Low antibody responsiveness is found to be associated with resistance to chemical skin tumorigenesis in several lines of Biozzi mice.

High and low antibody responder lines of mice from Selections I, III and G were assayed for two-step skin tumorigenesis using a protocol consisting in initiation with 7,12-dimethylbenz[a]anthracene (DMBA) and promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). Concordant results were obtained in the three selections: low antibody responder mice were shown to be significantly more resistant to tumor induction than the high responder counterparts. The difference was observed for all parameters: kinetics and percentages of tumor incidence and tumor multiplicity. The three bidirectional selective breeding experiments differed in several respects namely, the origin of the foundation populations, the antigens and immunization protocols used during the selection, as well as the breeding unit environments. Therefore, the consistent results relative to tumorigenesis strongly suggest that some of the alleles relevant to multispecific 'low' antibody production could contribute to the resistance to cutaneous chemical tumorigenesis.

Innate resistance to infection by intracellular bacterial pathogens differs in mice selected for maximal or minimal acute inflammatory response.

The intensity of nonspecific immune reaction and the host resistance to facultative intracellular pathogens are found to be associated in lines of mice selected for maximal (AIRmax) or minimal (AIRmin) acute inflammatory reactivity. AIRmax are more resistant than AIRmin mice to Salmonella typhimurium and Listeria monocytogenes infection, the differences between lines in LD50 being > 1000 and 100 times, respectively. This difference was shown to be related to the initial bacterial containment at the infectious focus, and to the control of bacterial multiplication in the spleen during the 1st week after s. c. inoculation of the bacteria. Specific immune responses were not deeply affected by the selective process: antibody production and delayed-type hypersensitivity were both of similar intensity in AIRmax and AIRmin mice. The differential susceptibility to infection seems independent of the Nramp-1 locus polymorphism; therefore, these two lines represent a powerful model for investigating the role of other genetic loci regulating the nonspecific immunity effectors in the course of infectious diseases.

Susceptibility and resistance to Leishmania amazonensis in H-2q syngeneic high and low antibody responder mice (Biozzi mice).

H-2 syngeneic H and L (Biozzi) mice provide a model to study Leishmania infections in which polar resistant and susceptible phenotypes are independent from H-2 differences. High-Ab-responder (H) and low-Ab-responder (L) mice syngeneic at the H-2 locus (H-2q) were, respectively, susceptible and highly resistant to Leishmania amazonensis infection. L-mice resistance was associated with high IFN-gamma and transient IL-4 production by lymph node (LN) cells, in contrast with sustained IL-4 and decreasing IFN-gamma production by susceptible H mice. IL-12 production could be detected only in LN from resistant mice. The cytokine production pattern was consistent with preferential progression to a Th1-type response in resistant L-mice, and to a Th2-type response in susceptible H-mice. We also investigated whether this shift towards Th1- or Th2-type cytokine responses was dependent upon H or L antigen presenting cells' (APC) intrinsic ability to preferentially stimulate either T-cell subset. To this end, LN-derived T-cell lines were grown from 12-day infected mice, when both strains produced IFN-gamma and IL-4. L-derived T-cell lines developed a Th2 cytokine pattern whereas H-derived T-cell lines produced IFN-gamma, IL-4 and IL-10 whatever the APC origin (H or L) used for their derivation. This work constitutes the first characterization of cellular immune responses to the intracellular parasite, L. amazonensis in H-2 syngeneic mice, an infection model in which polar resistant and susceptible phenotypes are determined by non-MHC genes.

Innate resistance to infection by intracellular bacterial pathogens differs in mice selected for maximal or minimal acute inflammatory response

The intensity of nonspecific immune reaction and the host resistance to facultative intracellular pathogens are found to be associated in lines of mice selected for maximal (AIRmax) or minimal (AIRmin) acute inflammatory reactivity. AIRmax are more resistant than AIRmin mice to Salmonella typhimurium and Listeria monocytogenes infection, the differences between lines in LD50 being > 1000 and 100 times, respectively. This difference was shown to be related to the initial bacterial containment at the infectious focus, and to the control of bacterial multiplication in the spleen during the 1st week after s. c. inoculation of the bacteria. Specific immune responses were not deeply affected by the selective process: antibody production and delayed-type hypersensitivity were both of similar intensity in AIRmax and AIRmin mice. The differential susceptibility to infection seems independent of the Nramp-1 locus polymorphism; therefore, these two lines represent a powerful model for investigating the role of other genetic loci regulating the nonspecific immunity effectors in the course of infectious diseases.

Functional differences in the specific B-cell compartment in high or low antibody responder mice.

The role of antigen-presenting cells (APC) in quantitative antibody synthesis regulation was studied in mice genetically selected for high (HI) or low (LI) antibody response. Irradiated spleen cells and enriched specific B cells from HI and LI mice co-isogenic at H-2s locus, were compared for their capacity to present chicken ovalbumin (OVA) to specific T-cell hybridomas. Minor differences were observed between HI and LI mice when three distinct hybridomas were stimulated in the presence of OVA and splenic macrophages APC. These differences were totally abolished when APC were pulsed with OVAxAb complexes. Looking at the B-cell compartment, hybridoma IL-2 responses were similar when TNP primed B cells were pulsed with OVA. However, when OVA was targeted on TNP-specific enriched B cells by pulsing with TNP-OVA, the IL-2 production by the T-cell hybridomas was stronger in the presence of HI B cells than in the presence of LI B cells. These results strongly suggest that an efficient Ag handling/processing by specific B cells is a major component of the high Ab responder status in Biozzi mice.

T-cell compartment involvement in two high antibody responder lines of mice (HI and HII Biozzi mice) respectively susceptible and resistant to collagen-induced arthritis.

The T-cell compartment was investigated in two high antibody responder lines of mice respectively susceptible (HI) and resistant (HII) to chicken collagen (CII)-induced arthritis (CIA). Previous data had shown that both lines were high anti-CII Ab producers, without any TCR V-beta gene defect or membrane expression impairment. The present studies demonstrate that anti-CII proliferation is much lower in HII than in HI. These results are confirmed by the limiting dilution analysis of anti-CII T-precursor frequencies (1/991 in HI and 1/12175 in HII). The percentage of CD8+ T cells is constitutively higher in HII mice, this difference increasing after CII immunization. This finding suggests a suppressive effect accounting for resistance to CIA. However, no restoration of specific response was achieved by in-vivo or in-vitro depletion of CD8+ T cells. T clones specific for Chicken CII could be obtained only from primed HI mice. Four of five clones with CD8+ phenotype proliferated in vitro to native and denatured CII and showed cytotoxic function in an anti-CD3 redirected assay. The CD4+ clone was shown to proliferate on both HI and HII-pulsed APC, which rules out a major CII processing/presentation defect in HII.

T-helper functions in lines of mice selected for high or low antibody production (Selection III): modulation by anti-CD4+ monoclonal antibody.

T-helper function was evaluated in mice genetically selected for high (H) or low (L) antibody (Ab) responsiveness to Salmonella flagellar antigen (Ag) (Selection III). In this Selection as opposed to what was demonstrated in Selections I, II and IVA, the interline difference was not proven to be based upon the modification of Ag processing and presentation at macrophage level. CD4+/CD8+ lymph node ratio is similar in HIII and LIII mice, both lines being equally susceptible to in vivo depletion of CD4+ T cells by GK 1-5 monoclonal antibody (mAb) treatment. Nevertheless, the Ab responsiveness of the two lines was differently modulated by GK 1-5 mAb: the inhibition of Ab responses to various Ag required lower mAb doses and was long lasting in LIII as compared to the transient effect of higher mAb doses observed in HIII. LIII mice were also refractory to Salmonella-induced reversion of GK 1-5 mAb inhibition. Moreover, in vitro specific I proliferation was constantly lower in LIII, though its IL-2 production was unexpectedly similar to that of HIII T cells. Results of in vivo and in vitro experiments are thus consistent with a defective response of T-helper cells to immunogenic challenge in LIII mice.

Isotypic distribution of antibody responses in lines of mice selected for high or low immunoresponsiveness.

1. The isotype distribution of antibody (Ab) responses to Salmonella antigens (Ag) was investigated in high (H) and low (L) Ab responder lines of mice from Selections III and IV carried out for responsiveness to flagellar (f) and somatic (s) Ag, respectively. 2. Primary immunization resulted in higher Ab titers of all isotypes in response to both Ag in H mice from both selections and was confirmed after booster injections. The interline difference (H-L) in response to the distinct isotypes ranged from 3.0 to 7.0 log2 to Ag f in Selection III and from 2.0 to 5.1 log2 to Ag s in Selection IV. 3. Comparison of isotype production to 3 Ag in Selections I, II, III and IV demonstrated that: 1) the highest responses in all mice are those against the selection Ag, 2) the isotypic pattern depends on both the Ag injected and the host's genetic constitution, and 3) the presence or lack of a multispecific effect is not due to isotype-restricted regulation.

Salmonella typhimurium infection in high and low antibody responder mice: inverse correlation between antibody responsiveness and resistance to infection.

Susceptibility to Salmonella typhimurium infection was compared in H (high Ab responder) and L (low Ab responder) mice obtained by several selective breeding experiments (Selections I, II, III, IV and IV A). H mice were always much more susceptible to infection than their L mice counterparts within a continuous LD 50 variation range. In three of the selections (I, II and IV A) the low responsiveness character is known to result mainly from rapid Ag degradation in L mice macrophages. It was hypothesized that resistance to multiplication of intracellular pathogens could be related to an increased catabolic activity towards Ag. This was actually demonstrated, in F2 segregant hybrids of selection IV A, by the significant inverse correlation between capacity for Ab production and resistance to infection.

Polygenic control of quantitative antibody responsiveness: restrictions of the multispecific effect related to the selection antigen.

Among the differences observed between the various high (H) and low (L) antibody responder lines of mice resulting from distinct bidirectional selective breedings, one of the most puzzling is the variation in the "multispecific effect," i.e., in the modification of antibody responses to antigens unrelated to those used during the selection. The best examples are the H and L lines of selection IV, selected on the basis of responses to somatic antigen of Salmonella which do not differ in their antibody responses to sheep erythrocytes (SE). However, a wide range of variability is observed in the responses of (HIV X LIV)F2 hybrids to this antigen, and it was therefore hypothesized that distinct groups of genes might regulate antibody responses to SE and the somatic antigen. Indeed, a new selection (IV-A) for anti-SE responsiveness started from these (HIV X LIV)F2 successfully produced a high and a low anti-SE responder line. The results of selection IV-A and the variance analysis of (HIV-A X LIV-A)F2 hybrids are reported. They are roughly similar to those in selection I, also carried out for anti-SE responsiveness. In vivo attempts to identify the major regulatory mechanism which contributes to the interline difference indicate that the efficiency of macrophage accessory function has been modified in selection IV-A, as was observed in selection I, whereas this function did not differ in HIV and LIV lines. Probably in relation to the involvement of macrophage function there is a notable increase of the multispecific effect in selection IV-A when compared with selection IV. The results of selection IV-A demonstrate that responsiveness to heterologous erythrocytes and to somatic antigen of Salmonella are under separate polygenic control operating through distinct regulatory mechanisms. The choice of the selection antigen and immunization procedure is of major importance for defining the gene interaction operating in each selective breeding experiment and the extent of its multispecific effect.

Multigenic control of specific and non-specific immunity in mice a review

Anti-infectious immunity is the main component in host-resistance, and the basis of vaccination efficacy. The lines of mice selected for high or low (H or L) immune responsiveness, in which opposite extreme potentialities have been clearly defined, offer an appropriate model for investigating the genetic factors of resistance to infections. In fact, post-immunization antibody titers happen to be more than 200 times higher in H than in L antibody responder mice, whatever the antigen specificity. This is owing to the additive effect of genes located at about 10 loci, one of which is the MHC. Innate resistance to intracellular pathogens is higher in L than in H mice, owing to the differences in the macrophages activity within the two lines: antigen catabolism is faster in L macrophages, which also prove more efficient in controlling early bacterial growth. The fact that these two functions are under a common genetic regulation has recently been brought to light. As was expected, H mice have a stronger innate and acquired resistance to all the infections that can be cleared by means of antibody production. Differences between H and L mice for innate and acquired immunity are quantitative, that is: time- and dose-dependent. The alternative advantages of H or L phenotype are discussed, as regards efficacy of the immune system in natural populations, and may stand for the persistence of polymorphism at loci endowed with high selective value.

Genetics of antibody responsiveness to bovine serum albumin and rabbit gamma-globulin. I. Genetic analysis of high and low responder lines of mice produced by selective breeding.

The selective breeding for antibody production against bovine serum albumin (BSA) and rabbit gamma-globulin (RGG) induced a large modification in responsiveness in the high (Hv) and low (Lv) responder lines at selection limit. The total response to selection (RT) was 9.0 log2 for BSA and 8.4 log2 for RGG. This gives an interline difference of 500-fold and 337-fold respectively in terms of passive agglutinin titres. For BSA responsiveness, there is, in F1 interline hybrids, an incomplete dominance effect of the low character (-0.41) and a marked maternal effect. Complete dominance effect of high character (1.08) without any maternal effect is observed for responses to RGG. The phenotypical variability of BSA responses in F2 segregants is due 60% to genetic factors and 40% to environmental effects. Such a distribution cannot be achieved for RGG responsiveness. Both responses to BSA and RGG are controlled by the additive effect of several independent loci (polygenic regulation). One of these genes is linked with the H-2 locus. The H-2 linked gene accounts for 29% of the total interline difference for response to BSA and only 11% for response to RGG. Experiments carried out to measure the reciprocal nonspecific effect of BSA and RGG responses failed to give clear-cut results. This important phenomenon will be the subject of the companion article.

Genetics of antibody responsiveness to bovine serum albumin and rabbit gamma-globulin. II. Evidence for a partially common genetic regulation of response to bovine serum albumin and rabbit gamma-globulin.

In order to measure the reciprocal nonspecific effect of the genetic regulation of antibody responsiveness to bovine serum albumin (BSA) and rabbit gamma-globulin (RGG), two independent bidirectional selective breedings for responses to these two antigens were carried out: selection V/BSA and selection V/RGG respectively. The total interline separation at selection limit (RT) was 5.3 log2 for selection V/BSA and 2.6 log2 for selection V/RGG. The sum of these two values (7.9 log2) is similar to the RT in selection V carried out by alternating these two antigens in consecutive generations. In selection V/BSA, the nonspecific effect for responsiveness to RGG was 72%. In selection V/RGG, the nonspecific effect for BSA responsiveness was 135%. The F1 hybrids between homologous lines of selection V/BSA and selection V/RGG presented a larger difference in antibody response to both antigens than their parental lines. This demonstrates an additive effect of the loci controlling the two responses.

Independent polygenic regulation of quantitative antibody responsiveness and expression of delayed-type hypersensitivity (DTH).

The intensity of delayed-type hypersensitivity (DTH) expression measured by footpad swelling was established in high and low antibody responder lines of mice produced by bidirectional selective breeding for antibody responsiveness to different antigens. These lines of mice presented a very large difference in antibody response to the antigens used in each selective breeding (selection Ags) and to several other unrelated Ags (nonspecific effect). The intensity of DTH reactivity to selection Ags and to unrelated Ags differed in the various lines investigated, but the intensity of DTH reactions was not correlated with antibody responsiveness. The results of the present article demonstrated that the expression of DTH reactivity and antibody responsiveness to the same antigens are polygenic characters subject to independent quantitative regulation.