High Responder Phenotype Research Articles

Monoclonal antibody characterization of a unique immune response control locus between H-2S and D.

The primary in vitro antibody response to the type 2 antigen, trinitrophenyl (TNP)-Ficoll, is controlled by two complementing loci in the H-2 region of the mouse major histocompatibility complex (MHC). High responder alleles at both loci are necessary for a high responder phenotype. Previous studies mapped one locus of control to the I-A subregion. In this report we demonstrate by recombinant analysis that the second locus of control is located between the H-2S and D regions. A comparison of responses in the B10.BAR6, B10.BAR10, and B10.BAR11 strains defined a locus controlling the response to TNP-Ficoll in a single haplotype, bounded on the left by the crossover event in the B10.BAR10 and on the right by the crossover event in the B10.BAR6 strain. A monoclonal antibody directed against this right-hand region of control has been produced (48.21.7) that blocks the response to TNP-Ficoll at the level of the antigen-presenting cell. The monoclonal antibody 48-21.7 is specific for the high responder b allele at the right-hand locus and did not inhibit responses to other protein antigens tested. The immune response to TNP-Ficoll was not inhibited by monoclonal antibodies that react with H-2Db or Qa-2 specificities, suggesting that the TNP-Ficoll response is controlled by a unique locus located between H-2S and D. Finally, 48-21.7 recognizes and precipitates a unique product of approximately 40,000 mol wt that is distinct from the H-2D region product recognized by the monoclonal antibody B22/249.

Changes in the Ir gene controlled response phenotypes in mice of the Ap and Ak family of alleles expressing naturally occurring variant molecules.

Several wild-derived H-2 haplotype mice were recently shown by serology and tryptic peptide fingerprinting to express I-region A molecules closely related to the Ap and Ak molecules of the laboratory strains. To determine if such naturally occurring minor structural variations in the A molecule alter Ir gene-controlled responsiveness, we examined the immune responses of these strains after primary immunizations to three synthetic polypeptide antigens: G60Phe40, GLPhe9, and GAT10. Inbred strains carrying the Ap (or Aq) allele are known to be high responders to G60Phe40 and GLPhe9 but low responders to GAT10. Strains expressing the Ak allele are classified as low responders to G60Phe40 and GLPhe9, but high responders to GAT10. Of seven strains examined belonging to the Ap family, one (B10.CAS2) failed to respond to either G60Phe40 or GLPhe9 as measured by antibody production and T cell proliferation. In addition, two strains (B10.STC90 and W12A) of the Ak family were found to be of responder phenotype to GLPhe9. Both GLPhe9 responses resulted from the introduction of new E beta genes into the I region through naturally occurring intergenic recombination between A beta A alpha and E beta. All strains of mice in the Ak family proved to be of the high responder phenotype in their responses toward GAT10. These results contrast strongly with known patterns of alloreactivity against the variant Ap and Ak molecules.

Arsonate-specific murine T cell clones. I. Genetic control and antigen specificity.

The antigen-induced proliferative response of lymph node cells (LNC) from mice sensitized to the monofunctional antigen L-tyrosine-p-azobenzenearsonate (ABA-Tyr) was used to monitor genetic control. All strains tested mounted significant responses, but those that were H-2(b) at both the I-A and I-E loci [B10., B6., B10.A(18R), A.BY, and C3H.SW] gave consistently weaker responses than other haplotypes. The F(1) progeny of matings between high and low responder phenotype parents (DBA/2 and B6, respectively) were high responders, establishing the dominance of the responder trait. Proliferative responses of LNC to ABA-Tyr were blocked by the appropriate anti-Ia monoclonal reagents. For example, B10.A(4R) LNCI (I-A(k), I-E(b)) were blocked by anti-I-A(k), whereas B10.A(3R) LNC (I-A(b), I-E(k)) were blocked by anti-I-E(k). Long-term cultures of T cell lines specifically reactive to ABA-Tyr were established from LNC of A/J mice immunized with ABA-Tyr and were cloned by limiting dilution. The proliferative responses to ABA-Tyr of 14 out of 15 clones tested were I-A restricted on the basis of activation by antigen-presenting cells from appropriate recombinant strains and the blocking activity of the monoclonal anti-Ia antibodies. The response of the other clone was I-E restricted. The fine antigen specificity of the clones was studied using structural analogs of the homologous antigen to induce proliferation. The clones could be divided into three types with respect to responsiveness to ABA-histidine (ABA-His). One group responded about equally well to ABA-His and ABA-Tyr. A second set responded less strongly to ABA-His than to ABA-Tyr, while the third showed no response above background to ABA- His. In all instances, the ABA-His-responding clones discriminated exquisitely between the 2-azo and 4-azo histidine isomers, responding only to the 4-azo compound. These T cell clones provide extremely useful tools for studies of T cell specificity, antigen recognition and lymphoid cell interaction systems.

"Allergic breakthrough" after antigen sensitization: height of IgE synthesis is temporally related to diurnal variation in endogenous steroid production.

Mice of the SJL strain normally display the low IgE responder phenotype. In the course of extensive studies on IgE responses with this strain of mice, it because apparent that the magnitudes of IgE antibody synthesis by SJL mice varied after antigen sensitization either alone or in conjunction with exposure to optimal enhancing doses of whole body irradiation. Analysis of the basis for this variability revealed that SJL respond to antigen sensitization by displaying cyclical IgE response curves determined by the time of day the sensitization is carried out. Moreover, when these low responders are converted to the high responder phenotype by low dose irradiation, the time of irradiation determines the magnitude of the IgE response obtained, whereas the time of antigen challenge determines the potential for generating a good response. The diurnal curve of IgE responses in both unirradiated and irradiation-enhanced low responder mice appears to be determined by normal variations in levels of endogenous corticosteroids, as indicated by the correlation between the normal diurnal curve of endogenous steroid production with the cyclical pattern of IgE responses. This apparent effect of endogenous steroid production on IgE antibody synthesis is supported by experiments demonstrating that the normal IgE pattern in both unirradiated and irradiated mice can be perturbed by administration of cortisone at the proper time of day. These results indicate that IgE antibody synthesis displays unique sensitivity to fluctuations in plasma steroid levels and may have significant implications for the role of steroids in the pathogenesis and management of allergic diseases.

Cell-mediated anti-tumor response in the RLmale 1 system. II. Control of the T killer cells by an H-2-linked Ir gene interfering with non-H-2 factors.

The generation of cytolytic T lymphocytes (CTL) during the immune response directed against the syngeneic X-ray-induced BALB/c RLmale 1 leukemia has been described previously: T killer cells react with a tumor antigen of RLmale 1 cells, and the H-2Dd molecules of the target cells play some role in the interaction. The study of anti-RLmale 1 responses of [(C57BL/6 x BALB/c) x BALB/c] mice shows that the major histocompatibility complex controls the anti-RLmale 1 CTL reaction at a second level, through an Ir gene, with dominant responsiveness, probably mapping to the right of I-B and controlling the high or low-CTL responder phenotype. Another non-H-2 associated gene interferes with the H-2 linked Ir gene, a responder allele at the non-H-2 locus, being necessary for the expression of the high-responder phenotype at the Ir gene locus.

Regulation of IgE antibody production by serum molecules. III. Induction of suppressive activity by allogeneic lymphoid cell interactions and suppression of IgE synthesis by the allogeneic effect.

Antibody responses of the IgE class are, like other immunoglobulin classes, regulated by a finely-tuned network of complex cellular and molecular interactions (1). Previous studies conducted in our laboratory (2, 3) have provided new insights into the differences in control mechanisms that result in individuals manifesting either the high (allergic) or low (nonallergic) IgE responder phenotype. These experiments have shown that certain manipulations (i.e. low dose X-irradiation) convert normally low responder mice to high IgE responders, apparently by diminishing a suppressor T-cell mechanism which normally dampens, rather selectively, IgE antibody production in such individuals. Similar findings have been made by Watanabe et al. (4). Recently, we have been studying the types of manipulations that could reverse the high IgE responsive state back to a low one. These studies (2, 3, 5, 6) have demonstrated that the high IgE responses induced in low responder mice can be substantially diminished, and even abolished, by passively transfusing serum or ascetic fluid from donor mice previously inoculated with mycobacterial-containing complete Freund's adjuvant (CFA). Because the suppressive activity of CFA-immune serum or ascitic fluid is so highly selective for IgE antibody production, we have recently termed these serum substances suppressive factors of allergy (SFA) (2, 3). The present study was undertaken to determine whether alternative means, particularly those that avoid administration of CFA, could be devised for the induction of SFA. Herein, we report the effectiveness of allogeneic lymphoid cell interactions in inducing SFA, both in vivo and in vitro, as well as the potent suppressive effects of an in vivo allogeneic effect on irradiation enhanced IgE antibody production in low responder mice.

Dence for a gene in rats affecting lymphocyte responsiveness to PHA

A gene controlling high responsiveness of lymphocytes to in vitro stimulation by PHA was transferred from the Lewis strain of rats to the BN background by ten generations of backcrossing. The high-responder phenotype was initially defined on the basis of incorporation of(3)H-thymidine, but we show that this trait also involves higher levels of mitotic activity than are observed with low responder lymphocytes. This gene is not closely linked to any histocompatibility locus which could be detected by skin grafting, and it does not appear to affect the proportion of T lymphocytes.

Regulation of IgE Antibody Production by Serum Molecules

Abstract IgE antibody production in mice of high and low IgE responder phenotypes, respectively, can be appreciably enhanced in magnitude after low dose whole body X-irradiation. Such enhanced responses, as well as adoptive secondary IgE responses, can be markedly suppressed by passive transfer of CFA-immune serum in low responder strains, but not in high responder strains. The studies presented here demonstrate that the suppressive activity of CFA-immune serum on IgE antibody production is strain specific. This is true even in reciprocal combinations of low IgE responder SJL and C57BL/6 mice, in which it was shown that serum capable of suppressing mice of the isologous strain was ineffective in diminishing IgE antibody production in the other low responder strain. Absence of suppressive activity in CFA-immune sera obtained from H-2 congenic donors which differed in their H-2 haplotypes while sharing many similarities in the background genome and, conversely, effective suppressive activity of H-2 congenic donor sera when H-2-identities between donor and recipient mice existed, strongly suggested a role, at least in part, of H-2 genes in dictating the strain specificity of such suppressive activity. Additional experiments provided evidence for a possible role of macrophages in catabolism of the active molecules in CFA-immune sera. These observations, together with those presented in the preceding paper, may provide valuable insight toward successful development of appropriate manipulations that could ultimately convert high IgE responder individuals into low responders.

Regulation of IgE Antibody Production by Serum Molecules

Abstract Exposure of mice to low doses of ionizing x-irradiation at or near the time of primary immunization with 2,4-dinitrophenyl (DNP)-Ascaris suum extract (ASC) results in substantial enhancement of IgE anti-DNP antibody responses; the IgG antibody responses of such mice do not increase after such manipulations. This selective enhancement of IgE antibody production occurs in mice of both high and low IgE responder phenotype, although the extent of enhancement compared to unmanipulated control animals is more striking in low IgE responder mice. The studies presented here demonstrate that the irradiation-enhanced IgE antibody responses of low responder SJL and C57BL/6 mice as well as of intermediate responder AKR mice can be effectively suppressed by passive transfer of CFA-immune serum obtained from isologous donor mice. Moreover, adoptive secondary IgE antibody responses in SJL recipients of primed syngeneic spleen cells can be totally abolished by passive transfer of CFA-immune serum or ascitic fluid from CFA-immune mice. The suppressive activity of CFA-immune serum can be diminished or eliminated by exposure of CFA-primed donor mice to low dose x-irradiation at an appropriate point during the priming regimen, after a single inoculation of CFA, and before collection of serum. Low dose x-irradiation was not effective in eliminating suppressive activity of CFA-induced ascites fluid obtained from donor mice inoculated repeatedly with CFA. In contrast to the capacity of CFA-immune serum from isologous donors to suppress irradiation-enhanced IgE responses of low responder mice, similar sera or ascites fluids were ineffective in suppressing irradiation-enhanced responses of high responder BALB/c or (SJL × BALB/c)F1 hybrid mice. This was true irrespective of whether such sera or ascites were obtained from isologous donors or from donor mice of the low IgE responder phenotype. The implications of these findings with respect to clarifying some of the previously confusing observations concerning 1) the effects of adjuvant administration on IgE antibody production, and 2) differences in the capacities of different mouse strains to produce IgE antibodies in high vs low quantities are discussed.

Genetic analysis of lymphocyte activation by lipopolysaccharide Endotoxin.

A genetic study of in vitro lymphocyte blastogenesis by the B-cell mitogen lipopolysaccharide endotoxin has been performed, using both low-responder C3H/HeJ mice and high-responder CBA/J mice. The crossbreeding of these strains produced F1 progeny that were intermediate in responsiveness. Responder types from the backcross of the F1 to the C3H/HeJ strain segregated into intermediate- and low-responder phenotypes, whereas intermediate- and high-responder phenotypes were produced from the backcross of the F1 to the CBA/J strain. The F2 generation consisted of all three responder phenotypes in frequencies that fit the classical Mendelian ratio of 1:2:1, indicating that the mitogenic response to lipopolysaccharide endotoxin in mice is most likely governed by a pair of autosomal co-dominant genes.