Histocompatibility-2 system in wild mice

Ninety-six wild mice trapped at 13 localities in the state of Texas were tested in the dye-exclusion cytotoxic test with a battery of 49 oligospecific H-2 antisera. The antisera detected 36 class I (K and D) and 10 class II (Ia) antigens. The phenotypic frequencies of private class I antigens ranged from 1 to 20%, the majority of them being in the range between 1 and 5%. At least some of the higher frequencies resulted from the presence of more than one antibody in the typing reagents, and from other factors complicating the typing. We estimate that the frequencies of most of the class I alleles among Texas wild mice are 1% or less. This estimate leads to the prediction that at least 200 alleles exist in Texas mice at theH-2K locus, and another 200 alleles exist at theH-2D locus. Frequencies of most of the class I public antigens were in excess of 20%. In the sample of 96 mice, 46 different phenotypic combinations of private class I antigens were found, and the frequency of blanks (mice unreactive with any of the antibodies to private class I antigens) was 27%. The frequencies of private class II antigens ranged from 5 to 15%. Some of the public class II antigens, in particular those controlled by theE region, occurred with frequencies of 80% or higher. The class II antigens were found in 26 phenotypic combinations. No striking linkage disequilibrium was found either between K and D antigens, or between class I and class II antigens. The polymorphism of theK, A, andD region appears to be higher than that of the corresponding regions of the human or rat major histocompatibility complex. The polymorphism of theE region is significantly lower than that of theA, K, andD regions. The polymorphism of theA region is extensive.     

Isolation and analysis of H-2 and Ia alloantigens from wild mouse strains.

Immunoprecipitation and SDS-PAGE analysis were used to examine H-2 and Ia antigens from mouse strains with wild-derived MHC haplotypes. Antisera raised against the wild-derived strains contained anti-H-2 and anti-Ia antibodies which precipitated antigen molecules readily distinguishable by a single assay procedure. The antibodies to wild strains were cross-reactive with standard laboratory haplotypes. Evidence supporting the similar genetic organization of wild and standard haplotypes was found, including isolation of separate H-2K and H-2D molecules from a wild-derived strain, and isolation of two separate Ia molecules from a wild-derived strain.     

Histocompatibility-2 system in wild mice. VI. Histogenetic analysis of sixteen B10.W congenic lines.

Sixteen B10.W congenic lines carrying wild derived H-2 haplotypes on C57BL/10Sn or B10 background were typed by the allogeneic cell-mediated lymphocytotoxicity (CML) assay; in addition, selected lines were also typed by the TNP-CML assay and by skin grafting. The analysis revealed similarity or identity of two strain pairs: SNA57 (H-2w21) ssems to carry a similar haplotype as B10.SM (H-2v), and STA10 and STA12 seem to share the same H-2K and H-2D alleles. All other B10.W strains were different from each other and from B10 congenic lines carrying inbred-derived H-2 haplotypes. These results agree with the results of the serologic typing with two exceptions: the KPA42, KPA132, and SNA57 lines, which were serologically indistinguishable from each other and from B10.SM, were distinguished by histogenetic typing. The presence among wild mice of a haplotype (H-2u21) that appears to be very similar to a haplotype (H-2v) carried by an inbred strain (B10.SM) has some interesting implications for considerations of H-2 gene mutability. The finding that haplotypes derived from different localities are different provides further evidence that the H-2 polymorphism is extensive, indeed.     

Ia antigens: markers of specific T suppressors immune to H-2 complex antigens

Two antisera to Ia antigens, products of the H-2 complex I-Cd and I-JkEk subregions, respectively, have been obtained by immunisation of the F1 hybrids of recombinant strains of mice. These antisera are shown to display the 50 per cent cytotoxic effect in vitro in the presence of complement upon lymphocyte populations immune to the H-2 complex antigens and enriched for specific suppressor T cells (SSC) by fractionation on the monolayer of target cells. The specificity of anti-Ia cytotoxins is shown by the cross antibody absorption with T- and B-cells of mice originated from the recombinant H-2 haplotypes and bearing either particular I-Cd, I-Jk and I-Ek antigens, or their combinations. Anti-I-Cd cytotoxins are found to react with both B and T cells at a different rate, and the anti-I-JkEk serum contains two antibody types directed to I-Ek and I-Jk products, respectively, the latter being able to react preferently with T cells. Although both antisera do inactivate the in vitro SSC function in the presence of complement at a similar degree, the inactivating action of the anti-I-Cd serum, but not that of the anti-I-JkEk serum, occurs without complement. SSC are established to bear both Ia-antigens, I-J and I-C on the same cell, as demonstrated by the cross antibody absorption and variation of the H-2 origin of SSC. These two markers are suggested to function differently in the SSC immune to the H-2 antigens and the I-C antigen expression on the SSC surface is presumed to be required for their interaction with the inhibited responder T cells proliferating in MLC.     

The histocompatibility-2 system in wild mice. XI. Ss and Slp properties of wild-derived H-2 haplotypes

In this study, 14 B10.W lines were examined for genetic traits associated with the Ss protein and Slp alloantigen. Three B10.W lines were found to possess low levels of serum Slp alloantigen. Of these three Slp-positive lines, expression of the alloantigen was sex-limited in two lines (B10.LIB55 and B10.STA12) and constitutive, i.e., found in both sexes, in the remaining line (B10.KPB128). Lines which were found to be Slp-negative were also tested for IA-controlled immune response to Slp alloimmunization. The finding that one of these Slp-negative lines produced no detectable anti-Slp indicates that nonresponder alleles exist in wild populations. Further, the discovery of an unexpected immune response to Slp in F1 hybrids involving lines B10.LIB55 and B10.STA12 suggests the existence of a variant form of Slp alloantigen in these lines.     

Enhanced in vitro cytotoxic anti H-2 responses in the presence of Ia antigens

Because surface Ig and Ia antigens cap independently, A.TH anti-A.TL serum combined with the indirect immunfluorescence technique could be used to test defined murine cell populations ofH-2 ^k haplotype for the presence of Ia antigens. Mitogen induced T- and B-cell derived blast cells, purified by velocity sedimentation at 1g, were tested for the expression of Ia^k antigens and then used both as stimulator cells and as target cells, in primary and secondary in vitro cytotoxic allograft responses. Fibroblasts, cortisone-resistant thymocytes, and nylon column purified splenic T cells were also included in these tests. Ia antigens were detected on 100% of LPS-induced blast cells, on 20%–30% of ConA-induced blast cells (100%Θ Thy-1 or antigen positive), but only to 5%–10% on PHA-blasts (100% Thy-1 antigen positive). Fibroblasts and nylon column purified splenic T cells were essentially Ia negative. Ia-positive allogeneic stimulator cells induced a far stronger in vitro cytotoxic T-cell response compared to Ia-negative stimulator cells; that is, there was a positive correlation between the expression of Ia antigens on the stimulator cells and the magnitude of cytotoxicity induced. This correlation was restricted to primary allograft responses. Ia antigens could not be detected as a target for killing in the cytotoxic effector phase, using both different target cells as well as the approach of “PHA dependent lysis” for detecting cytotoxic T lymphocytes.     

A study of histocompatibility-2 antigens in wild mice from Santiago,Chile

LyM-1 is the provisional designation given to a system of murine cell-surface alloantigens which are controlled by genes closely linked to those of theMls system. Formal genetic analysis has failed to disclose separation of genes determiningMls and LyM-1 antigens, but studies of the distribution of these antigens among inbred strains shows that the LyM-1 polymorphism is not primarily responsible for the MLR activity associated with Mls differences, and suggests that LyM-1 and Mls substances are products of genes at closely linked, but probably separate loci. Absorption analysis shows that strains whose cells react with anti-LyM-1.2 can be divided into at least two classes on the basis of the efficiency with which their cells remove anti-LyM-1.2 antibodies. This provides evidence for the existence of two LyM-1 alleles in addition to the one(s) possessed by nonreactive mouse strains.     

Macrophage Ia antigens. I. macrophage populations differ in their expression of Ia antigens

By indirect immunofluorescence and microcytotoxicity it was demonstrated that different populations of murine macrophages bear different amounts of Ia antigens on their membranes. At least three subpopulations could be distinguished: those that lack Ia antigens and predominate in peritoneal exudate; cells bearing I-A antigens that are the majority of splenic macrophages and a minor population in the peritoneum; and cells bearing I-C antigens that are a minor population in both spleen and peritoneum. Internal radioisotope labeling studies confirmed that the I region molecules are synthesized by the macrophages. It is suggested that these different macrophage subpopulations may play distinct roles in the immune response.     

The histocompatibility-2 system in wild mice

In this study, 14 B10.W lines were examined for genetic traits associated with the Ss protein and Slp alloantigen. Three B10.W lines were found to possess low levels of serum Slp alloantigen. Of these three Slp-positive lines, expression of the alloantigen was sex-limited in two lines (B10.LIB55 and B10.STA12) and constitutive, i.e., found in both sexes, in the remaining line (B10.KPB128). Lines which were found to be Slp-negative were also tested forIA-controlled immune response to Slp alloimmunization. The finding that one of these Slp-negative lines produced no detectable anti-Slp indicates that non-responder alleles exist in wild populations. Further, the discovery of an unexpected immune response to Slp in F₁ hybrids involving lines B10.LIB55 and B10.STA12 suggests the existence of a variant form of Slp alloantigen in these lines.     

The histocompatibility-2 system in wild mice

The I-region gene products of 29 wild-derivedH-2 haplotypes on a B10 background (B10.W congenic lines) were typed with alloantisera which detect 17 inbred I-region antigens. Five new I-region antigens were defined by expanding the inbred line panel ofH-2 haplotypes to includeH-2^u, H-2^v, andH-2^j. Based on serological analyses of the inbred and B10.W lines, the polymorphism of theIA gene (or genes) is estimated to be at a minimum of 15 alleles and theIE gene (or genes) at a minimum of 4 alleles. These results indicate that theIA subregion is more polymorphic than theIE subregion. By combining the I-region typing data with theH-2K andH-2D region typing data reported previously, a total of 11 new natural recombinants of inbredH-2 alleles were detected among the B10.W lines.     

Isolation of twelve monoclonal antibodies against Ia and H-2 antigens. Serological characterization and reactivity with B and T lymphocytes

The cell hybridization technique was used for the production of 12 monoclonal antibodies against H-2K^k, H-2D^b, I-A^k and I-E^k antigens. The strain distribution pattern indicated that three antibodies reacted with new H-2 and Ia determinants, respectively, while the majority of determinants defined by the monoclonal antibodies showed good correlation with H-2 and Ia determinants described by conventional alloantisera.Monoclonal Ia antibodies showed strong reactivity with about 90% of surface IgM positive B cells, but not with T cells. In double fluorescence studies, both I-A and I-E determinants were always found to be coexpressed on the same B cells. When the high sensitivity of the fluorescence activated cell sorter was utilized, about 30 to 40% of purified lymph node T cells were found to carry both I-A and I-E antigens, although in a much lower density than B cells. In conclusion, monoclonal Ia antibodies appear to display the same serological and cellular reactivity pattern as do conventional antisera.