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Biomedical subjects

R G Lynch

Publications and source records attributed to R G Lynch.

At least 73 records · Page 4Linked to original sources
PubMed1988

IgA:IgM and IgA:IgA hybrid hybridomas secrete heteropolymeric immunoglobulins that are polyvalent and bispecific.

Polyvalent bispecific antibodies were secreted by hybrid hybridoma cells when both parental clones expressed a naturally polymerizing immunoglobulin. Hybrid hybridomas made from IgA lambda 2 anti-trinitrophenyl (TNP) and IgA kappa anti-phosphocholine (PC) parental cells secreted polymeric IgA antibodies that bound both TNP and PC. Some of the TNP binding was dissociated from the PC binding under conditions of mild reduction and alkylation suggesting that the bispecific polymeric IgA contained disulfide-linked parental monomers as well as bispecific hybrid monomers. Hybrid hybridomas constructed from IgA lambda 2 anti-TNP and IgM kappa anti-ox erythrocyte parental cells secreted bispecific, polymeric immunoglobulin that contained mu-, alpha-, kappa-, and lambda 2-chains. The mu and kappa-chains dissociated from the alpha- and lambda 2-chains under conditions of mild reduction and alkylation, indicating that both parental monomers had been incorporated into the same polymeric immunoglobulin to form a heteropolymeric antibody molecule. Heterologous pairing of alpha and mu heavy chains in monomers was not detected. Hybrid hybridomas constructed from IgA lambda 2 and IgG3 lambda 2 or IgA lambda 2 and IgG1 kappa parents co-secreted both parental immunoglobulins, but the antibodies secreted by these clones did not form heteropolymers or exhibit heterologous heavy chain pairing. These findings establish that polyvalent, bispecific, polymeric immunoglobulin molecules can be produced by hybrid hybridomas when both parents express a naturally polymerizing class of heavy chain but not when only one parent does. Hybrid hybridomas that produce heteropolymeric immunoglobulins are sources of high avidity bispecific antibodies that may find a number of basic and practical applications. The hybridoma cells that produce these antibodies may provide useful tools for investigating the in situ determinants of immunoglobulin chain association and the regulation of antibody assembly and secretion.

Animals↗
PubMed1988

The contribution of constant region domains to the binding of murine IgM to Fc mu receptors on T cells.

IgM hybridoma constant region domain deletional mutants were used to investigate the domain requirements for binding of murine IgM to Fc mu receptors (Fc mu R) on normal murine T lymphocytes. Parental Sp 6:18 (mu, kappa; anti-trinitrophenyl) and its mutant proteins or their trinitrophenyl-antigen immune complexes were tested for their ability to inhibit the binding of pentameric IgM to Fc mu R on T lymphocytes. Inhibition was observed with ligands containing multiple copies of the third constant region domain. Inhibition did not occur with ligands missing the third constant region domain. In addition, a battery of rat monoclonal antibodies specific for individual murine IgM constant region domains was tested for the ability to inhibit the binding of pentameric murine IgM to Fc mu R on normal murine T lymphocytes. Total inhibition was observed with the antibodies directed to different epitopes located in C mu 3, but significant inhibition was not observed with antibodies directed to C mu 1, C mu 2, or C mu 4. Studies with domain deletional mutants and anti-domain antibodies have independently provided strong evidence that the C mu 3 domain plays a major role in the binding of IgM to Fc mu R on T lymphocytes and that C mu 1, C mu 2, and C mu 4 are not essential for binding. These studies have also provided evidence that valency and avidity influence the binding of IgM to T lymphocytes that express Fc mu R.

Antibodies, Monoclonal↗
PubMed1988

Analysis of human peripheral blood mononuclear cells bearing Fc receptors for the three major immunoglobulin isotypes.

Fluorochrome-labeled, human immunoglobulins (Igs) were used to characterize peripheral blood mononuclear cell (PBMC) subsets that had Fc receptors (FcRs) for the three major isotype classes, IgG, IgA and IgM. Relatively high concentrations of these Igs were needed to saturate FcRs. However, by a number of criteria, competition assays in particular, isotype-specific FcR binding was shown. In vivo bound Ig was directly measured by anti-Fc class-specific F(ab)2 antibodies. With the exception of monocytes, where a mean of 19% had in vivo bound IgG, only small percentages of other PBMC subsets had any type of in vivo bound Ig. Studies done on a large group of normal individuals revealed that approximately 27% of all PBMCs had gamma, 24% alpha, and 23% microFcRs. Dual fluorescence assays were performed to determine the gamma, alpha, and microFcR status on T, B, NK cells and on monocytes. The majority of staining for gamma, alpha, and microFcRs was accounted for by the numerically largest subfraction, the T cells. However, when FcR status was determined for each subset virtually all had either gamma, alpha, or microFcRs--94% of CD2+, 98% of CD4+, 73% of CD8+, 100% of CD19+, 100% of CDw19+, 92% of Leu 7+, and 61% of Leu 11+ cells. All three FcRs were expressed within each subset. Of interest, a small percentage of unfractionated and monocyte-depleted PBMCs coexpressed one of three pairs of FcRs, either alpha and gamma, alpha and mu, or mu and gamma.

Antigens, Differentiation↗
PubMed1988

Immunoglobulin (Fc) receptors on murine T- and B-lymphocytes: investigations using tumor models.

Lymphoid tumors are productive experimental models for the study of lymphocyte immunoglobulin receptors. Investigations with Fc receptor expressing lymphoid tumor cells have generated much useful information about: (a) the developmental expression of the different classes of Fc receptors on lymphoid cells of the T- and B-lineages; (b) the biochemical steps involved in the regulation of Fc receptor expression on lymphoid cells; (c) the structures of lymphoid cell Fc receptors and their genes; (d) the signals that induce alterations in the expression of Fc receptors on lymphoid cells; and (e) the molecular specificity of the binding of immunoglobulin to lymphoid cells Fc receptors. In addition, tumors that secrete immunoglobulins are providing useful models for analysis of the mechanisms by which B-cells influence Fc receptor expression and function on T-cells. An interesting, bi-directional immunoregulatory circuit involving Fc epsilon R+ host T-cells and IgE-secreting hybridoma cells has been identified that could prove useful in the analysis of the regulation of epsilon heavy chain expression. The studies discussed in this article and elsewhere in this volume serve to emphasize that, in addition to being clonal sources of key molecules such as Fc receptors and their messenger RNAs, lymphoid tumor cells that express Fc receptors are powerful and unique experimental models for investigating the developmental biology, regulation and function of lymphocyte Fc receptors.

Animals↗
PubMed1987

Thymus-dependent in vivo suppression of IgE synthesis in a murine IgE-secreting hybridoma.

In previous studies we demonstrated that BALB/c mice bearing ascitic tumors of the IgE-secreting hybridoma B53 (epsilon, kappa, anti-dinitrophenyl) developed large numbers of Lyt-1-2+ Fc epsilon R(+) T lymphocytes (T cells with membrane Fc receptors) in response to the elevated serum IgE concentration. The development of Fc epsilon R(+) T lymphocytes was followed by a progressive decrease in the levels of serum IgE in spite of continued proliferation of the hybridoma cells. This sequence of events suggested that the IgE-secreting hybridoma triggered a suppressive immunoregulatory circuit of the host that inhibited IgE expression by the hybridoma cells. The present studies were undertaken to investigate the basis for the subsequent decline in serum IgE levels in mice with B53 tumors and to identify host factors that might be involved in this process. We observed that ascitic B53 cells recovered at increasing time points from BALB/c mice exhibited a selective decline in steady state levels and rates of synthesis of epsilon-heavy chain protein and mRNA. The expression of kappa-light chain protein and mRNA appeared relatively unchanged. The decrease in epsilon-heavy chain gene expression did not occur when B53 tumors were passaged in nu+/nu+ mice or in BALB/c mice depleted of Lyt-2+ cells (suppressor/cytotoxic cell lineage), but did occur in nu+/nu+ mice reconstituted with neonatal BALB/c thymus and in BALB/c mice depleted of L3T4+ cells (helper/inducer cell lineage). That Fc epsilon R(+) T lymphocytes were directly involved in the inhibition of IgE expression was supported by the earlier and more pronounced inhibition of B53 IgE in mice infused with Fc epsilon R(+) T lymphocytes. We conclude from these findings that: 1) the decline in serum IgE levels that occurs toward the end of each generation of in vivo passage of the B53 hybridoma is due to decreased production of IgE by the hybridoma cells, 2) the decreased production of IgE is due to a selective loss of epsilon mRNA expression, 3) the decrease production of IgE by B53 cells is dependent on the presence of Lyt-2+ cells, and 4) Fc epsilon R(+) T lymphocytes participate in the mechanism by which IgE production is suppressed.

Animals↗
PubMed1987

Effect of B cell stimulatory factor-1 (interleukin 4) on Fc epsilon and Fc gamma receptor expression on murine B lymphocytes and B cell lines.

Culture of murine splenic B cells with interleukin 4 (IL-4) caused the up-regulation of the lymphocyte Fc receptor for immunoglobulin E (IgE) (Fc epsilon R) over a similar dose range as required for Ia up-regulation. However, the expression level of the Fc receptor for immunoglobulin G (Fc gamma R) did not increase, rather IL-4 caused a slight but consistent decrease in the Fc gamma R level on the B cells. Fc epsilon R+ B hybridoma cells also responded to IL-4 by exhibiting increased Fc epsilon R expression; with the hybridoma cells Fc gamma R levels were unaffected. IL-4 caused an increase in the number of Fc epsilon R per cell and the highest levels of expression were obtained by having both IgE and IL-4 present in the culture. The specificity of the increase was demonstrated by blocking IL-4-mediated actions with monoclonal anti-IL-4 (11B11). Experiments following the incorporation of [35S]methionine into the Fc epsilon R demonstrated that IL-4 increased the rate of Fc epsilon R biosynthesis; this provides an explanation for the IL-4-induced increase in Fc epsilon R expression. IL-4, unlike IgE, had no effect on the rate of degradation of the Fc epsilon R. Interferon-gamma (IFN-gamma) totally abrogated IL-4-mediated Fc epsilon R up-regulation; at the same concentration of IFN-gamma Ia up-regulation is also suppressed, although not as effectively. IFN-gamma was shown to directly suppress Fc epsilon R synthesis, thereby explaining the inhibitory action on Fc epsilon R levels. Finally, it was shown that 11B11 inhibited the increased expression of Fc epsilon R on B cells obtained from mice during the early, but not the late, stages of Nippostrongylus brasiliensis infection. This latter finding suggests that the high Fc epsilon R levels seen early in parasite infections are dependent upon IL-4. The results overall provide further insight into the biologic activities of IL-4.

Animals↗
PubMed1987

Immunoglobulin-specific suppressor T cells.

This article has considered evidence that supports the occurrence and functional importance of suppressor T cells that are directed to B cell targets. Cells with these features have been demonstrated in experimental animals and in humans. The designation "suppressor" comes from the serologic phenotype of these cells as well as from their functional property of noncytotoxic inhibition of B cell function. Distinct suppressor T cells with these properties have been identified that effect antigen-, idiotype-, isotype-, and allotype-specific suppression of B cell function. While such cells had been suspected from earlier studies of normal immune responses, the development of monoclonal B cell models using tumor cells has provided a means to readily detect these suppressor T cells and to investigate the mechanisms by which they mediate their effects. Tumor models have proved to be powerful tools in the effort to identify and analyze the elements that underlie the complexity of immune responses. Combined with the insights provided by molecular genetic approaches and flow cytometry, functional and responsive lymphoid tumor cells are being used with increasing frequency to address basic immunoregulatory issues. An important family of suppressor T cells with B cell targets are those that express surface Fc receptors, elaborate immunoglobulin-binding factors, and appear to participate in the regulation of immunoglobulin heavy chain class expression. In addition to their importance in the regulation of heavy chain class expression during normal immune responses, alterations in FcR+ T cells in a number of disease states may provide clues that will lead to a better understanding of disorders of immune regulation.

Animals↗
PubMed1986

Increased T epsilon cells in BALB/c mice with an IgE-secreting hybridoma.

Since it has been proposed that T cells with receptors for the Fc portion of IgE (T epsilon) in rats, mice and humans are IgE-specific regulatory cells, it was investigated whether mice with an IgE-secreting hybridoma might be a source of large numbers of T epsilon cells. BALB/c mice with ascitic tumors of the IgE hybridoma B53 (epsilon, kappa, anti-DNP) developed high serum concns of monoclonal IgE which was followed by the appearance of large numbers of Lyt1-2+ L3T4- T epsilon cells. In contrast, mice bearing a non-IgE producing variant of B53 failed to develop T epsilon cells. Mice infused with cell-free ascites fluid (containing high levels of monoclonal IgE) from the IgE-secreting B53 hybridoma developed high serum IgE levels and also developed high numbers of T epsilon cells. Mice infused with cell-free ascites obtained from the non-IgE-secreting variant ENP-1 (containing very low amounts of IgE) did not develop either high serum IgE levels or T epsilon cells. These findings suggest that high serum IgE concns induce large numbers of T cells that express phenotypic markers of suppressor cells and have surface IgE-Fc receptors. These studies extend to IgE the principle that hybridomas and plasmacytomas induce large numbers of immunoregulatory T cells that express Fc receptors specific for the heavy chain class of the secreted monoclonal immunoglobulin. Since T epsilon cells are normally present only in small numbers, their marked increase in mice with IgE-secreting hybridomas identifies a ready source of large numbers of T epsilon cells that can be used to investigate their regulatory properties.

Animals↗
PubMed1986

The physiology of B cells as studied with tumor models.

B-cell tumors have been extraordinary sources of information about antibodies, their genes, and the cells that express them. An important principle that has emerged from the study of lymphoid tumors is that the long-held view that malignant lymphoid cells are "frozen" at a fixed point in differentiation is not generally valid. Presentation of immunoregulatory signals to transformed B cells can profoundly influence their proliferation, morphology, differentiation, gene expression, and immunoglobulin synthesis. In addition to their responsiveness to immunoregulatory signals, some tumors of B lineage elaborate immunoregulatory signals. Until quite recently B-cell tumors were used primarily as monoclonal sources of molecules of immunological interest. While they continue to be important sources of receptors, growth and differentiation factors, differentiation antigens, and immunoregulatory factors, they are being used with increasing frequency to define the molecular events that occur in B cells subsequent to receipt of an immunoregulatory signal. While the use of tumor cells as models of normal cells is often viewed with some skepticism, it is difficult to find examples wherein tumors have been misleading. Quite to the contrary, B-cell tumors have regularly provided powerful tools for dissecting the molecular events that underlie B-cell development, function, and regulation.

Animals↗
PubMed1986

Components of immune function studied with lymphoid tumors.

Lymphoid tumors have proven to be productive models for investigating genetic, biochemical, physiological and developmental mechanisms of lymphoid cells. Lymphoid tumors of a given lineage exhibit a spectrum of phenotypes from clones whose features overlap extensively with their normal counterparts to clones whose features are not obviously represented in normal lymphoid populations. In addition to the phenotypic continuum represented in a library of related tumors, many clones exhibit phenotypic diversity spontaneously or in response to various stimuli. As laboratory technology has advanced many interesting biological features of lymphoid tumors have become amenable to precise analysis. Progress made in understanding the mechanisms that underly the phenotypic diversity and plasticity of lymphoid tumor cells has the dual impact of improving our knowledge of malignant and normal lymphoid cells.

B-Lymphocytes↗
PubMed1986

Increased T gamma and T mu cells in BALB/c mice with IgG and IgM plasmacytomas and hybridomas.

The results of previous studies in our laboratory have shown that mice bearing plasmacytomas and hybridomas that secrete IgA or IgE are accompanied by increased frequencies of Lyt-1-2+ T lymphocytes bearing Fc receptors (FcR) for IgA (T alpha) or IgE (T epsilon), respectively. The present study was undertaken to examine whether IgG- or IgM-secreting tumors influenced the frequency of T lymphocytes that express FcR for IgG or IgM. We studied mice bearing IgG- and IgM-secreting plasmacytomas and hybridomas. BALB/c mice injected subcutaneously with the IgG-secreting hybridoma HDP1 (gamma 1 kappa, anti-TNP) were sequentially examined for the frequencies and Lyt phenotypes of splenic lymphocytes bearing FcR for IgG (T gamma), IgM (T mu), and IgA (T alpha). A threefold increase in the frequency of T gamma lymphocytes that were Lyt-1-2+, L3T4- was seen. The frequencies of T mu and T alpha lymphocytes in these mice were not significantly altered. Similarly, mice injected subcutaneously with the IgM-secreting plasmacytoma MOPC 104E (mu lambda, anti-dextran) or the IgM-secreting hybridoma C1D1 (mu kappa, anti-ox RBC) were examined sequentially for the frequencies of T gamma, T mu, and T alpha lymphocytes. Mice with established IgM subcutaneous tumors showed a twofold increase in splenic, nylon wool-nonadherent T mu lymphocytes. This was associated with a relative increase in Lyt-2+ splenic T lymphocytes and a relative decrease in Lyt-1+ splenic T lymphocytes. No changes were observed in the frequencies of either T gamma or T alpha lymphocytes. These studies extend to IgG and IgM the observation that plasmacytomas and hybridomas secreting immunoglobulins of a specific isotype cause an expansion of T lymphocytes bearing FcR specific for the corresponding isotype. The expansion of FcR+ Lyt-1-2+ T lymphocytes likely represents an exaggerated, but otherwise normal, immunoregulatory response of the host. These cells may be an important element in the regulation of isotype expression.

Animals↗
PubMed1984

Idiotype-specific T cell suppression of light chain mRNA expression in MOPC-315 cells is accompanied by a posttranscriptional inhibition of heavy chain expression.

Earlier studies have shown that idiotype-specific T suppressor cells can directly inhibit the biosynthesis and subsequent secretion of immunoglobulin from the BALB/c myeloma, MOPC-315. This suppression is highly specific and does not affect nonimmunoglobulin protein synthesis in these cells. The selective character of this suppression suggested a transcriptional or translational mechanism of control. Therefore, we analyzed suppressed MOPC-315 cells for the expression of light and heavy chain mRNAs. Since the events that occur in a B cell subsequent to reception of suppressive signals from T cells are presently unknown, these experiments address a fundamental aspect of B cell regulation. Another important aspect of these studies is that they deal with a suppressor T cell that clearly operates directly upon the antibody-secreting cell. This system involves regulation of a B cell differentiative function without influencing the B cell clonal size. We have employed the trinitrophenyl (TNP)-specific myeloma, MOPC-315, because it provides a source of monoclonal B cells which in earlier studies have been shown to be responsive to idiotype-specific and TNP-antigen-specific immunoregulatory signals. The idiotype-specific T cells were generated by hyperimmunizing BALB/c mice with the myeloma protein bearing the 315 idiotype, in complete Freund's adjuvant. Poly A+ mRNA was isolated from MOPC-315 cells following coculture with either normal or idiotype-immune T cells. The mRNA was analyzed using hybridization techniques and cDNA probes specific for alpha or lambda 2 light chain constant regions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Suppressor T cell action inhibits the expression of an excluded immunoglobulin gene.

Cells of the murine plasmacytoid line MOPC-315 synthesize two distinct immunoglobulin light chains: a normal lambda II protein, which is incorporated into secretory and surface-bound immunoglobulin, and a truncated, nonfunctional lambda I protein found only in the cytoplasm. Idiotype-specific suppressor T lymphocytes selectively inhibit the expression of both lambda II- and lambda I-specific messenger RNA by MOPC-315 cells. This finding demonstrates that phenotypically excluded light chain genes can be subject to immunoregulatory control and suggests that the expression of divergent lambda isotypes may be coordinately regulated in immunoglobulin-secreting cells.

Animals↗
PubMed1983

Anti-idiotypic regulation of IgA expression in myeloma cells.

MOPC-315 (alpha lambda 2) is a BALB/c plasmacytoma that produces an anti-TNP antibody (M315). In addition to being secreted M315 is also expressed on the surface membranes of MOPC-315 cells. In the present studies an in vitro-adapted line of MOPC-315 was used to study the effect of affinity-purified, isologous anti-idiotypic antibodies (a-Id315) and idiotype-specific T cells on M315 expression. We observed that both monoclonal and polyclonal a-Id315 mediated a reversible clearance of surface membrane M315 but did not influence M315 secretion or MOPC-315 growth even when the myeloma cells were cultured in the continuous presence of a-Id315 for three weeks. Clearance of surface M315 was rapid, reversible, and a-Id315 dose-dependent. M315:a-Id315 complexes were shed from MOPC-315 cells in the form of microscopic membranous vesicles and a-Id315 was consumed in the process. Protein synthesis was required for re-expression of surface M315 only if a presynthesized internal pool of M315 had previously been depleted. In contrast, idiotype-specific T cells mediated specific inhibition of M315 secretion without influencing surface M315 expression. Although the anti-idiotypic antibodies and the anti-idiotypic T-cells are both directed to the surface membrane immunoglobulin on the cloned B cell, the anti-idiotypic antibodies regulate surface membrane expression of immunoglobulin while the anti-idiotypic T-cells regulate secretion of immunoglobulin. These observations support the view that in idiotype regulation, surface membrane immunoglobulin molecules function as a focusing device for regulatory effectors which actually determine the quality of the effect achieved.

Animals↗