RESUMEN

Galectin-3 (Gal-3) controls intercellular and cell-extracellular matrix interactions during immunological responses. In chronic inflammation, Gal-3 is associated with fibrotic events, regulates B cell differentiation and delays lupus progression. Gal-3 deficient mice (Lgals3-/-) have intense germinal center formation and atypical plasma cell generation correlated to high levels IgG, IgE, and IgA. Here, we used pristane (2,6,10,14-tetramethylpentadecane) to induce lupus-like syndrome in Lgals3-/- and Lgals3+/+ BALB/c mice. Mesentery and peritoneal cells were monitored because promptly react to pristane injected in the peritoneal cavity. For the first time, mesenteric tissues have been associated to the pathogenesis of experimental lupus-like syndrome. In Lgals3+/+ pristane-induced mice, mesentery was hallmarked by intense fibrogranulomatous reaction restricted to submesothelial regions and organized niches containing macrophages and B lymphocytes and plasma cells. In contrast, Lgals3-/- pristane-treated mice had diffuse mesenteric fibrosis affecting submesothelium and peripheral tissues, atypical M1/M2 macrophage polarization and significant DLL1+ cells expansion, suggesting possible involvement of Notch/Delta pathways in the disease. Early inflammatory reaction to pristane was characterized by significant disturbances on monocyte recruitment, macrophage differentiation and dendritic cell (DC) responses in the peritoneal cavity of pristane-induced Lgals3-/- mice. A correlative analysis showed that mesenteric damages in the absence of Gal-3 were directly associated with severe portal inflammation and hepatitis. In conclusion, it has suggested that Gal-3 orchestrates histological organization in the mesentery and prevents lupoid hepatitis in experimental lupus-like syndrome by controlling macrophage polarization, Notch signaling pathways and DC differentiation in mesenteric structures.

Asunto(s)

Galectina 3/metabolismo , Hepatitis/inmunología , Lupus Eritematoso Sistémico/inmunología , Macrófagos Peritoneales/inmunología , Mesenterio/patología , Animales , Modelos Animales de Enfermedad , Femenino , Fibrosis , Galectina 3/genética , Hepatitis/patología , Humanos , Inyecciones Intraperitoneales , Hígado/inmunología , Hígado/patología , Lupus Eritematoso Sistémico/inducido químicamente , Lupus Eritematoso Sistémico/complicaciones , Mesenterio/citología , Mesenterio/inmunología , Ratones , Ratones Noqueados , Terpenos/administración & dosificación , Terpenos/inmunología

RESUMEN

The peritoneum is a thin membrane that covers most of the abdominal organs, composed of a monolayer of mesothelial cells and subjacent submesothelial loose connective tissue. Cells from the peritoneal wall are correlated with peritoneal fibrosis and epithelial-to-mesenchymal transition. However, the distinct involvement of mesothelial or submesothelial cells in such phenomena is still not clear. Here, we propose a new strategy to obtain stromal cells from anterior peritoneal wall explant cultures. These cells migrated from peritoneal tissues and proliferated in vitro for 4 weeks as adherent fibroblast-like cells. Optical and electronic microscopy analyses of the fragments revealed a significant submesothelial disorganization. The obtained cells were characterized as cytokeratin- vimentin+ laminin+ α-smooth muscle actin+, suggesting a connective tissue origin. Moreover, at the third passage, these stromal cells were CD90+CD73+CD29+Flk-1+CD45-, a phenotype normally attributed to cells of mesenchymal origin. These cells were able to support hematopoiesis, expressing genes involved in myelopoiesis (SCF, G-CSF, GM-CSF, IL-7 and CXCL-12), and differentiated into osteogenic and adipogenic cell lineages. The methodology demonstrated in this work can be considered an excellent experimental model to understand the physiology of the peritoneal wall in healthy and pathological processes. Moreover, this work shows for the first time that submesothelial stromal cells have properties similar to those of mesenchymal cells from other origins.

Asunto(s)

Adipogénesis , Linaje de la Célula , Epitelio/metabolismo , Hematopoyesis , Osteogénesis , Peritoneo/citología , Animales , Movimiento Celular , Separación Celular , Técnicas de Cocultivo , Citometría de Flujo , Cinética , Masculino , Ratones Endogámicos BALB C , Mielopoyesis , Peritoneo/ultraestructura , Fenotipo , Células del Estroma/citología , Células del Estroma/metabolismo

RESUMEN

Galectin-3 (gal-3) is a ß-galactoside binding protein present in multivalent complexes with an extracellular matrix and with cell surface glycoconjugates. In this context, it can deliver a variety of intracellular signals to modulate cell activation, differentiation and survival. In the hematopoietic system, it was demonstrated that gal-3 is expressed in myeloid cells and surrounding stromal cells. Furthermore, exogenous and surface gal-3 drive the proliferation of myeloblasts in a granulocyte-macrophage colony-stimulating factor (GM-CSF)-dependent manner. Here, we investigated whether gal-3 regulates the formation of myeloid bone marrow compartments by studying galectin-3(-/-) mice (gal-3(-/-)) in the C57BL/6 background. The bone marrow histology of gal-3(-/-) mice was significantly modified and the myeloid compartments drastically disturbed, in comparison with wild-type (WT) animals. In the absence of gal-3, we found reduced cell density and diaphyseal disorders containing increased trabecular projections into the marrow cavity. Moreover, myeloid cells presented limited capacity to differentiate into mature myeloid cell populations in gal-3(-/-) mice and the number of hematopoietic multipotent progenitors was increased relative to WT animals. In addition, bone marrow stromal cells of these mice had reduced levels of GM-CSF gene expression. Taken together, our data suggest that gal-3 interferes with hematopoiesis, controlling both precursors and stromal cells and favors terminal differentiation of myeloid progenitors rather than proliferation.

Asunto(s)

Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Galectina 3/deficiencia , Animales , Diferenciación Celular , Galectina 3/genética , Galectina 3/metabolismo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Endogamia , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células del Estroma/citología , Células del Estroma/metabolismo

RESUMEN

Galectin-3 (gal-3), a beta-galactoside-binding animal lectin, plays a role in cell-cell and cell-extracellular matrix interactions. Extracellular gal-3 modulates cell migration and adhesion in several physiological and pathological processes. Gal-3 is highly expressed in activated macrophages. Schistosoma mansoni eggs display a large amount of gal-3 ligands on their surface and elicit a well-characterized, macrophage-dependent, granulomatous, inflammatory reaction. Here, we have investigated the acute and chronic phases of S. mansoni infection in wild-type and gal-3(-/-) mice. In the absence of gal-3, chronic-phase granulomas were smaller in diameter, displaying thinner collagen fibers with a loose orientation. Schistosoma-infected gal-3(-/-) mice had remarkable changes in the monocyte/macrophage, eosinophil, and B lymphocyte subpopulations as compared with the infected wild-type mice. We observed a reduction of macrophage number, an increase in eosinophil absolute number, and a decrease in B lymphocyte subpopulation (B220(+/high) cells) in the periphery during the evolution of the disease in gal-3(-/-) mice. B lymphopenia was followed by an increase of plasma cell number in bone marrow, spleen, and mesenteric lymph nodes of the infected gal-3(-/-) mice. The plasma IgG and IgE levels also increased in these mice. Gal-3 plays a role in the organization, collagen distribution, and mobilization of inflammatory cells to chronic-phase granulomas, niches for extramedullary myelopoiesis, besides interfering with monocyte-to-macrophage and B cell-to-plasma cell differentiation.

Asunto(s)

Diferenciación Celular , Galectina 3/genética , Ganglios Linfáticos/citología , Esquistosomiasis/inmunología , Enfermedad Aguda , Animales , Linfocitos B/citología , Linfocitos B/fisiología , Recuento de Células , Enfermedad Crónica , Cruzamientos Genéticos , Eosinófilos/citología , Eosinófilos/fisiología , Femenino , Granuloma/etiología , Granuloma/patología , Inmunoglobulina E/sangre , Inmunoglobulina G/sangre , Inmunohistoquímica , Cinética , Hígado/patología , Ganglios Linfáticos/fisiología , Linfopenia , Macrófagos/citología , Macrófagos/fisiología , Masculino , Mesenterio/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Monocitos/fisiología , Células Plasmáticas/citología , Schistosoma mansoni/inmunología , Schistosoma mansoni/patogenicidad , Esquistosomiasis/metabolismo

RESUMEN

In a previous study we monitored the distribution and phenotype expression of B1 cells during the evolution of experimental murine schistosomiasis mansoni and we proposed that the B1 cells were heterogeneous: a fraction which originated in the spleen and followed the migratory pathway to mesenteric ganglia, while the other was the resident peritoneal B1-cell pool. In the present study, we have addressed the question of whether these two B1-lymphocyte populations are involved in the production of the late Ig isotype IgE, which is present in high levels in schistosomal infection. Lymphocyte expression of surface markers and immunoglobulins were monitored by immunofluorescence flow cytometry. Both in the spleen and mesenteric ganglia, the B1 and B2 cells were induced to switch from IgM to IgE in the early Th2-dominated phase of the disease, with an increase of IgE in its later phases. Conversely, peritoneal B1-IgM+ switched to the remaining IgE+ present in high numbers in the peritoneal cavity throughout the disease. We correlated the efficient induction of the expression of late Ig isotypes by B1 cells with high levels of inflammatory cytokines due to the intense host response to the presence of worms and their eggs in the abdominal cavity. In conclusion, B1 cells have a different switch behavior from IgM to IgE indicating that these cell sub-populations depend on the microenvironment.

Asunto(s)

Linfocitos B/inmunología , Inmunoglobulina E/metabolismo , Esquistosomiasis mansoni/inmunología , Animales , Antígenos de Superficie/inmunología , Linfocitos B/metabolismo , Modelos Animales de Enfermedad , Femenino , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Inmunoglobulina E/inmunología , Activación de Linfocitos , Masculino , Ratones , Ratones Endogámicos C3H , Cavidad Peritoneal/citología

RESUMEN

In a previous study we monitored the distribution and phenotype expression of B1 cells during the evolution of experimental murine schistosomiasis mansoni and we proposed that the B1 cells were heterogeneous: a fraction which originated in the spleen and followed the migratory pathway to mesenteric ganglia, while the other was the resident peritoneal B1-cell pool. In the present study, we have addressed the question of whether these two B1-lymphocyte populations are involved in the production of the late Ig isotype IgE, which is present in high levels in schistosomal infection. Lymphocyte expression of surface markers and immunoglobulins were monitored by immunofluorescence flow cytometry. Both in the spleen and mesenteric ganglia, the B1 and B2 cells were induced to switch from IgM to IgE in the early Th2-dominated phase of the disease, with an increase of IgE in its later phases. Conversely, peritoneal B1-IgM+ switched to the remaining IgE+ present in high numbers in the peritoneal cavity throughout the disease. We correlated the efficient induction of the expression of late Ig isotypes by B1 cells with high levels of inflammatory cytokines due to the intense host response to the presence of worms and their eggs in the abdominal cavity. In conclusion, B1 cells have a different switch behavior from IgM to IgE indicating that these cell sub-populations depend on the microenvironment.

Asunto(s)

Animales , Femenino , Masculino , Ratones , Linfocitos B/inmunología , Inmunoglobulina E/metabolismo , Esquistosomiasis mansoni/inmunología , Antígenos de Superficie/inmunología , Linfocitos B/metabolismo , Modelos Animales de Enfermedad , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Inmunoglobulina E/inmunología , Activación de Linfocitos , Cavidad Peritoneal/citología

RESUMEN

Dipeptidyl peptidase IV (DPP-IV; CD26) (EC 3.4.14.5) is a membrane-anchored ectoenzyme with N-terminal exopeptidase activity that preferentially cleaves X-Pro-dipeptides. It can also be spontaneously released to act in the extracellular environment or associated with the extracellular matrix. Many hematopoietic cytokines and chemokines contain DPP-IV-susceptible N-terminal sequences. We monitored DPP-IV expression and activity in murine bone marrow and liver stroma cells which sustain hematopoiesis, myeloid precursors, skin fibroblasts, and myoblasts. RT-PCR analysis showed that all these cells produced mRNA for DPP-IV. Partially purified protein reacted with a commercial antibody to CD26. The K M values for Gly-Pro-p-nitroanilide ranged from 0.43 to 0.98 mM for the membrane-associated enzyme of connective tissue stromas, and from 6.76 to 8.86 mM for the enzyme released from the membrane, corresponding to a ten-fold difference, but only a two-fold difference in K M was found in myoblasts. K M of the released soluble enzyme decreased in the presence of glycosaminoglycans, nonsulfated polysaccharide polymers (0.8-10 æg/ml) or simple sugars (320-350 æg/ml). Purified membrane lipid rafts contained nearly 3/4 of the total cell enzyme activity, whose K M was three-fold decreased as compared to the total cell membrane pool, indicating that, in the hematopoietic environment, DPP-IV activity is essentially located in the lipid rafts. This is compatible with membrane-associated events and direct cell-cell interactions, whilst the long-range activity depending upon soluble enzyme is less probable in view of the low affinity of this form

Asunto(s)

Animales , Ratones , Ratas , Células de la Médula Ósea , Dipeptidil Peptidasa 4 , Células del Estroma , Línea Celular , Dipeptidil Peptidasa 4 , Expresión Génica , Sistema Hematopoyético , Immunoblotting , Hígado , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , ARN Mensajero

RESUMEN

Dipeptidyl peptidase IV (DPP-IV; CD26) (EC 3.4.14.5) is a membrane-anchored ectoenzyme with N-terminal exopeptidase activity that preferentially cleaves X-Pro-dipeptides. It can also be spontaneously released to act in the extracellular environment or associated with the extracellular matrix. Many hematopoietic cytokines and chemokines contain DPP-IV-susceptible N-terminal sequences. We monitored DPP-IV expression and activity in murine bone marrow and liver stroma cells which sustain hematopoiesis, myeloid precursors, skin fibroblasts, and myoblasts. RT-PCR analysis showed that all these cells produced mRNA for DPP-IV. Partially purified protein reacted with a commercial antibody to CD26. The K M values for Gly-Pro-p-nitroanilide ranged from 0.43 to 0.98 mM for the membrane-associated enzyme of connective tissue stromas, and from 6.76 to 8.86 mM for the enzyme released from the membrane, corresponding to a ten-fold difference, but only a two-fold difference in K M was found in myoblasts. K M of the released soluble enzyme decreased in the presence of glycosaminoglycans, nonsulfated polysaccharide polymers (0.8-10 micro g/ml) or simple sugars (320-350 micro g/ml). Purified membrane lipid rafts contained nearly 3/4 of the total cell enzyme activity, whose K M was three-fold decreased as compared to the total cell membrane pool, indicating that, in the hematopoietic environment, DPP-IV activity is essentially located in the lipid rafts. This is compatible with membrane-associated events and direct cell-cell interactions, whilst the long-range activity depending upon soluble enzyme is less probable in view of the low affinity of this form.

Asunto(s)

Células de la Médula Ósea/enzimología , Dipeptidil Peptidasa 4/fisiología , Microdominios de Membrana/enzimología , Células del Estroma/enzimología , Animales , Línea Celular , Dipeptidil Peptidasa 4/genética , Expresión Génica , Sistema Hematopoyético/enzimología , Immunoblotting , Hígado/citología , Hígado/enzimología , Ratones , ARN Mensajero/biosíntesis , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

Systemic production and mobilization of inflammatory cells and formation of hepatic periovular granulomas were studied in Schistosoma mansoni-infected mice with deficient interferon gamma (IFN-gamma) receptor (IFN-gammaR(o/o)). The impaired IFN-gamma signaling did not cause a significant modification of the overall kinetics of inflammatory cells, but mutant mice developed smaller hepatic periovular granulomas with a two-fold reduction in all the cell lineages. In granulomas of normal mice, the fully differentiated macrophages were progressively predominant, whilst in IFN-gammaR(o/o) mice, the granulomas contained a higher percentage of immature and proliferating monocytes. Granulomas of IFN-gammaR(o/o) mice had an enhanced and accelerated fibrotic reaction, corresponding to an increased content of proliferative and activated connective tissue cells. Simultaneously, their granulomas had an increased ratio of T over B cells, with an increase in CD8(+) and a reduction in CD4(+) T cells. The functional IFN-gamma receptor was not required for initial recruitment of monocytes and lymphocytes into granulomas, but it was necessary for the maturation of macrophages, upregulation of major histocompatibility class 2 (MHC-II) expression and consequent stimulation of lymphocyte subpopulations depending upon the MHC-II-mediated antigen presentation.

Asunto(s)

Granuloma/inmunología , Parasitosis Hepáticas/inmunología , Receptores de Interferón/deficiencia , Esquistosomiasis mansoni/inmunología , Animales , Células del Tejido Conectivo/citología , Células del Tejido Conectivo/inmunología , Citometría de Flujo , Granuloma/patología , Granuloma/fisiopatología , Parasitosis Hepáticas/patología , Parasitosis Hepáticas/fisiopatología , Linfocitos/citología , Linfocitos/inmunología , Macrófagos/citología , Macrófagos/inmunología , Ratones , Ratones Noqueados , Receptores de Interferón/genética , Receptores de Interferón/metabolismo , Schistosoma mansoni/crecimiento & desarrollo , Schistosoma mansoni/patogenicidad , Esquistosomiasis mansoni/patología , Esquistosomiasis mansoni/fisiopatología , Receptor de Interferón gamma

RESUMEN

Inflammatory granulomatous reactions in liver elicited by schistosomal infection have been shown to function as active extramedullar myelopoietic sites, producing potentially all the myeloid lineages. We have now addressed the question of the extramedullar B lymphopoiesis in these sites. We have shown the presence of early B cell precursors (pro-B cells) in the granulomas by immunophenotyping. Their total number in the liver was equivalent to the pro-B cells in the bone marrow of one femur. In agreement with their phenotype, the RT-PCR analysis showed that these cells expressed RAG-1 and lambda5 genes. However, the conversion of the pro-B to pre-B cells was not observed and no clonogenic B cell precursors could be detected in semi-solid cultures stimulated by IL-7. The granulomatous stroma was shown to produce IL-7 and express c-kit, and was able to sustain the full B lymphopoiesis in vitro. Conversely, the granuloma supernatant was shown to inhibit actively the development of B lymphocytes. We conclude that the granuloma environment elicits homing and proliferation of totipotent hematopoietic precursors, and that it is permissive for early commitment to the B cell lineage, but the full extramedullar production of B cell is abrogated by soluble factors produced inside the granulomas.

Asunto(s)

Linfocitos B/inmunología , Granuloma/inmunología , Hematopoyesis Extramedular/inmunología , Parasitosis Hepáticas/inmunología , Esquistosomiasis/inmunología , Animales , Linfocitos B/citología , Células de la Médula Ósea , Diferenciación Celular , Células Cultivadas , Femenino , Granuloma/patología , Hematopoyesis Extramedular/genética , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Inmunofenotipificación , Interleucina-7/metabolismo , Parasitosis Hepáticas/patología , Masculino , Ratones , Ratones Endogámicos C3H , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Esquistosomiasis/patología

RESUMEN

In schistosomiasis a systemic hyperplasia of the monomacrophagic cell lineage is associated with its mild modifications in myelograms and hemograms. We monitored the in vitro proliferation of myeloid precursors obtained from bone marrow, blood, spleen, and liver. The macrophage colony-forming unit (M-CFU) numbers were stable in bone marrow but increased progressively in spleen and in liver, reaching in each organ the values equivalent to one femur. The bone marrow had an increased production and enhanced capacity to release M-CFU. Their quantitative increase in blood and in peripheral tissues of schistosome-infected mice was associated with their qualitative modifications: augmented proliferative capacity, enhanced adhesion, and accelerated differentiation. The accelerated release of monomacrophage progenitors and their enhanced proliferation in peripheral tissues potentially account for the relatively low involvement of the bone marrow and for an efficient in situ production of phagocytes, which participate in host reactions to parasites.

Asunto(s)

Células de la Médula Ósea/citología , Células Madre Hematopoyéticas/citología , Leucopoyesis , Macrófagos/citología , Esquistosomiasis mansoni/sangre , Esquistosomiasis mansoni/patología , Animales , Recuento de Células , División Celular , Femenino , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C3H , Bazo/patología

RESUMEN

We studied the B-1 lymphocyte involvement in host reactions to parasites in the murine model of schistosomiasis. No modifications were observed in the prepostural phase of the disease. From the acute phase on, we observed sequentially an increase of Mac1- B-1 cells in the spleen, followed by their appearance in Peyer's patches and in mesenteric ganglia, suggesting that a fraction of splenic B-1 cells might follow this pathway of migration, acquiring progressively the Mac1 expression. These results are consistent with a primary activation of the splenic B cell compartment, with the subsequent mobilization of B-1 cells into the tissue involved by parasites. Conversely, we found no evidence of an increase of B-1 cells in the peritoneum, nor a mobilization of B-1 cells expressing the peritoneal phenotype (CD5lo, IgMhi) into the tissues involved by infection, despite the general inflammatory reactivity of peritoneal cells. In schistosomiasis, the peritoneal cavity B-1 cells on one side, and those involved in inflammatory reactions to parasites in the spleen, Peyer's patches, and mesenteric ganglia on the other, represent two distinct B-1 lymphocyte pools.

Asunto(s)

Subgrupos de Linfocitos B/inmunología , Esquistosomiasis mansoni/inmunología , Animales , Antígenos CD5/análisis , Femenino , Ganglios Simpáticos/inmunología , Inmunoglobulina M/análisis , Ganglios Linfáticos/inmunología , Recuento de Linfocitos , Masculino , Mesenterio/inervación , Ratones , Ratones Endogámicos C3H , Cavidad Peritoneal/citología , Ganglios Linfáticos Agregados/inmunología , Receptores de Antígenos de Linfocitos B/análisis , Bazo/inmunología

RESUMEN

Chronic inflammatory periovular granulomatous reactions elicited in liver by schistosomal infection are a site of active myelopoiesis. We quantified the colony-forming cells (CFCs) in granulomas and found that the whole liver contains a number of CFCs roughly equivalent to 50% of a femur. Clonogenic analysis showed the presence of committed as well as pluripotent and totipotent CFCs. Long-term Dexter-type cultures showed that the granuloma-derived totipotent CFCs do not have self-renewal capacity. Hence, they did not correspond functionally to haematopoietic stem cells, despite the fact that the stroma established by adherent cells harvested from granulomas had the capacity to sustain long-term proliferation of bone-marrow-derived haematopoietic stem cells. We conclude that myelopoietic cytokines produced by inflammatory reactions in schistosomiasis elicit mobilization of bone marrow CFCs into the circulation, which can settle in hepatic granulomas. This environment may induce their proliferation and differentiation, but not their self-renewal, sustaining temporary production of myeloid cell lineages which nevertheless depends upon cell renewal from the bone marrow pool of haematopoietic precursors.

Asunto(s)

Células de la Médula Ósea/citología , Granuloma/fisiopatología , Hematopoyesis Extramedular/fisiología , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Hepatitis Animal/fisiopatología , Animales , Células de la Médula Ósea/efectos de los fármacos , Diferenciación Celular , Células Cultivadas , Ensayo de Unidades Formadoras de Colonias , Eritropoyetina/farmacología , Femenino , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Granuloma/etiología , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/parasitología , Interleucina-3/farmacología , Masculino , Ratones , Ratones Endogámicos C3H , Óvulo/inmunología , Esquistosomiasis/complicaciones , Esquistosomiasis/fisiopatología

RESUMEN

1. Connective tissue cells isolated from hepatic granulomas (GR cells), induced in mouse liver tissue by schistosomal infection, are able to sustain myelopoiesis, while other connective tissue cells such as skin fibroblasts (SF) are not. 2. We compared the ability of SF and GR cells to sustain in vitro proliferation of the FDC-P1 myeloid cell line, dependent upon IL-3 or GM-CSF. 3. Only the GR stroma sustained the proliferation of co-cultured FDC-P1 cells. RT-PCR analysis showed that both cell lines expressed the message for GM-CSF, but not for IL-3. We showed that GM-CSF was produced by, and remained bound to the cell layer through heparan sulfate; this growth factor could be released by high-salt treatment in a biologically active form from both cell types. The same activity could be restored to NaCl-treated GR cells, but not to SF, by incubation with recombinant murine GM-CSF. 4. These results indicate that the ability of connective tissue cells to sustain myelopoiesis depends directly upon the capacity of their heparan sulfate-bearing molecules to bind and present the GM-CSF to the target cells in a biologically active form. Alternatively, a yet unidentified set of cell layer-associated molecules may be required for the positive or negative control of the membrane-bound GM-CSF.

Asunto(s)

Células de la Médula Ósea , Tejido Conectivo/patología , Fibroblastos/patología , Granuloma/patología , Animales , División Celular , Línea Celular , Tejido Conectivo/metabolismo , Fibroblastos/metabolismo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/biosíntesis , Granuloma/metabolismo , Proteoglicanos de Heparán Sulfato , Heparitina Sulfato/metabolismo , Interleucina-3/biosíntesis , Parasitosis Hepáticas/patología , Ratones , Ratones Endogámicos C3H , Proteoglicanos/metabolismo , Esquistosomiasis mansoni/patología

RESUMEN

1. Connective tissue cells isolated form hepatic granulomas (GR cells), induced in mouse liver tissue by schistosomal infection, are able to sustain myelopoiesis, while other connective tissue cells such as skin fibroblasts (SF) are not. 2. We compared the ability of SF and GR cells sustain in vitro proliferation of the FDC-P1 myeloid cell line, dependent upon IL-3 or GM-CSF. 3. Only the GR stroma susteined the proliferation of co-cultured FDC-P1 cells. RT-PCR analysis showed that both cell lines expressed the message for GM-CSF, but not for IL-3. We showed that GM-CSF was produced by, and remained bound to the cell layer through heparan sulfate; this growth factor could be released by high-salt treatment in a biologically active form from both cell types. The same activity could be restored to NaCl-treated GR cells, but not to SF, by incubation with recombinant murine GM-CSF. 4. These results indicate that the ability of connective tissue cells to sustain myelopoiesis depends directly upon the capapcity of their heparan sulfate-bearing molecules to bind and present the GM-CSF to the target cells in a biologically active form. Alternatively, a yet unidentified set of cell layer-associated molecules may be required for the positive or negative control of the membrane-bound GM-CSF

Asunto(s)

Ratones , Animales , Tejido Conectivo/metabolismo , Granuloma/metabolismo , Hematopoyesis , Parasitosis Hepáticas/metabolismo , Esquistosomiasis mansoni/metabolismo , Tejido Conectivo/patología , Técnicas de Cultivo , Fibroblastos/metabolismo , Fibroblastos/patología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Granuloma/patología , Heparitina Sulfato/metabolismo , Parasitosis Hepáticas/patología , Interleucina-3/metabolismo , Proteoglicanos/metabolismo , Esquistosomiasis mansoni/patología

RESUMEN

1. In schistosomal infection, the hyperergic acute phase of the disease evolves progressively into the chronic one, with establishment of a relative equilibrium between the parasites and the corresponding host responses. This down-regulation of host reactivity is considered to be under the control of T-lymphocyte circuits. 2. In the present study, we investigated lymphocyte populations in spleens of normal mice and the kinetics of the B-cell number increase in mice in the acute, chronic and late chronic phases of schistosomal infection, and we monitored their proliferation and activity in antibody isotype secretion. 3. We observed polyclonal B-cell activation and modulation of Ig isotype production, compatible with the alternate predominance of TH2 and TH1 lymphocyte subsets, in the acute and the chronic phases of the disease, respectively.

Asunto(s)

Linfocitos B/inmunología , Esquistosomiasis mansoni/inmunología , Linfocitos T/inmunología , Animales , Linfocitos B/metabolismo , Citocinas/biosíntesis , Femenino , Citometría de Flujo , Granuloma/inmunología , Isotipos de Inmunoglobulinas/biosíntesis , Isotipos de Inmunoglobulinas/inmunología , Activación de Linfocitos , Recuento de Linfocitos , Masculino , Ratones , Ratones Endogámicos C3H , Bazo/inmunología , Linfocitos T/metabolismo , Factores de Tiempo

RESUMEN

1. In schistosomal infection, the hyperergic acute phase of the disease evolves progressively into the chronic one, with establishment of a relative equilibrium between the parasites and the corresponding host responses. This down-regulation of host reactivity is considered to be under the control of T-lymphocyte circuits. 2. In the present study, we investigated lymphocyte populations in spleens of normal mice and the kinetics of the B-cell number increase in mice in the acute, chronic and late chronic phases of schistosomal infection, and we monitored their proliferation and activity in antibody isotype secretion. 3. We observed polyclonal B-cell activation and modulation of Ig isotype production, compatible with the alternate predominance of TH2 and TH1 lymphocyte subsets, in the acute and the chronic phases of the disease, respectively.

Asunto(s)

Animales , Femenino , Masculino , Ratones , Linfocitos B , Esquistosomiasis mansoni , Linfocitos T , Linfocitos B , Bazo/inmunología , Citocinas , Citometría de Flujo , Granuloma , Isotipos de Inmunoglobulinas , Recuento de Linfocitos , Linfocitos T , Factores de Tiempo , Activación de Linfocitos

RESUMEN

In previous research, we have observed that intrathoracic administration of platelet-activating factor-acether (PAF) promoted a delayed eosinophilia in the pleural cavity of rats that lasted for at least 96 h. We investigated the ability of pleural washings from rats previously injected with PAF (1 micrograms/cavity) to stimulate in vitro murine hematopoietic eosinophil proliferation. We observed that pleural fluid sustained eosinophil proliferation but not differentiation, under conditions in which PAF itself had no effect. The phenomenon lasted for 3 days and was maximal on the 1st day of culture. Treatment with neutralizing antibodies against interleukin (IL)-5, granulocyte-macrophage colony-stimulating factor (GM-CSF) or IL-3, alone or in combination, did not modify the eosinophil proliferation induced by PAF pleural fluid, suggesting that these cytokines may not be involved in the studied phenomenon. We conclude that the rat pleural fluid obtained 6 h after PAF administration induces eosinophil proliferation in vitro by a mechanism probably independent of IL-5, GM-CSF or IL-3.

Asunto(s)

Factores Biológicos/inmunología , Eosinófilos/citología , Factor de Activación Plaquetaria/inmunología , Pleura/inmunología , Animales , Factores Biológicos/biosíntesis , Médula Ósea/inmunología , Diferenciación Celular , División Celular , Células Cultivadas , Citocinas/inmunología , Femenino , Inyecciones , Masculino , Pleura/metabolismo , Ratas , Ratas Wistar

RESUMEN

In parasitic diseases, eosinophilia is controlled at the systemic level by soluble, circulating factors. In addition to their medullar production and migration to tissues involved by parasitosis, eosinophil populations in inflammatory infiltrates may be locally amplified by their in situ proliferation. In granulomas induced in liver tissue by eggs of schistosome worms, eosinophil proliferation and differentiation are observed. We have shown that they were under control of two cytokines, the activity of which can be demonstrated in supernatants of isolated granulomas maintained in culture for 24 hours. One of them has been identified as interleukin-5. The other one is secreted by adherent cells obtained from periovular granulomas, among which macrophages represent more than 99% of cells. It is considered to correspond to the previously described factor, secreted by inflammatory macrophages mobilized on intraperitoneal glass implants in mice with chronic schistosomiasis. In acute schistosomiasis, the activity of the interleukin-5 was predominant, whereas in the chronic phase of the disease, the stimulation of peripheral eosinopoiesis is taken over by the factor secreted by adherent cells. During the progression from the acute to the chronic phase of schistosomiasis, the immune reactivity of the host is down-regulated by T suppressor lymphocyte circuits. In addition, a redistribution of cellular controls of the host reaction to parasites may act as a complementary mechanism for establishment of the viable equilibrium between host and parasite.

Asunto(s)

Citocinas/fisiología , Granuloma Eosinófilo/patología , Eosinófilos/patología , Granulocitos/patología , Esquistosomiasis/patología , Animales , Diferenciación Celular , División Celular , Regulación hacia Abajo , Interleucina-5/fisiología , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL

RESUMEN

In chronic murine schistosomiasis mansoni, extramedullar myelopoiesis of eosinophils has been described, associated with tissue inflammatory infiltrates and periovular granulomas. It may be experimentally induced by intraperitoneal glass implants in mice with chronic schistosomiasis but not in normal mice or in mice with acute schistosomiasis. In vivo studies showed that this induction depended on macrophages adherent to the implants. In an in vitro test with bone marrow cells, we demonstrated that this eosinophil proliferation was mediated by a factor secreted by the mobilized macrophages. In contrast to murine interleukin-5 (IL-5), which induced both proliferation and differentiation of eosinophils, the studied monokine did not induce their maturation. A potent neutralizing monoclonal antibody for murine IL-5 did not abrogate the stimulatory activity of the monokine, indicating that it is a cytokine distinct from IL-5. These data, together with a quantitation of bone marrow, blood, and peripheral eosinophils, indicated that in chronic schistosomiasis, the systemic medullar supply of eosinophils is supplemented in tissues by their local proliferation, mediated by macrophages mobilized in local granulomatous and diffuse inflammatory reactions.

Asunto(s)

Eosinófilos/fisiología , Inflamación/metabolismo , Macrófagos/fisiología , Esquistosomiasis mansoni/sangre , Animales , Diferenciación Celular , División Celular , Enfermedad Crónica , Granuloma/metabolismo , Interleucina-5/farmacología , Recuento de Leucocitos , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL