Listeria antigen-binding cells in the spleens of normal and immunized mice resistant or susceptible to listeriosis

Normal mice of the Listeria-resistant C57Bl/6 strain contain in their spleens a higher number of cells that bind Listeria monocytogenes cell wall fraction antigen (LmA) than normal DBA/2 mice, which are more susceptible to infection. LmA-binding cells are probably B cells, nylon-wool adherent, and inhibited by anti-mouse immunoglobulin antibody but not sensitive to the action of monoclonal anti-mouse macrophage and anti-Thy.1.2 antibody. A single intraperitoneal injection of 10(8) Listeria monocytogenes causes a rapid increase in the number of LmA-binding cells in the spleens of C57Bl/6 mice, and this can be seen as early as 24 h. On the other hand, in DBA/2 mice an increase in these cells becomes evident only by the 4th day. Moreover, the increment in the number of LmA-binding cells in C57Bl/6 mice is more marked than in DBA/2 mice.     

Development of Amnesia in Different Mouse Strains

We studied passive avoidance retrieval after amnestic stimulation (arrest in unsafe section of the experimental setup) in C57Bl/6J, BALB/c, CBA/Lac, AKR/J, DBA/2J, C3H/HeJ, and ASC/Icg mice. We demonstrated resistance to amnestic stimulation in mice with high predisposition to freezing reaction (ASC/Icg) and memory deficit in other mouse strains.     

The relative difference in anti-Listeria resistance of C57BL/6 and A/J mice is not eliminated by active immunization or by transfer of Listeria-immune T cells.

In this study, we examined the effects of active and adoptive immunization on the anti-Listeria resistance of innately resistant C57BL/6 and innately susceptible A/J mice. Although active immunization with a sublethal dose of viable Listeria monocytogenes markedly enhanced the anti-Listeria resistance of both C57BL/6 and A/J mice, the 100-fold difference between the two strains in innate anti-Listeria resistance was not diminished. Following immunization with an equivalent sublethal dose (0.1 LD50) of L. monocytogenes, both C57BL/6 and A/J mice generated T cells that could transfer significant and comparable protection to syngeneic recipients that were challenged with up to a 10 LD50 dose of L. monocytogenes. When the absolute number of viable Listeria was compared, however, it was clear that T cells from immunized C57BL/6 mice were capable of transferring protection to syngeneic recipients at Listeria challenge doses that were more than 100-fold greater than could T cells from Listeria-immunized A/J mice. Both active immunization and adoptive transfer of syngeneic Listeria-immune T cells enhanced the accumulation of inflammatory neutrophils and macrophages in C57BL/6 and A/J mice. More inflammatory neutrophils were recovered from actively immunized C57BL/6 than from A/J mice, whereas more inflammatory macrophages were obtained from adoptively immunized C57BL/6 than from A/J mice. These results provide further evidence for the beneficial role of inflammation in genetically determined innate resistance and T-cell mediated resistance to listeriosis. These data also suggest that some mechanism in addition to inflammatory responsiveness may be responsible for limiting the expression of acquired anti-Listeria resistance in genetically susceptible A/J mice.     

Salmon calcitonin – a potent inhibitor of food intake in states of impaired leptin signalling in laboratory rodents

To compare the anorectic effectiveness of leptin and the amylin analogue salmon calcitonin (sCT), rodents were treated on 1 day with subcutaneous injections. In chow-fed C57Bl/6J mice, leptin and sCT reduced energy intake and acted additively. After C57Bl/6J mice had become leptin-resistant on being fed chocolate as a palatable high-caloric supplement to chow, their sCT-induced decrease in energy intake was more pronounced than in chow-fed mice with differential changes in the intake of chocolate (strong reduction) and chow (slight increase). Dose-response relationships for sCT-induced reductions in energy intake were analysed in chow-fed C57Bl/6J mice and two obese strains, ob/ob mice and melanocortin-4 receptor knockout (MC4-r-KO) mice, as well as in wild-type and fatty ( fa/fa ) rats. Compared to C57Bl/6J mice, reduction in food intake induced by sCT was attenuated in MC4-r-KO mice, and nearly absent in ob/ob mice, over the dose range investigated. Compared to C57Bl/6J mice, wild-type rats responded more sensitively to sCT and its efficiency was only slightly reduced in fatty ( fa/fa ) rats. Thus, while genetically induced failures of leptin signalling reduce the action of sCT, it effectively inhibits the intake of a palatable, high fat-high sugar diet even in states of diet-induced obesity with functional leptin resistance.     

A/J mice are susceptible and C57BL/6 mice are resistant to Listeria monocytogenes infection by intragastric inoculation

Previous studies demonstrated that the innate resistance of mice to Listeria monocytogenes infection by intravenous or intraperitoneal inoculation is regulated principally by the Hc locus on mouse chromosome 2. The A/J and C57BL/6 mouse strains were identified as prototype L. monocytogenes-susceptible and -resistant strains, respectively. In the present study, we compared the relative susceptibilities of A/J and C57BL/6 mice to intragastric (i.g.) inoculation with L. monocytogenes. The results of our study indicate that A/J mice are significantly more susceptible than C57BL/6 mice to an i.g. challenge with L. monocytogenes. This was reflected in the estimated 50% lethal doses for the two strains (10(6) and 10(8) CFU for A/J and C57BL/6 mice, respectively) and a more rapid and severe dissemination of the infection to the spleen and liver in A/J mice than in C57BL/6 mice. Histopathological examination of tissues from the infected mice confirmed the greater severity of disease in A/J mice. Clearance of a primary infection enhanced the resistance of both A/J and C57BL/6 mice to reinfection with L. monocytogenes via the gastrointestinal tract. However, the relative difference in susceptibility between the two strains was evident even after immunization. The A/J mouse holds promise as a model for investigating the pathogenesis of gastrointestinal listeriosis because of its ability to develop systemic infection following challenge with numbers of organisms similar to those recovered from some L. monocytogenes-contaminated food products.     

Genetically determined resistance to listeriosis is associated with increased accumulation of inflammatory neutrophils and macrophages which have enhanced listericidal activity.

The C57BL/6 and A/J inbred strains of mice differ markedly in their resistance to the facultative intracellular bacterium Listeria monocytogenes. One possible explanation for this genetically determined resistance is that phagocytes from Listeria-resistant strains of mice can kill L. monocytogenes more effectively than phagocytes from Listeria-susceptible strains of mice. We report here that inflammatory neutrophils and macrophages from Listeria-resistant mice (C57BL/6) exhibit a slight but significantly enhanced ability to kill L. monocytogenes in vitro as compared to inflammatory phagocytes from Listeria-susceptible mice (A/J). More importantly, however, Listeria-resistant mice recruited more inflammatory neutrophils and macrophages to the peritoneal cavity in response to i.p. injection of heat-killed Listeria than did Listeria-susceptible mice. These data suggest that genetically determined resistance to listeriosis is dependent on the enhanced inflammatory responsiveness of Listeria-resistant mice. Further support for this hypothesis was provided by experiments in which the passive transfer to A/J mice (C5-deficient) of plasma from C57BL/6 mice (C5-sufficient) enhanced the ability of the recipient A/J mice both to recruit inflammatory neutrophils to the peritoneal cavity in response to i.p. injection of heat-killed Listeria, and to clear L. monocytogenes from the spleen after a sublethal challenge of viable Listeria.     

Effect of cyclophosphamide treatment on the course of Mycobacterium lepraemurium infection and development of delayed-type hypersensitivity reactions in C57B1 and BALB/c mice.

Pre-treatment of Mycobacterium lepraemurium susceptible, BALB/c, and resistant, C57Bl, mice with cyclophosphamide markedly altered the development of delayed hypersensitivity during footpad infections with this organism. A tuberculin-type response demonstrated by untreated C57Bl mice was significantly intensified after week 3 in cyclophosphamide-pre-treated mice although this response had returned to normal levels by week 8. A Jones-Mote-type response demonstrated throughout experiments by untreated BALB/c mice was considerably increased in magnitude by week 3 in cyclophosphamide-pre-treated mice. By week 6 this response had become considerably protracted and was of the tuberculin-type. By week 8 however this response had started to diminish and by week 12 cyclophosphamide-treated and untreated BALB/c mice produced similar Jones-Mote-type responses when skin-tested. Cyclophosphamide pre-treatment had no effect on the growth of M. lepraemurium in C57Bl mice over 12 weeks. In BALB/c mice however cyclophosphamide-pre-treated mice demonstrated considerable resistance to infection at weeks 8 and 10 after infection but not thereafter. Whereas the magnitude of the delayed hypersensitivity response in C57Bl mice could not be correlated with resistance such a relationship could be demonstrated in BALB/c mice.     

PEEP-induced changes in epithelial permeability in inbred mouse strains

Inbred mouse strains have demonstrated a range of susceptibilities to inhaled environmental irritants. C57Bl/6J mice are highly susceptible while C3H/HeJ mice are resistant to ozone exposures, as assessed by lavaged protein. However, lavaged protein reflects a loss of both the endothelial and epithelial barrier. To determine whether basal differences exist in the epithelial barrier, we measured soluble tracer ((99m)technetium-diethylenetriamine pentaacetic acid, (99m)Tc-DTPA) clearance from the lung in spontaneously breathing, anesthetized mice and mice ventilated with increased lung volume with applied positive end-expiratory pressure (PEEP; 1, 6, or 10cmH(2)O). Both strains showed more rapid clearance during ventilation with 10cmH(2)O PEEP compared with other ventilation pressures (p<0.001). There was a substantial difference in clearance between the two strains during ventilation with 10cmH(2)O PEEP (mean half time for C57Bl/6J mice=19+/-4min versus 34+/-3min for C3H/HeJ mice; p<0.001). Thus, when lung volume is increased, the susceptible C57Bl/6J strain shows a greater change in epithelial barrier than the resistant C3H/HeJ strain. These results may reflect fundamental differences in lung architecture.     

T cell-mediated immune responses of lupus-prone BXSB mice and other murine strains.

Cellular-mediated immunity in the newly described BXSB strain of mice, which is prone to autoimmune disease, has been compared with that of two other strains, C57Bl/6 and 129/J. Quantificaiton of cytotoxic T cell responses to alloantigens and viruses (lymphocytic choriomeningitis and vaccinia virus) showed no difference in the kinetics of appearance and relative activity of cytotoxic T cells per spleen between the young and old BXSB and the control mice. The T cell-dependent primary footpad swelling after local injection with lymphocytic choriomeningitis virus was within the same range for all strains tested with respect to kinetics, but the size was greater by two-fold in C57Bl/6 mice. The susceptibility to systemic infection and subsequent induction of lymphocytes immune to Listeria monocytogenes were about equivalent in all strains. However, clearance of Listeria by the reticuloendothelial system and early non-immune bactericidal activity of the young and old BXSB were significantly lower than in the control strains. The results indicate that the cellular-mediated immunity (CMI) of BXSB mice compared favourably with that of other strains and that there is no apparent differences between CMI of BXSB mice before the onset of disease and during the course of disease. The role of the reduced reticuloendothelial function of BXSB mice in their autoimmune disease or in their high susceptibility to infection remains to be determined.     

Cytomegalovirus hepatitis: characterization of the inflammatory infiltrate in resistant and susceptible mice.

Mice susceptible and resistant to murine cytomegalovirus (MCMV) were infected with this virus and livers were harvested after 2-231 days. Cryostat sections were stained to visualize cells bearing CD4, CD8 or Mac-1 antigens. Mac-1+ cells were prevalent in inflammatory foci after 2 days. These cells persisted in susceptible BALB/c and A/J mice, but disappeared from livers of resistant C57Bl/6 and CBA/CaH mice by day 28. T cell inflammation peaked on days 7-11. This declined by day 56 in C57Bl/6 and CBA/CaH mice, but persisted in BALB/c and A/J mice for at least 231 days. Persistent CD8+ cells were dispersed throughout the parenchyma. More CD8+ cells were observed 7-14 days after infection in the livers of bg/bg (natural killer (NK) cell-deficient) C57Bl/6 and CBA mice, and in C57Bl/6 mice depleted of NK1.1 cells by MoAb. Thus, mice of strains susceptible to MCMV exhibit hepatitis characterized by persistence of dispersed CD8+ cells. This phenomenon may be limited by NK cells in resistant strains.     

Sendai virus infection in genetically resistant and susceptible mice.

The pathogenesis of Sendai virus infection in genetically resistant (C57Bl/6) and susceptible (DBA/2) nonimmune adult mice was investigated. Rising serum complement-fixation (CF) antibody titers were delayed in DBA/2 mice as compared with C57Bl/6 mice. C57Bl/6 mice developed descending desquamative endobronchiolitis, and DBA/2 mice developed descending proliferative endobronchiolitis and bronchogenic alveolitis. Peribronchiolar lymphoid cuffs that formed in C57Bl/6 mice were thicker and more densely populated than those of DBA/2 mice. Immunofluorescence demonstrated viral antigens confined to the epithelial lining of conducting airways in C57Bl/6 mice but extending to alveolar corner cells in DBA/2 mice. Studies with a transmission electron microscope confirmed that Type II pneumocytes were infected only in DBA/2 mice. IgG-containing cells selectively accumulated along the airways of both strains, but fewer were recruited by DBA/2 mice. These results suggest that genetic resistance to Sendai virus is expressed through the immune system.     

The relation between the T cells responsible for cell-mediated cytotoxic killing of mastocytoma cells and the helper-cell effect.

The relationship between T cells involved in cell-mediated immunity and antibody response of C57Bl/6J mice towards DBA/2 mastocytoma cells was investigated. Spleen cells primed with viable mastocytoma cells demonstrated marked cell-mediated cytotoxicity (CMC) in vitro, but antibody response of these cells to a hapten (TNP) conjugated to the allogeneic tumour cells in vitro was suppressed as compared with that of normal spleen cells. In contrast, spleen cells primed with frozen-thawed mastocytoma cells showed no CMC, but antibody response to the hapten in vitro of these cells was enhanced. C57Bl/6J mice primed with frozen-thawed mastocytoma cells produced more cytotoxic antibody than non-primed mice when immunized with viable mastocytoma cells. These results indicate that T cells involved in cell-mediated immunity and those involved in the antibody response to allogeneic mastocytoma cells are distinct.?222     

Lack of strain differences in spatial learning among seizure-prone and seizure-resistant mice

Learning rates were examined in the following inbred mice strains: DBA/2, C3H/He, C57Bl/6J, El, and ddY. DBA/2 mice become susceptible to audiogenic seizures after 2–3 weeks of age and El mice have generalized seizures in response to handling after 3 months of age, but the remaining three strains do not develop seizures. In this study, mice from all five strains underwent 32 training trials in a Morris water maze at 7–9 weeks of age. The seizure-prone DBA/2 and El mice, along with the nonepileptic ddY and C57Bl/6J mice, exhibited learning at similar rates, but the nonepileptic C3H/He mice were unable to learn the water maze task, probably due to visual difficulties. In the C57Bl/6J strain only, female mice learned the task significantly faster than males. There was no difference in the learning rate between the El strain and its parent ddY strain, or any correlation between spatial learning ability and kindling rates in these strains.     

Sex-limited protein: in vitro and in vivo functions.

Mouse complement component C4 exists in two isoforms, C4 and a protein with expression restricted to male animals called sex-limited protein (Slp). Although Slp is about 95% homologous to C4, it is generally believed to be non-functional, at least in conventional haemolytic complement assays. In a previous study, however, we showed that Slp is haemolytically active in a C1-inhibitor (C1INH)-regulated, EDTA-resistant mouse complement activation pathway. To study other possible implications of this finding, we generated constitutively expressing Slp-transgenic mice. The transgene was crossed into otherwise Slp-deficient C57Bl/6J and NZB mice. Members of the third backcross generation of C57Bl/6J mice were tested for functional Slp and classical and alternative complement pathway activities (CH50 and AP50 levels, respectively). Slp-transgenic C57Bl/6J mice showed enhanced CH50, but normal AP50 levels when compared with non-transgenic littermates. To discover a possible protective role for Slp in spontaneous systemic lupus erythematosus (SLE) in NZBxNZW (NZBxW) mice, the third backcross generation of Slp-transgenic NZB mice was mated with NZW mice and the development of SLE in the female offspring was followed. In these introductory experiments, Slp-transgenic NZBxW animals presented with a significantly extended life span. Our results imply that Slp is a mouse complement component with functions which partially resemble some of those of human C4A.     

Pregnancy reduces the genetic resistance of C57BL/6 mice to Listeria monocytogenes infection by intragastric inoculation

In this study, we compared genetically resistant C57BL/6 and susceptible A/J mice for their resistance to Listeria monocytogenes infection during pregnancy. Intragastric infection with modest numbers of bacterial cells (10⁵ CFU) caused reproducible fetal infection and abortion in both mouse strains. Bioluminescence imaging demonstrated dissemination of L. monocytogenes cells from maternal to fetal organs within 3 days of intragastric infection. Although non-pregnant C57BL/6 mice were significantly more resistant to infection than non-pregnant A/J mice, C57BL/6 and A/J mice had similar microbial loads (CFU) in maternal and fetal tissues during pregnancy. Inflammation and necrosis, however, were more severe in A/J mice as evaluated by semi-quantitative histopathology. Although the microbial load in fetal tissues was similar for all fetuses within a single uterus, inflammation and necrosis varied among individual fetuses and placentas. We also noted that the uterus is a target for L. monocytogenes infection in non-pregnant mice.     

Immune Response of Submissive and Aggressive Mice under Conditions of Opioid Receptor Activation

Experiments on the model of paired sensory contact demonstrated that stimulation of immune response in aggressive CBA and C57Bl/6J mice with 10- and 20-day experience of victories, respectively, was prevented by selective δ₂-opioid receptor agonists DSLET and κ-opioid receptor agonists rimorphin in a dose of 100 μg/kg. In C57Bl/6J mice with depression-like behavior, normalization (but not suppression) of the immune response under conditions of μ-opioid receptor stimulation (100 μg/kg) was observed. Selective modulation of activity of a certain type of opioid receptors can normalize the immune function modifi ed during the formation of a certain behavioral type.     

Local immune response to Mycobacterium lepraemurium in C3H and C57Bl/6 mice.

Subcutaneous footpad inoculation of living M. lepraemurium (L.MLM) induced, in high responder C57Bl/6 mice, a local granulomatous reaction associated with the production of effector cells which stopped the multiplication of bacilli in the draining popliteal node with the concurrent development of 24--48 hr delayed type hypersensitivity (DTH). The thymus-dependent local reaction did not occur after the injection of heat-killed M. lepraemurium (HK.MLM) or after the inoculation of L.MLM in nude mice. However, HK.MLM injection interfered with the onset of the local reaction and enhanced acid-fast bacteria (AFB) counts in the draining node. In low responder C3H mice, L.MLM produced a local and delayed footpad swelling but no restriction of bacilli multiplication in the draining lymph node was observed. This unresponsiveness was not due to an overloading of the inoculum dose since doses ranging from 3 x 10(4) to 3 x 10(7) MLM did not produce any granulomatous local reaction as in C57Bl/6 mice. The injection of dead bacilli in the contralateral footpad of subcutaneously (s.c.) infected C3H mice revealed Arthus-like and 18--24 hr delayed reactions. When 10(6) L.MLM per mouse were injected intravenously (i.v.), systemic infection, measured in the spleen, was found to be less restricted in C57Bl/6 than in C3H mice. Moreover, in C57Bl/6 mice low doses of L.MLM injected i.v. delayed the local reaction at first, then enhanced footpad swelling and AFB counts in the draining nodes, indicating some acquired defect of peripheral immunity. When a high dose of L.MLM (2 x 10(8)/mouse) was injected i.v., C57Bl/6 mice died sooner than C3H mice, indicating certain discrepancies between local resistance and systemic susceptibility.     

Comparative in vivo sister chromatid exchange induction by ethyl carbamate in maternal and fetal tissues of tumor-susceptible and -resistant murine strains

Murine susceptibility to ethyl carbamate-induced carcinogenesis is strain dependent. In vivo sister chromatid exchange (SCE) responses to ethyl carbamate were evaluated in bone marrow cells of gravid adenoma-susceptible (ICR/Jcl), and resistant (C57Bl/6J) and (DBA/2J) murine dams, as well as in liver cells of their respective ICR/Jcl, C57Bl/6J X DBA/2J (BDF1), and DBA/2J X C57Bl/6J (BDF), fetuses following a single intravenous injection of 1.1, 2.2, or 3.3 mmol/kg of ethyl carbamate on gestation day 13/14. Bone marrow tissues of C57Bl/6J and DBA/2J, but not ICR/Jcl dams, demonstrated greater sensitivity to SCE induction than liver cells of their respective fetuses. Furthermore, relative SCE responses in bone marrow among dams indicated greater sensitivity of the more tumor-susceptible ICR/Jcl and C57Bl/6J strains to SCE induction by ethyl carbamate relative to the more tumor-resistant DBA/2J strain. In addition, concurrent alterations (stimulation or inhibition) of bone marrow cell cycle kinetics by ethyl carbamate were consistent with hormone-related, strain-dependent hematopoietic stress during pregnancy.     

Non-dystrophic 129 REJ mice are susceptible to i.p. infection with Listeria monocytogenes despite an ability to recruit inflammatory neutrophils to the peritoneal cavity

In this study we compared the host response toListeria monocytogenesin 129 REJ mice with listeria-resistant (C57Bl/6j) and susceptible (Balb/c) mouse strains. In all experiments mice were inoculated by the i.p. route. 129 REJ mice and Balb/c mice were sensitive to listeriosis whilst C57Bl/6j mice were relatively resistant to i.p. infection. Relatively large numbers of viable bacteria could be detected in the spleens of 129 REJ mice as early as 6 h following i.p. inoculation suggesting that dissemination of listeria from the peritoneal cavity is rapid in this mouse strain. This contrasted with Balb/c mice which exhibited an early lag phase during which only low numbers of bacteria could be isolated from the spleens of infected animals. In response to both proteose peptone and live listeria, 129 REJ mice demonstrated a greater capacity to recruit neutrophils to the peritoneal cavity than Balb/c and C57Bl/6j mice. In addition, inflammatory phagocytes from 129 REJ mice were as bactericidalin vitroas phagocytes from the Balb/c and C57Bl/6j strains. However,in vivo, inflammatory neutrophils elicited by proteose peptone prior to i.p. infection withL. monocytogeneswere not protective in the three mouse strains tested. Despite the apparent inadequacy of peritoneal neutrophils in controlling early bacterial proliferation, depletion of neutrophils in 129 REJ mice severely exacerbated i.p. infection withL. monocytogenes. The results indicate that neutrophils provide an inefficient but essential means of controlling early outgrowth of listeria in the peritoneal cavity of 129 REJ mice. The excessive inflammatory response seen in 129 REJ mice may facilitate the early dissemination ofL. monocytogenesfrom the peritoneal cavity to peripheral sites.     

Hypersusceptibility of A/J mice to tuberculosis is in part due to a deficiency of the fifth complement component (C5)

Mycobacterium tuberculosis (MTB) causes tuberculosis in man, which occurs as an acute, chronic or dormant disease reactivating over several years. The mechanisms of persistence and reactivation are not well understood and there is a need for animal models. Moderate-dose, aerosol infection killed A/J mice earlier than partially resistant C57Bl/6 mice, whereas a low-dose, aerosol-induced chronic infection exacerbated earlier in A/J mice. A/J mice lethally infected with MTB but drug cured of disease underwent reactivation of tuberculosis at least 100 days before similarly infected C57Bl/6 mice. Because A/J mice were C5 deficient, congenic B10 mice sufficient and deficient for C5 were infected intravenously with MTB to define the role of C5. C5-deficient mice again showed enhanced growth of MTB in the lungs. MTB-infected macrophages from C5-deficient mice showed enhanced growth of MTB coinciding with a reduced secretion of both cytokines (TNF-alpha, IL-1beta, IL-6, IL-12) and chemokines (KC, MIP-2 and MIP-1alpha) in A/J and TNF-alpha and chemokines in C5-deficient mice. Because C5-deficient macrophages could be activated from extraneous C5 and TNF-alpha we suggest that both play a role in the macrophage-mediated killing as well as containment mechanisms in tuberculosis.