Biomedical subjects

S T Ju

Publications and source records attributed to S T Ju.

At least 55 records · Page 3Linked to original sources

PubMed1992

Intraperitoneal amyloid formation by amyloid enhancing factor--rich macrophages in ascitic fluid.

Although resident peritoneal cells from amyloidotic mice (amyloidotic peritoneal cells) are capable of processing the precursor protein of secondary amyloidosis, serum amyloid A (SAA) to amyloid fibrils, the peritoneum is a rare site for amyloid deposition. This is considered to be due to a deficiency of SAA in the peritoneum. To increase the supply of SAA to the peritoneum, ascitic fluid containing about the same protein constituents as in the serum was induced in mice. Amyloidotic peritoneal cells were packed in a microchamber which was shielded with filter membranes, and cultured in ascitic fluid supplemented with additional inflammatory factors. On the 7th day, Congo red-positive structures which showed green birefringence under polarized light were found inside and occasionally outside the chamber. By anti-AA or -SAA immunostaining, amyloid deposits and the cell surfaces of macrophages were positive. Immunologic depletion of T- and B-lymphocytes from the amyloidotic peritoneal cells did not adversely effect the amyloid formation in microchambers. These results suggest that either ascitic fluid containing sufficient amounts of SAA, or peritoneal macrophages with a high amyloid enhancing factor (AEF) activity are indispensable for AA amyloid fibrillogenesis in the peritoneum.

Amyloid↗

PubMed1991

A novel cytotoxicity of CD4+ Th1 clones on heat-shocked tumor targets. I. Implications for internal disintegration model for target death and hyperthermia treatment of cancers.

A novel cytotoxicity, which is normally hidden but unveiled upon interacting with heat-shocked tumor target, has been identified in CD4+ Th1 cells. When TNF-resistant, Ag-presenting tumor targets were heat shocked, the cytotoxicity by specific Th1 clones was significantly enhanced. Interestingly, the DNA of heat-shocked, unpulsed targets including Ia- tumor cells were also fragmented by Th1 clones. In contrast to the Ag-dependent, MHC-restricted cytotoxicity, the heat-shock-induced sensitivity to Th1 clones was a) Ag independent and MHC unrestricted, b) insensitive to CD4-mAb, c) resistant to actinomycin D and cycloheximide, and d) greatly enhanced by cholera toxin, PGE1, PGE2, and dibutyryl cAMP. This novel cytotoxicity was inhibited by mAb specific to lymphocyte function-associated Ag-1 and intercellular adhesion molecule-1. Upon culturing at 37 degrees C, mildly heat-shocked target cells gradually recovered, indicating that the heat-shock-induced sensitivity was transient and reversible. The implication of this study for a signal-induced internal disintegration mechanism for target death is discussed. The novel cytotoxicity of CD4+ Th1 cells may be an important mechanism of immune regulation under febrile conditions and an underlying mechanism for the hyperthermia treatment of cancers.

Animals↗

PubMed1991

Distinct pathways of CD4 and CD8 cells induce rapid target DNA fragmentation.

When activated with either Con A, a CD3-specific mAb, or Ag-pulsed B lymphoma (LK35.2) cells, CD4 (Th1) clones quickly induce DNA fragmentation in target cells followed by release of 51Cr-labeled intracellular materials. Both activated CD4 clones and CD8 (CTL) cells fragment target DNA into electrophoretically identical "ladder" pattern made of approximately 200 bp. The effect of various metabolic inhibitors on the ability of CD4 and CD8 cells to induce target DNA fragmentation was studied. Little effect was observed with the DNA synthesis inhibitor, mitomycin C. The RNA synthesis inhibitor, actinomycin D, and the protein synthesis inhibitor, cycloheximide, strongly inhibited the ability of CD4 cells, but not CD8 cells, to induce target DNA fragmentation. In contrast, target DNA fragmentation by CD8 cells, but not by CD4 cells, was inhibited by cholera toxin. Although cyclosporin A inhibited CD4 cells to fragment target DNA during the early phase (90 min) of E:T interaction, this inhibition was not sustained in the later phase (210 min) of the assay. Zinc ions inhibited the ability of both CD4 and CD8 cells to fragment target DNA. Treatment of effectors and targets with these inhibitors, followed by washings, demonstrated that the action of these inhibitors on effector cells alone is sufficient to inhibit target DNA fragmentation. The strong correlation among these parameters of DNA fragmentation and Cr-release assays supports the hypothesis of programed cell death. Although distinct cytolytic pathways are used by CD4 and CD8 cells to kill targets, both pathways deliver a signal that activates endonuclease(s), fragments target DNA, causes Cr-release, and lyses target cells. Taken together with our previous studies, the present findings demonstrate that activated cytolytic CD4 clones do not use perforin, serine proteases, and TNF as mediators for resistant target DNA fragmentation.

Animals↗

PubMed1991

The CD3-gamma and CD3-delta subunits of the T cell antigen receptor can be expressed within distinct functional TCR/CD3 complexes.

The T cell receptor for antigen (TCR) consists of two glycoproteins containing variable regions (TCR-alpha/beta or TCR-gamma/delta) which are expressed on the cell surface in association with at least four invariant proteins (CD3-gamma, -delta, -epsilon and -zeta). CD3-gamma and CD3-delta chains are highly homologous, especially in the cytoplasmic domain. The similarity observed in their genomic organization and their proximity in the chromosome indicate that both genes arose from duplication of a single gene. Here, we provide several lines of evidence which indicate that in human and murine T cells which expressed both the CD3-gamma and CD3-delta chains on their surface, the TCR/CD3 complex consisted of a mixture of alpha beta gamma epsilon zeta and alpha beta delta epsilon zeta complexes rather than a single alpha beta gamma delta epsilon zeta complex. First, a CD3-gamma specific antibody failed to co-immunoprecipitate CD3-delta and conversely, several CD3-delta specific antibodies did not coprecipitate CD3-gamma. Secondly, analysis of a panel of human and murine T cell lines demonstrated that CD3-gamma and CD3-delta were expressed at highly variable ratios on their surface. This suggested that these chains were not expressed as a single complex. Thirdly, CD3-gamma and CD3-delta competed for binding to CD3-epsilon in transfected COS cells, suggesting that CD3-gamma and CD3-delta formed mutually exclusive complexes. The existence of these two forms of TCR/CD3 complexes could have important implications in the understanding of T cell receptor function and its role in T cell development.

Animals↗

PubMed1990

Generation and use of site-specific antibodies to serum amyloid A for probing amyloid A development.

Serum amyloid A (SAA) is an acute phase reactant and the putative precursor of an amyloid fibril protein (AA). During amyloidogenesis in mice, the C-terminal portion of SAA that consists of 28 amino acids is cleaved off to produce the 75 amino acid AA. Of three known isotypes of SAA, SAA2 has been identified as amyloidogenic. For differential immunologic recognition of SAA and AA, antisera were generated in rabbits against AA protein and a synthetic peptide corresponding to the C-terminal (amino acids 84 to 103) of murine SAA2. The anti-AA antiserum reacted with AA and SAA, but the anti-peptide antiserum reacted with SAA only. Immunohistochemically anti-peptide antiserum predominantly stained the rims of murine amyloid deposits. In contrast, anti-AA antiserum generated an overall homogeneous staining of amyloid deposits. The data lend support to a view that SAA are concentrated in the vicinity of the surrounding cells (mostly macrophages) where they are processed to AA, or that SAA are deposited initially as components of the amyloid fibrils onto the outside margin of amyloid deposits and the C-terminus is cleaved off as maturation of the amyloid fibril progresses.

Animals↗

PubMed1990

Expression of two distinct cytolytic mechanisms among murine CD4 subsets.

A TNF (TNF-alpha and TNF-beta)-sensitive target, L929, and two TNF-resistant targets, P815 and LK were used to compare the cytolytic activity among subsets of CD4+ (Th) clones. Cytolytic activity was induced with either Con A, CD3-mAb, or Ag-pulsed LK cells. Six Th1 clones are strongly cytolytic against all three targets. In contrast, Th2 clones are either noncytolytic or weakly cytolytic. Although there is an apparent correlation between TNF production, killing of L929 cells, and the killing of TNF-resistant targets, an anti-TNF serum (capable of neutralizing both TNF-alpha and TNF-beta) selectively inhibits CD4 clones to lyse L929 cells, whereas the lysis of P815 or LK cells was unaffected. The continuous presence of noncytotoxic levels of Actinomycin D (AcD) and cycloheximide, but not mitomycin C, cyclosporin A (CsA), or cholera toxin (ChT) inhibits the lysis of Ag-pulsed, Ia-bearing LK cells; indicating a requirement for de novo synthesis of RNA and protein for cytolytic activity. Although pretreatment with AcD, CsA, or ChT strongly inhibits production of IL-2, TNF and IFN-gamma, only clones pretreated with AcD lose cytolytic activity against Ag-pulsed, Ia-bearing LK cells. These observations support a model of TNF-independent killing of TNF-resistant targets. The TNF-independent cytolytic activity does not correlate with serine esterase activity released into media upon activation of CD4 clones. Moreover, the effects of metabolic inhibitors on serine esterase release do not correlate with their effects on cytolytic activity. Collectively, the data demonstrate that activated CD4 cells express two distinct cytolytic activities; a TNF (and IFN-gamma)-mediated cytotoxicity and a TNF (and IFN-gamma)-independent cytolytic activity. Both pathways require de novo synthesis of RNA and protein and appear to be independent of granule enzyme release. Only the TNF-independent cytolytic activity is resistant to CsA and ChT inhibition.

Animals↗

PubMed1990

Metabolic inhibitors distinguish cytolytic activity of CD4 and CD8 clones.

The effect of various metabolic inhibitors on the expression of cytolytic activity of CD4 (TH1) and CD8 (CTL) clones was studied. The cytolytic activity of CD4 clones, but not CD8 clones, was sensitive to the RNA synthesis inhibitor actinomycin D and the protein synthesis inhibitor cycloheximide. Conversely, cholera toxin (CT) inhibited cytolytic activity of CD8, but not CD4 clones. Both mitomycin C, a DNA synthesis inhibitor, and cyclosporin A (CsA) failed to inhibit the cytolytic activity of either CD4 or CD8 clones. Although pretreatment with CsA or CT did not inhibit the cytolytic activity of CD4 clones, lymphokine (interleukin 2, IL2, interferon-gamma, IFN-gamma, and tumor necrosis factor, TNF) production was strongly inhibited. Similarly, pretreatment of a CD8 clone with actinomycin D or CsA inhibited lymphokine production without affecting cytolytic activity. The production of mRNA for TNF and IFN-gamma by concanavalin A-activated CD4 clones was also inhibited by CsA and CT. Moreover, perforin-specific mRNA was not detected in activated CD4 clones. Collectively, these observations demonstrated that de novo synthesis of RNA and protein is required for expression of cytolytic activity of CD4 clones, yet production of TNF, INF-gamma, IL 2 and perforin is not involved. In contrast, the cytolytic machinery of CD8 clones is present prior to activation and is quickly expressed following activation even when de novo synthesis of RNA, protein and lymphokines is blocked.

Animals↗

PubMed1990

Amyloid enhancing factor-loaded macrophages in amyloid fibril formation.

Amyloid enhancing factor (AEF) is believed to be a key agent that triggers the second (deposition) phase of amyloidogenesis. However, the target cells of AEF activation and their function after the activation have not yet been clearly identified. We found that peritoneal resident cells from amyloidotic mice contained very high AEF activity. With a simultaneous subcutaneous injection of 1.0 ml of the casein-adjuvant emulsion, an intravenous injection of 10,000 cells was consistently capable of inducing amyloidosis in a recipient mouse in 72 hours. After 2-hour cultures, the major AEF activity was found in the adherent cells (macrophages). An intravenous injection of 5 to 10 million of the live macrophages with the casein-adjuvant injection caused amyloid deposits in the recipient not only in the spleen and the liver but also in the lung (an extremely rare site of AA amyloid deposition). We have interpreted this finding to indicate that the injected AEF-loaded macrophages, while still residing in the lung and exposed to the blood stream, processed SAA to form amyloid. We further tested this postulate in an in vitro system. In a 4-day culture of the AEF-loaded macrophages in a medium containing SAA-rich mouse serum, small masses (less than 15 microns in diameter) of Congo red positive substance were observed scattered adjacent to or surrounded by the macrophages. The present observations lend strong credence to the conclusion that AEF-loaded macrophages are fully capable of processing SAA to AA and further to amyloid fibrils, and that they indeed play a role in the second phase of amyloidogenesis in vivo.

Amyloid↗

PubMed1989

Induction of antigen-specific antibody responses in primed and unprimed B cells. Functional heterogeneity among Th1 and Th2 T cell clones.

CD4+ T cells have been recently divided into two subsets. The functions of these subsets are thought to be distinct: one subset (Th1) is responsible for delayed type hypersensitivity responses and another (Th2) is primarily responsible for induction of antibody synthesis. To more precisely define the roles of both subsets in humoral immune responses, we examined the ability of a panel of nominal antigen specific Th1 and Th2 clones to induce anti-TNP specific antibody synthesis in TNP-primed or unprimed B cells. Four of nine Th1 clones induced little or no antibody synthesis with TNP-primed B cells. However, five other Th1 clones were very effective at inducing IgG anti-TNP plaque-forming cell (PFC) responses in primed B cells. One of these Th1 clones was analysed in detail and found to also provide helper function for unprimed B cells. Cognate B-T cell interaction was required for induction of both primary and secondary responses with this clone, indicating that a Th1 clone could function as a "classical" Th cell. The seven IL-4 producing Th2 clones examined were also heterogeneous in their ability to induce antibody secretion by TNP-primed B cells. Although four of the Th2 clones induced IgG and IgM anti-TNP PFC responses, two Th2 clones induced only IgM and no IgG antibody, and another clone failed to induce any anti-TNP PFC. All Th2 clones failed to induce any anti-TNP PFC. All Th2 clones produced high levels of IL-4, but "helper" Th2 clones produced significantly greater amounts of IL-5 than "non-helper" Th2 clones. These studies indicate that some IL-2- and some IL-4-producing T cell clones can induce TNP-specific antibody in cell clones can induce TNP-specific antibody in primed and unprimed B cells, and that Th1 and Th2 clones are heterogeneous in their ability to induce Ig synthesis. Therefore, although T cell clones can be classified as Th1 or Th2 types according to patterns of IL-2, IFN-gamma, or IL-4 synthesis, the functional capacity to induce antibody synthesis cannot be predicted solely by their ability to secrete these lymphokines.

Animals↗

PubMed1989

Production and immunoselection of IgM-IgA hybridomas: preparing immunoglobulins with dual binding specificity.

Fusion between the thioguanine resistant myeloma cell line MOPC-315 [which produces alpha, lambda-2 antibodies specific to the 2,4-dinitrophenyl (DNP) hapten] and a long term in vivo maintained hybridoma 6100.15 [which produces mu, lambda-1 antibodies specific to the 4-hydroxy-3-nitrophenyl acetyl (NP) hapten] resulted in the generation of 12 hybridomas. These hybridomas secrete a mixed family of immunoglobulins (Ig) that bind both DNP and NP and express both IgM and IgA serological determinants. Affinity purified molecules from NP, DNP, anti-mu, or anti-alpha immunosorbents react with both anti-mu and anti-alpha antisera, suggesting that these Ig represent IgM-IgA hybrid molecules. This conclusion was supported by idiotypic analyses. To determine the roles of individual immunoglobulin chains in determining antibody specificity this IgM-IgA hybridoma was used for immunoselection. Following lysis with specific anti-mu and anti-idiotype antibodies, an alpha+, mu- variant clone (A12) was identified, which secreted Ig that binds DNP but not NP. The DNP binding proteins were shown to express alpha, lambda-1 and lambda-2 chains. In contrast, the Ig which lack DNP binding activity only expressed alpha and lambda-1 determinants. The combined results demonstrate that the lambda-1 chain from 6100.15 hybridoma cannot replace lambda-2 of MOPC-315 for DNP binding activity. These data imply that these closely related lambda chains carry sites critical for antigen binding activity. An IgM-IgA hybridoma variant (MA2) which secretes Ig that binds to NP and DNP and expresses mu, alpha and lambda-2 chains was also characterized. This molecule lacked a lambda-1 chain. To determine if the Ig prepared with heterologous mu and lambda-2 chains had NP binding activity required immunoselection of a fourth clone (M2). M2 secretes homogeneous Ig bearing only mu and lambda-2 chains. In contrast to either parental Ig, the M2 antibody molecules express dual binding activity to both NP and DNP. Thus, critical amino acid substitutions in the MOPC-315 lambda-2 sequence are required for DNA binding specificity.

Animals↗

PubMed1988

Autoreactive T cells in MRL/Mpr-lpr/lpr mice. Characterization of the lymphokines produced and analysis of antigen-presenting cells required.

Lymph node cells from 4-wk-old MRL/Mp-lpr/lpr mice, but not from MRL/Mp-+/+ mice, when cultured in vitro for 5 to 7 days, will spontaneously proliferate and produce IL-2. We examined the expression of several cell surface Ag on lymph node cells from MRL/Mp-lpr/lpr mice before and after in vitro culture. There is an increase in the expression of Thy-1, L3T4, IL-2R, T cell activating protein, T cell receptor, and T3 complex on the surface of cultured cells. Cultured cells produced IL-3, IFN-gamma, and small but detectable amounts of IL-1 in addition to IL-2. Gamma irradiation of APC from young MRL/Mp-lpr/lpr mice or treatment of APC with a mAb (J11D) and C, completely abrogated their stimulatory capacity. These experiments suggest that B cells are the predominant APC responsible in the activation of autoreactive T cells in MRL/Mp-lpr/lpr mice. Lymph node cells from C57BL/6-lpr/lpr or C3H-lpr/lpr mice were unable to spontaneously proliferate or produce IL-2. Lymph node cells from (MRL/Mp-lpr/lpr x C57BL/6-lpr/lpr) F1 mice or (C3H-lpr/lpr x MRL/Mp-lpr/lpr) F1 mice did proliferate and produced IL-2 after in vitro culture. Using T cells from these F1 animals and APC from each parental haplotype, we found that APC from MRL/Mp-lpr/lpr mice induced more proliferation and greater amounts of IL-2, when compared to APC from F1 animals. APC from C57BL6-lpr/lpr mice or C3H-lpr/lpr were unable to induce spontaneous proliferation and IL-2 production. Therefore, B cells from MRL/Mp-lpr/lpr mice appear to possess unique features that enable them to activate autoreactive T cells more effectively than B cells from other mice bearing the lpr/lpr gene.

Animals↗

PubMed1988

Abnormal T cells from MRL/Mp-lpr/lpr mice do not respond to anti-T-cell-receptor antibody or tumor promoter and calcium ionophore A23187 despite the presence of the T-cell-receptor complex and protein kinase c.

Abnormal T cells from autoimmune MRL/Mp-lpr/lpr mice express T-cell-receptor complexes on their surfaces. These receptors are nonfunctional, since monoclonal anti-T-cell-receptor antibody-conjugated beads, which normally activate receptor-bearing T cells, were unable to activate these abnormal T cells. In addition, these abnormal T cells are unresponsive to the synergistic effect of phorbolmyristate acetate (PMA) and calcium ionophore A23187, as quantitated by proliferation, interleukin-2(IL-2) production, and the expression of IL-2 receptors. The failure of abnormal T cells to respond to PMA is not due to the absence of PMA receptors since Scatchard plot analysis reveals that there are 1-1.5 X 10(5) PMA-binding sites/cell with a Kd of 6-10 nM on these abnormal T cells. Similar to normal T lymphocytes, protein kinase c activity can be readily detected in the cytosolic fraction of these abnormal T cells. More importantly, in vitro culture of the abnormal T cells with PMA results in translocation of protein kinase c activity from the cytosolic fraction to the membrane fraction. From these experiments we concluded that the defect in the signal-transducing machinery in MRL/Mp-lpr/lpr T cells is not due to the lack of PMA receptors or the absence of protein kinase c activity, but may result from events which occur after the activation and translocation of protein kinase c. However, whether defects in response to a physiological stimulus (i.e., anti-receptor antibody) could occur in a step prior to protein kinase c activation remains to be determined.

Animals↗

PubMed1988

Cytolytic activity of Ia-restricted T cell clones and hybridomas: evidence for a cytolytic mechanism independent of interferon-gamma, lymphotoxin, and tumor necrosis factor-alpha.

Ten different helper T cell (Th) hybridomas that are specific to Ia or antigen plus Ia were found to express nonspecific cytolytic activity toward the cytotoxin (CT)-resistant P815 cells upon activation with either Con A or a monoclonal anti-T3 antibody (T3-mAb). In contrast to cytolytic Th1 clones which secrete high levels of interferon-gamma (IFN-gamma) and cytotoxin (CT) (lymphotoxin (LT, also known as TNF-beta) or tumor necrosis factor-alpha (TNF-alpha], these Th hybridomas produce low or undetectable levels of IFN-gamma and CT. No inhibitory activity of IFN-gamma and CT was observed in culture supernatants of activated Th hybridomas. Double-chamber experiments demonstrated that CT-sensitive L929 cells when physically separated from activated Th1 clones were killed by membrane-permeable CT. Under identical experimental conditions, lysis of P815 cells did not occur. Moreover, activation of Th hybridomas directly in wells containing the CT-sensitive L929 cells failed to induce target cell lysis. This confirms that these Th hybridomas produce little CT and argues against high local concentrations of CT being responsible for Th hybridoma-mediated killing of P815 cells. Finally, a polyclonal rabbit antiserum to rTNF-alpha, which strongly and specifically inhibited CT-mediated and Th1 clone-mediated killing of L929 cells, failed to inhibit P815 lysis by activated Th1 clones and Th hybridomas. These observations establish that a cytolytic mechanism independent of IFN-gamma, LT, and TNF-alpha is responsible for lysis of CT-resistant target cells.

Antibodies, Monoclonal↗

PubMed1987

An analogy between fetal haptoglobin and a potent immunosuppressant in cancer.

In spite of the numerous reports indicating the presence of humoral immunosuppressive factors in cancer patients, only a few of these factors have been biochemically identified. Furthermore, their role as effective immunosuppressors in vivo remains to be established. Our laboratory has attempted to isolate and identify the major immunosuppressive factor in the malignant effusions derived from ovarian and lung cancer patients. We have previously demonstrated that the Mr 52,000 immunosuppressive factor isolated from the ascites fluid of an ovarian cancer patient inhibited T-dependent immune responses in vivo and in vitro including the inhibition of E-rosetting. Thus, this immunosuppressive factor was named "suppressive E-receptor" (SER). Our current study demonstrates that this SER factor purified from malignant effusions derived from ovarian, lung, or head and neck cancer patients had a common component which dissociated equally into Mr 38,000-42,000 and 17,000-19,000 moieties on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under vigorous reducing conditions. Electroelution of these two components followed by a limited amino acid sequence determination revealed these two components to have N-terminal amino acid sequences identical to the beta and alpha 2 subunits of normal adult haptoglobin. Immunoelectrophoresis of SER using a polyclonal antiserum to neonatal cord blood demonstrated that SER, unlike normal haptoglobin, has slower electrophoretic mobility than the normal adult haptoglobin. Western blotting analysis of SER separated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under denaturing conditions failed to recognize a monoclonal antibody directed specifically to SER. However, this monoclonal antibody exclusively reacted with the SER separated by an analytical polyacrylamide gel electrophoresis gel under nondenaturing conditions while normal adult haptoglobins or purified but denatured haptoglobin obtained from the same malignant fluid as SER all failed to react with this antibody. Thus, SER appears to bear an additional epitope(s) that is absent in normal adult haptoglobin. Since the SER as well as the neonatal haptoglobin have at least 100 to 1000-fold more potent immunosuppressive activity than the normal adult haptoglobin, this additional epitope(s) present in SER may be responsible for the potent immunosuppressive property of SER.

Amino Acid Sequence↗

PubMed1987

Analysis of T cell responses to poly-L(GluLys) at the clonal level. I. Presence of responsive clones in nonresponder mice.

The synthetic random copolymer of L-glutamic acid and L-lysine (GL) is weakly or nonimmunogenic in all inbred strains of mice. Theories proposed to account for nonresponsiveness to GL include a deficient T cell repertoire, failure of antigen-presenting cells to present the antigen and/or the presence of suppressor cells. In this study we examine mechanisms for nonresponsiveness to GL. We demonstrate the existence of GL-reactive T cells which can be isolated with a relatively high frequency. These clones, which were derived following immunization of H-2d mice with poly(LGluLLysLTyr), also respond to several GL-containing polypeptides including the terpolymers of GL with phenylalanine, alanine (GLA) or leucine. Although recognition of GLA by heterogeneous T cell populations usually occurs in association with I-A determinants, these clones recognize GLA, as well as the other GL-containing polymers, in association with I-E determinants. Analysis of the antigen and alloreactivity patterns of these clones indicated that they expressed distinct antigen receptors. These studies imply that the T cell repertoire of "nonresponder" H-2d mice includes multiple GL-reactive T cell clones and that the antigen-presenting cells of these mice are effective in processing and presenting GL.

Animals↗

PubMed1987

Selective activation of helper and cytolytic T-cell functions of L3T4+ clones with either antireceptor antibody or phorbol ester and ionophore.

After activation with specific antigen and antigen presenting cells (APC) L3T4+ inducer T-cell clones can lyse Ia+ APC. The present study characterizes the mechanism of activation and specificity of L3T4+ inducer cell-mediated cytolytic function. Two methods that bypass the physiological stimulus of antigen presented on Ia+ APC were used to activate L3T4+ clones. The first method utilized an antireceptor monoclonal antibody (MAb), KJ16.133, to activate KJ16.133+ clones. The activated clones expressed nonspecific cytolytic activity, killing target cells irrespective of their H-2 haplotype or their ability to express cell surface Ia molecules. The crosslinking of bound KJ16.133 antibody greatly enhanced cytolytic activity. This activation is receptor specific because KJ16.133- clones were not activated under identical conditions. The second method of activation was provided by a synergistic action of phorbol-12-myristate-13-acetate (PMA) and ionophore A23187. These agents nonspecifically activated all L3T4+ clones tested. The simultaneous presence of the two agents is required for maximal activation. Again, the activated clones expressed potent nonspecific cytolytic activity. These observations demonstrated that L3T4+ inducer T-cell-mediated killing can be separated into two stages: an activation step, which can be specifically and nonspecifically triggered and an effector phase which causes nonspecific lysis of bystander targets. The induction of nonspecific cytolytic activity by antireceptor MAb was inhibited by anti-L3T4 MAb (GK1.5). In contrast, activation of nonspecific cytolytic activity by treatment with PMA plus A23187 was not inhibited by anti-L3T4 MAb. Under the above activation conditions, antireceptor MAb selectively induced the secretion of IL-3 and expression of nonspecific cytolytic activity. However, there was little or no concomitant proliferation and production of IL-2. In contrast, activation by PMA plus A23187 coordinately induces expression of nonspecific cytolytic activity, secretion of lymphokines (IL-3 and IL-2), and cell proliferation. Thus, the anticlonotypic activation preferentially induces certain functions whereas activation with PMA plus A23187 is not selective.

Animals↗

PubMed1986

Idiotype-specific helper cells (BH) express immunoglobulin markers and employ T cell function-associated molecules.

An Lyt-1+ population, distinct from T cell subsets, that helps expression of B cell responses to the 4-hydroxy-3-nitrophenyl acetyl (NP) hapten was characterized. This lymphoid population, called BH, is present in the spleens of normal and athymic mice and preferentially helps the expression of plaque-forming B cells that carry NPb idiotypic determinants. To define the mechanism by which this cell population functions, the roles of T and B lymphocyte function associated antigens were studied. The data indicate that BH cells express immunoglobulin receptor components, i.e., IgM, IgD heavy chain, and lambda light chain immunoglobulin markers as well as the J11d marker associated with immature B cells. BH cells may also express determinants identical to or cross-reactive with the T cell-associated antigens L3T4a, L3T4b, and LFA-1 as defined by treatment with monoclonal antibodies specific for these antigens. In addition, L3T4a- and LFA-1- but not Lyt-1-like antigens appear to be functionally involved in BH-dependent helper activity, since augmentation of NPb idiotypic PFC responses was blocked with anti-L3T4a or anti-LFA-1 monoclonal antibodies. Further analysis of BH-containing populations indicates that T cells are probably not involved in BH cell function and therefore are not responsible for the presence of Lyt-1, L3T4a, or L3T4b determinants in this T-independent system. The relationship of this helper cell subset to conventional T and B cell populations is discussed.

Animals↗

PubMed1986

Activation requirements of cloned inducer T cells. III. Need for two stimulator cells in the response of a cloned line to Mls determinants.

To gain insight into the nature of Mls determinants, we examined the stimulator cells responsible for the activation of inducer T cell clones by Mls determinants. Two types of clones responding to Mls determinants were identified. One type responded to purified B cells, but not to splenic adherent cells (SAC), from mice bearing Mls stimulatory determinants. The other type of Mls-reactive T cell clone, including the representative clone Ly1-N5, demonstrated a vigorous response to unfractionated spleen cells, but showed little or no response to B cells alone or to SAC alone from mice bearing the Mlsa or Mlsd stimulatory determinant. The response of these clones to Mls determinants required stimulation by two cell types. The failure of clone Ly1-N5 to respond to Mlsa-bearing B cells was reversed by the addition of SAC taken from mice bearing the Mlsa allele. In addition, SAC from mice bearing the nonstimulatory Mlsb allele could synergize with B cells from Mlsa-bearing animals. B cells were required to provide the Mlsa determinant, because the combination of Mlsa-bearing SAC and Mlsb-bearing B cells did not activate the clone. The response of clone Ly1-N5 to Mls is restricted by Ia determinants (shared by H-2b, H-2d, and H-2k haplotypes but not by the H-2q haplotype). The permissive H-2 alleles can be present either on the stimulator B cell or on the SAC. The optimal response of the clone was obtained by using B cells bearing Mlsa and the permissive Ia epitopes. However, a significant response of the clone to B cells bearing Mlsa but an inappropriate Ia (Iaq) was also seen in the presence of SAC bearing the nonstimulatory Mlsb allele but the permissive Ia epitopes.

Alleles↗