Geoffrey Haughton

Autoantibodies to phosphatidylcholine. The murine antibromelain RBC response.

The observation that murine B-cell populations can contain relatively large numbers of cells that produce IgM with the ability to lyse bromelain-treated mouse erythrocytes (BrMRBC), but not normal untreated MRBC, was made nearly 20 years ago. The major observations regarding the antigen specificity, the cells that produce this IgM, and the immunoglobulin V genes that encode them are summarized in this report. The epitope on BrMRBC that is recognized has been identified as the head group of phosphatidylcholine (PtC); B cells whose IgM has this specificity can be easily identified by their ability to bind fluorescent synthetic liposomes whose membrane contains PtC. The cells producing IgM specific for PtC all derive from the Ly-1 B-cell subset, and they use primarily two VH/VL gene pairs to encode the anti-PtC antibodies. The VH genes used describe two new VH gene families, VH11 and VH12. The genes encoding anti-PtC are unmutated and have characteristics and restricted VDJ constructions. The cells with this specificity, within individual mice, are polyclonal. These criteria are consistent with a primary antigen-driven clonal selection mechanism as the basis for the development of this immune specificity.

The genetic control of the immune response to murine histocompatibility antigens. I. The response to H-4 alloantigens

The genetic control of the immune response to H-4 histocompatibility alloantigens is described. The rejection of H-4.2-incompatible skin grafts is regulated by anH-2-linkedIr gene. Fast responsiveness is determined by a dominant allele at theIrH-4.2 locus. TheH-2b,H-2d, andH-2s haplotypes share the fast response allele;H-2a has the slow response allele. Through the use of intra-H-2 recombinants, we have mapped theIrH-4.2 locus to theI-B subregion of theH-2 complex; theH-2h4,H-215, andH-2t4 haplotypes are fast responder haplotypes. These observations suggest that the strength of non-H-2 histocompatibility antigens is ultimately determined by the antigen-specific recipient responsiveness.

Signaling to a B-cell clone by Ek but not Ak, does not reflect alteration of Ak genes

The mouse B-cell clone, CH12.LX (Iak, Ly-1+, μ+, δ+), can be induced to differentiate and secrete antibody in an antigen-specific, H-2-restricted manner. Induction requires two signals. One must be provided by the binding of specific antigen to the membrane IgM; the other is delivered by the binding of Ek-specific T-cell hybridomas to the Ek molecules of CH12.LX (Bishop and Haughton 1986). Previous studies demonstrated that Ek-specific monoclonal antibodies (mAbs) could substitute for T cells in delivering the second differentiative signal (Bishop and Haughton 1986). Although CH12.LX cells present Ak to Ak-restricted or alloreactive T-helper cells, neither T cells nor mAbs specific for Ak induce differentiation (Bishop and Haughton 1986). However, since the Akspecifc mAbs tested previously were β-chain-specific and the Ia epitope specificity of the T cells used was unknown, it is possible that the differentiative signal delivered to the CH12.LX class 11 molecule is chain-specific. Here we report the effects of ten additional Iak-specific mAbs upon the differentiation of CH12.LX. In addition, a cl)NA library was prepared from CHI 2.LX cells, clones corresponding to the α and β chains of the Ak molecule were isolated, and their nucleotide sequences were determined. Finally, the Ak and Ek molecules of CH12.LX and H-2k spleen cells were compared by two-dimensional gel electrophoresis to examine possible post-translational differences in the Iak molecules of CH12.LX.

Genetics, the immune response and oncogenesis.

The immune surveillance theory states that a major driving force in the evolution of the immune response has been the necessity to recognize and eliminate clones of neoplastic cells which arise either spontaneously or under the influence of external oncogenic agents. Be it true or false, it is a good theory in that a number of testable predictions derive from it. These include the following: (a) All, or at least the vast majority of neoplastic cells bear altered cell-surface components which can be recognized by the immune system of the primary host; (b) Recognition of the altered cell-surface components leads to mobilization of an immune effector mechanism which eliminates, causes reversion to a non-neoplastic state or causes long-term suppression of the neoplastic cells; (c) Immunosuppression adequate to prevent the rejection of transplanted tumors should lead to decreased latency and increased incidence of spontaneous tumors and of induced tumors following a standard oncogenic insult; (d) Inherent immunodeficiency should be associated with a high incidence of malignancy and a high susceptibility to oncogenesis; and (e) Polymorphic immunoregulatory genes (Ir genes) which influence susceptibility to oncogenesis through regulation of response to tumor-assodated transplantation antigens (TATA) might be expected to exist. The best candidates would probably be genes afEecting susceptibility to specific viral oncogenesis shown to be linked either to the major histocompatibility complex (MHC), or to immunoglobulin heavy chain (Ig-H) allotype. Resistance to oncogenesis would be expected to be dominant over susceptibility.

Antigenic cross reactivity between benign prostatic hyperplasia and adenocarcinoma of prostate

Tissue cultures were established from biopsy specimens of adenocarcinoma of the prostate (ACP) and benign prostatic hyperplasia (BPH). Generally, peripheral blood lymphocytes from BPH and ACP patients were cytotoxic to both ACP and BPH cells, but not normal fibroblasts nor cells cultured from other types of malignant tissue. Peripheral blood lymphocytes from normal control patients or from patients with other types of cancer were not cytotoxic to ACP- or BPH-derived cells. These findings are consistent with a cross reactive autoimmune response in ACP and BPH patients, directed against a common antigen(s) present on both ACP and BPH cells.

Specific immunosuppression by minute doses of passive antibody. III. Reversal of suppression by peritoneal exudate cells from immune animals

Abstract Thioglycollate stimulated peritoneal exudate, harvested from mice immunized against sheep erythrocytes and transferred to normal syngeneic recipients, reversed the immunosuppression caused by passively administered anti-sheep cell antibody. Reversal was antigen specific. The degree of reversal was directly related to the number of cells transferred and was independent of the dose of antibody used for immunosuppression. The ability to reverse suppression was a property of cells present in high frequency in peritoneal exudate, but in low frequency in suspensions of lymph node cells. Long lasting reversal of suppression only occurred when the donor cells were histocompatible for the recipients. There was no reversal when peritoneal cells were transferred across an H-2 barrier, but transient reversal was seen when such cells were transferred across multiple non-H-2 barriers.

Isotype Switching in CD5 B Cells

Antibodies are diverse both with respect to antigen specificity and with respect to the biological consequences that follow antigen binding. The former results from the structure of the assembled V, D, and J heavy chain and V and J light chain genes, and the latter depends upon which particular heavy chain constant region gene is expressed. Isotype switching is the process by which mature IgM-expressing B cells, having selected a particular V,-D-J, and V,-J,-C, combination for immunological relevance, rearrange the ZgH locus, C region DNA sequences to express those selected V-region and light-chain combinations on IgG, IgE, or IgA constant-region genes. This process allows the enormous diversity of antigen-combining sites generated by V-D-J recombination, heavy-light chain associations, and somatic hypermutation to be expressed on immunoglobulin (Ig) molecules with a variety of functional capabilities. In this report we will summarize our work with the murine CD5+ B-cell lymphoma CHlZLX, which switches spontaneously in vitro from the expression of IgM to any one of several IgG subclasses or IgA. We began with the observation that variants of CH12,LX, expressing isotypes other than IgM, could be isolated in vitro by fluorescence-activated cell sorting. I We than asked three related questions about this process: (1) Can we develop a method by which the spontaneous isotype-switch frequency from IgM to each of the available isotypes (encoded by C, genes located 3’ of C p ) can be quantified? (2) Do clones of CH12.LX expressing IgG3, IgGl, or IgG2b retain the ability to undergo further isotype switching, and if so, to which isotypes can they switch? and (3) Can we use this method of enumerating switch frequency to determine the effect of T cell-derived interleukins, cytokines, mitogens, antigen, or other factors on (a) isotype-switch frequency and (b) the relative frequency of switching to each of the available isotypes? There have been several studies with conventional splenic B cells that have been interpreted as showing that interleukin-4 (IL-4)2*3 and transforming growth factor-@ (TGF-@r regulate isotype switching by directing IgM-expressing B cells to switch to particular downstream isotypes. Such conclusions are difficult to support when working with a highly heterogeneous population of B cells in a variety of stages of differentiation, because the soluble factor@) may, in fact, be selecting a small subpopulation of B cells precommitted to switching to a particular isotype. We hoped that by working with a clone of in vitro-adapted B cells with defined antigen specificity, we could establish an in vitro model of switching that would enable us to address these and other questions about switch regulation.

Production, Testing, And Utility Of Double Congenic Strains Of Mice

SUMMARY The production and testing of “double congenic” strains of mice are described. BIO. A and B10.D2/o females were mated with B10.129(21M) males, and their Ft offspring were intercrossed. The F2 progeny were selected on the basis of coat color, tumor challenge, skin grafting, and hemagglutination testing for the genotypes H-2a H-4bp/H-2a H-4bp and H-2d lI-4bp//-2d Il-4bp. The resulting strains wore designated B10-//-.T H-4bpWts and B10-//-2* H-4bp/W ts. Skin grafts were transplanted orthotopically from BW-II-2a//-//p and B10-H-2a11-4b p donors to BIO. A and B10.D2/O recipients, respectively. The speed of rejection of grafts incompatible at the H-4 locus (11-4b-11-4°) depended upon the H-2 haplotype. On the H-2d background, grafts were rejected with a mean survival time (MST) of 40.4 days. Grafts were rejected more slowly on H-2h and H-2a backgrounds with MSTs of 48.il days and >S8.1 and >70.4 days (two groups for H-2a), respectively. The utility of double congenic strains in investigating regulation of the immune response to non-H-2 histocompatibility antigens by the H-2- linked immune response region is discussed.

H-2 control of expression of an idiotype shared by normal B cells and a B-cell lymphoma

The spleens of normal B10,H-2aH-44bp/Wts (2a4b) mice; contain cells which, in response to mitogen stimulation, secrete hemolytic antibody specific for a determinant present on both sheep and bromelain-treated mouse erythrocytes. These cells were found to be Ly-1 positive. Approximately 50% of these cells bear surface immunoglobulin (sIg) with the same idiotype as the sIg of a 2a4b-derived B-cell lymphoma, CH12. Backcross analysis revealed H-2 control of the frequency of the idiotype-positive B cell. The regulatory gene did not correlate with the Igh-1 allotype, and analysis of 22 inbred mouse strains mapped the gene to the I-E subregion. Surprisingly, only strains homozygous for Eαkexpressed the idiotype, and expression was a recessive trait. Possible mechanisms for this control of idiotype expression and its relation to lymphomagenesis are discussed.

The production, testing, and utility of double congenic mouse strains. II. B10-h-2ah-7b/wts and b10-h-2dh-7b/wts.

Two new double congenic strains, B10-H-2aH-7b/Wts and B10-H-2dH-7b/Wts, were selected to differ from B10.A and B10.D2/o, respectively, at theH-7 locus. The survival time ofH-7-incompatible skin grafts is dependent upon theH-2 haplotype of recipient and donor.