Gregory W. Siskind

Immunological studies of aging: Normal B-cell repertoire in aged mice: Studies at a clonal level

As previously reported, old mice produce lower avidity plaque-forming cells (PFC) after immunization with 2,4,6-trinitrophenyl-Ficoll (TNP-F) than do young mice. However, if spleen cells from TNP-F-immunized old mice are incubated with hapten to elute auto-anti-idiotype antibody then high avidity PFC, comparable to those in young mice, are detected. To further evaluate the effect of age on the B-cell repertoire anti-2,4,6-trinitrophenyl-bovine gamma globulin (TNP-BGG) hybridomas were prepared from young (6 to 8 weeks old) and old (18 to 24 months old) mice which had been primed and boosted with TNP-BGG. The monoclonal antibodies (MoAbs) were TNP-specific. Spleens from old and young mice were comparable with respect to the incidence of immunoglobulin-secreting hybridomas obtained, the incidence of TNP-BGG-specific hybridomas obtained, and the isotype distribution of the anti-TNP-BGG hybridomas. The avidities for TNP-BGG of the IgG1 anti-TNP-BGG MoAbs obtained from old and young donors were also comparable. The overall results thus suggest that old and young mice have similar B-cell repertoires and that differences in the antibodies produced are due to regulatory influences.

Distribution of antibody affinities: Technique of measurement

Abstract A computational method has been developed to describe, by means of a histogram, the distribution of affinities in an antibody sample. It was shown that the distribution of affinities was generally not symmetrical and that the binding data could not be adequately, approximated by a Gaussian or Sipsian distribution. Early (day 7) after immunization there is an approximately symmetrical distribution of affinities. This is followed by a progressive skewing of the population towards the high affinity end of the distribution so that by day 90 there is a markedly asymmetric distribution consisting predominantly of high affinity subpopulations of relatively restricted heterogeneity. Low affinity antibodies persist in approximately constant amounts from day 7 to 1 yr after immunization. The methods for computing affinity distributions from binding data are described. The procedure is justified by its ability to adequately approximate the distribution of affinities in a series of hypothetical populations for which theoretical binding was calculated.

Lower antibody response to tetanus toxoid associated with higher auto-anti-idiotypic antibody in old compared with young humans

The production of anti‐tetanus toxoid (TT) antibody and anti‐anti‐TT (auto‐anti‐Id) antibody has been measured in three young and three old persons. Both antibodies were measured using ELISA. We found that: (i) the anti‐TT response of old subjects was lower than that of young subjects, (ii) scrum auto‐anti‐Id antibody concentration was higher in old compared with young humans before boosting with TT, and (iii) anti‐TT antibodies from different humans shared Id cross‐reactivity when tested with a rabbit anti‐Id antibody. Thus, the serum anti‐TT antibody response to boosting was inversely correlated with the serum level of auto‐anti‐Id. This conclusion is consistent with the view that the higher level of auto‐anti‐antibody in older subjects contributes to their impaired antibody response to TT.

Peripheral T Cells Select the B‐Cell Repertoire in Old Mice

These studies have shown that the alterations in the repertoire of antibody produced by old mice is not due to an intrinsic defect in the bone marrow or in the B-lymphocyte population arising from the bone marrow but rather to a selective downregulation by auto-anti-idiotypic antibody and idiotype-anti-idiotype interactions, shifting the idiotype distribution in the peripheral B-cell population. Thus, the clonal distributions of B cells generated by bone marrow of old and young mice are very comparable. The age-related differences in antibodies expressed by young and old mice are, to a great extent, determined by the activity of a peripheral regulatory immune network. This immune cellular network operates prior to exposure to antigen, presumably on the basis of an idiotype-anti-idiotype network between T and B lymphocytes. After exposure to antigen, a network of idiotype-anti-idiotype antibody interactions also contributes to differences in the immune responses of old and young mice to foreign antigens. If the expressed repertoire of antibody reflects down-regulation of auto-anti-idiotypic antibody, comparable repertoires of B-cell clones would be expected to be recovered from old and young mice if B cells from old mice were rescued from selective peripheral downregulatory influences active in old mice. Support for this hypothesis has been obtained by generating B-cell hybridomas from young and old mice immunized with TNP bovine gamme globulin (Marcenario et al. 1989). The same number of anti-TNP hybridomas and a comparable number of IgG and high-affinity antibody-producing clones were recovered from the spleens of young and old mice. Thus, the actual B-cell clonal repertoires of young and old mice appear to be similar although the expressed repertoires of antibody-producing lymphocytes from old and young mice are very different. This conclusion has considerable impact on strategies that could be employed to reverse the senescence of humoral immunity. Strategies to counter downregulatory influences which constrain the expression of the B-cell population should be more effective than attempts to reconstitute the repertoire of B lymphocytes in aged individuals. Finally, the mechanisms underlying these age-associated shifts in the expressed humoral antibody response can be attributed to life-long interactions with self and foreign antigens. The overall shift may be described as a decreased reactivity to foreign antigens and a complementary increase in reactivity with self antigens.(ABSTRACT TRUNCATED AT 400 WORDS)

Production of auto-anti-idiotypic antibody during the normal immune response: IX. Characteristics of the auto-anti-idiotype antibody and its production

Using hapten-reversible inhibition of plaque formation as an assay for auto-anti-idiotype antibody (anti-Id) and as a means for following idiotype (Id) expression, we have obtained evidence that following immunization with trinitrophenyl (TNP) conjugates (a) there are differences in Id expression in the anti-TNP antibody response to different TNP conjugates although there is some overlap; (b) different strains, although showing some differences in Id expression, tend to produce cross-reactive Ids, thus no obvious allotype linked inheritance of Id expression is observed in this heterogeneous immune response; (c) the auto-anti-Id produced following immunization with TNP-Brucella abortus or TNP-Ficoll tends to be of the IgG2a and IgG2b isotypes.

Determination of antibody avidity at the cellular level by the plaque inhibition technique: effect of valence of the inhibitor.

Inhibition of plaque formation by multivalent and univalent ligands was compared as an assay of avidity of antibody produced by PFC. Multivalent ligands are much more effective as inhibitors and their use tends to impart an appearance of lack of heterogeneity and high avidity to the PFC populations being studied. It is thus probably generally advisable to employ univalent ligands in such studies.

A comparison of the Farr technique with equilibrium dialysis for measurement of antibody concentration and affinity

The Farr technique has been compared with equilibrium dialysis in regard to its ability to measure antibody concentration and affinity. An excellent qualitative agreement between the two techniques was obtained. Quantitatively, the Farr technique tended to underestimate antibody concentration and overestimate antibody affinity. It is likely that these observations are due to the failure of some low affinity antibodies to bind hapten under the conditions of the Farr assay.

Distribution of antibody affinities—II. Fractionation of antibody with respect to its hapten binding affinity☆

Abstract Antibodies from individual sera were fractionated on the basis of their binding affinity by use of a specific immunoadsorbent. The distribution of affinities in the original serum and in each of the series of 10 fractions derived from it was computed. The antibodies were highly heterogeneous and generally not distributed in a symmetrical manner. The distribution of affinities varied with different immunizing conditions. It is clear from the data presented that while values for ‘average affinity’ generally reflect accurately the overall binding properties of an antibody sample, they do not provide a complete picture of the actual distribution of affinities present. It was found that antibody samples exist which are indistinguishable with regard to ‘average affinity’ but differ significantly with respect to the actual distribution of the antibody affinities. Detailed analysis of the binding curves indicated that even animals forming mainly high affinity antibody had considerable amounts of very low affinity antibodies present in their serum.

Tolerance induction by a poorly arthritogenic collagen II can prevent collagen-induced arthritis

Collagen type II (CII)-induced arthritis (CIA) can be induced in 78% of B10.RIII mice (H2r) by intradermal (id) immunization with CII of bovine origin in complete Freunds adjuvant (CFA), whereas immunization with CII of chick origin induces arthritis in less than 5% of these mice. Nevertheless, tolerization of B10.RIII mice with intravenously injected chick CII renders the animals resistant to induction of CIA by immunization with bovine CII. Such tolerization can be achieved either by intravenous injection of 500 micrograms chick CII 1 week prior to immunization with bovine CII in CFA or by such an intravenous injection of chick CII 2 weeks after immunization with bovine CII in CFA. Postimmunization treatment results in a significant decrease in the concentration of antibody to bovine CII. Preimmunization administration of chick CII causes a marked decrease in the antibody reactive with chick CII without a significant effect on the anti-bovine CII antibody concentration. In DBA/1 mice, a strain in which both bovine CII and chick CII can induce a high incidence of the disease, intravenous injection of bovine CII can also prevent arthritis induced by chick CII, even when given 7 or 14 days after immunization. The fact that chick CII as tolerogen is quite effective in preventing arthritis in B10.RIII mice, while as immunogen it is very ineffective in inducing arthritis in this strain, may be interpreted as evidence for interaction between different epitopes on CII in the pathogenesis of CIA.

Effect of age on ease of B-cell tolerance induction☆

Abstract It was shown in intact mice, and in lethally irradiated, thymectomized recipients of peripheral B cells from donors of various ages, that the ease of B-cell tolerance induction with the dinitrophenyl derivative of the copolymer of d -glutamic acid and d -lysine decreases with age. That is, the B-cell population of 6- to 7-month-old BALB/c mice is more resistant to tolerance induction than is the B-cell population of 1 1 2 - to 2-month-old mice and the B-cell population of 10- to 11-month-old mice is highly resistant to tolerance induction. This age-related resistance of the peripheral B-cell population to tolerance induction might, at least in part, account for the increased incidence of autoantibodies in aged animals. However, the increased resistance to tolerance induction appears to occur at an earlier age than the increased incidence of autoantibodies. This suggests that changes in ease of tolerance induction may not be the sole factor responsible for the increase in autoantibodies in aged animals. Evidence is also presented for an age-related decrease in the function of the B-cell population with respect to the response to a T-dependent antigen.