Regulation on murine T cell responses to autologous antigens by alpha-fetoprotein

Murine α-fetoprotein (AFP), a major component of fetal and newborn sera, was shown to exert potent immunosuppressive effects on autologous mixed lymphocyte reactions (AMLR) in vitro. Thus, the relatively vigorous proliferative response of newborn CBA/J thymocytes reacting in mixed cultures against adult syngeneic spleen cells was almost totally abrogated by 200 and 100 μg/ml AFP over the 6-day time course studied, with significant suppression still evident in the presence of 10 μg/ml AFP. In contrast, the maximum achievable suppression of parellel allogeneic MLRs was only 40 to 60%. The newborn thymocyte anti-adult syngeneic spleen AMLR was shown to be mediated by an Lyt 1⁺23^? T-cell subset reacting against Ia⁺ adult non-T stimulator cells. Newborn and adult AMLRs resulting from autochthonous T responder/non-T stimulator cell mixtures from individual animals were also found to be highly sensitive to AFP-mediated suppression. The fact that fetal-derived AFP could be shown to efficiently inhibit neonatal thymocyte responses to autologous antigens when tested in vitro in amounts 20 to 50 times lower than the levels present in fetal and newborn sera suggests a potentially important role for endogenous AFP in the regulation of autosensitization during ontogeny.     

Celiac disease association with CD8+ T cell responses: identification of a novel gliadin-derived HLA-A2-restricted epitope

One of the diagnostic hallmarks of the histological lesions associated with celiac disease is the extensive infiltration of the small intestinal epithelium by CD8(+) T cells of unknown Ag specificity. In this study, we report recognition of the gliadin-derived peptide (A-gliadin 123-132) by CD8(+) T lymphocytes from celiac patients. A-gliadin 123-132-specific IFN-gamma production and cytotoxic activity were detected in PBMCs derived from patients on gluten-free diet, but not from either celiac patients on gluten-containing diet or healthy controls. In contrast, A-gliadin 123-132-specific cells were isolated from small intestine biopsies of patients on either gluten-free or gluten-containing diets. Short-term T cell lines derived from the small intestinal mucosa and specific for the 123-132 epitope recognized human APC pulsed with either whole recombinant alpha-gliadin or a partial pepsin-trypsin gliadin digest. Finally, we speculate on a possible mechanism leading to processing and presentation of class I-restricted gliadin-derived epitopes in celiac disease patients.     

Genetic analysis of gliadin components in winter wheat using two-dimensional polyacrylamide gel electrophoresis

Summary Blocks of gliadin components found both in a number of varieties and in single F₂ grains of winter wheat intervarietal hybrids have been studied by two-dimensional electrophoresis combining electrophoresis in acidic aluminium-lactate buffer (pH3.1) and SDS-electrophoresis. Gliadin components (spots) have been shown to be inherited as linked groups (blocks), codominantly and in accordance with a gene dosage in triploid endosperm. Blocks include components differing in their electrophoretic mobility and molecular weight. Some allelic variants of blocks differ only in presence of few additional components or in the electrophoretic mobility of components with similar molecular weights; other variants may contain no similar components. Apparently, in the course of evolution, mutations in individual genes of gliadin-coding loci and processes changing the number of expressing genes and the sizes of their structural part occurred.     

Blocks of gliadin components in winter wheat detected by one-dimensional polyacrylamide gel electrophoresis

Summary Inheritance of gliadin components in winter wheat has been studied by one-dimensional polyacrylamide gel electrophoresis. Single F₂ grains from 36 intervarietal hybrid combinations have been analysed. The genetic analysis has revealed blocks, including 1–6 gliadin components, which are inherited as individual mendelian traits. About 80 variants of blocks have been detected. On the basis of the allelism test they are grouped into 6 series in accordance with the number of known gliadin-coding loci located on chromosomes of the homoeologous groups 1 and 6. Each series includes 8–18 blocks controlled by different alleles of one gliadin-coding locus. Blocks of components have been confirmed to be inherited codominantly in accordance to the gene dose in the triploid endosperm. The highest similarity between members of one series is observed in groups of blocks controlled by chromosomes ID and 6D. On the contrary, many blocks controlled by chromosomes 1A and 1B have no bands in common. The presented catalogue of blocks of components may be used to make up gliadin genetic formulae and to compare electrophoregrams obtained by different authors. Blocks of gliadin components are suitable genetic markers for use in revealing and studying heterogeneity of wheat varieties, in tracing their origin, in identifying recombinations, translocations and substitutions of the genetic material and in solving many other problems of the origin, evolution and selection of hexaploid wheat.     

T cell regulation of antibody responses: an I-A-specific, autoreactive T cell collaborates with antigen-specific helper T cells to promote IgG responses

In earlier studies we showed that hapten-specific inducer T cell clones specifically induce B cells from immunized donors to secrete IgM antibodies. However, IgG responses were not observed, suggesting that an additional signal(s) was required. In this report, we show that an autoreactive T cell clone produces a factor(s) that collaborates with antigen-specific inducer T cells to promote specific IgG responses. This factor is not restricted by antigen or MHC determinants and promotes IgG production both in vivo and in vitro. These findings suggest that autoreactive cells may play an important role in the regulation of isotype expression.     

Identification of the components of the murine T cell antigen receptor complex

In addition to the alpha and beta chains of the MHC class II restricted antigen receptor, monoclonal anti-receptor antibodies coprecipitate four polypeptides that appear to be noncovalently associated with the alpha-beta dimer of murine T cells. Included in the murine T cell antigen receptor complex are two glycoproteins of 25 kd (gamma) and 21 kd (delta) and two nonglycosylated polypeptides of 26 kd (epsilon) and 16 kd (zeta). The epsilon chain appears to possess an intrachain disulfide bond and zeta exists in the complex as a disulfide-linked homodimer. The delta chain is phosphorylated on a serine residue in response to T cell activation with antigen. In contrast, both delta and epsilon are phosphorylated in response to treatment of the T cells with phorbol 12-myristate 13-acetate. These polypeptides may play a role in the transduction of the signal(s) in T cell activation.     

T cell responses to Listeria monocytogenes

Owing to its unique intracellular biology that allows it to gain access to the host cell cytosol, Listeria monocytogenes induces potent, protective CD8 responses. The study of these responses has served as a paradigm to understand cell-mediated immunity to microbial pathogens. The availability of mutants specifically defective in unique aspects of the intracellular biology of this pathogen has greatly aided these studies. During the past few years, progress has been made to understand the contribution of the innate immune system and CD4 T cells in the generation of robust, long lasting CD8 responses to L. monocytogenes.     

T cell responses to Chlamydia trachomatis

Chlamydia trachomatis is the most common cause of bacterial sexually transmitted disease in the United States, as well as the leading cause of preventable blindness worldwide. Immunity to C. trachomatis requires a variety of cell types, each employing an array of effector functions. Recent work has demonstrated that both CD4+ and CD8+ T lymphocytes play a major role in protective immunity to C. trachomatis, predominantly through their secretion of interferon-gamma. This review describes the generation of acquired immunity to C. trachomatis and focuses on how T cells contribute to both protection and immunopathology.     

Four distinct patterns of memory CD8 T cell responses to chronic murine cytomegalovirus infection

CMVs are beta herpesviruses that establish lifelong latent infection of their hosts. Acute infection of C57BL/6 mice with murine CMV elicits a very broad CD8 T cell response, comprising at least 24 epitopes from 18 viral proteins. In contrast, we show here that the CD8 T cell response in chronically infected mice was dominated by only five epitopes. Altogether, four distinct CD8 T cell kinetic patterns were evident. Responses to some epitopes, including M45, which dominates the acute response, contracted sharply after day 7 and developed into stable long-term memory. The response to m139 underwent rapid expansion and contraction, followed by a phase of memory inflation, whereas the response to an M38 epitope did not display any contraction phase. Finally, responses against two epitopes encoded by the immediate early gene IE3 were readily detectable in chronically infected mice but near the limit of detection during acute infection. CD8 T cells specific for the noninflationary M45 epitope displayed a classic central memory phenotype, re-expressing the lymph node homing receptor CD62L and homeostatic cytokine receptors for IL-7 and IL-15, and produced low levels of IL-2. Responses to two inflationary epitopes, m139 and IE3, retained an effector memory surface phenotype (CD62L(low), IL-7Ralpha(-), IL-15Rbeta(-)) and were unable to produce IL-2. We suggest that immunological choices are superimposed on altered viral gene expression profiles to determine immunodominance during chronic murine CMV infection.     

Immunity to leprosy. II. Genetic control of murine T cell proliferative responses to Mycobacterium leprae

T cell proliferative responses to Mycobacterium leprae were measured after immunization of mice at the base of the tail with antigen and challenging lymphocytes from draining lymph nodes in culture with M. leprae. This T cell response to M. leprae has been compared in 18 inbred strains of mice. C57BL/10J mice were identified as low responder mice. The congenic strains B10.M and B10.Q were found to be high responders, whereas B10.BR and B10.P were low responders. F1 (B10.M X C57BL/10J) and F1 (B10.Q X C57BL/10J) hybrid mice were found to be low responders, similar to the C57BL/10J parent, indicating that the low responsive trait is dominant. Whereas B10.BR mice were shown to be low responders to M. leprae, B10.AKM and B10.A(2R) were clearly high responders, indicating that the H-2D region influences the magnitude of the T cell proliferative response. Gene complementation within the H-2 region was evident. Genes outside the H-2 region were also shown to influence the response to M. leprae. C3H/HeN were shown to be high responder mice, whereas other H-2k strains, BALB.K, CBA/N, and B10.BR, were low responders. Gene loci that influence the T cell proliferation assay have been discussed and were compared to known background genes which may be important for the growth of intracellular parasites. Because mycobacteria are intracellular parasites for antigen-presenting cells, genes that affect bacterial growth in these cells will also influence subsequent immune responses of the host.     

Polymorphism and inheritance of gliadin components controlled by chromosome 6A of spring durum wheat

The gliadin composition of 78 spring durum wheat varieties has been studied by one-dimensional (Al-lactate, pH 3.1) and two-dimensional (first dimension, Al-lactate, pH 3.1; second dimension, sodium dodecyl sulfate-polyacrylamide gel) electrophoresis. Analysis of hybrids has shown that all components of the alpha zone of gliadin spectra are inherited together as blocks and are, probably, coded for by a cluster of tightly linked genes located on chromosome 6A. Fourteen variants of gliadin blocks have been identified, which can be classified into five families on the basis of component composition. All families but one have analogues among chromosome 6A-controlled blocks of bread wheat. The results indicate that some of the genome A diploid genotypes that were ancestors of durum wheats were also ancestors of bread wheats and that polyploid wheats were produced by repeated allopolyploidization events, as has been suggested earlier.     

A dominant role for the immunoproteasome in CD8+ T cell responses to murine cytomegalovirus

Murine cytomegalovirus (MCMV) is an important animal model of human cytomegalovirus (HCMV), a β-Herpesvirus that infects the majority of the world's population and causes disease in neonates and immunocompromised adults. CD8(+) T cells are a major part of the immune response to MCMV and HCMV. Processing of peptides for presentation to CD8(+) T cells may be critically dependent on the immunoproteasome, expression of which is affected by MCMV. However, the overall importance of the immunoproteasome in the generation of immunodominant peptides from MCMV is not known. We therefore examined the role of the immunoproteasome in stimulation of CD8(+) T cell responses to MCMV - both conventional memory responses and those undergoing long-term expansion or "inflation". We infected LMP7(-/-) and C57BL/6 mice with MCMV or with newly-generated recombinant vaccinia viruses (rVVs) encoding the immunodominant MCMV protein M45 in either full-length or epitope-only minigene form. We analysed CD8(+) T cell responses using intracellular cytokine stain (ICS) and MHC Class I tetramer staining for a panel of MCMV-derived epitopes. We showed a critical role for immunoproteasome in MCMV affecting all epitopes studied. Interestingly we found that memory "inflating" epitopes demonstrate reduced immunoproteasome dependence compared to non-inflating epitopes. M45-specific responses induced by rVVs remain immunoproteasome-dependent. These results help to define a critical restriction point for CD8(+) T cell epitopes in natural cytomegalovirus (CMV) infection and potentially in vaccine strategies against this and other viruses.     

Baculovirus activates murine dendritic cells and induces non-specific NK cell and T cell immune responses

We previously reported that the baculovirus induced a strong host immune response against infections and malignancies. Among the immune cells, the dendritic cells were most strongly infected and activated by the baculovirus, although the exact mechanism remained unclear. Here, we evaluated the non-specific immune responses of bone marrow-derived dendritic cells (BMDCs) after infection by a wild-type baculovirus. MHC class I and II molecules and co-stimulation molecules (CD40, CD80, and CD86) on BMDCs were up-regulated by baculovirus infection. At the same time, the BMDCs produced pre-inflammatory cytokines (IL-6, IL12p70, and TNF-α) and IFN-α. NK cells showed IFN-γ production, CD69 up-regulation, and enhanced cytotoxicity when they were co-cultured with baculovirus-infected BMDCs. T cells showed IFN-γ production, CD69 up-regulation, and cell proliferation. Ex vivo analysis performed in vitro produced similar results. These findings suggested that baculovirus-infected dendritic cells induce non-specific immune responses and can be used as an immunotherapeutic agent against viral infections and malignancies, together with present therapeutic drug regimens.     

Antigen-specific CD8+ T cell responses in intestinal tissues during murine listeriosis

Infection of mice with Listeria monocytogenes induces a strong CD8+ T cell response, which is critical for the control of bacteria and for protection against re-infection. We analyzed the CD8+ T cell response in different intestinal tissues following oral and intravenous (i.v.) L. monocytogenes infection. After oral infection, bacterial titers in small intestine and large intestine, and the listeria-specific CD8+ T cell response in the mucosa of both parts of the intestine, were highly correlated. Oral infection of CD28-deficient mice revealed that this response was strictly dependent on CD28 costimulation. Significant listeria-specific CD8+ T cell responses also occurred in all intestinal tissues analyzed after i.v. infection or after DNA vaccination, indicating that the accumulation of listeria-specific CD8+ T cells in these tissues only partially depends on local antigen presentation and inflammation.     

VSL#3 probiotic preparation has the capacity to hydrolyze gliadin polypeptides responsible for Celiac Sprue

The native structure and distribution of gliadin epitopes responsible for Celiac Sprue (CS) may be influenced by cereal food processing. This work was aimed at showing the capacity of probiotic VSL#3 to decrease the toxicity of wheat flour during long-time fermentation. VSL#3 (10(9) cfu/ml) hydrolyzed completely the alpha2-gliadin-derived epitopes 62-75 and 33-mer (750 ppm). Two-dimensional electrophoresis, immunological (R5 antibody) and mass spectrometry analyses showed an almost complete degradation of gliadins during long-time fermentation of wheat flour by VSL#3. Gliadins non-hydrolyzed during fermentation by VSL#3 were subjected to peptic-tryptic (PT) digestion and analyzed by CapLC-ESI-Q-ToF-MS (Capillary Liquid Chromatography-Electrospray Ionization-Quadrupole-Time of Flight-Mass Spectrometry). Search for several epitopes showed the only presence of alpha2-gliadin-fragment 62-75 at a very low concentration (sub-ppm range). Compared to IEC-6 cells exposed to intact gliadins extracted from the chemically acidified dough (control), VSL#3 pre-digested gliadins caused a less pronounced reorganization of the intracellular F-actin which was mirrored by an attenuated effect on intestinal mucosa permeability. The release of zonulin from intestinal epithelial cells treated with gliadins was considerably lower when digested with VSL#3. Agglutination test on K 562 (S) cells showed that the PT-digest of wheat flour treated with VSL#3 increased the Minimal Agglutinating Activity of ca. 100 times. Wheat proteins were extracted from doughs and subjected to PT digestion. Compared to PT-digest from chemically acidified dough, celiac jejunal biopsies exposed to the PT-digest from the dough fermented by VSL#3 did not show an increase of the infiltration of CD3(+) intraepithelial lymphocytes. Proteolytic activity by probiotic VSL#3 may have an importance during food processing to produce pre-digested and tolerated gliadins for increasing the palatability of gluten-free products.     

Chromosomal control of wheat gliadin protein epitopes: analysis with specific monoclonal antibodies

Summary The genetic relationships between small clusters of monomeric alcohol-soluble wheat (Triticum aestivum L.) grain storage proteins (gliadins) were studied using a panel of monoclonal antibodies and immunoblotting, ELISA, and RIA methods. Use of Chinese Spring nullisomic-tetrasomic lines showed that several narrow-specificity antibodies bound specifically to gliadins encoded by genes located on a single chromosome. In at least one case, antibodies bound to genetic blocks of gliadins, indicating that these block members have structural homology. However, often not all gliadins of a block were recognized by an antibody. For broad-specificity antibodies and some narrow-specificity antibodies, structural genes on several chromosomes were important. Studies with several primitive wheat species indicated that, while antibodies usually bound gliadins from the same genome in bread and primitive wheats, antibodies sometimes bound proteins of quite differing mobilities in the two wheat types. Use of antibodies to identify gliadin blocks is simpler than block analysis based on performing crosses, and should be of value in monitoring genotype/end-use quality relationships.     

High selectivity of human tissue transglutaminase for immunoactive gliadin peptides: implications for celiac sprue.

Celiac Sprue is an HLA DQ2 (or DQ8)-associated autoimmune disorder of the human small intestine that is induced by dietary exposure to wheat gliadin and related proteins from barley, rye, and possibly other food grains. Recently, tissue transglutaminase (tTGase)-catalyzed deamidation of gliadin peptides has been shown to increase their potency for activating patient-derived, gliadin-specific T cells, suggesting that tTGase plays a causative role in the onset of an inflammatory response to toxic food grains. To dissect the molecular recognition features of tTGase for gluten derived peptides, the regioselectivity and steady-state kinetics of tTGase-catalyzed deamidation of known immunogenic peptides were investigated. The specificity of recombinant human tTGase for all immunogenic peptides tested was comparable to and, in some cases, appreciably higher than the specificity for its natural substrate. Although each peptide was glutamine-rich, tTGase exhibited a high degree of regioselectivity for a particular glutamine residue in each peptide. This selectivity correlated well with Q --> E substitutions that have earlier been shown to enhance the immunogenicity of the corresponding gliadin peptides. The specificity of tTGase toward homologues of PQPQLPY, a sequence motif found in immunodominant gliadin peptides, was analyzed in detail. Remarkably, the primary amino acid sequences of wheat-, rye-, and barley-derived proteins included many single-residue variants of this sequence that were high-affinity substrates of tTGase, whereas the closest homologues of this sequence found in rice, corn, or oat proteins were much poorer substrates of tTGase. (Rice, corn, and oats are nontoxic ingredients of the Celiac diet.) No consensus sequence for a high-affinity substrate of tTGase could be derived from our data, suggesting that the secondary structures of these food-grain peptides were important in their recognition by tTGase. Finally, under steady-state turnover conditions, a significant fraction of the tTGase active site was covalently bound to a representative high-affinity immunogenic gliadin peptide, suggesting a common mechanism by which cells responsible for immune surveillance of the intestinal tract recognize and generate an antibody response against both gliadin and tTGase. In addition to providing a quantitative framework for understanding the role of tTGase in Celiac Sprue, our results lay the groundwork for the design of small molecule mimetics of gliadin peptides as selective inhibitors of tTGase.     

Activation and inactivation of antiviral CD8 T cell responses during murine pneumovirus infection

Pneumonia virus of mice (PVM) is a natural pathogen of mice and has been proposed as a tractable model for the replication of a pneumovirus in its natural host, which mimics human infection with human respiratory syncytial virus (RSV). PVM infection in mice is highly productive in terms of virus production compared with the situation seen with RSV in mice. Because RSV suppresses CD8 T cell effector function in the lungs of infected mice, we have investigated the nature of PVM-induced CD8 T cell responses to study pneumovirus-induced T cell responses in a natural virus-host setting. PVM infection was associated with a massive influx of activated CD8 T cells into the lungs. After identification of three PVM-specific CD8 T cell epitopes, pulmonary CD8 T cell responses were enumerated. The combined frequency of cytokine-secreting CD8 T cells specific for the three epitopes was much smaller than the total number of activated CD8 T cells. Furthermore, quantitation of the CD8 T cell response against one of these epitopes (residues 261-270 from the phosphoprotein) by MHC class I pentamer staining and by in vitro stimulation followed by intracellular IFN-gamma and TNF-alpha staining indicated that the majority of pulmonary CD8 specific for the P261 epitope were deficient in cytokine production. This deficient phenotype was retained up to 96 days postinfection, similar to the situation in the lungs of human RSV-infected mice. The data suggest that PVM suppresses T cell effector functions in the lungs.     

棉铃虫雌蛾对小麦花挥发性气味的触角电位反应

测定了棉铃虫Helicoverpa armigera处女雌蛾和交配雌蛾对小麦花中单一或多个挥发性物质的EAG反应。在单组分或多组分的测定中,棉铃虫处女蛾和交配蛾在P=0.05水平上并不存在显著性差异,但在混合的绿叶气味物质中加入含量为1×10^-5～5×10^-5（体积比）的正庚醛时,交配蛾对混合物的触角电位反应显著高于处女蛾的反应（P<0.05）。