bluuebunny/crossref_metadata_2025 huggingface dataset:
Early appearance of "natural" mucosal IgA responses and germinal centers in suckling mice developing in the absence of maternal antibodies.
Authors: Kramer D R , Cebra J J
Published: December 2022

We have examined the role of passively transferred maternal Abs in the ontogeny of "natural" mucosal IgA responses before weaning of suckling mice by comparing the immune status of gut-associated lymphoid tissue (GALT) (Peyer's patches, mesenteric lymph nodes, and lamina propria) in 7- to 25-day-old F1 severe combined immunodeficient (scid)/+ mice generated through reciprocal crosses of C.B17 scid/scid and normal congenic (+/+) adult mice. We have also examined the ability of prenatal vs postnatal transfer of maternal immunity to forestall the development of natural neonatal mucosal IgA responses by swapping litters of F1 scid/+ pups at birth between +/+ and scid/scid mothers. Our results demonstrate that F1 scid/+ pups born to or nursed by scid/scid mothers undergo an accelerated development of natural IgA responses that include germinal center reactions in both Peyer's patches and mesenteric lymph nodes. These early IgA responses are evident as: 1) increased frequencies of IgA-producing GALT organ cultures; 2) increased mean IgA output by GALT organ cultures; 3) increased frequencies (> 1 log) of IgA-secreting cells from GALT detected by ELISPOT at 16 days of age; and 4) germinal center development by 17 days of age detected by in vivo bromodeoxyuridine incorporation. Finally, FACS analyses of enteric bacteria isolated from F1 scid/+ pups and stained for the presence of surface-bound mouse IgA demonstrate that the bacterial flora is a major target of both maternal secretory IgA and of the earliest IgA Abs produced in the neonatal GALT of pups deprived of maternal immunity.
Murine minor histocompatibility antigens detected by helper T cells. Recognition of an endogenous peptide.
Authors: Wettstein P J
Published: December 2022

Minor histocompatibility (H) Ags, encoded by autosomal and sex-linked genes, have classically been identified by the cytolytic T lymphocyte response to class I-bound minor H Ags. A limited number of studies have identified minor H Ags (defined by congenic strains) that stimulate Th cells. We have selected a panel of minor H Ag-specific, CD4+ Th hybrids by the fusion of a BW5147 variant with a C57BL/6 anti-BALB.B T cell line. Studied hybrids secrete IL-2 following stimulation with BALB.B spleen cells and stimulation is blocked with anti-I-Ab Ab. These Th hybrids recognize a minimum of six different Ags (helper T target (HTT)) encoded by independently segregating genes as defined by stimulator cells from 11 CXB recombinant inbred strains. The CXB strain distribution patterns of detected Ags did not match the distribution pattern of the BALB/c MTV6 viral genome indicating that none of the detected Ags were known viral superantigens. Four tested HTT Ags appeared not to be acquired by stimulators as exogenous Ags as BALB.B spleen cells but not a mixture of BALB/c spleen cells and C57BL/6 APC stimulated IL-2 production by the respective Th hybrids. To determine whether Th hybrids recognized peptides bound to I-Ab, I-Ab molecules were immunoprecipitated from the LB27.4 cell line that stimulated the A6.5.5 hybrid (HTT-4 specific). Peptides were extracted from I-Ab molecules, ultrafiltered, and separated by HPLC. A single fraction (with flanking peaks of lower activity) sensitized syngeneic APC to stimulate A6.5.5 hybrid cells; sensitization was observed at pH 5.0 but not pH 7.0, consistent with HTT-4 peptide:I-Ab binding occurring in an endosomal compartment. These observations indicate that at least one class II-restricted minor H Ag is recognized by Th cells as a peptide bound to class II molecules.
Recombination potential of the human DIR elements.
Authors: Moore B B , Meek K
Published: December 2022

The human DIR genes are putative DH elements that are GC rich and are found between DN and DM DH gene segments. The DIR genes each have consensus recombination signal sequences (RSS) that could conceivably generate DIR coding regions of about 126 nucleotides. These RSS should allow for DH-DH rearrangements that do not violate the 12/23 recombination rule. Several Ig CDR3 sequences have been assigned to DIR usage; however, there are frequent gaps and mismatches associated with these assignments. In some instances these CDR3 sequences might be better explained by GC rich N segment addition. This report analyzes the recombination potential of the human DIR elements by PCR. Though DH-JH rearrangement of the DH genes flanking the DIR regions (DM and DN) are easily demonstrated, very little evidence of DIR-JH rearrangement could be documented. Furthermore, in amplifications that should concurrently detect both DH-DIR rearrangements (which maintain the 12/23 recombination rule) and DH-DH rearrangements (which violate the 12/23 recombination rule), DH-DH rearrangements predominate. We conclude that the DIR-associated RSS participate minimally in both DH-DH and DH-JH rearrangement. In addition, we describe several conventional DH-DH rearrangement intermediates, demonstrating unequivocally that this phenomena occurs during human Ig rearrangement.
Characterization of RFM mouse T lymphocyte anti-oncofetal antigen immunity in apparent tumor-free, long-term survivors of sublethal X-irradiation by limiting dilution T lymphocyte cloning.
Authors: Rohrer J W , Culpepper C , Barsoum A L , Coggin J H
Published: December 2022

Fractionated sublethal x-irradiation induces thymic lymphomas in up to 60% of RFM mice by 6 mo after irradiation, but no thymomas arise after 6 mo. All radiation-induced tumors expressed oncofetal Ag (OFA) and thymic OFA expression significantly preceded detectable thymomas. To determine whether lymphoma-free, radiation survivor mice had anti-OFA T cell immunity, we analyzed their clonable 5T lymphoma-reactive T lymphocytes and determined the resistance of such mice to challenge with RFM lymphoma cells. RFM mice that were irradiated 6 to 6.5 mo earlier, but which had no apparent tumors, had no more resistance to OFA+ 5T lymphoma cell challenge than nonirradiated age-matched mice. These mice actually developed maximal tumor incidence significantly faster. Analyses of 5T lymphoma-reactive T cell clones established from RFM mice 11 mo after irradiation, however, suggest that activation of anti-OFA immunity occurred subsequent to irradiation. Previously irradiated RFM mice yielded 257% more tumor-reactive T cell clones/mouse than non-irradiated controls. All clones from both sets of mice specifically proliferated to purified OFA. Each irradiated mouse yielded some clones that proliferated to fivefold lower doses of OFA than did any T cell clones from nonirradiated mice. Only these low-dose OFA-reactive clones responded equally to irradiated 4T and 5T cells by proliferation, IFN-gamma secretion, and target lymphoma cell killing. 4T cells express less OFA/cell than 5T cells. Some noncytotoxic CD8 T cells that inhibited cytotoxic T cell function were cloned only from irradiated RFM mice.
Activation of cAMP-dependent pathways in human airway smooth muscle cells inhibits TNF-alpha-induced ICAM-1 and VCAM-1 expression and T lymphocyte adhesion.
Authors: Panettieri R A , Lazaar A L , Puré E , Albelda S M
Published: December 2022

Asthma is a disease of airway inflammation and hyper-reactivity associated with lymphocytic infiltration in the bronchial submucosa. We recently demonstrated that human airway smooth muscle (ASM) cells express the cell adhesion molecules ICAM-1 and VCAM-1, which are up-regulated by cytokines such as TNF-alpha, and which mediate binding of activated T lymphocytes. In this study, we examined whether an increase in [cAMP]i, presumably via activation of cAMP-dependent protein kinase, modulates TNF-alpha-induced ICAM-1 and VCAM-1 on ASM. We found that treatment of ASM with either forskolin, which directly activates adenylyl cyclase, or with cholera toxin, which activates the heterotrimeric GTP-binding protein, Gs, inhibited TNF-alpha-induced cell adhesion molecule expression. In addition, treatment with either isoproterenol or prostaglandin E2, which activates receptors coupled to Gs and increases [cAMP]i, also inhibited TNF-alpha-induced expression of ICAM-1 and VCAM-1 on ASM. Furthermore, adhesion of activated T cells to TNF-alpha-stimulated ASM was inhibited by treating the ASM cells with either forskolin or PGE2. These data suggest that cAMP-dependent protein kinase activation decreases cytokine-induced expression of cell adhesion molecules on ASM cells, modulates T cell binding to airway myocytes and, thus, suggests novel therapeutic approaches to airway inflammation.
Bone marrow-derived murine mast cells migrate, but do not degranulate, in response to chemokines.
Authors: Taub D , Dastych J , Inamura N , Upton J , Kelvin D , Metcalfe D , Oppenheim J
Published: December 2022

We have determined that several chemokines induce mast cell migration in vitro. This directed migration is dependent on the presence of particular extracellular matrix proteins and the activation status of the cells. Mast cell haptotactic responses were observed in response to various chemokines on vitronectin-, laminin-, and fibronectin-coated filters. Unstimulated mast cells were chemoattracted only by monocyte chemotactic protein-1 and RANTES on vitronectin-coated and, to a lesser extent, laminin-coated filters, whereas IgE-activated mast cells migrated in response to monocyte chemotactic protein-1, regulated on activation normal T expressed and secreted, platelet factor-4, and macrophage inflammatory protein-1 alpha on all three matrix proteins. No significant migration was observed on collagen type IV-coated or uncoated filters. Mast cell migration in response to chemokines on extracellular matrices and its enhancement by IgE-dependent activation provide a mechanism by which cells may be drawn to sites of inflammation. Chemokine-induced mast cell recruitment may be particularly relevant in host defense responses to parasitic infections, allergic reactions, Jones-Mote reactions, and in wound healing.
Glomerular binding activity in MRL lpr serum consists of antibodies that bind to a DNA/histone/type IV collagen complex.
Authors: Bernstein K A , Valerio R D , Lefkowith J B
Published: December 2022

To clarify the pathogenesis of lupus nephritis, we developed an assay that defines a glomerular binding activity (GBA) in both murine and human lupus sera highly correlated with nephritis. In the current study, we used a cross-adsorption strategy to establish that the GBA in MRL lpr serum binds to the glomerular basement membrane (GBM). We subsequently observed that this binding to the GBM was competitively inhibited by either exogenous DNA or histone, abrogated by pretreatment of the GBM with DNAse, and restored after DNase treatment with DNA/histone in a synergistic fashion. GBM binding was also completely inhibited by pretreatment of GBM with collagenase but not heparatinase. The effect of collagenase was not reversed by the subsequent addition of DNA, but was restored by the sequential re-addition of type IV collagen and DNA. By using purified basement membrane components, we found that MRL lpr serum bound avidly to DNA coated on type I collagen but less well (or not at all) to DNA coated on type IV collagen, laminin, or fibronectin. Histone pretreatment of type IV collagen before DNA addition, however, synergistically enhanced binding in a fashion similar to that seen with native GBM. Thus, the GBA in MRL lpr serum seems to be comprised of autoantibodies that bind to histones and/or DNA that adhere to type IV collagen within the GBM. These data support the planted Ag hypothesis as the principal pathogenic mechanism in lupus nephritis and suggest that multiple autoantibodies may contribute to this disorder.
Mucosal imbalance of IL-1 and IL-1 receptor antagonist in inflammatory bowel disease. A novel mechanism of chronic intestinal inflammation.
Authors: Casini-Raggi V , Kam L , Chong Y J , Fiocchi C , Pizarro T T , Cominelli F
Published: December 2022

The etiology and pathogenesis of inflammatory bowel disease (IBD) are unknown. Increasing evidence supports the theory that chronic IBD is the result of dysfunctional immunoregulation manifested by an inappropriate production of mucosal cytokines. The aim of the present study was to test the hypothesis that a specific mucosal imbalance of IL-1 and IL-1 receptor antagonist (IL-1ra) production plays an important role in the perpetuation and chronicity of intestinal inflammation. Total IL-1, IL-1ra, and the IL-1ra/IL-1 ratio were measured in freshly isolated intestinal mucosal cells, as well as in mucosal biopsies obtained from control, Crohn's disease, and ulcerative colitis patients. IL-1 alpha, IL-1 beta, and IL-1 ra were measured by specific non-cross-reacting radioimmunoassays and ELISA. A markedly significant decrease in the intestinal mucosal IL-1ra/IL-1 ratio was found in both Crohn's disease and ulcerative colitis patients when compared with control subjects (p < 0.01). The IL-1ra/IL-1 ratio correlated closely with the clinical severity of disease (r = -0.7846, p < 0.001). Furthermore, the observed decrease in the IL-1ra/IL-1 ratio was specific for IBD because a decreased IL-1ra/IL-1 ratio was not found in patients with self-limiting colitis. These results support the hypothesis that an imbalance between IL-1 and IL-1ra production is of pathogenic importance in chronic inflammatory diseases, including IBD.
Breaking tolerance leads to autoantibody production but not autoimmune liver disease in hepatitis B virus envelope transgenic mice.
Authors: Wirth S , Guidotti L G , Ando K , Schlicht H J , Chisari F V
Published: December 2022

Hepatitis B virus (HBV) transgenic mice containing the HBV envelope open reading frame under the transcriptional control of the mouse albumin promoter express hepatitis B surface Ag (HBsAg) in all of their hepatocytes and secrete HBsAg (10 to 40 ng/ml) into the circulation. Because these transgenic mice show no signs of spontaneous liver cell injury or autoimmunity toward the viral (self-) Ag, we asked whether the state of self-tolerance could be reversed by the induction of an acute necroinflammatory liver disease or by immunization with HBV envelope proteins, with the aim of creating a transgenic model for chronic, immune-mediated hepatitis. Our studies indicate that repetitive administration of bacterial LPS, IFN-gamma, or HBsAg-specific CTL, all of which were previously shown to cause liver cell injury and inflammation, does not break tolerance at the T or B cell level, suggesting that the intrahepatic lymphomononuclear cell infiltrate induced by these agents consists of HBsAg-nonspecific cells. The adoptive transfer of HBsAg-primed nontransgenic CD4+ T cells into transgenic mice did not induce anti-HBs autoantibody production by transgenic B cells, even though transgenic B cells were fully responsive to immunization with HBsAg when appropriate T cell help was provided in a nontransgenic environment. Immunization of transgenic mice with purified HBsAg in CFA and repetitive infection with rHBV envelope vaccinia virus led to production of T cell-dependent anti-HBs autoantibodies that cleared HBsAg from the serum, but not to activation of HBsAg-specific CTL. We conclude that HBV envelope transgenic mice are largely tolerant to the transgene product at the T cell but not at the B cell level, and that the activation of an anti-HBs response was not sufficient to induce an autoimmune liver disease in this HBV envelope transgenic mouse model.
Hybrid and allogeneic resistance to T cell grafts mediated by murine NK and CD8+ T cells.
Authors: Davenport C , Haile A , Kumar V , Bennett M
Published: December 2022

Lethally irradiated mice can reject H-2 allogeneic or parental strain stem cells in bone marrow cell (BMC) grafts within 48 h after transplantation. This rapid rejection of BMC grafts occurs without prior sensitization and is mediated by NK1.1+ NK cells. One hypothesis to account for the ability of host NK cells to mediate acute rejection of allogenic and parental stem cells is that these effector cells recognize hemopoietic histocompatibility (Hh-1) Ags on the donor stem cells. T cells present in the donor BMC can prevent NK cell-mediated rejection. However, lethally irradiated mice can also reject T cells present in lymph node cell preparations that respond to alloantigens of the host. T cell grafts from H-2k/Hh-1k, H-2r/Hh-1null, and H-2ia1/Hh-1null donors were rejected by CD8+ NK1.1- T cells. Ags other than Hh-1 Ags appeared to be recognized by these CD8+ T cells. In contrast, host NK cells rejected H-2d/Hh-1d T cell grafts, whereas both NK cells and CD8+ TCR-alpha beta + T cells rejected H-2b/Hh-1b T cell grafts. Therefore, both CD8+ TCR-alpha beta T cells and NK cells mediate allogeneic and hybrid resistance to T cell grafts.
Distinct differentiative stages of CD4+CD8+ thymocyte development defined by the lack of coreceptor binding in positive selection.
Authors: Dutz J P , Ong C J , Marth J , Teh H S
Published: December 2022

Cortical CD4+CD8+ thymocytes mature into CD4+ or CD8+ thymocytes through a process termed positive selection. To better define differentiative stages of CD4+CD8+ thymocyte development in positive selection, we performed a phenotypic analysis of CD4+CD8+ thymocytes from H-Y mice mated to various genetic backgrounds. We have previously shown that coordinate binding of the H-Y TCR and the CD8 coreceptor to the restricting Db MHC class I molecule is required for the efficient positive selection of this TCR. In this study we have used TCR, CD5, and CD45 expression levels as markers for thymocyte maturation. Lack of CD8/Db interaction was achieved by introducing a mutation that abrogates CD8 binding in the alpha 3 domain of Db. We found that the absence of coreceptor ligation prevented TCR up-regulation in CD4+CD8+ thymocytes and resulted in a developmental arrest characterized by low levels of TCR and CD45. We have previously shown that deletion of CD4+CD8+ thymocytes expressing the H-Y TCR is facilitated by CD8 coreceptor ligation. Here we show that expression of the deleting ligand in the absence of coreceptor ligation caused CD5 up-regulation without concomitant TCR or CD45 up-regulation in CD4+CD8+ thymocytes. In a beta 2-microglobulin null background, introduction of the H-Y TCR caused the majority of CD4+CD8+ thymocytes to express an unusually low level of of the CD5 activation marker, suggesting that a low-affinity or noncognate TCR/MHC interaction may be required for initial CD5 up-regulation to intermediate levels. Collectively, these observations favor a maturational process in positive selection in which CD5 up-regulation precedes CD45 and TCR up-regulation.
Molecular cloning of a cDNA clone encoding a phosphoprotein component related to the Ig receptor-mediated signal transduction.
Authors: Inui S , Kuwahara K , Mizutani J , Maeda K , Kawai T , Nakayasu H , Sakaguchi N
Published: December 2022

We have shown previously that a 52-kDa phosphoprotein (p52) co-precipitated with Ig receptor (IgR)-associated MB-1 protein was inducibly phosphorylated by the stimulation with 12-O-tetradecanoyl-phorbol-13-acetate. By immunizing the 52-kDa protein co-precipitated with MB-1, we prepared a mAb (19-14), which can immunoprecipitate the p52 and the associated kinase molecule. Immune complex kinase assay with the 19-14 mAb showed that the p52 is associated with a novel kinase molecule and is involved in IgR-mediated signal transduction. Here we isolated a cDNA clone (alpha 4) from a murine bone marrow cDNA library in lambda gt-11 vector by the 19-14 mAb. The alpha 4 cDNA encodes a novel protein of 340 amino acids with multiple potential phosphorylation sites. The 1.4-kb alpha 4 mRNA is expressed in various cells including cell lines of B lineage, T lineage, monocytic, fibroblast, and normal organs such as liver, spleen, thymus, and brain. To study the molecule encoded by the alpha 4 cDNA, we produced a chimeric protein of the glutathione S transferase (GST)-alpha 4 in pGEX-3X expression vector. The 19-14 mAb binds to the GST-alpha 4 fusion protein. Rabbit anti-alpha 4 Ab prepared by immunizing the GST-alpha 4 fusion protein immunoprecipitated a 52-kDa phosphoprotein from WEHI-231 cells. The 52-kDa phosphoprotein immunoprecipitated by the anti-alpha 4 Ab is tightly associated with kinase activity that is similarly observed with the p52 protein reported previously. Moreover, the 52-kDa phosphoprotein immunoprecipitated with the anti-alpha 4 Ab is identical to the p52 by the protein comparison on non-equilibrium pH gradient gel electrophoresis/SDS-PAGE, isoelectric focusing/SDS-PAGE, and V8 protease mapping. The 52-kDa phosphoprotein is associated with functional components that are inducibly phosphorylated by anti-IgM stimulation. These results suggested that the alpha 4 gene-encoded molecule is functionally involved in the IgR-mediated signal transduction in B cells.
Activation of IFN-alpha, IFN-gamma, MxA, and IFN regulatory factor 1 genes in influenza A virus-infected human peripheral blood mononuclear cells.
Authors: Ronni T , Sareneva T , Pirhonen J , Julkunen I
Published: December 2022

Mononuclear blood cells have an important role in immunity as they produce different cytokines in response to microbial infections. We infected primary human mononuclear blood cells with a pathogenic influenza A virus (A/Beijing/353/89) and studied the activation of IFN-alpha, IFN-gamma, IRF-1, and MxA genes. IFN-alpha and IFN-gamma steady state mRNA levels peaked at 6 to 9 h after infection and declined rapidly thereafter. Only a modest (twofold) increase in IRF-1 mRNA was seen. MxA gene expression, normally strictly regulated by IFN-alpha/beta, had expression kinetics similar to those of IFN mRNA. Infection experiments done in the presence of cycloheximide showed that influenza virus infection could induce all genes studied in the absence of detectable protein synthesis. Pretreatment with IFN-alpha, but not with IFN-gamma, caused a dose-dependent inhibition of influenza virus replication in PBMC, and this inhibition correlated with increasing levels of MxA protein. Influenza virus replication was also inhibited in a stably transfected, MxA-expressing promonocytic U937 cell line. The results suggest that MxA protein significantly contributes to IFN-mediated host defence mechanisms against influenza A virus.
Identification of complement receptor type 1-related proteins on primate erythrocytes.
Authors: Nickells M W , Subramanian V B , Clemenza L , Atkinson J P
Published: December 2022

The purpose of this study was to characterize the structure and function of the immune adherence receptor (CR1, CD35, C3b/C4b receptor) of primates. Western blotting, immunoprecipitation, ELISA, and affinity chromatography with homologous C3b and C4b were utilized. The major cross-reactive E membrane protein of ten species of primates tested was lower in m.w. than was human CR1 and fell into two size groups of 55 to 75 and 130 to 165 kDa. There was 10- to 100-fold more CR1 per primate E than human E. Five species also expressed lesser quantities of a protein similar in m.w. (approximately 200 kDa) to human CR1. In contrast to E, the major cross-reactive protein on PBMC was similar in size to human CR1. Four species also expressed lesser amounts of a lower m.w. protein on their PBMC of the same M(r) as that found on their E. Affinity chromatography demonstrated that the approximately 200-kDa form, if present, was recovered with a similar efficiency to that of human CR1. Three patterns of binding, however, were identified among the lower m.w. proteins: 1) C3b > or = C4b; 2) C4b > C3b; and C3b only or predominantly. The fact that these E proteins cross-react with Ab to human CR1, bind homologous C3b and, in most cases, C4b, and for some species represent the only such protein expressed on their E identifies them as immune adherence receptors. The 70-kDa CR1 of the chimpanzee E seems to arise by alternative splicing of the mRNA encoding the 200-kDa protein. These data raise interesting questions relative to the evolution of CR1 in primates and provide a basis for analysis of structure-function relationships among these size forms of CR1.
Molecular basis for a familial defect in phagocyte expression of IgG receptor I (CD64).
Authors: van de Winkel J G , de Wit T P , Ernst L K , Capel P J , Ceuppens J L
Published: December 2022

Four individuals have been identified, within a single family, who lack phagocyte expression of the high affinity type I IgG receptor (CD64). As a result, their monocytes are unable to support mouse IgG2a anti-CD3-induced T cell mitogenesis (nonresponder individuals). Southern blotting proved all three human Fc gamma receptor I (hFc gamma RI) genes to be present in nonresponders without major structural changes. Nucleotide sequencing showed identical hFc gamma RIA promoter regions in all individuals. At the message level, a distinct difference was noted between monocytes from control (responder) donors and from nonresponders. Both a 1.7- and 1.6-kb message were found in responders, whereas in nonresponders only the 1.6-kb species was detectable. Reverse transcriptase-PCR analyses showed the hFc gamma RIa transcript (encoding a receptor with three extracellular Ig-like domains) to be present at a approximately 15- to 20-fold lower level in nonresponder monocytes. Importantly, we found a single nucleotide difference (C --> T) within the extracellular domain exon 1-encoding region of hFc gamma RIA in nonresponders, resulting in the change of codon 92 (encoding an arginine) into a termination codon. This change likely affects mRNA stability and, thereby, leads to undetectable expression of phagocyte-hFc gamma RIa. Despite this defect, these individuals are apparently healthy, suggesting that hFc gamma RIa is dispensable for phagocyte functioning.
TCR-stimulated naive human CD4+ 45RO- T cells develop into effector cells that secrete IL-13, IL-5, and IFN-gamma, but no IL-4, and help efficient IgE production by B cells.
Authors: Brinkmann V , Kristofic C
Published: December 2022

It is currently believed that IgE production by B cells is induced by activated CD4+ Th2-like cells that produce increased amounts of IL-4 but low levels of IFN-gamma. We found that "naive" CD4+ 45RO- T cells primed and restimulated in vitro via the TCR developed into effector cells that produced large amounts of IL-13, IL-5, and IFN-gamma, but no IL-4. Such CD4 T cells induced surface IgD+E- B cells to produce large amounts of IgE. The IgE response could be blocked completely by neutralizing anti-IL-13 Abs, whereas anti-IL-4 had no effect. Addition of exogenous IL-4 during priming of CD4 cells suppressed clonal expansion and the development of Th cells including helpers for IgE, but increased endogenous production of IL-4 and IL-5, and suppressed production of IFN-gamma. Addition of exogenous IFN-alpha, IFN-gamma, or neutralizing anti-IFN-gamma mAbs affected neither priming of CD4 T cells nor B cell help. In contrast to CD4+ 45RO- naive T cells, CD4+ 45RO+ "memory" T cells primed and restimulated in vitro secreted IL-4 in addition to IL-13, IL-5, and IFN-gamma, and the IgE response induced by such cells could be blocked only by a combination of anti-IL-4 plus anti-IL-13 Abs. Unlike CD4 cells, CD8 cells could not be primed to help IgE production. The results indicate that TCR/CD3 cross-linking on naive human CD4 T cells is sufficient to induce the development of potent IgE helper cells, which secrete large amounts of IL-13, IL-5, and IFN-gamma, but no IL-4. Such Th cells may exacerbate atopic inflammation, because all by themselves they could drive IL-13-dependent IgE production, IL-5-dependent eosinophilia, and IFN-gamma-dependent macrophage activation, but could not suppress IFN-gamma production by other T cells via IL-4.
Expression and induction of nuclear factor-kappa B-related proteins in thymocytes.
Authors: Sen J , Venkataraman L , Shinkai Y , Pierce J W , Alt F W , Burakoff S J , Sen R
Published: December 2022

Thymocytes mature in response to the cues from the thymic micro-environment, which regulate stage-specific gene expression during development. We find that several proteins that bind the kappa B sequence in vitro are constitutively activated in freshly isolated thymocytes. These include the rel-related p50 homodimers, p50/p65 heterodimers, low levels of c-rel, and two other factors that may be thymus specific. Disruption of the thymic micro-environment resulted in loss of DNA-binding, suggesting that lymphocyte-stromal cell interactions induce and maintain these proteins in a DNA-binding form. Phorbol ester and ionomycin treatment induced p50, p65, and p68 c-rel kappa B DNA-binding activity. Expression of p68 c-rel protein, but not p50 or p65, was suppressed by the immunosuppressive drug FK506. Because FK506 specifically inhibits the appearance of mature single-positive thymocytes, gene expression regulated by p68 c-rel may play a role in selection and maturational signals involved in the double-positive to single-positive transition.
Translational regulation of murine complement factor B alternative transcripts by upstream AUG codons.
Authors: Garnier G , Circolo A , Colten H R
Published: December 2022

Factor B (Bf), a constituent of the alternative pathway of complement activation, is encoded by a single gene that is located within the MHC. In murine kidney and intestine, two Bf transcripts (Bf short and Bf long), generated from distinct transcriptional initiation sites, are expressed in approximately equal amounts. In the liver, > 95% of Bf mRNA is the short transcript. To ascertain the biologic consequences of this tissue-specific mRNA polymorphism, we quantitated the effect of structural differences between the two transcripts on net Bf protein synthesis. Cell-free translation of Bf mRNA in vitro revealed that the rate of translation of Bf short is about twice that of Bf long. The 5' extension of Bf long includes four short open reading frames upstream of the authentic translational initiation codons. Mutation of all four upstream AUGs generates a Bf long transcript with a translational rate about equal to that of Bf short. This effect was primarily accounted for by mutation of the second AUG from the 5' end. Similar studies of Bf expression in vivo showed an approximately twofold difference in translational rate between Bf long and the transcript mutated at the upstream AUGs. Because systemic and local inflammation in kidney alters the ratio of Bf long and short, net production of complement protein Bf in extrahepatic tissues is regulated by both transcriptional and translational control mechanisms.
A novel phosphocholine antigen protects both normal and X-linked immune deficient mice against Streptococcus pneumoniae. Comparison of the 6-O-phosphocholine hydroxyhexanoate-conjugate with other phosphocholine-containing vaccines.
Authors: Fischer R T , Longo D L , Kenny J J
Published: December 2022

A novel form of phosphocholine (PC), p-nitrophenyl-6-(O-phosphocholine)hydroxyhexanoate (EPC) coupled to keyhole limpet hemocyanin (KLH) has been compared with unencapsulated, avirulent Streptococcus pneumoniae (R36a) and with the traditional thymus-dependent form of PC, diazophenylphosphocholine (DPPC)-conjugated KLH for its vaccine potential against virulent S. pneumoniae. Immunization with any of these three PC-containing Ags protects normal mice against a lethal challenge with 10(4) S. pneumoniae, whereas only EPC-KLH provides total protection to Xid mice. DPPC-KLH and unencapsulated S. pneumoniae confer less than 40% protection in Xid mice. Passive transfer of a PC-specific hybridoma Ab made from EPC-KLH-immunized Xid mice also provided protection against lethal challenge with S. pneumoniae. Protective anti-PC Ab were capable of binding to the surface of virulent bacteria, whereas anti-PC Ab incapable of binding to the bacterial surface failed to protect. Furthermore, serum Ab from EPC-KLH immunized and protected mice bound to S. pneumoniae, whereas secondary Abs from DPPC-KLH- or R36a-immunized mice failed to bind to the bacteria. EPC-KLH is potentially a vaccine candidate for pneumococcal prophylaxis in settings of immune compromise.
Analysis of clonal CD8+ T cell expansions in normal individuals and patients with rheumatoid arthritis.
Authors: Fitzgerald J E , Ricalton N S , Meyer A C , West S G , Kaplan H , Behrendt C , Kotzin B L
Published: December 2022

In the course of studying the circulating TCR repertoire in humans, we noted several individuals with an increase in the percentage of CD8+ T cells expressing a particular V region. In some cases, these CD8 expansions were dramatic, occupying over 40% of the total CD8 repertoire. Using a panel of mAbs to different TCR V regions, we found that over 30% of healthy adults (> 35 years of age) harbor an expansion that alters the peripheral blood CD8 TCR repertoire. A wide range of V regions were expressed by these expansions. Considering that the mAbs used cover only a portion of the V beta repertoire, the data suggest that over 70% of adults are likely to harbor such expansions. Junctional region sequencing showed that the CD8 subset expansions were clonal, and serial studies as long as 4 years showed that they persisted indefinitely. Expansions were not identified in the CD4 population. Discordant expression of one large V beta 6.7+ clone was found in one identical twin set, suggesting the possibility that an environmental exposure is involved in their generation and/or expansion. In one large family, we found five family members with a large CD8 subset expansion. Remarkably similar usage of J beta regions was noted, and two individuals demonstrated V beta 3-expressing clones with homologous CDR3 regions, differing by only one major substitution. The repertoire data from this family suggest that the T cell clones have arisen in response to a common Ag. Studies of patients with rheumatoid arthritis found a significantly increased frequency of circulating CD8 subset expansions that expressed a different V region repertoire compared with the healthy individuals studied. Overall, our results emphasize a frequent alteration in the human CD8 TCR repertoire, most likely related to an environmental exposure, in both healthy individuals and patients with rheumatoid arthritis. The presence of these expansions will be important to consider in any study of human TCR repertoire, and their implication for health and disease will be important to understand.
Regulated expression of Ly-6A.2 is important for T cell development.
Authors: Bamezai A , Palliser D , Berezovskaya A , McGrew J , Higgins K , Lacy E , Rock K L
Published: December 2022

Ly-6A.2 is a surface protein on T cells that may play a role in lymphocyte activation. The regulation of Ly-6A.2 expression during T cell lymphopoiesis has been intriguing. It is one of the earliest markers expressed on pluripotent hemopoietic stem cells and is present on both primitive and mature T cells, but its expression is extinguished in the thymus during key developmental stages. To determine whether Ly-6A.2 is active on developing T cells, as well as the significance of its developmental regulation, Ly-6A.2 was expressed throughout T cell development under control of the T cell-specific human CD2 enhancer in transgenic mice. The constitutive overexpression of Ly-6A.2 in vivo led to a marked impairment in the generation of thymocytes. Development was arrested at the time in thymic development when Ly-6A.2 expression is normally turned off. These results indicate that the regulated expression of Ly-6A.2 in thymocytes may be important for normal development. Moreover, these findings demonstrate that Ly-6A.2 is active in the thymic microenvironment.
IL-10 prevents naturally occurring fetal loss in the CBA x DBA/2 mating combination, and local defect in IL-10 production in this abortion-prone combination is corrected by in vivo injection of IFN-tau.
Authors: Chaouat G , Assal Meliani A , Martal J , Raghupathy R , Elliott J F , Elliot J , Mosmann T , Wegmann T G
Published: December 2022

CBA x DBA/2 placentae are quantitatively or qualitatively deficient in their production of the anti-inflammatory Th2-type cytokines IL-4 and IL-10 compared with the nonresorption-prone CBA x BALB/c mating combination. Wastage in this mating combination is accompanied by increased levels of local inflammatory cytokines. In addition, alloimmunization enhances the placental production of IL-4 and IL-10 in CBA x DBA/2 matings. Furthermore, rIL-10 by itself completely reverses the high incidence of fetal resorption after i.p. injection. Conversely, anti-IL-10 increases the resorption rate, but only in CBA x DBA/2 matings. On the other hand, injecting either anti-IFN-gamma or pentoxifillin (an anti-TNF agent) partially reduces the resorption. When given together, they produce a synergistic remission of fetal loss. Finally, we report that recombinant ovine trophoblast protein, an IFN-tau which is known to influence reproductive outcome in ruminants, can also counteract increased CBA x DBA/2 fetal resorption. It simultaneously induces increased placental IL-4 and IL-10 production in this mating combination. These results indicate that the placentally produced anti-inflammatory cytokines can play a vital role in the survival to term of the fetal allograft, by counteracting deleterious inflammatory cytokines.
Flow cytometric analysis for cytokine production identifies T helper 1, T helper 2, and T helper 0 cells within the human CD4+CD27- lymphocyte subpopulation.
Authors: Elson L H , Nutman T B , Metcalfe D D , Prussin C
Published: December 2022

Using three-color flow cytometric analysis for the detection of intracellular cytokines, we have been able to determine the exact combination of cytokines produced by individual T lymphocytes. Because CD4+CD27- lymphocytes have been shown to produce more IL-4 and IL-5 than CD4+CD27+ lymphocytes, cells from normal individuals (n = 4) and helminth-infected patients (n = 4) were sorted magnetically for the CD4+CD27+ and the CD4+CD27- subpopulations. Intracellular staining for IL-4, IL-5, and IFN-gamma subsequent to mitogen stimulation for 6 h revealed that although almost no CD4+CD27- lymphocytes produce both IL-5 and IFN-gamma (0.03-1.4%), a distinct proportion produce both IL-4 and IFN-gamma (0.1-8.0%), and 66% to 84% of IL-5-producing cells also produce IL-4. Patients and normal individuals had the same functional T cell subsets, but the CD4+CD27- lymphocytes from patients had higher frequencies of cells producing IL-4 (geometric mean (GM), 24.3% vs 16.4%) or IL-5 (GM, 10.2% vs 2.9%), whereas those of normal individuals had higher frequencies of cells producing IFN-gamma (GM, 44.5% vs 17.2%; p = 0.043). These analyses also revealed that the CD4+CD27- population included significantly higher frequencies of cells that were IL-5+IFN-gamma- (GM, 4.9% vs 1.5%; p = 0.025), IL-4+IFN-gamma- (GM, 13.8% vs 3.5%; p = 0.025), and IFN-gamma+IL-4-IL-5- (GM, 27.3% vs 12.0%; p = 0.011) than the CD4+CD27+ population. Thus, we have clearly demonstrated Th1, Th2, and Th0 cell subsets within the CD4+-CD27- population of human lymphocytes.
Fas ligand-mediated cytotoxicity is directly responsible for apoptosis of normal CD4+ T cells responding to a bacterial superantigen.
Authors: Ettinger R , Panka D J , Wang J K , Stanger B Z , Ju S T , Marshak-Rothstein A
Published: December 2022

Exposure of naive CD4+ T lymphocytes to superantigens such as staphylococcal enterotoxin B (SEB) induces a strong proliferative response. Prolonged exposure or subsequent restimulation of the responding T cell population with SEB leads to the apoptotic events of activation-induced cell death (AICD). However, T cells derived from either Fas-deficient lpr or Fas ligand-deficient gld autoimmune mouse strains, fail to undergo AICD under these conditions. Instead, these autoimmune T cells mount a vigorous proliferative response, suggesting a critical role for Fas/FasL interactions in this form of autoapoptosis. In the current study, we found that SEB-induced AICD was tied to the rapid induction of FasL expression in cells constitutively expressing high levels of Fas. Furthermore, the addition of soluble Fas-IgG fusion protein to the SEB-restimulated cultures blocked AICD and resulted in a 2 degrees proliferative response that was comparable in magnitude and kinetics to that of the lpr and gld T cells. The rapid onset of apoptosis in normal T cells subsequent to restimulation with SEB was in direct contrast to the proliferative response of the initial cultures, even though comparable levels of Fas and FasL RNA were found in T cells after 1 degree and 2 degrees challenge. The clonal expansion of the normal T cells responding to the initial SEB stimulation was, however, dramatically compromised when the normal cells were cocultured with an MRL-lpr responder population; addition of soluble Fas-IgG rescued the normal component of the response. Together, these data demonstrate first, that Fas/FasL interactions are intimately tied to superantigen-induced AICD, a form of autocrine cell death, and second, that FasL-mediated cytotoxicity is responsible for the disappearance of normal CD4+ T cells in lpr cocultures.
Regulation of L-selectin mRNA in Jurkat cells. Opposing influences of calcium- and protein kinase C-dependent signaling pathways.
Authors: Kaldjian E P , Stoolman L M
Published: December 2022

L-Selectin initiates leukocyte attachment to venular endothelium during lymphocyte recirculation through lymph nodes, leukocyte recruitment into sites of inflammation, and the hematogenous spread of lymphoid malignancies. The density of L-selectin at the cell surface is a major determinant of binding activity and entry into tissues. Post-transcriptional shedding is one control mechanism; however, the extent and physiologic relevance of pre-translational regulation has not been defined. The current study shows that mitogen-/IL-2-driven proliferation of human T cells first increased then markedly decreased the expression of L-selectin on the blast population. The prevalence of specific mRNA showed parallel changes, implying that receptor density is controlled, in part, at the pretranslational level. We used the IL-2-independent Jurkat cell line to determine whether signaling through C-type protein kinases and intracellular calcium regulated L-selectin mRNA directly. Selective pharmacologic activation of these pathways with phorbol esters and calcium ionophore, respectively, resulted in opposite effects on both L-selectin density and mRNA levels. Phorbol esters induced receptor shedding followed by progressive increases in L-selectin density and steady state levels of mRNA. Addition of a calcium ionophore with the phorbol ester blocked both the reexpression of surface receptor and the increase in mRNA. Treatment with ionophore alone resulted in a steady decline in L-selectin expression and mRNA levels. Cyclosporin A, a specific inhibitor of calcineurin, blocked the impact of ionophore on both basal and phorbol-induced levels of L-selectin mRNA. Ionophore alone did not induce apoptosis, significantly alter cell cycle kinetics, or increase transcription of the IL-2 gene under conditions that suppressed L-selectin. Thus, calcineurin seems to be a proximal enzyme in a novel regulatory cascade that suppresses L-selectin expression independent of its known effects on proliferating T cells. In light of the findings in Jurkat, we propose that the protein kinase pathway up-regulates L-selectin mRNA and surface expression early in mitogen-driven T cell proliferation. Chronic elevation of intracellular calcium in repeatedly stimulated T cells then down-regulates expression at the pretranslational level through prolonged activation of calcineurin.
Regulatory role of gamma delta T cells in uterine intraepithelial lymphocytes in maternal antifetal immune response.
Authors: Suzuki T , Hiromatsu K , Ando Y , Okamoto T , Tomoda Y , Yoshikai Y
Published: December 2022

To elucidate the potential roles of the gamma delta T cells in uterine intraepithelial lymphocytes (IEL) in the regulation of maternal antifetal immune response during pregnancy, we examined the kinetics and function of gamma delta T cells in uterine IEL obtained from (C3H/He x AKR/J) pregnancy. The number of gamma delta T cells increased in the uterine IEL in (C3H/HexAKR/J) pregnancy more than those in (C3H/HexC3H/He) pregnancy and much more than in nonpregnant C3H/He mice. The uterine IEL in (C3H/HexC3H/He) pregnancy significantly proliferated in response to AKR/J stimulator cells. In contrast, the uterine IEL in (C3H/HexAKR/J) pregnancy showed little, if any, proliferation in response to the same stimulator cells. gamma delta T cell depletion from the uterine IEL in (C3H/HexAKR/J) pregnancy restored their responsiveness against AKR/J stimulator cells. Both gamma delta T cell-enriched fraction and alpha beta T cell-depleted fraction, but neither gamma delta T cell-depleted fraction nor alpha beta T cell-enriched fraction in the uterine IEL exhibited suppressive activity against allogeneic responses of nonpregnant C3H/He LN cells. This suppressive activity was shown by transferring the supernatant of culture medium in the uterine IEL stimulated with AKR/J cells that contained a large amount of TGF-beta, and the suppressive activity was significantly blocked by addition of anti-TGF-beta mAb to the culture. Taken together, our results suggest that gamma delta T cells in the uterine IEL suppress the maternal antifetal immune response at the maternal-fetal interface at least in part through TGF-beta production to prevent a rejection of the fetus.
Tissue distribution of natural peptides derived from a ubiquitous dehydrogenase, including a novel liver-specific peptide that demonstrates the pronounced specificity of low affinity T cell reactions.
Authors: Wu M X , Tsomides T J , Eisen H N
Published: December 2022

The peptides recognized by CD8+ CTL normally arise by proteolysis of intracellular proteins. To learn whether these peptides are generated similarly in diverse cell types, we examined the variety and abundance of naturally processed peptides that derive from a ubiquitous enzyme, alpha-ketoglutarate dehydrogenase, and are recognized in association with the class I MHC protein, Ld, by a CTL clone (2C). A characteristic set of three peptides was found in diverse tissues, but their abundance varied greatly, apparently unrelated to differences in class I MHC expression, e.g., they were surprisingly abundant in liver. We also found in liver a fourth naturally processed peptide (p2Ca-Y4, LSPYPFDL) that differs by one oxygen atom from a previously characterized natural peptide (p2Ca, LSPFPFDL). CTL discrimination between these peptides in association with the same class I MHC protein, Kb, demonstrates the striking specificity that can be exhibited by low affinity T cell reactions.