Innate resistance to experimental African trypanosomiasis: differences in cytokine (TNF-alpha, IL-6, IL-10 and IL-12) production by bone marrow-derived macrophages from resistant and susceptible mice

Resistance to African trypanosomiasis is under multigenic control. BALB/c mice are highly susceptible while C57Bl/6 mice are relatively resistant. Macrophages eliminate opsonized trypanosomes from the bloodstream and are involved in immunosuppression. We therefore investigated the production of a number of cytokines (IL-10, IL-6, TNF-alpha and IL-12) by bone marrow-derived macrophages (BMDM) from C57Bl/6 and BALB/c mice following challenge with either Trypanosoma congolense or Trypanosoma brucei. BMDM from C57Bl/6 mice, upon challenge with whole cell extracts (WCE) of T. congolense or T. brucei, produced significantly more TNF-alpha and IL-12 than those from BALB/c mice. The production of these cytokines was significantly enhanced by pretreatment of the cells with IFN-gamma. BMDM from BALB/c mice, however, produced significantly more IL-6 and IL-10 than those from C57Bl/6 mice. In contrast to LPS stimulation, simultaneous treatment of cells with WCE and IFN-gamma enhanced IL-10 synthesis by BMDM from BALB/c mice. These results indicate that cytokine genes are differentially regulated in macrophages from trypanosome-susceptible and -resistant mice and are consistent with our previous findings wherein retrovirus-immortalized macrophage cell lines from BALB/c and C57Bl/6 mice produce differential amounts of cytokines after phagocytosis of trypanosomes.     

Dysregulation of the inflammatory response to the parasite, Toxoplasma gondii, in P2X7 receptor-deficient mice

The P2X(7) receptor (P2X(7)R) is a two transmembrane receptor that is highly expressed on the surface of immune cells. Loss of function polymorphisms in this receptor have been linked to increased susceptibility to intracellular pathogens. P2X(7)R gene knockout (P2X(7)R(-/-); on a C57Bl/6J background), C57Bl/6J and BALB/c mice were infected with the avirulent ME49 strain of the intracellular parasite, Toxoplasma gondii, and susceptibility determined by monitoring weight loss. P2X(7)R(-/-) mice lost significantly more weight than C57Bl/6J mice from day 8p.i. C57Bl/6J, in turn, lost significantly more weight than BALB/c mice. Thus, by day 10p.i., P2X(7)R(-/-) mice had lost 5.7 ± 0.7% of their weight versus 2.4 ± 0.6% for C57Bl/6J mice, whereas BALB/c mice had gained 1.9 ± 0.5%; by day 12p.i., P2X(7)R(-/-) mice had lost 15.1±0.6%, C57Bl/6J had lost 10.1±0.8% and BALB/c had lost 4.8 ± 0.8% of their weight. Neither parasite burden nor liver pathology was greater in the P2X(7)R(-/-) mice than in C57Bl/6J mice but BALB/c mice had significantly smaller numbers of parasites and less pathology in their livers than these strains. Absence of the P2X(7) receptor did not affect IFN-γ, IL-12, IL-1β, monocyte chemoattractant protein-1 (MCP-1) or TNF production. However, both P2X(7)R(-/-) and C57Bl/6J mice produced more IL-1β and TNF than BALB/c mice. There was one important point of differentiation between the P2X(7)R(-/-) and C57Bl/6J mice, namely the significantly enhanced and prolonged production of nitric oxide, accompanied by delayed production of IL-10 in the P2X(7)R-deficient mice.     

Innate resistance to Trypanosoma congolense infections: differential production of nitric oxide by macrophages from susceptible BALB/c and resistant C57Bl/6 mice.

BALB/c and C57B1/6 mice differ in resistance to Trypanosoma congolense infections. Evidence suggests that macrophages play a central role in the resistance to trypanosomiasis. Nitric oxide (NO) produced by macrophages in response to various stimuli or pathogens is one of the important arms of nonspecific immunity. We investigated the production of NO by the peritoneal macrophages and bone marrow-derived macrophages (BMDM) from trypanosome-resistant C57B1/6 and -susceptible BALB/c mice following stimulation with T. congolense whole cell extract (WCE) or following phagocytosis of T. congolense mediated by anti-variant surface glycoprotein (VSG) antibodies of IgM or IgG2a isotype. C57B1/6 peritoneal macrophages as well as BMDM produced significantly more NO than similar BALB/c macrophages in response to T. congolense lysate and IFN-gamma. In both BALB/c and C57B1/6 BMDM cultures, phagocytosis of T. congolense mediated by anti-VSG antibodies of IgG2a isotype in the presence of IFNgamma induced two- to ninefold more NO than phagocytosis mediated by IgM antibodies and C57B1/6 BMDM produced significantly higher amounts of NO than BALB/c BMDM under these conditions. NO produced by BMDM was found to exert trypanostatic effect on T. congolense in vitro, but was not found to influence the in vivo infectivity of these treated parasites under the conditions used in this study.     

B cell response during infection with the MAT a and MAT alpha mating types of Cryptococcus neoformans

In the present study, we compared the B cell response of BALB/c and C57Bl/6 mice during Cryptococcus neoformans infection. This response was investigated using virulent serotype D forms of mating types alpha and a (MAT alpha and MAT a). C57Bl/6 mice showed massive (mainly cerebral) infection by both types, while BALB/c were resistant to infection. Some resistance of C57Bl/6 mice was induced by previous immunization with the capsular polysaccharide from MAT alpha. Passive immunization of C57Bl/6 mice with purified antibody (Ab) obtained from capsular polysaccharide-immunized mice also increased resistance to infection. Both mouse strains showed comparable low IgM response to the capsular polysaccharide from MAT alpha, and only C57Bl/6 mice produced IgM to the polysaccharide of MAT a. Comparable levels of different immunoglobulin (Ig) isotypes against capsular components of MAT alpha and MAT a were detected, and the response of C57Bl/6 mice was higher when compared to that of BALB/c mice. FACS analysis indicated an increase in the percentage of a high-granulosity (side-scatter) splenic subpopulation and in the percentage of splenic Gr-1+ cells in infected C57Bl/6 mice. In addition, the percentage of follicular splenic B cells was decreased after C. neoformans infection of C57Bl/6 mice. This response was more pronounced when we investigated infection induced by the MAT a mating type. Taken together, our results indicate that capsular polysaccharide derived from MAT alpha and MAT a types of C. neoformans have a stimulatory effect upon B cells but that there is no correlation between resistance of BALB/c mice and Ab production. However, the increase in resistance of C57Bl/6 mice parallels the production of Abs and a major change in splenic cell populations.     

Poorer NF-kappa B signaling by microfilariae in macrophages from BALB/c mice affects their ability to produce cytotoxic levels of nitric oxide to kill microfilariae

Upon activation with microfilariae (mf), macrophages from C57Bl/6 mice showed higher nuclear factor-kappa B (NF-kappa B) but lower activating protein 1 DNA-binding activity as compared to BALB/c macrophages. The C57Bl/6 macrophages produced cytotoxic levels of nitric oxide (NO) to kill Setaria cervi mf as compared to BALB/c macrophages. Inhibition of the NF-kappa B signal by pyrrolidine dithiocarbamate (PDTC) blocked NO production and microfilaricidal activity of C57Bl/6 macrophages and inclusion of the exogenous NO generator (SNP) in the PDTC treated C57Bl/6 macrophage cultures induced mf cytotoxicity. These results underscore that the NF-kappa B signal (induced in response to mf) is important for the NO-mediated microfilaricidal activity of macrophages.     

Cytokines regulate the pattern of rejection and susceptibility to cyclosporine therapy in different mouse recipient strains after cardiac allografting

We determined the role of cytokines in regulating the pattern of rejection and recipient susceptibility to cyclosporine (CsA) in a mouse cardiac allograft model. Hearts from C3H mice transplanted into untreated BALB/c (Th2-dominant) and C57BL/6 (Th1-dominant) mice showed different patterns of rejection. C3H allografts in BALB/c mice showed typical acute vascular rejection (AVR) with strong intragraft deposition and high serum levels of anti-donor IgG with predominant IgG1, while C3H allografts in C57BL/6 mice showed typical acute cellular rejection (ACR) with massive intragraft infiltration of CD4(+) and CD8(+) lymphocytes and low serum levels of anti-donor IgG with predominant IgG2a. Elevated intragraft mRNA expression of IL-2, IFN-gamma, and IL-12 mRNA was present in C57BL/6 recipients, whereas allografts in BALB/c mice displayed increased IL-4 and IL-10 mRNA levels. CsA therapy completely inhibited ACR and induced indefinite allograft survival in C57BL/6 recipients, while the same therapy failed to prevent AVR, and only marginally prolonged graft survival in BALB/c recipients. In contrast, rapamycin blocked AVR, achieving indefinite survival in BALB/c recipients, but was less effective at preventing ACR in C57BL/6 recipients. The disruption of the IL-12 or IFN-gamma genes in C57BL/6 mice shifted ACR to AVR, and resulted in concomitant recipient resistance to CsA therapy. Conversely, disruption of IL-4 gene in BALB/c mice markedly attenuated AVR and significantly prolonged allograft survival. These data suggest that the distinct cytokine profiles expressed by different mouse strains play an essential role in regulating the pattern of rejection and outcome of CsA/rapamycin therapy.     

Analysis of the specific 3H-diazepam binding in the brain of inbred mice with differing emotionality

The influence of sodium chloride and gamma-aminobutyric acid (GABA) on H3-diazepam binding in CNS membrane fraction were investigated in C57Bl/6, BALB/c mice and their F1 hybrids. The addition of GABA (1, 10 and 100 microM) elevated the level of radioligand binding with CNS membranes in all the inbred mice in similar manner. The higher stimulating effect of sodium chloride (50, 100 and 150 mM) on the H3-diazepam reception was found in BALB/c mice and F1 hybrids, as compared to C57Bl/6 mice. It has been suggested that membrane-dependent conformational reconstructions of supramolecular receptor complex are the cause of genetic differences in the regulation of H3-diazepam reception by Cl(-)-ionophore.     

Mechanisms of self-cure from Trypanosoma congolense infection in mice

The mechanism(s) of resistance to African trypanosomiasis caused by Trypanosoma congolense was investigated by using the Dinderesso/80/CRTA/3 isolate to which C57B1/6 are resistant (low parasitemia and self-cure) and BALB/c sensitive (high parasitemia and death). The resistance of C57B1/6 is similar to that found in some natural hosts of African trypanosomes such as certain indigenous West African cattle and wild Bovidae. The antibody response to epitopes exposed on the variant surface glycoprotein of a clone obtained from the Dinderesso/80/CRTA/3 isolate was measured by a complement-mediated lysis assay in C57B1/6 and BALB/c. After infections with 10(4), 10(5), or 10(7) motile organisms, antibody appeared in C57B1/6 4 to 8 days earlier than in BALB/c. Peak antibody titers were similar in both strains but were reached about 4 days earlier in C57B1/6. In this strain, antibody appeared during and controlled the first wave of parasitemia, whereas in BALB/c, parasitemia reached a plateau above 10(8) organisms per ml before antibody could be detected, and at this time the animals were dying. At peak antibody response, the proportion of immunoglobulin (Ig) M and IgG antibody was the same in both strains. The antibody response had the same kinetics in C57B1/6 and BALB/c after injection of 10(4), 10(5), and 10(7) lethally irradiated but intact parasites, but the peak titers were 10(3) to 10(4) times lower than after live challenge. The response to nonirradiated trypanosomes appeared to be T cell independent, because the antibody titers were the same in congenitally athymic nu/nu and normal C57B1/6, and no evidence for the induction of T cell activity could be demonstrated in the infected nude mice. A role for trypanolytic serum factors in resistance could not be demonstrated. The extent of immunosuppression after infection with nonirradiated organisms was compared in the two strains by measuring the in vitro response of their splenic lymphocytes to concanavalin A, pokeweed mitogen, and allogeneic cells and their ability to mount an in vivo response to an unrelated trypanosome challenge. Both strains were partially immunosuppressed during rising parasitemia, but as C57B1/6 controlled parasitemia, immunosuppression was gradually reversed, whereas in BALB/c it became worse. Several explanations might account for the resistance of C57B1/6 to the Dinderesso/80/CRTA/3 isolate of T. congolense. It appears that an early immune response is a decisive factor in this resistance.     

Vascular extracellular adenosine metabolism in mice correlates with susceptibility to atherosclerosis

Abstract

Animal models are widely used in atherosclerosis research. The most useful, economic and valid is mouse genetic model of this pathology. Purinergic signaling is an important mechanism regulating processes involved in the vascular inflammation and atherosclerosis. The aim of this study was to measure vascular activities of nucleotide and adenosine-degrading ecto-enzymes in different strains of mice and to compare them to atherosclerotic susceptibility.

The vascular extracellular nucleotide catabolism pathway was analyzed in 6-month-old male genetically unmodified mouse strains: FVB/NJ, DBA/2J, BALB/c, C57Bl/6J and mouse knock-outs on C57Bl/6J background for LDLR (LDLR-/-) and for ApoE and LDLR (ApoE-/-LDLR-/-). LDLR-/- mice were a model of moderate hypercholesterolemia, while ApoE-/-LDLR-/- mice, a model of severe hypercholesterolemia with advanced atherosclerosis.

FVB/NJ, DBA/2J and BALB/c mice showed high rates of vascular extracellular AMP hydrolysis and low activity of adenosine deamination. In turn, all mice with the C57Bl/6J background expressed diminished activity of vascular AMP hydrolysis. Mice with genetically-induced hyperlipidemia and atherosclerosis on the C57Bl/6J background revealed increased ecto-adenosine deaminase activity.

Mouse strains that were resistant to atherosclerosis (FVB/NJ, DBA/2J, BALB/c) exhibited a protective extracellular vascular ecto-enzyme pattern directed toward the production of anti-inflammatory and anti-atherosclerotic adenosine. In turn, mice with genetically induced hypercholesterolemia and atherosclerosis expressed disturbed activities of ecto-5’nucleotidase and ecto-adenosine deaminase related to decreased production and increased degradation of extracellular adenosine.     

Susceptibility and resistance to Trypanosoma congolense infections

We have put emphasis on recent findings in experimental Trypanosoma congolense infections in highly susceptible BALB/c and relatively resistant C57Bl/6 mice. Based on various analyses, it has been shown that a major difference in resistance to T. congolense infections is expressed early in infection at the macrophage level. A novel plastic-adherent Thy1.2(+) suppressor lymphocyte, which in absolute synergy with a Thy 1.2(-) cell exerts its suppression via interleukin-10 and interferon-gamma opens up an exciting new field of research.     

Genetic control of heat sensitivity and activity level of murine arylsulfatase B

BALB/c male mice possess twofold higher kidney p-nitrocatechol-SO4 arylsulfatase B than do A/HeJ male mice; however, their liver arylsulfatase activities are comparable. Twentyfold-purified kidney arylsulfatases B from these two strains have similar Michaelis constants, electrophoretic mobilities, pH optima, and inhibitor profiles; however, the BALB/c enzyme is more heat stable than the A/HeJ enzyme. BALB/c, C3H/HeJ, DBA/2J, and SWR/J mice share an autosomal allele, As-1a, which apparently determines the heat-stable arylsulfatase B, while A/HeJ and C57BL/6J mice possess the As-1b allele, which determines the heat-sensitive enzyme. A second autosomal locus, Asr-1, determines liver arylsulfatase B activity. C57BL/6J mice carry the Asr-1a allele, which results in high liver activities, while C3H/HeJ mice are homozygous for the low-activity allele, Asr-1b. Male mice generally have 30-40% higher kidney activities than females; however, female kidney arylsulfatase activities rise and actually surpass those of males during late pregnancy and lactation.     

Effect of Salmonella enteritidis 11RX infection on two-stage skin carcinogenesis in mice.

S. enteritidis 11RX infection inhibits the growth of a number of transplantable tumours in mice. In addition, oral infection of mice with S. enteritidis 11RX inhibits colon carcinogenesis by 1,2 dimethylhydrazine. This study has examined the effect of S. enteritidis 11RX infection on two-stage skin carcinogenesis in mice using 7,12 dimethylbenz(a)anthracene (DMBA) as initiator and croton oil as promoter. No protection was observed in either LACA or (BALB/c x C57Bl/6J)F1 mice when live 11RX was repeatedly administered i.v. during promotion. When a protein antigen extract from S. enteritidis 11RX was administered i.v. to previously immunised mice during skin carcinogenesis, significant protection was observed both in terms of the number of mice with papillomas and the number of papillomas per mouse. However, the protection was weak and transient. LACA mice were much more susceptible to skin carcinogenesis by DMBA and croton oil than were (BALB/c x C57B1/6J)F1 mice. A preliminary study indicated that BALB/c, C57B1 and CBA mice were also relatively resistant to skin carcinogensis.     

Protection of mice against influenza virus infection: Enhancement of nonspecific cellular responses by Corynebacterium parvum

Groups of C57Bl/6J, BALB/c, BALB/c nu⁺/nu⁺ mice, inoculated intranasally with Corynebacterium parvum (350 μg/mouse) were protected from death by an otherwise lethal dose of influenza virus, A/WSN (H1N1) inoculated 3 days later. The lungs of C. parvum-trented, virus-infected C57Bl/6J, BALB/c, or BALB/c nu⁺/nu⁺ mice contained significantly less infectious virus than did controls, and this reduction was apparent as soon as 24 hr after virus inoculation. The maximum protective effect correlated with increased lung interferon levels. C. parvum treatment caused an increase in the lung cell number which was in part due to a large increase (ca. 10-fold) in macrophage content, and the natural killer cell activity was also enhanced, though not as markedly as occurred 3 days after infection. Most (>85%) of the resident macrophages in normal lungs were susceptible to infection by virus (as indicated by hemadsorption), whereas most of those recovered from the lungs of C. parvum-treated mice resisted infection. Despite the increase in macrophage content, the level of specific immune responses to infection, such as cytotoxic T-cell activity, DTH reaction, and antihemagglutinin antibody, remained unchanged by C. parvum treatment so that the major if not only effect of this treatment was on the level of the less-specific components of the immune system.     

Comparison of survival rates in four inbred mouse strains under different housing conditions: effects of environmental enrichment

Housing conditions can affect the well-being of laboratory animals and thereby affect the outcomes of experiments. The appropriate environment is essential for the expression of natural behavior in animals. Here, we compared survival rates in four inbred mouse strains maintained under three different environmental conditions. Three mouse strains (C57BL/6J, C3H/HeN, and DBA/2J) housed under environmental enrichment (EE) conditions showed improved survival; however, EE did not alter the survival rate of the fourth strain, BALB/c. None of the strains showed significant differences in body weights or plasma corticosterone levels in the three environmental conditions. For BALB/c mice, the rates of debility were higher in the EE group. Interestingly, for C57BL/6J and C3H/HeN mice, the incidence of animals with alopecia was significantly lower in the EE groups than in the control group. It is possible that the enriched environment provided greater opportunities for sheltering in a secure location in which to avoid interactions with other mice. The cloth mat flooring used for the EE group was bitten and chewed by the mice. Our findings suggest that depending on the mouse strains different responses to EE are caused with regard to health and survival rates. The results of this study provide basic data for further studies on EE.     

Genetic differences in BCG-induced resistance to Schistosoma mansoni are not controlled by genes within the major histocompatibility complex of the mouse.

Induction of nonspecific resistance to Schistosoma mansoni infection after the i.v. injection of viable BCG was investigated in outbred mice and a panel of inbred and H-2 congenic strains. Significant protection was induced in CF1, A/J, C57BL/6, C57BL/10, DBA/2, C57BR, and SJL mice. BALB/c mice were not protected whereas CBA and C3H mice expressed intermediate degrees of protection. Expression of the protective phenomenon is not controlled by genes within the MHC as shown by the marked differences in response between BALB/c and DBA/2 (H-2d) as well as between C57BR and C3H (H-2k) mice. H-2 congenic strains with C57BL/10 background (B10.A and B10.D2) were high responders. BALB.B10 mice carrying the high responder (B10) MHC on the nonresponder (BALB/c) background were not protected. The degree of splenic hypertrophy did not correlate with the expression of nonspecific resistance. These results demonstrate that, in addition to controlling specific immune responses, genetic differences influence the nonspecific protective phenomena related to BCG administration as well.     

Genetic analysis of strains C57BL/6J and BALB/cJ for Ath-1, a gene determining atherosclerosis susceptibility in mice

We previously reported that mice have at least one major gene determining atherosclerosis susceptibility, Ath-1. Susceptible alleles of Ath-1 are found in strain C57BL/6J and are associated with relatively low levels of high-density lipoprotein cholesterol (HDL-C) when these mice are fed an atherogenic diet. Resistant alleles of Ath-1 are found in strains C3H/HeJ and BALB/cJ and are associated with relatively high levels of HDL-C. Data reported earlier from the set of seven recombinant inbred (RI) strains, derived from C57BL/6By and BALB/cBy, showed that these parental strains differed at Ath-1. However, due to the limited number of RI strains, it was not possible to determine with certainty whether Ath-1 was the only major gene determining atherosclerosis susceptibility in these two strains or to determine its map position accurately. In this report, examination of F1, F2, and backcross progeny from a cross between C57BL/6J and BALB/cJ demonstrates that Ath-1 is the major gene determining atherosclerotic lesion formation and HDL-C levels in female mice. The data from male animals suggest that environmental factors or modifying genes also influence male HDL-C levels and thus partly obscure the Ath-1 phenotype. HDL-C levels in F1 progeny resemble the BALB/c parent. The data from the cross provide confirmatory evidence that Ath-1 is linked to Alp-2 on chromosome 1 with a map distance of 4.8 +/- 2.3 (SE). Combining these data with a previous cross between strain C57BL/6 and strain C3H/HeJ gives a map distance between Ath-1 and Alp-2 of 4.9 +/- 1.8 based on 7 crossovers found among 144 tested chromosomes.     

The effects of genetic variation in N-acetyltransferases on 4-aminobiphenyl genotoxicity in mouse liver

Inbred, congenic and transgenic strains of mice were characterized for acetylation of p-aminobenzoic (PABA) and the carcinogen 4-aminobiphenyl (4ABP). C57Bl/6 mice have the NAT2*8 allele, A/J mice have NAT2*9 and congenic B6.A mice have NAT2*9 on the C57Bl/6 background. The first transgenic strain with human NAT1, the functional equivalent of murine NAT2, was also tested. The murine NAT2*9 allele correlated with a slow phenotype measured with the murine NAT2 selective substrate PABA. The two strains having this allele also had a lower capacity to acetylate 4ABP. A line with five copies of the human NAT1 transgene was bred for at least five generations with either C57Bl/6 or A/J mice. There was no significant change in PABA NAT activity on the C57Bl/6 background but a 2.5-fold increase was seen in hNAT1:A/J compared with A/J. The effect of variation in NATs on 4ABP genotoxicity was assessed in these strains. Twenty-four hours after exposure to a single oral dose of 120 mg 4ABP/kg, hepatic 4ABP-DNA adducts were detected by immunofluoresence in all strains. Nuclear fluorescence intensities (mean+/-S.D.) were 41.1+/-3.6 for C57Bl/6, 37.9+/-1.11 for A/J and 36.3+/-2.44 for B6.A. There was no correlation between murine NAT2 alleles and 4ABP-DNA adduct levels. Similar results were seen with the transgenic strains. The data indicate that the range of variation present in these strains of mice was insufficient to alter susceptibility to 4ABP genotoxicity. The impact of these relatively modest differences in the acetylation of the activation of 4ABP may be masked by other competing biotransformation reactions since 4ABP is a substrate for both NAT1 and NAT2. Mouse models with variation in both isoforms are needed to adequately assess the role of variation in NATs in susceptibility to 4ABP genotoxicity.     

Induction of xanthine oxidase and depression of cytochrome P-450 by interferon inducers: genetic difference in the responses of mice

Interferon and interferon inducing agents depress hepatic cytochrome P-450 systems. They also induce hepatic xanthine oxidase activity. It has been suggested that free radicals produced by xanthine oxidase may cause the loss of P-450. High titers of serum interferon are induced by poly IC (poly riboinosinic acid.polyribocytidylic acid) in both C57Bl/6J and C3H/HeJ mice; Newcastle disease virus (NDV) induces a high titer of interferon in C57Bl/6J mice but not in C3H/HeJ mice. The induction of xanthine oxidase activity by NDV in C3H/HeJ mice was less than half that seen in C57Bl/6J mice, thus demonstrating a relationship between the induction of xanthine oxidase, the depression of P-450 and a genetically determined difference in responsiveness of mice to interferon inducers.