Ehrlich-tumor-bearing mice.

Sialoadhesin (sheep erythrocyte receptor, SER) is a macrophage-restricted adhesion molecule that binds certain sialylated ligands. It is borne by bone marrow stromal macrophages, promoting the interaction with developing myeloid cells, and by a subset of tissue macrophages involved in antigen presentation and activation of tumor-reactive T cells. The expression of sialoadhesin on SER+ macrophages is not constitutive but requires the continuous supply of a sialoadhesin-inducing serum factor. Tumor growth is often associated with marked alterations of myelopoiesis and impairment of T cell activation; yet the expression of sialoadhesin in macrophages derived from tumor bearers has not been addressed. The aim of this study was to assess whether Ehrlich tumor (ET) - a murine mammary carcinoma - growth may modify the sialoadhesin expression by bone marrow macrophages and/or sialoadhesin-inducing activity in ET-bearing sera. Moreover, putative functional sialoadhesin inhibitors produced by ET cells were tested. The results indicate that bone marrow cells from ET bearers show a seven- to eight-fold decrease in SER+ cells as detected by flow cytometry. This is accompanied by an overall decrease in sheep erythrocyte binding to tumor-bearer-derived bone marrow cells, but also by lower numbers of plastic-adherent cells. Functional sialoadhesin expression is preserved at the single-cell level and no inhibitors are found in ET-bearing sera or ET cell culture supernatants. Tumor progression does not impair the sialoadhesin-inducing activity of ET-bearing sera, or the ability of SER- macrophages (e.g. peritoneal macrophages) to respond to such an induction. In conclusion, while SER+ macrophages are greatly decreased in bone marrow from ET bearers, this is not due to a down-regulation of sialoadhesin expression, nor to an impairment of sialoadhesin-inducing factor or to the presence of sialoadhesin-binding moieties of tumor origin, but, more likely, to a decrease of fully mature macrophages.

Establishment and characterization of murine bone marrow stromal macrophage line

The authors established a murine bone marrow stromal cell line QXMSC1 by discarding the suspensible hematopoietic cells and passaging the adherent stromal cells many times in long-term bone marrow culture. QXMSC1 cells have been passaged 65 times for 15 months and immortalized. The mean number of QXMSC1 chromosomes is 66 ± 4. The appearance of the cells is elliptical. There are many pseudopods and mononuclei under the optical microscope. Many lisosomes and phagosomes in cytoplasm exist under the transmission electron microscope. There are no desmosome junction between the cells, no lipid drops, no intermediate filament. Vimentin is positive and keratin is negative by stain of immunocytochemistry. Nonspecific lipase staining is positive. These results indicate that QXMSC1 cells are murine bone marrow macrophages.

Role for interleukin-1 (IL-1) in benzene-induced hematotoxicity: inhibition of conversion of pre-IL-1 alpha to mature cytokine in murine macrophages by hydroquinone and prevention of benzene-induced hematotoxicity in mice by IL-1 alpha

Chronic exposure of humans to benzene (BZ), a myelotoxin, causes aplastic anemia and acute leukemia. The stromal macrophage that produces interleukin-1 (IL-1), a cytokine essential for hematopoiesis, is a target of BZ's toxicity. Monocyte dysfunction and decreased IL-1 production have been shown to be involved in aplastic anemia in humans. Hydroquinone (HQ), a toxic bone marrow (BM) metabolite of BZ, causes time- and concentration-dependent inhibition of processing of the 34-Kd pre-interleukin-1 alpha (IL-1 alpha) to the 17-Kd mature cytokine in murine P388D1 macrophages and BM stromal macrophages, as measured by Western immunoblots of cell lysate proteins using a polyclonal rabbit antimurine IL-1 alpha antibody. HQ over a 10-fold concentration range had no effect on the lipopolysaccharide (LPS)-induced production of pre- IL-1 alpha precursor or on cell viability or DNA and protein synthesis. Stromal macrophages obtained from the femoral BM of C57Bl/6 mice exposed to BZ (600 or 800 mg/kg body weight) for 2 days were incapable of processing the 34-Kd pre-IL-1 alpha to the mature 17-Kd cytokine when stimulated in culture with LPS. Stromal macrophages from mice coadministered BZ and indomethacin, a prostaglandin H synthase (PHS) inhibitor that has been shown to prevent BZ-induced myelotoxic and genotoxic effects in mice when coadministered with benzene were able to convert the pre-IL-1 alpha to mature cytokine. Administration of recombinant murine IL-1 alpha (rMuIL-1 alpha) to mice before a dose of BZ that causes severe depression of BM cellularity completely prevents BM depression, most probably by bypassing the inability of the stromal macrophage in BZ-treated animals to process pre-IL-1 alpha to the mature cytokine.

Role for Interleukin-1 (IL-1) in Benzene-Induced Hematotoxicity: Inhibition of Conversion of Pre-IL-lα to Mature Cytokine in Murine Macrophages by Hydroquinone and Prevention of Benzene-Induced Hematotoxicity in Mice by IL-lα

Chronic exposure of humans to benzene (BZ), a myelotoxin, causes aplastic anemia and acute leukemia. The stromal macrophage that produces interleukin-1 (IL-1), a cytokine essential for hematopoiesis, is a target of BZ's toxicity. Monocyte dysfunction and decreased IL-1 production have been shown to be involved in aplastic anemia in humans. Hydroquinone (HQ), a toxic bone marrow (BM) metabolite of BZ, causes time- and concentration-dependent inhibition of processing of the 34-Kd pre-interleukin-1 alpha (IL-1 alpha) to the 17-Kd mature cytokine in murine P388D1 macrophages and BM stromal macrophages, as measured by Western immunoblots of cell lysate proteins using a polyclonal rabbit antimurine IL-1 alpha antibody. HQ over a 10-fold concentration range had no effect on the lipopolysaccharide (LPS)-induced production of pre-IL-1 alpha precursor or on cell viability or DNA and protein synthesis. Stromal macrophages obtained from the femoral BM of C57Bl/6 mice exposed to BZ (600 or 800 mg/kg body weight) for 2 days were incapable of processing the 34-Kd pre-IL-1 alpha to the mature 17-Kd cytokine when stimulated in culture with LPS. Stromal macrophages from mice coadministered BZ and indomethacin, a prostaglandin H synthase (PHS) inhibitor that has been shown to prevent BZ-induced myelotoxic and genotoxic effects in mice when coadministered with benzene were able to convert the pre-IL-1 alpha to mature cytokine. Administration of recombinant murine IL-1 alpha (rMuIL-1 alpha) to mice before a dose of BZ that causes severe depression of BM cellularity completely prevents BM depression, most probably by bypassing the inability of the stromal macrophage in BZ-treated animals to process pre-IL-1 alpha to the mature cytokine.

Isolation and immunocytochemical characterization of human bone marrow stromal macrophages in hemopoietic clusters.

Stromal macrophages (M phi) have been localized in situ and isolated within erythroid clusters from human marrow. Stromal M phi arborize in an extensive network uniformly distributed throughout marrow interstitium, and express the phenotype CD4+, CD11a+, CD11c+, CD13+, CD14+, CD16+, CD18+, CD31+, CD32+, FcRI+, HLA-DR+, and CD35-, transferrin receptor-negative, and CD11b (weak). They express endocytic receptor antigens, but show significant differences in myeloid antigen expression compared with freshly harvested or cultured monocytes. Human stromal M phi are therefore specialized mature marrow M phi that are accessible for further investigations in infectious, storage, or hemopoietic disorders.