The presence of MOMA-2+ macrophages in the outer B cell zone and protection of the splenic micro-architecture from LPS-induced destruction depend on secreted IgM

The role secretory IgM has in protecting splenic tissue from LPS-induced damage was assessed in mice incapable of secreting IgM but able to express surface IgM and IgD. Within seconds after LPS challenge, 99% of the (131)I-labeled LPS was found in the liver and the spleen of both sIgM-deficient and wild-type mice. In the spleen FITC-labeled LPS was found on the surface of 2F8(+) scavenger receptor macrophages localized in the outer marginal zone, while none of the labeled LPS could be detected on marginal zone ER-TR9(+) and MOMA-1(+) macrophages. An additional population of macrophages, MOMA-2(+), were capable of producing C3 locally in the T and B cell zone after LPS challenge. Local C3 production was regulated, as no C3 was found in splenic tissue of unchallenged mice. Interestingly, in the absence of circulating and locally produced secretory IgM, MOMA-2(+) macrophages of the T and B cell zone failed to establish an additional ring of C3-producing macrophages in the outer B cell zone close to the marginal zone upon LPS challenge. The consequence was a massive destruction of the microarchitecture of the spleen where marginal zones disorganized, lymphoid follicles and T cell zones disrupted and follicular DC (FDC) networks disappeared.     

Use of mice tolerant to lipopolysaccharide to demonstrate requirement of cooperation between macrophages and lymphocytes to generate lipopolysaccharide-induced colony-stimulating factor in vivo

Injection of lipopolysaccharide (LPS) into mice was followed by a rapid elevation of colony-stimulating factor (CSF) in the serum. A second, challenging injection of LPS given 3 to 4 days later failed to induce elevated levels of CSF in the serum. Such mice tolerant to LPS were used as an experimental tool to identify the CSF-producing cells which respond to LPS. We observed that generation of LPS-induced CSF in mice tolerant to LPS could be restored by an intraperitoneal injection of spleen cells 24 h before the challenging injection of LPS. Depletion of the adherent cells from the spleen cells reduced the ability of the splenic lymphocytes to restore the capacity of the mice tolerant to LPS to generate serum CSF. Reconstitution of the splenic lymphocytes with 5% thioglycolate-elicited peritoneal macrophages, however, reestablished the restorative capacity of these cells, whereas almost no restoration was observed after direct injection of elicited peritoneal macrophages. These data suggest that the spleen cells are active in generating CSF, provided that macrophages are present and can interact with the splenic lymphocytes to generate LPS-induced CSF in the serum.     

Depletion of liver and splenic macrophages reduces the lethality of Shiga toxin-2 in a mouse model.

The haemolytic uraemic syndrome (HUS) is a clinical syndrome consisting of haemolytic anaemia, thrombocytopenia, and acute renal insufficiency. HUS is the most frequent cause of acute renal failure in childhood. It has been previously suggested that the presence of Shiga toxin (Stx) is necessary but not sufficient for HUS development, and cytokines such as tumour necrosis factor-alpha (TNF-alpha) and IL-1beta appear to be necessary to develop the syndrome. Since the mononuclear phagocytic system (MPS) is the major source of these cytokines, macrophages might be one of the relevant targets for Stx action in the pathophysiology of HUS. In this study our objective was to examine the role of the hepatic and splenic macrophages in a mouse model of HUS induced by injection of Shiga toxin type-2 (Stx2) or Stx2 plus lipopolysaccharide (LPS). For this purpose, depletion of mice macrophages by liposome-encapsulated clodronate (lip-clod), followed by injection of STx2 or Stx2 plus LPS, was assayed. In this study we show that depletion of hepatic and splenic macrophages by clodronate treatment induces a survival of 50% in animals treated with Stx2 alone or in presence of LPS. This maximal effect was observed when lip-clod was injected 48-72 h before Stx2 injection. Biochemical and histological parameters show characteristics of the lesion produced by Stx2, discarding non-specific damage due to LPS or lip-clod. In addition, we determined that the toxic action of Stx2 is similar in BALB/c and N:NIH nude mice, indicating the T cell compartment is not involved in the Stx2 toxicity. Briefly, we demonstrate that macrophages play a central role in the pathophysiology of HUS, and that the systemic production of cytokines by liver and/or spleen is for Stx2 to manifest its full cytotoxic effect. In addition, the toxicity of Stx2 alone, or in presence of LPS, is independent of the T cell compartment.     

Carrageenan Primes Leukocytes To Enhance Lipopolysaccharide-Induced Tumor Necrosis Factor Alpha Production

We have previously reported that pretreatment with carrageenan (CAR) enhances lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-alpha) production in and lethality for mice. Whole blood cultured in vitro was used to show that CAR pretreatment results in about a 200-fold increase in LPS-induced TNF-alpha production. CAR by itself did not induce TNF-alpha production. However, CAR-treated cultured medium sensitized whole blood to make more LPS-induced TNF than did saline-treated cultured medium in vitro. It was also demonstrated that CAR pretreatment increases TNF-alpha mRNA levels of both blood cells and peritoneal exudate cells, but not of bone marrow cells. Immunoelectron microscopic analysis revealed that polymorphonuclear leukocytes and macrophages are TNF-alpha-producing cells in CAR-treated mice. In CAR-treated mice, TNF-alpha was seen early after LPS injection in leukocytes in hepatic sinusoids and on the surfaces of endothelial cells. TNF-alpha was also detected late after LPS injection in hepatocytes which become edematous. These results suggest that CAR primes leukocytes to produce TNF-alpha in response to LPS and that they play an important role in the pathogenesis of liver injury.     

Peritoneal Lavage Reduces Lipopolysaccharide-Induced Elevation of Serum TNF-α and IL-6 and Mortality in Mice

The contribution of peritoneal cells to lipopolysaccharide (LPS)-induced elevation of serum TNF- and IL-6 levels and mortality has been studied. Peritoneal lavage performed before LPS administration reduced serum cytokine levels by approximately 50% and mortality from 50 to 100%. The effect of peritoneal lavage is due to the removal of peritoneal cells as reinjection of peritoneal cells eliminated the protective effect of lavage on LPS-induced mortality. A special role of peritoneal macrophages in the systemic response to LPS was suggested by the finding that LPS-induced an increase in intracellular TNF- and IL-6 in peritoneal macrophages but in neither splenic nor bone marrow macrophages. Intraperitoneal injection of thioglycollate broth 4 days prior to lavage increased the number of peritoneal cells removed by lavage and increased protection from LPS mortality. Peritoneal lavage performed 30 to 120 minutes after the LPS administration completely protected all mice from LPS-induced mortality, suggesting the possibility that such treatment may offer a novel therapeutic approach to septic shock.     

Modulation of macrophage Ia expression by lipopolysaccharide: stem cell requirements, accessory lymphocyte involvement, and IA-inducing factor production.

The mechanism of induction of murine macrophage Ia expression by lipopolysaccharide (LPS) was studied. Intraperitoneal injection of 1 microgram of LPS resulted in a 3- to 10-fold increase in the number of IA-positive peritoneal macrophages (flow cytometry and immunofluorescence and a 6-to 16-fold increase by radioimmunoassay. The isolated lipid A moiety of LPS was a potent inducer of macrophage Ia expression. Ia induction required a functional myelopoietic system as indicated by the finding that the response to LPS was eliminated in irradiated (900 rads) mice and reinstated by reconstitution with bone marrow cells. Comparison of LPS-induced Ia expression in normal and LPS-primed mice revealed a faster secondary response to LPS. The memory response could be adoptively transferred to normal mice with nonadherent spleen cells prepared 60 days after LPS injection. Spleen cells prepared 5 days after LPS injection caused Ia induction in LPS-nonresponder mice; such induction was not observed in irradiated (900 rads) recipients. The cell responsible for this phenomenon was identified as a Thy-1+, immunoglobulin-negative nonadherent cell. The biosynthesis and expression of Ia were not increased by direct exposure of macrophages to LPS in vitro. Small amounts of LPS inhibited Ia induction by gamma interferon. LPS showed positive regulatory effects on Ia expression by delaying the loss of Ia expression on cultured macrophages and by stimulating the production of Ia-inducing factors. Supernatants from cultured spleen cells stimulated with LPS in vitro contained antiviral and Ia-inducing activity that was acid labile, indicating that the active factor is gamma interferon. We conclude that induction of Ia expression by LPS in vivo is a bone-marrow-dependent, radiation-sensitive process which involves the stimulation of a gamma interferon-producing accessory lymphocyte and a delay in Ia turnover.     

Increased Ferroportin-1 Expression and Rapid Splenic Iron Loss Occur with Anemia Caused by Salmonella enterica Serovar Typhimurium Infection in Mice

The Gram-negative intracellular bacterium Salmonella enterica serovar Typhimurium causes persistent systemic inflammatory disease in immunocompetent mice. Following oral inoculation with S. Typhimurium, mice develop a hematopathological syndrome akin to typhoid fever with splenomegaly, microcytic anemia, extramedullary erythropoiesis, and increased hemophagocytic macrophages in the bone marrow, liver, and spleen. Additionally, there is marked loss of iron from the spleen, an unanticipated result, given the iron sequestration reported in anemia of inflammatory disease. To establish why tissue iron does not accumulate, we evaluated multiple measures of pathology for 4 weeks following oral infection in mice. We demonstrate a quantitative decrease in splenic iron following infection despite increased numbers of splenic phagocytes. Infected mice have increased duodenal expression of the iron exporter ferroportin-1, consistent with increased uptake of dietary iron. Liver and splenic macrophages also express high levels of ferroportin-1. These observations indicate that splenic and hepatic macrophages export iron during S. Typhimurium infection, in contrast to macrophage iron sequestration observed in anemia of inflammatory disease. Tissue macrophage export of iron occurs concurrent with high serum concentrations of interferon gamma (IFN-γ) and interleukin 12 (IL-12). In individual mice, high concentrations of both proinflammatory (tumor necrosis factor alpha [TNF-α]) and anti-inflammatory (IL-10) cytokines in serum correlate with increased tissue bacterial loads throughout 4 weeks of infection. These in vivo observations are consistent with previous cell culture studies and suggest that the relocation of iron from tissue macrophages during infection may contribute to anemia and also to host survival of acute S. Typhimurium infection.     

Role of interleukin-18 (IL-18) during lethal shock: decreased lipopolysaccharide sensitivity but normal superantigen reaction in IL-18-deficient mice

Lethal shock can be associated with excessive secretion of cytokines such as tumor necrosis factor (TNF) and gamma interferon (IFN-gamma). IFN-gamma mediates macrophage activation and appears to be controlled by interleukin (IL)-12 and IL-18. To investigate the role of IL-18 in vivo, we generated IL-18-deficient mice by gene targeting. IL-18(-/-) mice showed decreased sensitivity towards lipopolysaccharide (LPS)-induced shock. LPS-induced IFN-gamma production was abrogated, yet induction of IL-12 and TNF was not affected. Both wild-type and IL-18-deficient mice succumbed to LPS-induced lethal shock after sensitization with D-galactosamine. However, in marked contrast to LPS, the bacterial superantigen Staphylococcus aureus enterotoxin B (SEB) induced comparable serum levels of IFN-gamma in IL-18(+/+) and IL-18(-/-) mice, accompanied by an upregulation of cell surface markers CD14, CD122 (IL-2Rbeta), and CD132 (IL-2Rgamma) on peritoneal macrophages. Moreover, SEB injection rendered IL-18-deficient mice sensitive for subsequent challenge with LPS. The degree of sensitization was comparable to that in wild-type controls with respect to lethality. However, LPS-induced TNF levels in serum were significantly reduced in SEB-sensitized IL-18-deficient mice. These results imply that IL-18 plays an important role in induction of IFN-gamma and lethality in response to LPS.     

Surfactant protein A inhibits lipopolysaccharide-induced in vivo production of interleukin-10 by mononuclear phagocytes during lung inflammation

We previously demonstrated that resident alveolar macrophages from naive mice do not synthesize interleukin (IL)-10, whereas mononuclear phagocytes (MP) recruited during the lung inflammatory process are transiently competent for IL-10 production when exposed to lipopolysaccharide (LPS) in vitro. As surfactant protein A (SP-A), a member of the collectin family, inhibits LPS-induced in vitro IL-10 formation by bone marrow-derived macrophages, we studied its effect on MP under in vivo inflammatory conditions. When mice with LPS-induced inflamed lungs were given a second intranasal LPS administration, IL-10 concentration recovered in the bronchoalveolar lavage fluids varied as a function of the time interval between the two LPS doses. Thus, IL-10 concentration increased with the number of MP up to Day 3, and then decreased to undetectable values within 24 h, despite a continued increase in the number of MP. Analysis of IL-10 mRNA from purified MP indicated that gene expression correlated with the IL-10 level in the bronchoalveolar lavage fluid. In contrast to IL-10 production, SP-A concentrations during LPS-induced inflammation decreased with a nadir at Day 3, and then increased significantly within 24 h. Furthermore, intranasal administration of exogenous SP-A to mice with LPS-induced inflamed lungs led to a repression of the IL-10 production. In summary, this study demonstrates for the first time an in vivo inhibitory role of SP-A on the anti-inflammatory activity of MP, through inhibition of IL-10 production.     

Overexpression of Interleukin-15 increases susceptibility to lipopolysaccharide-induced liver injury in mice primed with Mycobacterium bovis bacillus Calmette-Guerin

Mice primed with Mycobacterium bovis bacillus Calmette-Guérin (BCG) are highly sensitive to lipopolysaccharide (LPS)-induced liver injury and lethality. We found that interleukin-15 (IL-15) transgenic (Tg) mice primed with BCG were more susceptible to LPS-induced liver injury than non-Tg mice. The numbers of CD44+ CD8+ T cells expressing intracellular gamma interferon (IFN-gamma) significantly increased in the livers of BCG-primed IL-15 Tg mice after LPS injection, and the depletion of CD8+ T cells from BCG-primed IL-15 Tg mice completely abolished the susceptibility to LPS-induced lethality. Liver T cells from BCG-primed IL-15 Tg mice produced IFN-gamma in vitro in response to LPS, which was inhibited by the addition of anti-IL-12 monoclonal antibody (MAb). In vivo treatment with anti-IL-12 MAb inhibited the appearance of CD44+ CD8+ T cells expressing intracellular IFN-gamma after LPS injection. These results suggest that the overexpression of IL-15 increases susceptibility to LPS-induced liver injury in BCG-primed mice via bystander activation of CD8+ T cells.     

In vivo effects of LPS on B lymphocyte subpopulations. Migration of marginal zone-lymphocytes and IgD-blast formation in the mouse spleen

The aim of the present study was to analyze the effect of LPS on the localization and differentiation of splenic B lymphocytes. Therefore, we used a double immunoperoxidase technique which enabled us to detect both the IgM+ IgD- marginal zone lymphocytes and the IgM+ IgD+ follicular lymphocytes in the same tissue section. Next to a dramatic disappearance of the predominantly IgM+ IgD- lymphocytes in the marginal zone shortly after an intravenous injection of LPS, an increased number of these cells could be found in the splenic follicles. The present results strongly suggest that the IgM+ IgD- cells in the splenic follicles represent immigrating marginal zone lymphocytes, and not differentiating follicular B cells, because no IgM+ IgD- cells could be observed in the follicles of draining lymph nodes shortly after a subcutaneous injection of a similar amount of LPS. These observations support the suggestion that LPS induces a migration of marginal zone lymphocytes into the follicles. The present results also showed the formation of IgD plasmablasts in the inner PALS and around the terminal arterioles of the spleen after LPS administration. The induction of IgD plasmablasts appeared to be a specific effect of LPS which may be related to its toxic properties.     

Progressive Accumulation of Bacterial Lipopolysaccharide In Vivo During Murine Acute Graft-Versus-Host Disease

In a previous report the authors demonstrated that acute graft-versus-host disease (GVHD) was associated with pathologic amounts of tumour necrosis factor alpha (TNF-α) and the appearance of lipopolysaccharide (LPS) in the blood of GVH reactive mice just prior to death. In this study the authors have investigated the kinetics of LPS accumulation in different organs during the course of acute GVHD using a murine model. Unirradiated C57BL/6×AF1 (B6AF₁) mice were transplanted with C57BL/6 (B6) lymphoid cells and killed at predetermined times after transplantation for LPS analysis. Control animals were injected with either 60×10⁶ B6AF1 lymphoid cells (syngeneic) or 60×10⁶ irradiated (2000 rad) CBA lymphoid cells (allogeneic). Lipopolysaccharide began to appear in the liver and the spleen of GVH reactive mice from day 2 post-transplant and by day 10 all GVH reactive mice tested positive for hepatic and splenic LPS. Low levels of LPS were detected in some control mice from days 2 to 10 post-transplant but LPS was never detected after day 10 in control groups. Total hepatic and splenic LPS in acute GVH reactive mice peaked at a time coincident with the appearance of LPS in the serum and with the onset of mortality. These results demonstrate that tissue levels of LPS increase throughout the course of acute GVHD and are sufficient to trigger the release of pathologic amounts of TNF-α from primed macrophages resulting in the cachexia and mortality associated with acute GVHD in this model.     

Role of CLIC4 in the host innate responses to bacterial lipopolysaccharide

Chloride intracellular channel (CLIC) 4 has diverse functions in membrane trafficking, apoptosis, angiogenesis and cell differentiation. CLIC4 is abundantly expressed in macrophages, but its role in innate immune functions is unclear. Here, we show that primary murine macrophages express increased amounts of CLIC4 after exposure to bacterial lipopolysaccharide (LPS). Endogenous CLIC4 level was significantly elevated in the brain, heart, lung, kidney, liver and spleen after LPS injection of mice. Stable macrophage lines overexpressing CLIC4 produced more TNF, IL-6, IL-12 and CCL5 than mock transfectants when exposed to LPS. To explore the role of CLIC4 in vivo, we generated CLIC4-null mice. These mice were protected from LPS-induced death, and had reduced serum levels of inflammatory cytokines. Upon infection with Listeria monocytogenes, CLIC4-deficient mice were impaired in their ability to clear infection, and their macrophages responded to Listeria by producing less inflammatory cytokines and chemokines than the WT controls. When challenged with LPS in vitro, deletion of clic4 gene had little effect on MAPK and NF-κB activation, but led to a reduced accumulation of phosphorylated interferon response factor 3 (IRF3) within macrophages. Conversely, overexpression of CLIC4 enhanced LPS-mediated IRF3. Thus, these findings suggest that CLIC4 is an LPS-induced product that can serve as a positive regulator of LPS signaling.     

Endotoxin-induced gamma interferon production: contributing cell types and key regulatory factors

Gamma interferon (IFN-gamma) is an important mediator of endotoxin (lipopolysaccharide [LPS])-induced immune responses. However, the specific cell types that produce IFN-gamma in response to LPS and the cellular factors that regulate LPS-induced IFN-gamma production have not been fully determined. The present studies were undertaken to characterize the cell populations that produce IFN-gamma after LPS challenge in the spleens of mice and to determine the regulatory factors that modulate LPS-induced production of IFN-gamma. Our studies show that the levels of splenic IFN-gamma mRNA and protein production peak at 6 and 8 h, respectively, after systemic LPS challenge. Approximately 60% of IFN-gamma-producing cells are natural killer (NK) cells (CD3(-)DX5(+)) and 25% are NKT cells (CD3(+)DX5(+)). Most of the remaining IFN-gamma-producing cells are T cells (CD3(+)DX5(-)), macrophages, and dendritic cells. Functionally, interleukin-12 (IL-12) is the major IFN-gamma-stimulating factor after LPS challenge, with costimulation provided by IL-15, IL-18, and B7 proteins. IL-10 is a major inhibitor of LPS-induced IFN-gamma production. Unlike intact heat-killed gram-negative and gram-positive bacteria, the class II major histocompatibility complex did not play a functional role in LPS-induced IFN-gamma production. LPS is a potent stimulus for splenic IL-10, IL-12 p40, and IL-15 mRNA expression, whereas IL-12 p35 and IL-18 mRNAs, as well as B7 proteins, are constitutively expressed in the mouse spleen. Of the factors studied, IL-18 serves as the most potent costimulus with IL-12 for IFN-gamma production, followed by IL-15 and B7 proteins. These data demonstrate that NK cells and NKT cells are the most abundant IFN-gamma-producing cells in the mouse spleen after LPS challenge and that IL-10 and IL-12 are key functional regulators of LPS-induced IFN-gamma production.     

Clearance and organ localization of particles and soluble complexes in mice with circulating complexes.

The clearance and organ localization of a number of substances cleared by either Fc-dependent or -independent mechanisms was studied in normal mice and in mice with endogenously produced persistent circulating complexes. Clearance of covalent dimers of mouse IgG, chicken IgG and ovalbumin were no different between the two groups of mice. By contrast, hepatic and splenic uptake of dimeric mouse IgG (but not of chicken IgG or ovalbumin dimer) was impaired in the mice with persisting complexes. Surprisingly the rate of clearance of sheep red blood cells (SRBC) was increased in mice with persisting complexes as was hepatic uptake of polyvinyl pyrrolidone. It is suggested that the mononuclear phagocytes of mice with persistent circulating complexes are non-specifically stimulated while their ability to take up soluble complexes by Fc-dependent attachment is selectively impaired.     

Ultrastructural observations on cytotoxic effector cells infiltrating pancreatic islets of low-dose streptozocin treated mice

Summary The aim of this study was to observe the ultrastructural events, during the onset of diabetes mellitus in the low-dose streptozocin (LDS)-treated mouse model with emphasis on the infiltrating elements. Forty male C57 BL/6J mice were given 40 mg/streptozocin on 5 consecutive days and killed 5, 6, 7, 8, 9, 10, 15, and 18 days after the first injection. Results demonstrated that islet infiltration occurring in LDS-treated mice is characterized by a very early pre-infiltration state in which mononuclear phagocytes in islet capillary vessels were considerably increased in number. A new histopathological time sequence for the early insulitis is described, in which attraction of blood mononuclear phagocytes into the islet capillary lumen is the first step. During the successive stage, occurring on days 6–8 we observed that mononuclear phagocytes migrate through capillary and venule walls into the islet parenchyma, where they differentiate into tissue macrophages. It was only later (step 3) that these macrophages acquired novel properties, typical of their activated state and started to phagocytose islet beta-cell debris. These data suggest that during the pre-infiltration and early insulitis the mononuclear phagocyte system plays a key role in the onset of LDS diabetes.     

Interleukin-1 gene expression in rabbit vascular tissue in vivo.

Cultured human vascular endothelial and smooth muscle cells express interleukin-1 (IL-1) genes when exposed to bacterial lipopolysaccharides (LPS) and a variety of inflammatory mediators. Local production of IL-1 may contribute to the pathogenesis of various vascular diseases. Therefore the ability of intact vascular tissue to accumulate IL-1 mRNA and synthesize de novo biologically active IL-1 protein was examined. Escherichia coli LPS (10 micrograms/kg) was administered intravenously to adult rabbits and total RNA was isolated from aortic tissue at various times after LPS injection. In saline-injected rabbits, RNA extracted from the thoracic aorta contained little or no IL-1 message detected by Northern analysis using IL-1 alpha and beta cDNA probes cloned from an LPS-stimulated rabbit splenic macrophage library. Lipopolysaccharide treatment promptly induced transient accumulation of mRNA for IL-1 alpha and IL-1 beta within the aorta (maximal 1-hour after injection). Short-term organoid cultures of rabbit aorta exposed to LPS in vitro synthesized immunoprecipitable IL-1 alpha protein. Extracts of aortic tissue excised 1.5 to 3.0 hours after intravenous LPS administration contained immunoreactive and biologically active IL-1 alpha. Anti-rabbit IL-1 alpha antibody neutralized the biologic activity (more than 90%). Microscopic and immunohistochemical studies did not disclose adherent or infiltrating macrophages in rabbit aorta at the time of maximal IL-1 mRNA accumulation after LPS administration (1.5 hours), indicating that intrinsic vascular wall cells rather than mononuclear phagocytes probably account for the IL-1 activity induced by LPS. In addition, aortic tissue from rabbits fed an atherogenic diet showed an enhanced ability to accumulate IL-1 alpha and beta mRNA and produce immunodetectable protein in response to LPS administration. These studies demonstrate inducible IL-1 gene expression in rabbit vascular tissue in vivo and support a local role for this cytokine in vascular pathophysiology.     

Accelerated rate of mononuclear phagocyte production in vitro by splenocytes from autoimmune motheaten mice.

The rapidly fatal autoimmune disease accompanied by pneumonitis in the mutant mouse known as motheaten is caused by an autosomal recessive gene. The proliferative capacity and production rate of splenic mononuclear phagocytes at different maturational stages, defined by morphologic criteria, were examined by two in vitro tritiated thymidine administration protocols and radioautography. The replicative rate of splenic promonocytes from 3-week-old normal mice was found to approximate that of adult bone marrow cells. Monocytes, macrophages, and previously described macrophagelike cells from motheaten mice had an accelerated rate of maturation in vitro, compared with similar cells from normal mice. These differences in the production rates could be attributed to the shorter period of time that promonocytes from the mutant mice are in the S phase of the cell cycle. Evidence, also, verified that the macrophagelike cells were derived from promonocytes via the monocyte-macrophage compartment. The unusual in vitro growth characteristics of the splenic mononuclear phagocytes from motheaten mice could be a manifestation of an in vivo abnormality of this lineage that could contribute to the development of pulmonary disease in these animals.     

Population kinetics of peritoneal LPS-reactive B lymphocytes

The dynamic behaviour of isolated populations of peritoneal LPS-reactive (LPSr) B lymphocytes was studied upon transfer of peritoneal cells (PerC) from C57BL/6 LPS responder into C57BL/10ScCr LPS non-responder mice. We have followed the persistence and life-span of the transferred LPSr donor B cells in the spleen and peritoneal cavity of both intact and X-irradiated adult hosts after i.v. or i.p injection and neonatal 1-day-old recipients after i.v. transfer. We have found that lymphocyte life-spans can be influenced by local host environments, as the transferred PerC LPSr cells showed different kinetics according to their route of injection, organ localization, and age or state of the recipients. Thus, while in intact hosts most of the transferred peritoneal LPSr cells decayed with time, following transfer into X-irradiated recipients the same cells were able to expand and replenish the lymphoid tissues of the host. Moreover, upon transfer into intact hosts, the kinetic properties of peritoneal LPSr cells from adult mice differ from splenic LPSr cells of age-matched animals, but mimic those of spleen cells from young, 1-to 2-week-old donors. These findings may reflect the different phenotype composition of adult spleen cells (poor in Ly1 B cells) and peritoneal and neonatal spleen cells (both rich for Ly1 B cells), or may be the result of selective events leading to the peritoneal accumulation of cells with different population dynamics.     

Pro-inflammatory effects of matrix metalloproteinase 7 in acute inflammation

Matrix metalloproteinase 7 (MMP7) is a member of the MMP family. In the small intestine, MMP7 is responsible for activating α-defensins, which are broad-spectrum anti-microbial peptides produced by the Paneth cells. We report that MMP7^−/− mice are resistant to LPS-induced lethality and that this resistance is correlated with reduced levels of systemic cytokines. LPS induced the upregulation and activation of MMP7 in the small intestine, degranulation of the Paneth cells, and induction of intestinal permeability in MMP7^+/+ mice. In MMP7^−/− mice, both LPS-induced intestinal permeability and consequent bacterial translocation to the mesenteric lymph nodes were reduced. Based on gene expression analysis and evaluation of intestinal damage, we attribute the protected state of MMP7^−/− mice to reduced intestinal inflammation. Interestingly, we found that different α-defensins, namely Crp1 (DEFA1) and Crp4 (DEFA4), can stimulate IL-6 release in macrophages and ileum explants in a TLR4 independent way. We conclude that absence of MMP7 protects mice from LPS-induced intestinal permeability and lethality, and suggest that MMP7-activated α-defensins, in addition to their previously recognized bactericidal and anti-inflammatory roles, may exhibit pro-inflammatory activities in the intestine by activating macrophages and amplifying the local inflammatory response in the gut, leading to intestinal leakage and subsequent increase in systemic inflammation.