bluuebunny/crossref_metadata_2025 huggingface dataset:
Purified lymphocyte function-associated antigen-3 (LFA-3) activates human thymocytes via the CD2 pathway.
Authors: Denning S M , Dustin M L , Springer T A , Singer K H , Haynes B F
Published: December 2022

Defining the cellular and molecular mechanisms of interaction of developing thymocytes with nonlymphoid cells of the thymic microenvironment is critical for understanding normal thymus function. We have previously shown that the CD2/LFA-3 adhesion pathway is important in the interaction of thymocytes with a variety of LFA-3+ nonlymphoid thymic microenvironment cell types. Moreover, T cell activation via the CD2 (alternative, Ag independent) pathway is considered an important mechanism for intrathymic T cell proliferation. To study the relevance of CD2/LFA-3 interactions to human thymocyte activation, we have used purified LFA-3 Ag in several in vitro assays of thymocyte proliferation. Whereas LFA-3 Ag alone did not induce thymocyte proliferation, LFA-3 Ag in combination with the anti-CD2 antibody, CD2.1, and rIL-2 induced marked thymocyte proliferation. Additionally, the anti-CD28 antibody, Kolt2, could substitute for rIL-2, resulting in thymocyte activation induced by LFA-3 Ag in combination with antibodies CD2.1 and Kolt2. In both triggering systems, LFA-3 induced thymocyte activation was dependent upon the concentration of LFA-3 Ag. LFA-3 Ag-dependent thymocyte activation was directed primarily toward CD1-, mature thymocytes. Finally, intact SRBC that express the sheep homolog of LFA-3, T11TS, in combination with antibody CD2.1 and rIL-2 could also induce thymocyte activation. These data suggest that interaction of LFA-3 molecules with thymocyte CD2 molecules may provide a component of the stimulus for normal intrathymic thymocyte activation leading to thymocyte proliferation.
Specific immune regulation of chronic-relapsing experimental allergic encephalomyelitis in mice.
Authors: Kennedy M K , Dal Canto M C , Trotter J L , Miller S D
Published: December 2022

These studies were designed to examine immunologic means of regulating the clinical course of murine chronic-relapsing experimental allergic encephalomyelitis (R-EAE). We asked whether induction of specific immune tolerance to the major CNS myelin proteins, myelin basic protein (MBP) and proteolipid protein (PLP), could inhibit the development of R-EAE. Neuroantigen-specific tolerance was induced in SJL/J mice in a dose-dependent manner by the i.v. injection of mouse spinal cord homogenate-coupled syngeneic splenocytes (MSCH-SP) on day -7 relative to immunization on days 0 and +7. Sham-tolerized controls developed significant MBP- and PLP-specific DTH responses before the onset of clinical R-EAE. In contrast, MSCH-SP tolerized mice exhibited a dramatically reduced incidence of clinical and histologic signs of disease which correlated with the failure to develop MBP- and PLP-specific DTH responses. In 10 separate experiments, 118/149 (79%) of control mice, but only 22/137 (16%) of tolerized mice developed clinical R-EAE. Tolerance took time to develop and lasted at least 4 wk as mice injected with Ag-coupled splenocytes on day -1 relative to immunization remained susceptible to R-EAE, whereas mice injected on days -7, -14, or -28 were resistant. Tolerance induction required neuroantigens as injection of splenocytes coupled with a syngeneic mouse kidney homogenate failed to significantly alter the incidence of R-EAE or the development of neuroantigen-specific DTH responses. Thus, induction of R-EAE can be specifically and significantly regulated after the i.v. injection of splenocytes coupled with a crude, heterogeneous mixture of neuroantigens (i.e. MSCH).
Development of an antigen-specific CD8 suppressor effector clone in man.
Authors: Takeuchi T , Schlossman S F , Morimoto C
Published: December 2022

A long-term cultured IL-2-dependent keyhole limpet hemocyanin (KLH)-specific CD8 (T8) suppressor clone (5B9) was generated from a healthy donor hyperimmunized with KLH. The 5B9 clonal population suppressed in vitro anti-KLH antibody response but did not suppress anti-TT antibody response or PWM-driven IgG synthesis. Moreover, 5B9 cells could not suppress anti-TT antibody response even in the presence of free KLH. 5B9 cloned cells suppressed the anti-KLH antibody response of B cells cultured with CD4+4B4+ cells without requiring the presence of CD8+ cells. This KLH-specific CD8 suppressor clone is an effector type rather than an inducer type of suppressor T cell. The cloned cells expressed alpha- and beta-TCR proteins (defined by WT-31 antibody) on their cell surface. More importantly, the CD3:TCR complex was functionally important in the suppression induced by this clone, because after CD3 antigen modulation from its cell surface, the suppressor effector function was abolished.
Modulation of experimental allergic encephalomyelitis with anti-T suppressor factor antibodies.
Authors: Perry L L , Barzaga M E , Ferguson T A
Published: December 2022

mAb reactive with T suppressor factors (TsF) were used to alter the course of myelin basic protein-induced experimental allergic encephalomyelitis in (SJL/J x PL/J)F1 mice. In vivo administration of mAb 14-12, reactive with effector TsF, exacerbated the clinical expression of encephalomyelitis as evidenced by prolonged periods of total limb paralysis in affected animals. This aggravation of disease signs is probably related to the inhibition of effector Ts function by mAb 14-12 thus allowing T cell autoreactivity to proceed unchecked. Disease course was influenced more favorably by i.v. administration of mAb 14-30 reactive with a subset of inducer TsF. Ten days of treatment with this mAb resulted in a reduction in the incidence and severity of disease, noted as the development of minimal limb weakness but no paralysis in the majority of affected animals. Adoptive transfer experiments revealed the presence of Ag-specific Ts in mAb 14-30-treated mice that inhibited recipient Lyt-1+ responses to myelin basic protein, the immunizing autoantigen. Suppression by transferred Ts was revealed only by treatment of the donor population with anti-Lyt-1.2 plus C, however, indicating a role for contrasuppressor activity in the regulation of autoimmune T cell function. Results are considered relevant to the potential for immunotherapeutic management of multiple sclerosis in man.
The selection of effector T cell phenotype by contrasuppression modulates susceptibility to autoimmune injury.
Authors: Kelly C J , Mok H , Neilson E G
Published: December 2022

The genetic susceptibility to murine alpha TBM disease is a dominant trait that maps to H-2K. In previous studies we have shown that the critical difference between susceptible (SJL) and nonsusceptible (B10.S(8R] mice is the phenotype of the tubular Ag-specific effector T cells (TDTH). In SJL mice, these TDTH are Lyt-2+, whereas in B10.S(8R) mice the TDTH are L3T4+. These phenotypic differences have an important functional correlate: Lyt-2+ TDTH are nephritogenic, whereas L3T4+ TDTH are typically not nephritogenic. Both mouse strains have the potential to differentiate both L3T4+ and Lyt-2+ TDTH. The preferential selection of a single TDTH phenotype in each is the result of differential T cell regulation. In the present studies, we have examined the contribution of suppressor and contrasuppressor T cells in the regulation of TDTH phenotype selection. Our studies show that in both SJL and B10.(8R) mice, after exposure to Ag, a suppressor T cell subpopulation functions to inhibit the nephritogenic Lyt-2+ TDTH. In SJL, but not B10.S(8R) mice, this suppression is counterbalanced by Lyt-2+, Vicia Villosa lectin-adherent T cells. Such contrasuppressor function is mediated through a T cell-derived soluble protein (TcsF), which is Ag-binding and recognized by alpha I-JS antisera. This functional TcsF activity maps, as does susceptibility to disease, to H-2K. In the presence of genetically compatible TcsF, the TDTH phenotype in nonsusceptible mice switches to that of susceptible mice. These Lyt-2+ TDTH from nonsusceptible mice are fully capable of inducing tubulointerstitial nephritis following adoptive transfer. Our studies describe a new role for Tcs cells and augment our understanding of their etiopathogenetic role in autoimmunity.
Production of human bone marrow-derived suppressor factor. Effect on antibody synthesis and lectin-activated cell proliferation.
Authors: Mortari F , Singhal S K
Published: December 2022

Natural suppressor cells resident in normal human bone marrow (BM) exert potent suppressor activity on in vitro antibody responses and other immune functions. A suppressor-enriched population of BM cells can constitutively produce a soluble mediator with similar suppressor activity and kinetics as the suppressor cells. This novel BM-derived suppressor factor (BDSF) suppresses human in vitro primary antibody responses as well as lectin-activated proliferative responses. The mediator (BDSF) has a Mr of less than 1.5 kDa, contains a lipid component, and is insensitive to indomethacin treatment. The BM cells producing the factor bear the HNK-1 surface marker but not T, B, or macrophage markers. The ability of BDSF to suppress Ag-dependent IgM responses during the inductive phase makes it an ideal molecule with the potential to regulate early immune and hemopoietic events within the BM compartment.
Production of B cell-stimulating factors by B cells in patients with systemic lupus erythematosus.
Authors: Tanaka Y , Saito K , Shirakawa F , Ota T , Suzuki H , Eto S , Yamashita U
Published: December 2022

The production of B cell-stimulating factors (BSF) by B cells in patients with systemic lupus erythematosus (SLE) was studied in vitro. B cells from SLE patients markedly proliferated and differentiated into Ig-producing cells by in vitro culture without any stimulation. The culture supernatant of these B cells contained BSF activity that stimulated Staphylococcus aureus Cowan I-treated normal B cells to proliferate and differentiate into Ig-producing cells. By a Percoll gradient density centrifugation, BSF-producing cells were enriched in the higher density fraction, but were reduced in the lower density fraction. The BSF also stimulated the proliferation and the differentiation of SLE B cells. By a Percoll gradient density centrifugation, SLE B cells responsive to the BSF were enriched in the higher density fraction, but were reduced in the lower density fraction. The Mr of the BSF was estimated as about 18,000 Da by Sephacryl S-200 column chromatography. The BSF fraction did not possess IL-2 and IFN activity, but possessed IL-1 activity, which stimulated murine thymocyte proliferative responses. The BSF activity was partially, but not completely, absorbed by an anti-IL-1 alpha antibody. Furthermore, the BSF possessed IL-4 activity, which induced not only the proliferative responses of normal B cells stimulated with B cell mitogens, but also the expression of low affinity Fc epsilon R/CD23 on normal B cells. The BSF also possessed IL-6 activity, which induced the proliferative responses of IL-6-dependent hybridoma cells, MH-60 BSF2. Moreover, human rIL-1, rIL-4, and rIL-6 stimulated SLE B cells. These results suggest that SLE B cells spontaneously produce the BSF such as IL-1 alpha, IL-4, and IL-6 and express their receptors on their surface, and the interaction between the BSF and their receptors stimulates SLE B cells to spontaneously proliferate and differentiate into Ig-producing cells as an autocrine mechanism.
Natural cytotoxic cell-specific cytotoxic factor produced by IL-3-dependent basophilic/mast cells. Relationship to TNF.
Authors: Richards A L , Okuno T , Takagaki Y , Djeu J Y
Published: December 2022

The murine IL-3-dependent mast cell line, PT18-A17, and the rat basophilic leukemia cell line, RBL-2H3, were found to mediate natural cytotoxic (NC) activity via the release of a soluble factor which specifically lysed NC-sensitive WEHI-164 but not NK-sensitive YAC-1 tumor cells. The release of this NC cell-specific cytotoxic factor was enhanced by triggering of both types of cells via IgE receptor bridging. This factor had activity on TNF-sensitive but not TNF-resistant cell lines and could be neutralized by two independently produced polyclonal anti-mouse TNF antisera. It was not neutralized by antibodies against mouse IFN-alpha/beta or IFN-gamma. Moreover, it was not neutralized by a monoclonal or a polyclonal anti-human TNF, demonstrating that the rodent TNF differed antigenically from human TNF. These results indicate that the cytotoxic factor released from a murine IL-3-dependent mast cell line and from a rat basophilic leukemia cell line is immunologically and functionally related to murine TNF.
The IL-4 gene maps to chromosome 11, near the gene encoding IL-3.
Authors: D'Eustachio P , Brown M , Watson C , Paul W E
Published: December 2022

IL-4/B cell stimulatory factor 1 (IL-4) is a potent mediator of the growth and differentiation of cells of most hemopoietic lineages. IL-4 is one of a number of lymphokines produced by T cells after activation with Ag or mitogen. In order to map the chromosomal location of the IL-4 gene, Chinese hamster-mouse somatic cell hybrids were used in Southern blot analyses with an IL-4 cDNA probe. These results suggested that the IL-4 gene was located on chromosome 11. In contrast, the gene encoding IL-2 was localized to either chromosome 1 or 3. The identification of a strain-specific Bgl II restriction enzyme polymorphism in the IL-4 gene was used to map the IL-4 gene to a position on mouse chromosome 11 within 1 centimorgan of the gene encoding IL-3.
Detection of IL-1 alpha and IL-1 beta gene expression by in situ hybridization. Tissue localization of IL-1 mRNA in the normal C57BL/6 mouse.
Authors: Takács L , Kovacs E J , Smith M R , Young H A , Durum S K
Published: December 2022

IL-1 is a cytokine with a wide variety of effects on cells involved in inflammatory and immune responses, hemopoiesis, and bone formation. Many cell types have been shown to produce IL-1 in vitro; however, very little is known about the source and role of IL-1 in vivo. By using in situ hybridization, we examined the tissue distribution of cells containing IL-1 mRNA in normal C57BL/6 mice. The results show that many organs contain IL-1 mRNA-positive cells, but the highest frequency was found in lymphoid organs. The distribution and localization of these cells suggest that many of the IL-1 mRNA-producing cells are tissue macrophages. Organs exposed to environmental Ag and microbial products (lymph nodes, liver, intestine, lung, and uterus) had high frequencies of IL-1 mRNA-producing cells, suggesting that IL-1 is produced in local inflammatory or immune responses in vivo. The production of IL-1 mRNA in the thymus and in the bone marrow suggests that IL-1 is available to play physiologic roles in T cell differentiation and in hemopoiesis.
HLA-DQ molecules form alpha-beta heterodimers of mixed allotype.
Authors: Kwok W W , Schwarz D , Nepom B S , Hock R A , Thurtle P S , Nepom G T
Published: December 2022

Retroviral vectors with an internal cytomegalovirus major immediate-early gene enhancer/promoter regulating HLA class II gene expression were used to transfer HLA cDNA into human EBV-transformed B-lymphoblastoid cell lines. HLA-DQ2 beta and DQ3.2 beta cDNA were transferred into DQ3.2 and DQ2 homozygous lymphoblastoid cell lines, respectively. Serologic analysis of the infected cell lines with allospecific mAb demonstrated surface expression of these exogenous DQ molecules implying that DQ alpha-chains from DR3, DQ2-positive cells can pair with DQ3.2 beta-chains and, similarly, DQ alpha-chains from DR4, DQ3.2-positive cells can pair with DQ2 beta-chains. Immunoprecipitation of the introduced DQ3.2 beta molecule resulted in co-purification of the allotype-mismatched endogenous DQ2 alpha polypeptide. We also show that vectors with a cytomegalovirus major immediate-early gene enhancer/promoter result in higher levels of expression of the transduced gene compared to previously described HLA vectors with either the SV-40 early enhancer/promoter or the lymphotropic papovavirus-enhanced SV-40 promoter. Although deletion of HLA cDNA did sometimes occur in the process of generating virus-producing clones, the HLA cDNA is stably maintained in virus-producing clones, once it is generated. This retroviral expression system is a highly efficient way to study HLA gene function.
Inhibition of human natural killer cell activity by the protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine is an early but post-binding event.
Authors: Steele T A , Brahmi Z
Published: December 2022

Recent evidence has demonstrated a protein kinase C (PKC)-dependent step in cytotoxic T lymphocyte activation. Here, we examined the influence of PKC in the lytic response of human NK cells to K562, an NK-sensitive tumor target cell. We used the known protein kinase inhibitors 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7) and HA1004. H-7 caused a dose-related inhibition of NK cell-mediated cytolysis (CMC) when the inhibitor was present throughout the course of the 3-h chromium release assay. The 50% inhibitory concentration for H-7 was 7 microM. In contrast, HA1004, which exerts a greater inhibitory effect on cyclic nucleotide-dependent protein kinases than PKC, had no effect on NK-CMC. The suppression of NK-CMC by H-7 was not due to inhibition of binding of the effector cells to target cells and could be reversed by the addition of PMA. H-7 was most effective in abrogating NK-CMC when added to the assay within the first 30 min and treatment of the effector and target cells with H-7 resulted in no loss of NK-CMC. Because nearly 50% of the normal NK lytic activity had taken place by 30 min, this suggested that H-7 inhibited an early event. H-7 exerted a dose-related suppression of antibody-dependent cell-mediated cytotoxicity (ADCC) suggesting that NK-CMC and ADCC share the utilization of PKC, however, HA1004 did not inhibit ADCC. Treating NK cells with IL-2 or IFN-beta did not overcome the inhibition of NK-CMC by H-7. In this study, we have thus demonstrated the presence of a PKC-dependent step in NK-CMC and ADCC.
Inhibition of lipopolysaccharide activation of 7OZ/3 cells by anti-lipopolysaccharide antibodies.
Authors: Kirkland T N , Ziegler E J , Tobias P , Ward D C , Michalek S M , McGhee J R , Macher I , Urayama K , Appelmelk B J
Published: December 2022

We have investigated the ability of mAb against LPS to inhibit LPS-induced activation of 7OZ/3 pre-B cells. The fine specificity and relative affinity of these mAb for lipid A and LPS were also determined. We found that antibodies inhibited only the activity of glycolipids which they bound with relatively high affinity. However, two high affinity antibodies binding to non-lipid A epitopes did not block cellular activation. Some, but not all, relatively high affinity antibodies binding to the lipid A region of the LPS molecule inhibited biologic activity. The inhibitory antibodies bound to at least two distinct epitopes within the lipid A region. These data suggest that LPS interacts with 7OZ/3 cells in a highly specific fashion.
Influence of recombinant IL-4, IFN-alpha, and IFN-gamma on the production of human IgE-binding factor (soluble CD23).
Authors: Delespesse G , Sarfati M , Peleman R
Published: December 2022

This study documents the influence of rIL-4, IFN-gamma, and IFN-alpha on the production of IgE-BF and the expression of lymphocyte receptor for IgE or CD23 Ag (Fc epsilon R II) by human mononuclear cells. IL-4 increases the secretion of IgE-binding factor (BF) by highly purified B lymphocytes, adherent cells, and U937 monoblastic cells. The effect of IL-4 on purified B cells is augmented by costimulating the cells with F(ab')2 anti-IgM. IFN-gamma, IL-2, IL-1-alpha, or IL-1 beta and the low m.w. B cell growth factor have no effect on IgE-BF production by purified B cells even when they are used in combination with anti-IgM. Stimulation of purified T cells with IL-4 or IL-4 plus PMA leads to the production of very small amounts of IgE-BF that might well be derived from the contaminating non-T cells. IFN-gamma increases IgE-BF synthesis by unfractionated PBMC, T cell-depleted PBMC, adherent cells, and U937 cells suggesting that it induces monocytes to release IgE-BF, IFN-gamma suppresses the IL-4-induced Fc epsilon R II expression and IgE-BF production by highly purified B cells but not by PBMC or their T cell-depleted fractions. IFN-alpha inhibits IgE-BF production by IFN-gamma-stimulated PBMC and by IL-4-stimulated cells suggesting that it exerts its effect on B cells and on monocytes. Moreover IFN-alpha suppresses the IL-4-induced expression of Fc epsilon R II on B cells. Both IFN-alpha and IFN-gamma suppress the synthesis of IgE by PBMC in response to IL-4. Taken collectively the results indicate that: 1) IL-4 induces IgE-BF production by both B cells and monocytes, 2) IFN-gamma stimulates IgE-BF synthesis by monocytes but suppresses its production by IL-4-stimulated B cells, and finally 3) IFN-alpha inhibits IgE-BF synthesis in response to either IFN-gamma or IL-4.
Expression of IL-1 receptors on human peripheral B cells.
Authors: Tanaka Y , Shirakawa F , Oda S , Eto S , Yamashita U
Published: December 2022

The expression of IL-1R on human peripheral B cells was analyzed by the binding assay with 125I-labeled human rIL-1 alpha and by the flow cytofluorometry by the use of FITC-conjugated IL-1 alpha. The proliferation and the differentiation of B cells stimulated with Staphylococcus aureus Cowan I (SAC) in the presence of T cell-derived factors were dependent on IL-1. By the binding experiment with 125I-labeled IL-1 alpha, B cells expressed only few IL-1R without any stimulations. When they were stimulated with SAC, IL-1R on B cells began to increase by only 1 h, reached the maximum level at 6 h. The binding of 125I-labeled IL-1 alpha to B cells was inhibited by the addition of either cold IL-1 alpha or IL-1 beta suggesting that IL-1R on B cells reactive for IL-1 alpha and IL-1 beta were identical. By Scatchard plot analysis, the existence of two classes of IL-1R on B cells was found. A major class of IL-1R (320 molecules/cell) has a lower affinity (Kd = 3.8 x 10(-10) M) and a minor class of IL-1R (70 molecules/cell) has a higher affinity (Kd = 4.4 x 10(-12) M). When B cells were stimulated with SAC, both lower and higher affinity IL-1R were increased to 1960 molecules/cell and 300 molecules/cell, respectively. Furthermore, IL-1R on B cells were also detected with FITC-conjugated IL-1 alpha by a flow cytofluorometer. Only 3 to 5% of B cells expressed IL-1R without any stimulations. When B cells were stimulated with SAC, IL-1R-positive B cells were increased to 20%. The addition of anti-class II antibodies inhibited B cell proliferation and differentiation induced with SAC, IL-1, and T cell-derived factors. Anti-class II antibodies also inhibited the number of IL-1R on B cells. These results suggest that the expression of IL-1R was induced as the initial stage of B cell activation and that class II Ag play an important role for the expression of IL-1R on B cells.
Lymphocytes stimulated by allogeneic B cell lines cleave the third component of complement and fix C3 fragments. Their nonspecific lytic capacity is elevated against complement receptor type 2-carrying targets.
Authors: Ramos O F , Algarra I , Sármay G , Yefenof E , Gergely J , Klein E
Published: December 2022

Human blood lymphocytes stimulated in mixed cultures by allogeneic B cell lines were shown to cleave C3 molecules. The B cell lines were derived from Burkitt lymphoma patients: 1) established from their EBV-negative lymphoma, 2) the EBV-positive sublines converted in vitro, and 3) lymphoblastoid cell lines (LCL) i.e., B lymphocytes transformed in vitro by EBV. These cell lines differed considerably in their capacity to stimulate allogeneic lymphocytes. The split products of C3 were detected in the supernatants and on the surface of the activated lymphocytes at levels which correlated with the strength of stimulation. Lymphocytes cultured with LCL had the highest levels of thymidine incorporation blast transformation, C3 cleavage, and C3 fragment fixation. Lymphocytes exposed to the EBV-negative Burkitt lymphomas were stimulated weakly and their C3-activating capacity was low. Irrespective of the efficiency of lymphocyte stimulation induced in the cultures, 60 to 70% of the blasts were found to fix C3 fragments. The majority of the lymphocytes which fixed C3 fragments were T blasts that carried the CD3 marker and expressed IL-2R (CD25). CD4 and CD8 cells were represented with equal frequency in the C3-fragment fixing and C3-fragment negative populations. Pre-exposure of the MLC-activated lymphocytes to human serum increased their cytotoxic capacity toward CR type 2-carrying targets. The enhanced lysis was abrogated by F(ab)2 rabbit anti-human C3d or rabbit anti-CR type 2 antibodies. The results suggest that the C3 fragments fixed on the lymphocytes bind to CR on the targets and elevate the avidity of binding between the two interacting cells. This was also indicated by an increase in the frequency of lymphocyte-target conjugates.
Antigen-triggered membrane potential changes in IgE-sensitized rat basophilic leukemia cells: evidence for a repolarizing response that is important in the stimulation of cellular degranulation.
Authors: Labrecque G F , Holowka D , Baird B
Published: December 2022

We have studied Ag-induced membrane potential changes of rat basophilic leukemia cells by using the potential-sensitive dye, bis-(1,3-diethylthiobarbiturate)trimethineoxonol. A rapid membrane depolarization is triggered by a multivalent Ag, and it has a bell-shaped dose dependence that parallels the degranulation response but not the extent of cross-linking of the IgE-receptor complexes. As the temperature is reduced from 37 degrees C, this depolarization response slows and decreases in magnitude until complete inhibition is observed at 15 degrees C, similar to the temperature dependence previously observed for the Ag-stimulated rise in cytoplasmic Ca2+ and for degranulation. The results imply that a highly temperature-dependent step subsequent to Ag binding and cross-linking is necessary for the depolarization response. A partial return to the resting potential is seen to follow the depolarization response to Ag. This repolarization process is inhibited by quinidine.HCl and Ba2+ in parallel with an inhibition of the degranulation response. Repolarization is not affected by 4-aminopyridine or by the absence of K+ in the external buffer. These data suggest that the repolarization is caused by a previously uncharacterized K+ channel.
A virus-specific CD4+ cell-mediated cytolytic activity revealed by CD8+ cell elimination regularly develops in uncloned human antiviral cell lines.
Authors: Bourgault I , Gomez A , Gomard E , Picard F , Levy J P , Gomrad E
Published: December 2022

Antiviral HLA class II-restricted cytotoxic CD4+ clones have been relatively well characterized in vitro but their significance in the immune response remains unknown. Here anti-influenza A and anti-EBV CD4+ CTL have been studied by using permanent cell lines either untreated or depleted of CD8+ cells. In bulk cultures, HLA class I-restricted anti-viral CD8+ CTL account for all of the detectable killer cell activity, whereas after elimination of CD8+ cells an HLA class II-restricted killer activity mediated by CD4+/2H4-/4B4+ cells was consistently observed. The CD4+ CTL were fully differentiated in all of the cultures tested from the third in vitro passage because they could be demonstrated immediately after elimination of CD8+ cells. These CD4+ killer cells were equivalent to the CD8+ cells in terms of their lytic capacity. The absence of any class II-restricted antiviral activity in bulk cultures seems to be related to the very small numbers of CD4+ cells present in these antiviral cell lines. However, CD4+ cytolytic activity could not be detected during the first two in vitro passages, even when limiting dilution analysis of the CTL precursors were performed, showing that the killer function of Th cells differentiate only after several in vitro stimulations.
A novel gamma delta T cell receptor for antigen adds limited diversity to the gamma delta repertoire in adult thymus.
Authors: Marusić-Galesić S , Saito T , Tentori L , Zuñiga-Pflucker J , Raulet D H , Allison J P , Kruisbeek A M
Published: December 2022

The surface expression of CD3-associated TCR chains on hybridoma cell lines derived from adult gamma delta thymocytes was analyzed. These cell lines were unusual, in that a) they expressed a surface heterodimer consisting of a 40- and a 42-kDa chain, i.e., comprised of chains different from any previously reported gamma delta-TCR all of which express C gamma 1- or C gamma 2-encoded gamma-chains; b) their CD3-associated TCR could not be categorized as alpha beta-TCR dimers, despite the similarities in m.w. of the TCR chains, because full size 1.3-kb beta-chain mRNA capable of encoding a functional beta-chain could not be detected in these cells; c) neither of the receptor chains could be precipitated with anti-C gamma 1C gamma 2-peptide antisera. Biochemical analysis demonstrated that the 42-kDa delta-chain is a novel chain, which differs from any reported delta-chains in size, charge and number of glycosylation sites. Collectively, the data on analysis of the 40-kDa chain strongly suggest that it represents a gamma-chain encoded for by the C gamma 4 locus, protein products of which have not yet been reported in the thymus. This gamma-chain was also unique, in that its isoelectric point was much lower than that of other gamma-chains. The gamma- and delta-chains on these C gamma 4-expressing hybridomas were indistinguishable from one another in size and charge (as determined by nonequivalent pH gradient electrophoresis/SDS-PAGE analysis and analysis after endoglycosidase treatment). Because the cell lines were randomly chosen from large panels of hybridomas, these results may well imply strikingly nonrandom pairing of thymocyte-derived C gamma 4 chains and the delta-chains reported here. Thus, only limited additional gamma delta repertoire diversity may be generated by availability of this gamma delta-TCR in the thymus.
The common acute lymphoblastic leukemia antigen gene maps to chromosomal region 3 (q21-q27).
Authors: Barker P E , Shipp M A , D'Adamio L , Masteller E L , Reinherz E L
Published: December 2022

Complementary DNA and genomic clones corresponding to the gene for the common acute lymphoblastic leukemia antigen (CALLA) were used to investigate the genetic structure and location of the CALLA locus. The gene, which encodes a 100-kDa type II transmembrane glycoprotein, appears to be a single copy locus of greater than 45 kb which is not rearranged in malignancies expressing cell surface CALLA. Cell hybrid analysis indicates that the CALLA-related DNA sequences are found on human chromosome 3. In situ hybridization studies reveal the regional location of the CALLA locus to be 3q21-27.
Restricted assembly of MHC class II molecules in transgenic mice.
Authors: Le Meur M , Waltzinger C , Gerlinger P , Benoist C , Mathis D
Published: December 2022

Studies on cell lines transfected with MHC class II genes have revealed important limitations on the assembly of haplotype-mismatched A alpha:A beta complexes. These findings led to the speculation that pairing restrictions, if applied in a cell type-specific fashion, might be involved in various autoimmune phenomena. We have investigated pairing restrictions in vivo by analyzing transgenic mice that carry an Ak alpha chain, an Ak beta chain, or the Ak alpha:Ak beta complex on an H-2b or H-2s background. Our conclusion is that the assembly of haplotype-mismatched A alpha:A beta complexes is limited in vivo, and that this is equally true for all cell types examined, regardless of their role in the immune response.
Studies of c-myb gene regulation in MRL-lpr/lpr mice. Identification of a 5' c-myb nuclear protein binding site and high levels of binding factors in nuclear extracts of lpr/lpr lymph node cells.
Authors: Mountz J D , Steinberg A D
Published: December 2022

Lymph node T cells of MRL-lpr/lpr mice are characterized by the production of very large amounts of c-myb mRNA. To study the control of c-myb expression, a search was made for sites on the 5' c-myb gene which could bind regulatory proteins. DNase I digestion of nuclear chromatin uncovered four DNase I hypersensitive sites in the first intron of the c-myb gene, and a single site approximately 300 bp 5' to the initiation codon. Lambda exonuclease digestion of a 5'-myb fragment in the presence of nuclear extracts from either MRL-lpr/lpr PLN or EL-4 thymoma revealed stop sites approximately 300 bp 5' (-271 to -322) to the ATG initiation codon. DNase I footprint analysis demonstrated a guanine-cytosine enriched region of potential binding sites (-274 to -319) in the region of the stop sites and a fifth potential binding site closer to the initiation codon (-163 to -168). Specific gel shift bands were detected by a 5'-myb fragment (-346 to -155) with extracts from a number of different lymphoid cell lines and the appropriate specific and non-specific competitor DNA. The DNA giving rise to these gel shift bands encompassed the region defined by the stop site and footprinting studies. To determine whether or not the protein binding to the 5' c-myb gene at -274 to -319 was associated with increased c-myb mRNA, we studied nuclear extracts of several cell lines and compared the amount of binding to the amount of c-myb mRNA found on Northern analyses. Among the cell lines, there was a correlation between c-myb expression and the amount of the 5'-myb DNA binding protein. In addition, MRL-lpr/lpr lymph node cells had high c-myb expression and large amounts of the 5'-myb binding protein. This result suggests that the binding may play some role in the c-myb expression. Moreover, the most immature cell lines had the greatest amount of the binding factor, suggesting that its regulatory effect on c-myb expression might be important in early differentiation events.
Induction of the H-2 D antigen during B cell activation.
Authors: Southern S O , Swain S L , Dutton R W
Published: December 2022

Mitogenic activation causes increased expression of class I Ag of the MHC in mouse B cells. The increased expression was seen in flow cytometry analysis for both K and D in k as well as d haplotypes. A more detailed molecular analysis was carried out for H-2Dd. Increased expression (10- to 20-fold) of the H-2 Dd gene was detected at both protein and messenger RNA levels, and the time course for the accumulation of H-2 Dd protein on the cell surface parallels the increase in the steady-state messenger RNA levels. The increase in H-2 Dd expression in small B cells stimulated with LPS is detectable after 10 h of culture. The present data provide molecular and serologic evidence about alterations in the expression of the H-2 Dd Ag, previously identified as a B cell activation antigen B7.2. Our results indicate a new significance for the function and regulation of the MHC during immune responses, and suggest that the class I molecules may serve some role in the B cell activation process.
Qa-2 expression in the adult murine thymus. A unique marker for a mature thymic subset.
Authors: Vernachio J , Li M , Donnenberg A D , Soloski M J
Published: December 2022

The MHC Ag, Qa-2, is expressed on all peripheral T cells, a subset of bone marrow cells, and to a lesser extent on B cells. The Qa-2 Ag is also expressed on 5 to 6% of normal adult murine thymocytes. Through the use of flow cytometry, counterflow centrifugal elutriation and acridine orange staining, we have analyzed the cell surface phenotype, cell size, and cell cycle status of this thymic population. Our studies indicate that Qa-2+ thymocytes are large, non-mitotic, G1 cells which have the cell surface phenotype of CD5+, CD3+, J11dLO and lack receptors for peanut agglutinin. This population can be further subdivided into three categories; CD4+/CD8-, CD4-/CD8+, and CD4-/CD8-. These data indicate that Qa-2 surface expression can only be detected on thymocytes in the final stages of differentiation. The Qa-2 Ag can be used as a cell surface marker to identify a unique subset of mature thymocytes.
Modulation of IL-2-induced human B cell proliferation in the presence of human 50-kDa B cell growth factor and IL-4.
Authors: Vazquez A , Mills S , Maizel A
Published: December 2022

Several human T cell derived factors capable of stimulating human B cells to synthesize DNA have been previously described. One such factor exhibits an apparent m.w. of 50,000 Da and has been termed 50-kDa-B cell growth factor (BCGF). In this report, we show that a human B cell proliferation pathway based on the sequential action of anti-mu antibody, 50-kDa-BCGF and IL-2 is inhibited in the presence of human rIL-4. Although IL-4 itself is capable of triggering B cell DNA synthesis as measured at 72 h, this lymphokine inhibits, in a dose-related manner, the 50-kDa-BCGF driven response of B cells to IL-2 when such proliferation is determined after 144 h. This inhibition takes place at an early step of the B cell activation and does not require the presence of IL-4 during the whole culture period. Such inhibitory activity of IL-4 is specific to the IL-2-induced B cell proliferation because DNA synthesis measured in the presence of semi-purified human 12-kDa-BCGF is not affected by the presence of IL-4. Our results suggest that a particular pathway of human B cell activation leading to the proliferation of these cells in the presence of IL-2 could be either up- or down-modulated by 50-kDa-BCGF and IL-4, respectively.
Course of transplanted Heymann nephritis kidney in normal host. Implications for mechanism of proteinuria in membranous glomerulonephropathy.
Authors: Makker S P , Kanalas J J
Published: December 2022

Heymann nephritis [model of membranous glomerulonephropathy (MGN)] kidneys (n = 20) with proteinuria were transplanted into unilaterally nephrectomized normal syngeneic Lewis recipient rats to study the course of established MGN lesion in normal milieu. Concurrent with the loss of C3 staining from the MGN lesion at 2 to 4 wk after transplantation, the proteinuria decreased from the transplanted kidney (p less than 0.02). Thereafter, from 2 to 28 wk no further decrease was noted in proteinuria which stabilized at a lower but still abnormal level. No appreciable decrease in IgG deposits in MGN lesion was noted up to 12 wk but at 28 wk the deposits had decreased significantly (p less than 0.005). However, the deposits did not resolve completely even at 40 wk. The results indicate that in MGN one component of proteinuria is due to ongoing activation of C with the deposition of new antibody and the other is perhaps due to structural damage to glomerular filter. The former appears reversible and the later irreversible. Although the resolution of IgG deposits in the lesion is very slow significant improvement can occur with time (several months).
A novel cell type responsible for marrow graft rejection in mice. T cells with NK phenotype cause acute rejection of marrow grafts.
Authors: Yankelevich B , Knobloch C , Nowicki M , Dennert G
Published: December 2022

Acute rejection of allogeneic and semiallogeneic marrow grafts has long been considered to be a function of the natural immune system because it shares many features with NK activity in mice. With the use of a recently developed in vivo adoptive transfer assay in which spleen cells are transferred from mice able to reject a particular marrow graft into mice that fail to do so, we show that the cells responsible for induction of marrow graft rejection indeed display the phenotype of NK cells: they lack the T cell Ag CD4 and CD8 but express the NK Ag NK1 and ASGM1. The rejection induced by adoptively transferred cells is exquisitely specific--a feature that points to a specific recognition process by the transferred cells. To elucidate what the recognition structure on these cells may be we found that they express CD3 and most likely the beta-chain of the TCR. Highly purified responder cells with the NK1+, CD3+, CD4-, CD8- phenotype, when transferred into nonresponder recipients, cause specific marrow graft rejection. We conclude that the acute rejection of bone marrow grafts is caused by a cell that expresses NK phenotype but is of T cell lineage. This may suggest the specificity of acute marrow graft rejection is caused by a specific recognition process that involves TCR.
Comparative effect of recombinant IL-1, -2, -3, -4, and -6, IFN-gamma, granulocyte-macrophage-colony-stimulating factor, tumor necrosis factor-alpha, and histamine-releasing factors on the secretion of histamine from basophils.
Authors: Alam R , Welter J B , Forsythe P A , Lett-Brown M A , Grant J A
Published: December 2022

Most cytokines possess multiple biologic activities. This study was undertaken to investigate the effect of rIL-1 beta, -2, -3, -4 and -6, IFN-gamma, TNF-alpha, and granulocyte-macrophage (GM)-CSF on basophils from 16 donors and the amount of histamine released was compared with that by partially purified mononuclear cell-derived histamine-releasing factor (HRF) and anti-IgE. We found that only IL-3 and GM-CSF at relatively high doses (50 to 500 ng/ml) released small amounts of histamine (3 to 14%) from two allergic donors. In contrast, both HRF and anti-IgE released significant amounts of histamine from all donors. Other cytokines did not release any measurable quantity of histamine. Simultaneous addition of several cytokines to the basophils also failed to release histamine. IL-3, GM-CSF, and IL-1 can also release histamine at lower concentrations (less than 5 ng/ml) when incubated with basophils in the presence of D2O. Basophils from 6 out of 13 allergic donors released histamine in response to IL-3, whereas three donors responded to IL-1 beta and two responded to GM-CSF. The results of this study demonstrated that although IL-3 and GM-CSF release small amounts of histamine only from a select group of allergic patients, mononuclear cell-derived HRF is more potent in their action and release histamine from normals as well as allergic patients.
Expression of a cross-reactive idiotope on naturally occurring murine antibodies against 3-fucosyllactosamine, levan, and dextran.
Authors: Kimura H , Umeda M , Marcus D M
Published: December 2022

We recently identified a cross-reactive Id (6C4) that is expressed on the H chain of many BALB/c mAb against the 3-fucosyllactosamine (3-FL) determinant, Gal(beta 1-4) (Fuc(alpha 1-3] GlcNAc-R. The VH segments of seven mAb that we recently sequenced are encoded by VH441, which also encodes VH segments of antibodies against galactan, levan, and dextran. To analyze the expression of the 6C4 Id on naturally occurring anti-carbohydrate antibodies, we isolated 6C4+ antibodies by affinity chromatography from pools of normal BALB/c serum. Approximately 20 to 30% of antibodies against 3-FL and levan, and all antibodies against dextran, were removed from the sera by passage over a column containing mAb 6C4. Absorption of the eluate with 3-FL beads removed anti-3-FL antibodies but not anti-dextran or anti-levan. The expression of a cross-reactive Id on naturally occurring antibodies against several carbohydrate Ag suggests that these antibodies may participate in an Id network. We also reported previously that BALB/c mice have naturally occurring anti-3-FL antibodies and respond well to immunization against this determinant, whereas C57BL/6 mice do neither. To examine the role of the Igh-C allotype in the regulation of the anti-3-FL response, we studied congenic strains of BALB/c and C57BL/6 mice. Both congenic strains produced anti-3-FL antibodies in response to immunization, but only C.B-20 mice exhibited naturally occurring antibodies. These data suggest that the naturally occurring and elicited antibody responses against 3-FL are differentially regulated.
Cessation of neutrophil influx in C5a-induced acute experimental arthritis is associated with loss of chemoattractant activity from the joint space.
Authors: Haslett C , Jose P J , Giclas P C , Williams T J , Henson P M
Published: December 2022

Neutrophil emigration is a critical component of the inflammatory process and is generally thought to play a role in host defense as well as in the tissue injury that often accompanies inflammation. Most inflammatory reactions exhibit a sequence of emigrating cell types, thus clearly demonstrating that the neutrophil influx eventually ceases and that the neutrophils are then removed from the lesion. It has been our premise that in order to understand the processes that lead to the progressive inflammatory reactions that underly so many disease processes, it is important to determine the mechanism by which the "normal" inflammatory response resolves. The purpose of this study was to identify the time of cessation of neutrophil influx in experimental arthritis induced by the injection of C5 fragments (C5f) and to investigate mechanisms underlying the cessation process. The migration of i.v. delivery pulses to inflamed joints was assessed by lavage of the joint space and by external scintigraphy. We found no evidence for the development of inhibitory systems against chemotactic factors or "desensitization" of the inflamed site, because a second injection of C5f into joints which had been injected previously with C5f resulted in enhancement rather than inhibition of migration. Neither was evidence found for altered tissue barriers to migration or for desensitization of neutrophils as possible explanations for cessation of influx. The major mechanism appeared to be a loss of chemoattractant activity in the joint space between 2 h and 6 h after C5f injection which was detected by transfer into a fresh joint. Radiolabeled C5a des-Arg had a t1/2 of disappearance from the joint of less than 1 h, which suggested that the transferred chemoattractant must, in part, have been due to the generation of new chemotaxins by C5f injection. These observations suggest that continued generation of chemoattractants or failure of their subsequent removal may be mechanisms leading to persistent neutrophil influx in chronic inflammation.
Stimulation of degranulation from human eosinophils by platelet-activating factor.
Authors: Kroegel C , Yukawa T , Dent G , Venge P , Chung K F , Barnes P J
Published: December 2022

Platelet-activating factor (PAF) is a highly active mediator which has been implicated in allergic inflammation and bronchial asthma, possibly by interacting with eosinophils. We have examined the effect of PAF on activation of purified human eosinophils as measured by degranulation (eosinophil peroxidase, eosinophil cationic protein, arylsulfatase B, beta-glucuronidase, and alkaline phosphatase) and oxidative metabolism (superoxide anion production). PAF induced enzyme release at concentrations ranging from 1 pM to 10 microM in a rapid (t1/2 5 to 8 min), Ca2+-dependent and noncytotoxic manner from both the specific and small granules, whereas its biologic precursor and metabolite, lyso-PAF, had no effect. For all enzymes, maximal enzyme release occurred at 100 nM PAF with a mean ED50 value of 1.47 +/- 0.4 nM. At this concentration the mean percentage of total enzyme release by PAF from specific granules was 20.3 +/- 1.6% (17.9% for eosinophil peroxidase, 20.6% for beta-glucuronidase, 22.4% for alkaline phosphatase) and 28.8 +/- 2.2% from small granules (arylsulfatase B). Calcium ionophore A23187, PMA, and opsonized zymosan also induced eosinophil degranulation but their peak effect after 10-min incubation with maximal release 14.7%, 12.9%, or 14.1%, respectively, was lower when compared with PAF. Incubation of eosinophils with the PAF-antagonist WEB 2086 led to a parallel shift of the dose-response curve to the right, indicating a competitive antagonism. PAF also caused generation of superoxide anions by human eosinophils but this occurred at higher concentrations of PAF (1 microM to 30 microM) with an ED50 of 8.4 +/- 0.9 microM. Again, this effect was competitively inhibited by WEB 2086. These studies demonstrate that PAF activates human eosinophils to release granule constituents and generate superoxide anions. Since both PAF and eosinophil products are associated with pathogenesis of bronchial asthma our findings may be of particular pathophysiologic relevance.