Mycobacterium lepraemurium and the Ity/Lsh/Bcg mouse gene.
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Animals↗
Biomedical subjects
I N Brown
Publications and source records attributed to I N Brown.
Interferon-gamma as an adjuvant in immunocompromised mice.
We have compared interferon-gamma (IFN-gamma) with saponin and interleukin-1 (IL-1) as adjuvants for a blood-stage malaria vaccine in mice with various immunological abnormalities. IFN-gamma was particularly effective in Biozzi low antibody responder mice, mice selectively bred to produce antibody of low affinity, and mice depleted of CD4+ T cells. IFN-gamma and other cytokines may be safe adjuvants for use in human immunodeficiency states.
Adjuvants, Immunologic↗
Derivation and biochemical characterization of an enterovirus group-specific monoclonal antibody.
This paper describes the derivation and characterization of monoclonal antibodies reactive with the group-specific epitopes of the VP1 peptide shared by enteroviruses. When tested against a wide range of the prototype strains of Coxsackie viruses A and B, polioviruses, and echoviruses, the monoclonal antibodies reacted with all of them in two or more assay systems. The virus of hepatitis A was the only enterovirus tested which was not recognized by these monoclonal antibodies. This finding is consistent with recent reports demonstrating genomic dissimilarity between this virus and the other enteroviruses. Although monoclonal antibodies have frequently been found to be assay specific, these particular antibodies were found to be equally effective in different immunochemical and biological assay systems. Biochemical characterization of one of these monoclonal antibodies is described and its potential usefulness for diagnosis is discussed.
Antibodies, Monoclonal↗
Clinical and research application of an enterovirus group-reactive monoclonal antibody.
Diagnostic methods employed in enterovirus laboratories are generally laborious, slow and expensive. This is largely because type-specific neutralization tests still play the major role in identification and diagnostic serology. In the companion paper we describe the derivation of monoclonal antibodies against epitopes of the VPI peptide which are shared by all of the enteroviruses tested to date, with the exception of hepatitis A virus. This study describes the application of one of these monoclonal antibodies in several research and diagnostic procedures, illustrating a special utility in a wide variety of assay systems. This monoclonal antibody has proved particularly useful in the detection of enterovirus antigens in circulating immune complexes, and in identifying field isolates of this group of viruses. Immunohistochemistry, previously almost impossible in enterovirus diagnosis and research due to the large number of serotypes, is now shown to be practical and informative when this monoclonal antibody is used.
Animals↗
The Ity/Lsh/Bcg gene significantly affects mouse resistance to Mycobacterium lepraemurium.
Mouse resistance to infection with Mycobacterium lepraemurium was measured by counting the total number of intact acid-fast bacilli in the spleen 8 weeks after i.v. injection of a standard inoculation. The effect of Ityr on resistance to M. lepraemurium was confirmed and the results extended to two Ityr strains of mice, A and C57L, not previously tested. Resistance to M. lepraemurium was also examined in the F1, backcross and F2 generations of BALB/c X CBA crosses, and in the congenic strain B10.LLshr that is Ityr. In all experiments the results were consistent with the view that resistance to M. lepraemurium is significantly affected by a gene close to or identical to the Ity/Lsh/Bcg gene on mouse chromosome 1. Sex had a marked effect on resistance to M. lepraemurium, so that the males of some genetically resistant strains were almost as susceptible as some genetically susceptible females.
Animals↗
The effect of oral Mycobacterium vaccae on subsequent responses of mice to BCG sensitization.
It has been postulated that previous exposure to mycobacteria in the environment may be a contributing factor to the variable efficacy of BCG vaccination in protecting against human tuberculosis or leprosy, in different geographical regions. To test this hypothesis mice were given Mycobacterium vaccae in their drinking water for 3 weeks immediately, 27 days, or 54 days before they were injected subcutaneously with BCG. Spleen cells were examined, 50 days after injection of the mice, for ability to proliferate in vitro in response to killed mycobacteria added to the cultures. The results show that, depending on the timing of the exposure of the mice in vivo to M. vaccae before BCG injection and the dose of the mycobacterial challenge in vitro, oral administration of this species of environmental mycobacteria can either enhance, mask or interfere with the expression of sensitization by BCG. Thus, our data from mice support the hypothesis that the results of BCG vaccination for a particular human population are influenced by exposure to indigenous mycobacteria.
Animals↗
Rapid methods for counting mycobacteria--comparison of methods for extraction of mycobacterial adenosine triphosphate (ATP) determined by firefly luciferase assay.
A comparison of 5 different methods of extraction of adenosine 5'-triphosphate (ATP) from mycobacterial cells was carried out using Mycobacterium bovis, BCG as a model. ATP was measured using the luciferin-luciferase bioluminescence reaction. Boiling buffer extraction was the best method. The amount of ATP extracted correlated with the number of colony forming units over a wide range of count. Although great sensitivity in terms of number of bacilli detectable was not achieved the method was rapid and appears suitable for drug sensitivity testing of tubercle bacilli.
Adenosine Triphosphate↗
Macrophage functions in Biozzi mice.
The faster degradation of antigen by macrophages in Biozzi low (L) responder mice, compared to Biozzi high (H) responder mice, is thought to be responsible for their lower antibody response. We have measured four functions associated with macrophages to see whether macrophages from L mice were generally more active than those from H mice. Peritoneal macrophages obtained from normal mice were compared with those from groups of mice given Mycobacterium bovis BCG or Propionibacterium acnes. Cells from normal H mice gave a stronger oxidative burst when triggered with phorbol myristate acetate, and were more cytotoxic for tumour cells than cells from L mice. Cells from all mice injected with BCG or P. acnes gave a stronger oxidative burst, and were more cytotoxic for tumour cells; again, both responses were higher in H mice than in L mice. By contrast, when groups of mice that had received P. acnes were given endotoxin and bled, higher titres of tumour necrosis factor were found in the sera of L mice. Spleen cells from both lines of mice released similar levels of interleukin-1, both spontaneously and in response to lipopolysaccharide. Our results suggest that these various macrophage responses are expressed independently in H and L mice.
Animals↗
Mycobacterium vaccae and immune responses: implications for leprosy control.
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Animals↗
Inbred mouse strain resistance to Mycobacterium lepraemurium follows the Ity/Lsh pattern.
Inbred mouse strains and their F1 hybrids infected intravenously with Mycobacterium lepraemurium showed different mean survival times (MST). BALB/c and C57BL mice were particularly susceptible, whereas C3H, CBA and DBA/2 mice were relatively resistant. Resistance as judged by MST was dominant in the F1 hybrids. A similar ranking order was obtained by comparing the doubling time of the bacillus in the bone marrow, the increase in spleen weight between 4 and 12 weeks after infection, and the pathology of the liver during infection. The general pattern suggests that mouse resistance to M. lepraemurium is, at least in part, controlled by a gene with the same strain distribution as the genes for resistance to Salmonella typhimurium (Ity') and Leishmania donovani (Lsh') and the gene controlling resistance to Mycobacterium bovis BCG (Bcg). Ity, Lsh and Bcg are all known to be on chromosome 1, suggesting a centre controlling reactions to intracellular infections.
Animals↗
Mycobacterium bovis, BCG, modulation of murine antibody responses: influence of dose and degree of aggregation of live or dead organisms.
Mycobacteria have the ability to enhance or depress immune responses. This paper describes experiments designed to investigate the parameters determining the direction of modulation. It has been shown previously that 10(8) liver Mycobacterium bovis BCG depress the ability of mouse spleen cells to produce a primary antibody response in vitro to SRBC 2-3 weeks after i.v. injection, whereas the same number of dead organisms enhance this response. Using the same growth medium for the BCG (Glaxo glycerol-free medium), we now find that decreasing the BCG dose to mice from 10(8) to 10 (6) liver organisms results in enhanced responses and increasing the dose to more than 10(8) dead organisms results in depressed responses. It thus appears that bacterial load is the important factor determining whether depression or enhancement of the primary antibody response will occur, rather than the viability of the organisms per se. However, when the BCG was grown in Middlebrook 7H9 broth, doses as high as 4 X 10(9) dead BCG/mouse failed to depress although depressed responses were found if sufficient live organisms (7 X 10(8)) were injected. In view of the known growth characteristics of BCG in these 2 bacteriological media, it is suggested that the degree of aggregation of the injected suspension may also be of importance in determining whether or not depression will occur. A comparison of the effects of BCG injected untreated or after dispersion of bacterial aggregates supports this idea. Some degree of splenomegaly was always found in mice with depressed splenic responses but a large spleen did not necessarily yield cell suspensions with depressed responses.
Animals↗
BCG, Corynebacterium parvum or Mycobacterium leprae added to cultures of BCG-primed mouse spleen cells cause an enhanced primary antibody response in vitro.
A few weeks after mice were injected i.v. with 10(8) live Mycobacterium bovis, BCG, the antibody response of their spleen cells to SRBC in vitro was comparable with the response of cells from untreated mice. Addition of BCG organisms to the culture vessels resulted in enhanced antibody-forming cell (AFC) responses by the primed cells but not by the cells from the untreated mice. No evidence was found for a direct stimulation of B cells and cell depletion experiments suggested macrophages were directly involved. BCG added to the cultures up to 68 h after they were set up, but not later, still caused enhancement. No enhancement was found when DNP-Ficoll was used as antigen. The ability to stimulate the anti-SRBC response was not restricted to the organism used for priming. Enhancement was also found if C. parvum or M. leprae were added to BCG-primed cells and if BCG was added to C. parvum-primed cells. The relevance of the results to the search for a leprosy vaccine is discussed.
Animals↗
Active suppression masks an underlying enhancement of antibody production in vitro by spleen cells from BCG-infected mice.
The depressed antibody responses resulting from the administration of live BCG i.v. to mice have been investigated. The antibody response of spleen cells to SRBC or DNP-Ficoll in vitro was followed using Marbrook culture vessels. Depressed responses were also found in vivo confirming the results obtained in vitro. The response in vitro of normal spleen cells was suppressed by the addition of spleen cells from mice injected with BCG but not by the medium in which they had been growing for 2 days. The response of the normal spleen cells was also not suppressed by freeze/thaw disrupted BCG spleen cells, suggesting that the depressed responses in the mice injected with BCG are due to an active suppression by intact cells. This was confirmed by the cell-depletion experiments. Removal of cells from the BCG-primed cell populations using carbonyl iron or adherence to plastic not only abrogated the depressed responses but revealed an underlying enhancement of the immune response. The data suggest that the suppressive cell might be a macrophage.
Animals↗
Phagosome/lysosome fusion: a possible prerequisite for the enhancement of antibody responses in vitro by BCG, Mycobacterium leprae and Corynebacterium parvum.
Primary in vitro antibody responses to SRBC were suppressed in cultures prepared from the spleens of CBA mice injected i.v. 20 days previously with 10(8) liver BCG. In contrast, cultures prepared from mice injected with dead BCG showed enhanced responses. In vitro spleen cell responses of the mice had returned to normal levels 4--6 weeks after their injection, but if dead BCG, M. leprae or C. parvum was added to the cultures, responses were enhanced. The enhancing effect of the added bacteria could be removed by adding also suramin, a drug known to inhibit in vitro fusion of lysosomes with phagosomes. It is suggested that the different in vivo effects of live and dead BCG may relate to differences in their handling by macrophages and more especially that the enhanced antibody forming cell response seen in the restimulated cultures of spleen cells from BCG primed mice, depends upon efficient intracellular fusion of lysosomes with the phagosomes containing the added dead bacteria.
Adjuvants, Immunologic↗
Suppressed or enhanced antibody responses in vitro after BCG treatment of mice: importance of BCG viability.
Mycobacterium bovis, BCG, is known to be capable of either enhancing or suppressing various immune responses. Using a standard technique and number of organisms, some of the parameters predetermining whether enhancement or supression will occur have been investigated. Dead BCG given intravenously into mice caused an enhancement of the antibody response in vitro to sheep erythrocytes. In contrast, the same number of viable organisms caused suppression if given intravenously but enhancement if given subcutaneously. The inclusion of 25% or more killed organisms in an intravenous inoculum of fully viable organisms changed suppression to enhancement. Treatment of BCG infected mice with streptomycin lessened the suppression but did not change it to enhancement. The possible causes of suppression are discussed.
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Antibody response in vitro of spleen cells from Plasmodium yoelii-infected mice.
The primary antibody response to sheep erythrocytes and dinitrophenylated Ficoll of spleen cells from mice infected with Plasmodium yoelii was studied in vitro. The response to sheep erythrocytes was enhanced between 2 and 4 days after infection and depressed at later intervals. Cell fractionation experiments carried out at the time of immunosuppression indicated a defect of macrophage function. At the same time the response to dinitrophenylated Ficoll was normal.
Animals↗
Enhancement of the antibody response in vitro by adherent cells from mice infected with Mycobacterium lepraemurium.
Spleen cells from mice systemically infected 4 to 6 weeks previously with Mycobacterium lepraemurium gave an enhanced primary antibody response in vitro to sheep erythrocytes, but responded normally to dinitrophenylated polyumerized flagellin. The ability to enhance the response was associated with the glass-adherent spleen cell population and with peritoneal cells. Similar cells obtained from infected mice depleted of T lymphocytes failed to enhance the antibody response. These studies suggest that macrophages that became activated during the development of cell-mediated immunity to infection can also stimulate antibody responses to thymus-dependent antigens.
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Immunologically mediated macrophage aggregation in monolayers of peritoneal cells from BCG-sensitized mice.
Aggregation of cultured macrophage monolayers derived from BCG-sensitized mice was produced if nonadherent cells and specific antigen (tuberculin) were present, particularly if the antigen was renewed in the form of tubercle bacilli. The evidence indicated that antigen-stimulated BCG-sensitized lymphocytes in these cultures produced a soluble factor, which in the presence of the renewed supply of antigen caused the aggregation. The phenomenon was irreversible and followed by death of the macrophages. The 'overlays' of aggregated monolayers would aggregate normal macrophages, provided that the recipient cultures contain-d their own (normal) lymphocytes as well as antigen; this suggested that cultures of BCG-sensitized peritoneal cells produced a factor able to effect aggregation via the activity of normal lymphocytes. Overlays from aggregated monolayers were able also to inhibit the migration of normal mouse macrophages; this and other evidence suggested a similar origin for the inhibition factor, but the latter's identity with the aggregation factor remains undecided. We conclude that the aggregation factor depends upon the presence of specific antigen both for its formation and its expression.
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