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In order to examine how rat erythrocytes stimulate erythrocyte autoantibody production at the molecular level, we have identified rat erythrocyte antigens by immunoprecipitation and western blotting using monoclonal antibodies and antisera. A novel non-radioactive immunoprecipitation technique was used, which employed biotin as a label and a luminescent detection system. The new method was validated by comparison with conventional immunoprecipitation using 125I. Glycophorins of relative molecular mass (Mr) 81,000 and 38,000 were found to be the major antigenic components of rat erythrocytes, while band 3 (the most abundant erythrocyte membrane protein) was not recognized by rat-specific antibodies. The same surface antigens were recognized by sera from mice producing erythrocyte autoantibodies and by sera from mice in which autoantibody production was suppressed. Nine other minor rat-specific antigens were identified by blotting, ranging in Mr from 23,000 to 147,000. Analysis of the integral membrane proteins of rat and mouse erythrocytes by sodium dodecyl sulphate (SDS) electrophoresis followed by silver or periodic acid-Schiff (PAS) stains revealed differences between the glycophorins, but not between rat and mouse band 3. Thus, the major antigenic differences correspond to discernible biochemical differences between rat and mouse erythrocyte sialoglycoproteins.

MHC class I molecules on the cellular surface display peptides that either derive from endogenous proteins (self or viral), or from endocytosis of molecules, dying cells or pathogens. The conventional antigen-processing pathway for MHC class I presentation depends on proteasome-mediated degradation of the protein followed by transporter associated with antigen-processing (TAP)-mediated transport of the generated peptides into the endoplasmic reticulum (ER). Here, peptides are loaded onto MHC I molecules before transportation to the cell surface. However, several alternative mechanisms have emerged. These include TAP-independent mechanisms, the vacuolar pathway and involvement of autophagy. Autophagy is a cell intrinsic recycling system. It also functions as a defence mechanism that removes pathogens and damaged endocytic compartments from the cytosol. Therefore, it appears likely that autophagy would intersect with the MHC class I presentation pathway to alarm CD8 + T cells of an ongoing intracellular infection. However, the importance of autophagy as a source of antigen for presentation on MHC I molecules remains to be defined. Here, original research papers which suggest involvement of autophagy in MHC I antigen presentation are reviewed. The antigens are from herpesvirus, cytomegalovirus and chlamydia. The studies point towards autophagy as important in MHC class I presentation of endogenous proteins during conditions of immune evasion. Because autophagy is a regulated process which is induced upon activation of, for example, pattern recognition receptors (PRRs), it will be crucial to use relevant stimulatory conditions together with primary cells when aiming to confirm the importance of autophagy in MHC class I antigen presentation in future studies., (© 2020 The Authors. Scandinavian Journal of Immunology published by John Wiley & Sons Ltd on behalf of The Scandinavian Foundation for Immunology.)

This paper describes a procedure for analysing multiple antibody reactivities that explores a commercially available immunoblot system, and is based on a double staining of nitrocellulose membranes, revealing both antibody reactivities and the migration position of the blotted proteins in the membrane. Quantification of both stainings by densitometry allowed the accurate superposition of the immunoreactivity and total protein profiles of each lane. Moreover, the protein stainings of the different lanes could be adjusted with a simple-scale transformation algorithm, correcting for possible distortions during electrophoretic migration, and allowing for the precise comparison of the immunoreactivity profiles in different lanes. The procedure is discriminatory enough to identify unique reactivity patterns in random pools of 104 activated B cells, and to define strain-specific natural antibody repertoires. The utility of this immunoblot method as an assay for simultaneously scoring multiple reactivities to hundreds of antigens in complex mixtures of antibodies, and thus defining antibody repertoires in a global manner, is discussed. [ABSTRACT FROM AUTHOR]

Protective immunity against mycobacteria is dependent on antigen-specific T cells. The antibodies induced upon immunization with mycobacteria have no apparent role in host protection, Serological techniques have detected some antigens that are also recognized by human T cells but may fail to recognize others. Potentially, there may be differences in the epitopes seen by the T and B cell anti-mycobacterial antigen repertoires. We have screened the different components of sonicated BCG or Mycobacterium leprae that were separated according to their molecular weight (MW) by SDS-PAGE and then electroblotted on nitrocellulose paper. The blots were cut into squares and tested directly in a T cell proliferation assay. Our results indicate that peripheral T cells of healthy leprosy patient contacts respond preferentially to the lower MW (<70,000) and not the higher MW fractions of M. leprae and BCG, in contrast to the humoral response of these same individuals. The most important fractions in inducing a lymphoproliferative response were in the regions of 11-16 kDa of BCG and M. leprae and to the 22-26 kDa region of M. leprae. These fractions appeared to represent molecular weight regions that were in some instances clearly distinct from previously defined antigens. It was further shown that lymphoproliferation in response to mycobacterial fractions correlated with the production of gamma interferon, a lymphokine required for macrophage activation and elimination of mycobacteria. These studios allow the direct assessment of antigens involved in protective T cell-mediated immunity, and should be helpful in selecting relevant antigens for skin testing and immunization. [ABSTRACT FROM AUTHOR]

Two strains of mice were genetically selected for susceptibility (TS-Ab/HetS strain) or resistance (TR-Ab/HetS strain) to oral tolerance of the humoral response by using ovalbumin (OVA). The progressive interstrain divergence produced by bi-directional selective breeding during 15 generations demonstrated the polygenic nature of oral tolerance. This paper shows the humoral and delayed-type hypersensitivity (DTH) responses, after intragastric administration of OVA and subsequent immunization with that immunogen in complete Freund adjuvant (CFA). Only the TS-Ab/HetS mice were tolerant for immunoglobulin (Ig)G production with its tolerance degree being the same as that obtained when Al ((OH)(3)) was employed. The DTH reactivity was not correlated to the antibody responsiveness, because both the TS-Ab/HetS and TR-Ab/HetS strains had their DTH reactions suppressed. The cyclophosphamide (Cy) pretreatment prevented DTH suppression on TR-Ab/HetS but do not in TS-Ab/HetS mice. Interstrain difference was also observed for the splenic index in the Cy-treated groups, although the number of splenocytes was the same. Flux cytometry cell analysis showed the recovery of CD3(+) cell numbers in both strains, but only the TR-Ab/HetS mice had their CD4/CD8 pattern restored. These results suggest: firstly, the independent control of DTH and humoral tolerance responsiveness; secondly no support for the clonal anergy concept; and thirdly the matrix proteins neo-synthesis after Cy treatment may facilitate the tolerance abrogation.

The clinical manifestations of human Chagas disease are associated with distinct and complex host-parasite interactions that directly involve the host's immune system. In this study, we analysed the relationship between the production of intracytoplasmic cytokines after in vitro stimulation with the recombinant antigens CRA (cytoplasmatic repetitive antigen) or FRA (flagellar repetitive antigen) from Trypanosoma cruzi and the chronic cardiac or indeterminate clinical forms of Chagas disease. The chagasic patient groups consisted of 39 individuals, selected at the Chagas Disease Unit of the Oswaldo Cruz University Hospital, whom presented either a cardiac form without cardiac dilatation (CARD 1), cardiac form with cardiac dilatation (CARD 2) or indeterminate form (IND). Blood samples were obtained from these patients and cultured in the presence of CRA or FRA. The cytokines produced by lymphocytes and monocytes after antigen stimulation were analysed by flow cytometry. Our results showed that the IFN-γ and TNF-α, produced by CD8+ T lymphocytes after in vitro stimulation with CRA, differed among chagasic patients with CARD 1, CARD 2 or IND. We propose that these cytokines could be utilized as immunological markers for clinical cardiac forms of Chagas disease. In a prospective study of patients presenting IND and CARD 1, the assay performed in this paper could serve as a tool to monitor therapeutic interventions, thus improving the patient's quality of life. [ABSTRACT FROM AUTHOR]

In this paper, the authors show that liquid-liquid partition chromatography in an aqueous two-phase system offers unique possibilities of comparing the overall surface properties of intact antibodies in solution before and after binding of antigen. The authors demonstrate that the surface properties of antigen-antibody complexes are dependent on the variable regions of the antibodies, the nature of the antigen and/or possible conformational changes induced by antigen binding. Thus, each antigen-IgG antibody pair formed one type of complex with respect to the exposed dominant surface. The antigen-binding sites of IgG antibodies were exposed and dominant even after binding of hapten or hapten-carrier. In contrast, the antibody-combining sites were concealed upon protein binding and the exposed surfaces of the protein-antibody complexes were related mainly to those of the antigen. IgA1, IgA2, IgE and IgM formed, in comparison to the IgG, hapten-antibody complexes which exhibited surface properties that could be related to both the antigen-binding sites and Fc parts of the antibodies. Moreover, the results indicated that antigen-induced conformational changes occurred in either IgA1, IgA2, IgE, or IgM, but not in IgG1, -2, -3 and -4, making the surfaces of their heavy chain constant regions more similar.

Tuberculosis (TB) is the largest single infectious cause of human mortality. The incidence of TB has remained high in most of the developing world and the disease has recently re-emerged as a public health problem in industrialized countries. The development of a new improved TB vaccine is a highly prioritized international research area, which has been further stimulated by the appearance of multi-drug resistant strains of Mycobacterium tuberculosis. The present status of the attempts to characterize the protective immune response to TB will be reviewed with special emphasis on recent progress in the identification and characterization of target molecules recognized by protective cells. This paper will focus on proteins released from live bacteria and discuss their role in the host-pathogen interaction and the ongoing attempts to use these molecules in TB subunit vaccines. [ABSTRACT FROM AUTHOR]

In a previous work the authors demonstrated that immunization with Schistosoma mansoni 28-kDa glutathion-S-transferase (Sm28GST) was able to reduce hepatic damage in infected mice and that the adoptive transfer of Sm28GST-specific T cells reproduced the protective effect obtained with the recombinant molecule. In the present paper, the authors show that Sm28GST is also able to stimulate an antigen-specific, cytotoxic T-cell response against Sm28GST-pulsed P815 target cells in normal mice and that effector cells induced in vivo were classical Class I MHC-restricted CD8+ lymphocytes. The authors found no spontaneous CTL activity against Sm28GST-pulsed target cells during the course of the infection by S. mansoni although Sm28GST is expressed at different developmental stages of the parasite. It was observed, however, that immunization with Sm28GST is sufficient to elicit a significant level of CTL response for 6 weeks in infected mice. The role of these Class I MHC-restricted CD8+ lymphocytes in the protection observed precisely at the same period in immunized mice remains to be elucidated. The authors also observe that immunization with the lipopeptidic form of the C-terminal peptide of the molecule (190-211 peptide) led to a CTL activation comparable to that observed after immunization with the whole molecule demonstrating the feasibility of using a synthetic lipopeptide as immunogen for a CTL response against Sm28GST epitopes. Moreover, like Sm28GST-specific CTLs, 190-211 lipopeptide-specific cells were also Class I MHC-restricted lymphocytes. [ABSTRACT FROM AUTHOR]

The mechanisms associated with the modulation of immune response in the chronic phase of human schistosomiasis mansoni infection are complex and involve many cell types. In the present paper the authors demonstrate that antigenic stimulation of peripheral blood mononuclear cells (PBMC) from chronic-intestinal schistosomiasis mansoni patients with polyacrylamide beads (PB) conjugated to Schistosoma mansoni soluble egg antigens (PB-SEA) or adult worm antigen preparation (PB-SWAP) were able to induce a statistically significant increase on the in vitro multinucleated giant cell (MGC) formation after the 15th day in culture. A correlation between an increase in the number of MGC and a decrease in in vitro granuloma formation index to PB-SEA and PB-SWAP was observed. Moreover, the authors demonstrated a down-regulation of lymphocyte proliferative responses to S. mansoni antigens, during the differentiation pathway of monocytes towards MGC formation, due to a decrease in the antigen-presenting capacity of these cells. These phenomena also correlate with a concomitant decrease in the expression of HLA-DR and CD54 adhesion molecules on the surface of MGC. The results suggest that differentiation of monocytes to MGC may be one of the immunoregulatory mechanisms involved in the down-regulation of the granuloma reaction against S. mansoni eggs. [ABSTRACT FROM AUTHOR]

In this paper, the authors report the use of liquid-liquid partition chromatography (LLPC) in an aqueous polyethylene glycol (PEG)/dextran two-phase system to compare the surface properties (partition properties) of human antibodies and fragments thereof. The surface properties of all the monoclonal antibodies of different classes and subclasses investigated were within the same broad range as that observed for the polyclonal antibodies and no relationship was found between the exposed surfaces of the immunoglobulins (Ig) and their heavy chain isotype. Moreover, Fc fragments from various IgG1, 2 and 4 myeloma proteins were found to exhibit similar surface properties. Employing chimeric antibodies with identical variable regions the authors found that intact IgG1, 2 and 4 displayed identical surface properties, while the corresponding IgA1, IgA2, IgG3, IgE and IgM antibodies differed both from each other and from the IgGs. The surface properties of chimeric IgG3 could be made similar to those of the IgG1, 2 and 4 chimers by partially reducing the length of the hinge section, but new differences in surface properties appeared when their hinges were of similar length. Thus, LLPC can be used to detect differences or similarities in the surface properties of the antigen-binding regions as well as the Fc part in the various isotypes. This can shed light on biological activities such as antigen binding and effector function. [ABSTRACT FROM AUTHOR]

Recent studies have provided considerable insight into the mechanism of BCR-mediated B-cell activation, but the inhibitory signals transferred by FcγRs leading to down-regulation of BCR-activated B lymphocytes are not clarified yet. In the present paper the authors give an overview on new findings regarding BCR structure and signal transduction mechanisms induced by the B-cell antigen receptor complex and outline, partly based on their own observations, the possible mechanisms resulting in FcγR-mediated inhibition of B cells. [ABSTRACT FROM AUTHOR]

Given that the associative recognition of antigen (two signal) model permits a certain level of 'autoreactivity'. Faro & Carneiro argue that this unavoidable "autoreactivity" is required as part of the regulation of the general physiology of the animal. This paper gives the author's reply pointing out two facets of the functioning of the immune system. (I) Under an associative recognition of antigen model, the scenarios of B-cell activation postulated by them are ruled out. (2) There is no way to use a destructive ridding effector function to regulate the self/non-self discrimination because autoimmune reactivity would accompany any postulated regulatory mechanism. [ABSTRACT FROM AUTHOR]

The Ro/SSA and La/SSB antigens are common targets for autoantibodies found in the sera of patients with Sjögren's syndrome and SLE. The anti-Ro/SSA and anti-La/SSB antibodies often appear together but are not cross-reactive. This paper describes the humoral autoimmune response to the Ro/SSA 60 kDa protein moiety with respect to the presence of IgM and IgG1-4 antibodies. IgM antibodies to the Ro 60 kDa protein coexisted with IgG anti-Ro 60 kDa antibodies in nearly half of the sera. A similar fraction also contained IgM anti-La/SSB antibodies. The frequency of sera with IgM antibodies of both specificities was that expected from random overlap. A predominating IgG1 anti-Ro 60 kDa response was found in all patients, but anti-Ro 60 kDa antibodies of the other IgG subclasses were present also in a high number of sera. This is in contrast to the reported IgG subclass distribution of anti-La/SSB antibodies. Mapping of IgM and IgG1-4 antibody recognition of different parts of the Ro 60 kDa protein was also performed. IgM and IgG1-4 antibodies of all sera reacted with the central part of the Ro 60 kDa protein, encompassing amino acid residues 181-320. [ABSTRACT FROM AUTHOR]

Brown Norway (BN) rats are poor responders to T-cell mitogens and alloantigens when compared to Lewis (LEW) rats. This is dependent partly upon a defect in IL-2 production. The TH2-mediated immune abnormalities observed in BN rats injected with mercuric chloride (HgCl2) are self-limited and it is probable that this regulation phase involves TH1-like cells. This paper reports on a study of the ability of lymph node cells (LNC) from normal BN and LEW rats and from HgCl2-injected BN rats to produce IL-2 and to proliferate when stimulated in vitro by Con A or alloantigens in mixed lymphocyte reaction (MLR), as well as to develop a cytotoxic T lymphocyte (CTL) response to alloantigens. This study will confirm that LNC from BN rats proliferate less than LNC from LEW rats, that the former produce less IL-2 than the latter, and that the proliferative response is restored partially after addition of IL-2. In addition, it is shown (1) that the CTL response is defective in normal BN rats when compared to that of normal LEW rats, and (2) that, after the second week of HgCl2, injections, the proliferative responses to Con A and alloantigens are improved as well as IL-2 production, and a complete restoration of CTL function is observed. These results show that normal BN rats are deficient in the induction of TH1-like cells and that, from the second week of HgCl2 injections, these TH1 functions improve. [ABSTRACT FROM AUTHOR]

Two main ideas have been put forward lo explain the unexpectedly low anti-hapten antibody titres which can result from pre-priming a mouse with carrier before hapten-carrier immunization. The first involves the interaction of a network of idiotype-specific suppressor T cells, the second instead arguing for the role of intrinsie B-cell anergy. This paper proposes that the data available can equally be interpreted as reflecting the suboptimal interaction between T and B cells at differing stages of maturity, provided that memory B cells can be divided into two subsets. Further, it is suggested that these considerations must be taken into account in the analysis of B-cell anergy in receptor transgenic mice. [ABSTRACT FROM AUTHOR]

The heavy and light chain nucleotide sequences of 17 monoreactive and polyreactive rheumatoid factors largely derived from the inflamed synovial tissue of two patients with rheumatoid arthritis are described. Some of these sequences have been the subject of a previous report from our laboratories. Additionally, a few rheumatoid factors from the peripheral blood of patients with systemic lupus erythematosus and Sjogren's syndrome as well as a normal individual are included. A review of our previous results as well as the new data provided within this paper lead to the following major conclusions: (1) Rheumatoid factors and polyreactive antibodies derive from a diverse array of VH and VL gene segments; (2) While many rheumatoid factors and polyreactive antibodies are direct or nearly direct copies of germline genes, some show clear evidence of somatic mutation; (3) The CDR3 of all of these antibodies is extraordinarily diverse in length and composition. Certain `restrictions' do appear in this very large sample: (a) the polyreactive antibodies are exclusively lambda, and (b) there seems to be a preponderance of a particular subset of VH3 genes beyond that one would expect based on random utilization. [ABSTRACT FROM AUTHOR]

Antigen 60 (A60), a member of the thermostable macromolecular antigen family (TMA) and main component of old tuberculin and purified protein derivative (PPD), has been purified from the cytoplasm of Mycobacterium bovis BCG; its structure and metabolism have already been described. In the present paper, the action of A60 on humoral immunity has been analysed by an ELISA type immunoassay, and that on cellular immunity by the mouse footpad swelling test. Injection of very low A60 doses into unprimed mice produced an undetectable level of anti-A60 antibodies: the effect or a booster inoculation was not appreciable in the absence of incomplete Freund's adjuvant, but was evident when the latter was added. Higher doses of the antigen produced an appreciable primary response, and a sharp and long-lasting secondary response, which had a 10-fold higher intensity in the presence of incomplete adjuvant. No detectable delayed hypersensitivity reactions were observed in unprimed mice after footpad injection of A60, whereas clear responses were elicited in primed mice. This effect was more pronounced when the footpad was injected after a secondary response than after a primary response, and it was invariably magnified by incomplete adjuvant. It is concluded that A60 is a powerful immunogen, which is able to induce primary and secondary responses and delayed hypersensitivity reactions, effects that are adjuvant-modulated and develop concurrently. [ABSTRACT FROM AUTHOR]

We have developed a new assay for quantitative detection of all helper T cells that can induce normal B lymphocytes to proliferation and Ig secretion. To establish the optima] assay conditions, we have used cloned I helper cells of defined specificities that had previously been shown to activate normal B lymphocytes expressing the specific antigen(s) on direct cellular interactions. As shown in this paper, "irrelevant" B lymphocytes-that is, those that do not express either antigen or restriction elements recognized by the effector helper T cells-can also be induced in the presence of appropriate concentrations of pokeweed mitogen which are not mitogenic for the "target" B lymphocytes. 'Nonspecific" plaque-forming cell responses are of the same magnitude as those provided to specifically triggered targets and equal or better than those induced by lipopolysaecharide. The assay is highly sensitive and enables "semiquantitative' detection of less than 20-30 effector T cells per culture. Since effeetor helper T cells can be detected regardless of the clonal specificity, the assay appears useful for quantitative studies of various populations of T helper cells displaying mixed specificities and for the classification of cells with unknown functions. [ABSTRACT FROM AUTHOR]

The differentiation of Ig+ B cells into plaque-forming cells is dependent on antigen and factors produced by T cells and/or macrophages. We describe here the production of T-cell factors termed lymphocyte promotor factors (LPF). A foetal calf serum-specific T-cell line and its clones synthesize LPF, which is defined as factors that polyclonally stimulate normal spenic T cells to differentiate into cytotoxic T lymphocytes (T-LPF) and normal splenic B cells to differentiate into plaque-forming cells into (PFC) (B-LPF) in the apparent absence of specific antigen. The proliferation of and the B-LPF production of all T-cell clones tested were foetal calf serum-specific and I-Bb-restricted. Some of these clones produced only T-LPF, some clones produced only B-LPF, and some clones produced both T-LPF and B-LPF. B-LPF stimulate the polyclonal differentiation of Ig+ B cells into PFC without the apparent need for helper T cells, is different from T-LPF, and induces almost exclusively IgM PFC. The B-LPF described in the present paper are compared with previously described T-cell factors, which stimulate antigen-specific B-cell responses or bystander B-cell responses. The conclusion is that B-LPF are probably different from B-cell growth factors, T-cell replacing factors, allogeneic effector factors, and interleukin 2. [ABSTRACT FROM AUTHOR]

Cellular cytotoxicity reactions can be studied in a manner analogous to that used lo measure enzyme activity. This approach yields two parameters : Vmax, the maximal rate of target cell lysis that can be achieved by the lymphocyte preparation tested, and KMapp, the apparent Michaelis constant. By analogy to many enzyme-catalysed reactions, KMapp values for cytotoxicity reactions have generally been interpreted in terms of dissociation constants for the interaction of receptor sites on effector cells with antigens on the target cells. In this paper we demonstrate that experimentally determined KMapp values for natural or antibody dependent cytotoxicity reactions arc approximately equal to the concentration of NK or K effector cells in the lymphocyte preparation tested. This result makes possible the simultaneous determination of both effector cell frequency and lytic activity in a given lymphocyte preparation. [ABSTRACT FROM AUTHOR]

Immune responsiveness of guinea pigs to dinitrophenyl-poly-L-lysine and to the lysine rich random co-polymer of L-glutamic acid and L-lysine are both controlled by a single gene, the `poly-L-Iysine gene'. This paper reviews recent experiments which demonstrate that these two antigens specifically compete with one another for being presented to T cells by the same antigen-presenting cells. This finding is interpreted to mean that antigens to which responsiveness is controlled by the same single gene compete for the Ir gene product of antigen-presenting cells. The review discusses if the products of the immune response genes-presumably the Ia antigens-may constitute a third specific antigen recognition system. It further speculates if this idea may help to provide insight into the phenomenon of histocompatibility-restriction and into the nature of the mixed leucocyte reaction. [ABSTRACT FROM AUTHOR]

This paper reviews the present knowledge of B-L antigens encoded by the chicken B complex as regards to the following aspects: (1) identification and cellular expression, (2) structural studies, (3) evidence for two distinct populations of B-L antigens, (4) mapping of B-L loci of the B complex, (5) B-L and immune response, and (6) the role of the B-L antigens for the control of mixed lymphocyte reactions (M LR) and graft-versus-host (GVH) reactions. It is concluded that B-L antigens of the chicken exhibit extensive homology with mammalian La antigens. A genetic map of the B complex is presented. [ABSTRACT FROM AUTHOR]

Optimal proliferative response of T lymphocytes to purified protein derivative of tuberculin (PPD) in vitro requires that antigen be presented by autologous macrophages or allogeneic macrophages sharing HLA-D/DR determinants with the T cell donor. In some cases, however, T cells may respond to a limited extent to PPD in association with macrophages expressing different HLA-D/DR determinants. In this paper experiments are presented where various combinations of T cells and HLA-D/DR disparate macrophages were stimulated with PPD. We often found a strong PPD-specific response in HLA-D/DR-incompatible combinations in which the macrophages carried HLA-DR antigens known from serological studies to be cross-reactive with those of the T cell donor than in combinations in which this was not the case. Thus, the cross-reactions detected by serology may sometimes be reflected on a functional level in T lymphocyte/macrophages cooperation. [ABSTRACT FROM AUTHOR]

Polymyxins are known to be inhibitors of certain polyclonal B cell activators such as lipopolysaccharide and dextransulphate. However, increased specific responses to hapten-coupled mitogens have been reported after the addition of polymyxins to superoptimal conjugate doses. In this paper we have studied the effect of polymyxin B on superoptimal polyclonal doses of lipopolysaccharide. Results similar to those reported for hapten-lipopolysaccharide conjugates were obtained. Polymyxin B was also found to exert adjuvant properties for a primary immune response to sheep erythrocytes and to be a thymus-independent antigen. [ABSTRACT FROM AUTHOR]

The amino acid sequence of the COOH-terminal fragment TM2 of cod allergen M has been determined by the dansyl-Edman procedure. The fragment consists of 38 residues (mol. wt. 3,854), and the sequence shows great homology with the sequence of the corresponding fragment of white muscle albumin from carp, hake, and pile. Unique for the TM2 (and allergen M) from cod is the presence of a tryptophyl residue at position 5 from the COOH-terminal end. [ABSTRACT FROM AUTHOR]

Lung transplant (LTX) patients are at high risk of cytomegalovirus (CMV) infection, which is often associated with high mortality and morbidity. Reactivation of CMV causes cell injury due to the cytopathic effect of viral replication and triggering of T cell immunity. The aim of this study was to compare expression of immune checkpoints (ICs) (PD‐1, CTLA‐4, LAG‐3 and TIGIT) in CD4, CD8 and CD56 and activation markers CD137, CD154 and CD69 of end‐stage patients awaiting lung transplant. Eighteen pre‐LTX positive for anti‐CMV IgG titres and 18 healthy subjects were enrolled. IC and activation markers have been evaluated through flow cytometric analysis in HC and pre‐LTX patients. Reactive (QF+) and unreactive (QF−) patients were stratified according to QuantiFERON‐CMV assays. ICs' and activation markers' expression were determined before and after in vitro stimulation with pp‐65 and IE‐1 antigens. Lower expression of PD‐1 was observed in CD4 and CD8 cells of pre‐LTX patients than controls, whereas CTLA4 appeared upregulated in CD56 and CD8 cells. TIGIT is increased on the surface of CD4, CD8 and NK cells after peptide stimulation in QF‐negative patients and PD‐1 is only downregulated after stimulation in the QF‐positive patients. This study provides new evidence of immune dysregulation in patients with end‐stage lung disorders, particularly in relation to immune checkpoint cell biology. The change in QF+ mostly happens on cytotoxic cells NK and CD8, while the changes in QF− were observed in adaptive immune cells, including CD4 and CD8. [ABSTRACT FROM AUTHOR]

Presents several papers at an international workshop on the role of dendritic and Langerhans cells in immunoregulation in Oslo, Norway. Initiation and regulation of immune responses; Importance of class II antigens for the restriction of local immune responses; Advances in the basic understanding of local immune reactions.

Familial clustering of the skin disease primary hyperhidrosis suggests a genetic component to the disease. The human leucocyte antigen (HLA) is implicated in a range of diseases, including many comorbidities to hyperhidrosis. No study has investigated whether the HLA genes are involved in the pathogenesis of hyperhidrosis. We, therefore, compared HLA alleles in individuals with and without hyperhidrosis in this study of 65 000 blood donors. In this retrospective cohort study, we retrieved information on individuals with and without hyperhidrosis using self‐reported questionnaires, the Danish National Patient Registry and the Danish National Prescription Registry on participants recruited to the Danish Blood Donor Study between 2010 and 2019. Association tests using logistic regression were conducted for each HLA allele corrected for sex, age, body mass index, smoking and principal components. Overall, 145 of 65 795 (0.2%) participants had hospital diagnosed hyperhidrosis. Similarly, 1379 of 15 530 (8.9%) participants had moderate‐severe self‐reported hyperhidrosis, of whom 447 (2.9%) had severe self‐reported hyperhidrosis. Altogether, 28 participants had both hospital diagnosed and moderate‐severe self‐reported hyperhidrosis. Severe self‐reported hyperhidrosis was associated with HLA‐A*80:01 (adjusted odds ratio 26.97; 95% confidence interval 5.32‐136.70; n = 7; P <.001). Moderate‐severe self‐reported hyperhidrosis and hospital diagnosed hyperhidrosis were not associated with any HLA. The association between hyperhidrosis and HLA‐A*80:01 was based on a very small number of cases and not replicated in other patient subsets, and therefore likely a chance finding. Thus, this study suggests that genes other than the HLA are involved in the pathogenesis of hyperhidrosis. [ABSTRACT FROM AUTHOR]

The murine T-cell clone E11 isolated from a primary H-2k histocompatible one-way mixed lymphocyte culture of B10.BR anti C3H/Tif spleen cells was used to study multidirectional interactions in strong stimulatory Mls disparate responses. Several parameters have recently been studied, and proliferation of T cells upon stimulation by macrophages or B cells of Mls-disparate stimulator cells, induction of differentiation of B cells of stimulator strains but inhibition of their macrophage differentiation, and the inhibition by E11 T cells of the production of a mitogenic mediator by Mls-disparate spleen cells have been found. As shown in this paper, these phenomena can be explained by an Mlsa, d, e specific induction of gamma-interferon (IFN-gamma) production in the responder B10.BR (Mlsb) E11 T cells. It is suggested that IFN-gamma, as a regulator of feedback mechanisms plays a critical role in Mls disparate T-cell stimulation.

Intrasplenic immunization with antigen immobilized on nitrocellulose (NC) paper results in an immune response with nanogram amounts of antigen without the use of Freund's adjuvant. Comparing the sensitivity of the intrasplenic immunization with intraperitoneal or subcutaneous administration, we found that the intrasplenic method resulted in more positive animals with higher titres than did the other techniques with the smallest amount of antigen (70 ng of bovine serum albumin). With larger amounts of albumin the intraperitoneal method yielded the highest titres. No loss of antigenicity was observed when we immunized with equal amounts of protein separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), immunoblotted and stained with either Coomassie brilliant blue R 250 or Amido black 10 B. [ABSTRACT FROM AUTHOR]

The cloned murine cell line BK-BI-2.6.C6 has previously been shown to exhibit T cell charac- teristics, to synthesize and express MHC class II molecules, and to present protein antigens to antigen-dependent T cell clones. As a more definitive proof of the T-cell nature of these cells, transcripts of the rearranged F cell antigen receptor (TcR) β gene were assessed by Northern blot analysis. BK-BI-2.6.C6 cells constitutively transcribe mRNA for the light chain of TcR and express the disulphide-linked α,β TcR heterodimer at the cell surface. In addition mRNA for the polymorphic MHC class II subunits Aα and Aβ as well as for the invariant γ chain were detected. BK-BI-2.6.C6 T cells effectively stimulated bovine insulin-reactive T hybridoma cells to lymphokine production in the presence of this antigen. Since the antigen-presenting and the responding T cell populations arc maintained in culture in the absence of feeder cells, contamination by conventional accessory cells is excluded. These data unequivocally demonstrate that cloned routine Ia-expressing T cells can act as antigen-presenting accessory cells. [ABSTRACT FROM AUTHOR]

Skin biopsies were investigated with two different immunohistochemical techniques, thus revealing HLA-DQ antigens on HLA-DR-expressing keratinocytes in the late skin manifestations of a Borrelia spirochete infection. In the early skin lesions only HLA-DR antigens were present on the keratinocytes. The invariant γ chain of class II transplantation antigens was observed on keratinocytes in 1:5 of the late cases. Upon penicillin treatment detectable HLA-DR and HLA-DQ antigens disappeared completely from the keratinocytes. Furthermore, the mononuclear cell infiltrates dominated by anti-Leu 1 and anti-Leu 3a-reaetive cells and containing many cells with markers for activation (HLA-DR, HLA-DQ, transferring, and interleukin 2 receptors) diminished markedly. The possibility that the expression of different class II transplantation antigens on keratinocytes might reflect separate functional demands of these cells or an altered immunological reactivity in the host, is discussed. The precise functional role of the temporary expression of the class II antigens on non-lymphoid cells, however, remains an enigma. [ABSTRACT FROM AUTHOR]

Suppressor cells obtained from spleens of normal A mice, or factor extracted from these suppressor cells, abolished the syngeneic delayed type hypersensitivity (syn-DTH) response of X-irradiated A mice injected with trinitrophenyliited spleen cells and challenged with syngeneic lymphoblasts. Some of the physical, chemical and biological properties of the suppressive factor (SF) were characterized. The SF was relatively temperature-stable and its activity was destroyed by pronase (but not with RNase or DNase). The activity of the SF was absorbed on concanavalin A and anti-I-Jk Sepharose columns, suggesting that the factor is a glycoprotein-bearing I-Jk product. The approximate molecular weight of the factor is 50,000-60,000. The SF was absorbed on plastic adherent cells (but not on non-adherent cells). Adherent cells that absorbed the SF abrogated the ability of primed T cells to transfer the syn-DTH to naive X-irradiated recipients. In contrast, SF that was presented directly to the primed T cells failed to abolish their ability to transfer DTH. These findings suggest that the adherent cells serve as mediators, transferring the SF from factor-producing cells (Lyt-1+2+3+, I-Jk+- T cells) to target cells (Lyt-1+ primed T cells). [ABSTRACT FROM AUTHOR]

It must be for almost ten years now that I have been stubbornly holding the view that the strength of transplantation antigens is essentially a direct function of the number of immunologically competent (antigen sensitive) cells which can recognize the antigen as foreign and react against it. (Morten Simonsen, 1970 (131.) [ABSTRACT FROM AUTHOR]

A simple rapid spot immunoprecipitate assay (SIA) is presented. The antigen antibody reaction takes place in a field a field of alternating current; small amounts of specific antisera are used. and the influence of diffusion or electromobility of the antigen is avoided. Determinations of IgG, IgA and IgM in sera from healthy individuals correlated highly significantly with radial immuno-diffusion (RID) results. The reaction conditions in SIA, particularly the availability of a sufficiency of specific antibodies, may permit more accurate estimates of monoclonal forms of Ig than in RID, where the combination depends on diffusion of one reactant, The precision of SIA quantitation of IgO and IgA was within 10% and 13% for IgM. Working ranges of the standard curves were learn about 25-150 ng for G and A and 75-300 ng in a IgM; least squares regressiorn coefficients for four sets of standards on three SIA plates were 0.995 for IgG, 0.969 for IgA, and 0.986 for IgM, IgA at 7.5 ng in a 2.5 mg/I dilution was at the lower limit of demonstrability. Photometric registrations and estimates made by visual inspection compared highly significantly (P<0.001), differing by + 3%. [ABSTRACT FROM AUTHOR]

Dinitrophenyl (DNP)-lysine-polymethylmethacrylate and DNP-cellulose conjugates do not irreversibly inactivate anti-DNP antigen-sensitive cells, regardless of the dose (up to 10 mg) or persistence of the stimulation (up to 2 weeks). Since these conjugates constitute pure hapten presentations, it is concluded that the pure hapten presentation to B lymphocyte does not irreversibly inactivate them. When murine spleen cells are culture with Escherichia coli lipopolysaccharide (LPS) and (non-immunogenic) DNP-lysine-polymethylmethacrylate or (non-immunogenic) DNP-cellulose conjugates, an anti-DNP immune response occurs. However, replacement of DNP-lysine-polymethylmethacrylate with polymethylmethacrylate, or DNP-cellulose with cellulose, also results in a similar anti-DNP response. It is consequently concluded that the anti-DNP response elicited by DNP-Ficoll is, upon exhaustive testing, carrier-dependent. This implies that the mechanism of DNP-Ficoli immunogenicity is not two cooperative signals passed on to B lymphocytes via the hapten DNP. These results argue against any two-signal model of B-lymphocyte activation. [ABSTRACT FROM AUTHOR]

Primed LD typing cells were prepared against single HLA baplotypes within families and used to type a random panel of 48 individuals. PLT cells could be grouped on the basis of highly correlated responses with the panel test cells; the 21 different PLT cells could be used to define 6 different PL antigens. One of these, PL 3, was split by the responses of two of the 21 PLT cells which measured an antigen called PL 3.1. [ABSTRACT FROM AUTHOR]

Targeting of antigen to dendritic cells ( DCs) increase the efficiency of immunization procedures and may facilitate the development of more effective vaccines. Several surface molecules on DCs have shown to be useful for antigen targeting, but many more deserves investigation for their efficacy in this respect. With this end in mind, a simple in vitro assay for screening of optimal targets for antigen-delivery to murine DCs was established. Splenocytes from mice immunized with rat Ig G were targeted in vitro with a panel of different rat monoclonal antibodies (m Abs) directed against surface markers on murine DCs. The resulting T-cell activation was analysed by determining the number of IFN- γ and IL-4 secreting cells by ELISPOT. A positive effect of targeting was evident with several of the m Abs. Thus, mAbs against CD11c, CD36, CD205 and Clec7 A all induced IFN- γ responses that were significantly higher than those induced by non-targeting control m Abs. Anti- CD36 also induced IL-4 responses that were significantly higher than the control. The assay described here allows simultaneous analysis of a large number of potential target structures and facilitates direct comparison between the different targets regarding the strength of the T-cell responses induced by the targeted DCs. The assay could be useful as a first-line screening of potential target structures on murine DCs. [ABSTRACT FROM AUTHOR]

The advent of major histocompatibility complex ( MHC) tetramer technology has been a major contribution to T cell immunology, because tetramer reagents permit detection of antigen-specific T cells at the single-cell level in heterogeneous populations by flow cytometry. However, unlike MHC class I tetramers, the utility of MHC class II tetramers has been less frequently reported. MHC class II tetramers can be used successfully to enumerate the frequencies of antigen-specific CD4 T cells in cells activated in vitro, but their use for ex vivo analyses continues to be a problem, due in part to their activation dependency for binding with T cells. To circumvent this problem, we recently reported the creation of a new generation of reagents called MHC class II dextramers, which were found to be superior to their counterparts. In this review, we discuss the utility of class II dextramers vis-a-vis tetramers, with respect to their specificity and sensitivity, including potential applications and limitations. [ABSTRACT FROM AUTHOR]

The purpose of this study was to understand the autoimmune response and immunogenicity of a tumour-associated antigen IMP2/p62 in breast cancer. Autoantibody responses to IMP2/p62 were evaluated by enzyme-linked immunosorbent assay ( ELISA), Western blotting and indirect immunofluorescence assay in sera from patients with breast cancer, benign breast tumour and normal human individuals. Immunohistochemistry ( IHC) study with breast cancer tissues was also performed to analyse protein expression of IMP2/p62. The results have demonstrated that IMP2/p62 can induce a relatively higher frequency of autoantibody response in breast cancer (14.3%, 7/49) compared to patients with benign breast tumour (5.6%, 2/36) and normal individuals (2.2%, 1/44). The frequency of IMP2/p62 expression in breast cancer tissues was significantly higher than that in normal tissues ( P < 0.01). The data suggest that autoantibody against IMP2/p62 may be a useful serum biomarker for early-stage breast cancer screening and diagnosis. [ABSTRACT FROM AUTHOR]

It is well recognized that the physiological/pathological consequences of an immune response, against a foreign or a self-antigen, are often critically dependent on the class of immunity generated. Here we focus on how antigen interacts with the cells of the immune system to determine whether antigen predominantly generates Th1 or Th2 cells. We refer to this mechanism as the 'decision criterion' controlling the Th1/Th2 phenotype of the immune response. A plausible decision criterion should account for the variables of immunization known to affect the Th1/Th2 phenotype of the ensuing immune response. Documented variables include the nature of the antigen, in terms of its degree of foreignness, the dose of antigen and the time after immunization at which the Th1/Th2 phenotype of the immune response is assessed. These are quantitative variables made at the level of the system. In addition, the route of immunization is also critical. I describe a quantitative hypothesis as to the nature of the decision criterion, referred to as the Threshold Hypothesis. This hypothesis accounts for the quantitative variables of immunization known to affect the Th1/Th2 phenotype of the immune response generated. I suggest and illustrate how this is not true of competing, contemporary hypotheses. I outline studies testing predictions of the hypothesis and illustrate its potential utility in designing strategies to prevent or treat medical situations where a predominant Th1 response is required to contain an infection, such as those caused by HIV-1 and by Mycobacterium tuberculosis, or to contain cancers. [ABSTRACT FROM AUTHOR]

Human plasma lactose-binding immunoglobulin ( LIg) isolated by affinity chromatography on lactose- Sepharose was largely Ig G with significant Ig A and Ig M contents. LIg-mediated agglutination of desialylated human RBC was inhibited equally by the α- and β-anomers of methyl galactoside. Recognition of either the terminal α-galactose ( TAG)-containing glycans of bovine thyroglobulin or the N-acetyl lactosamine ( Lac NAc)-terminating glycans of asialofetuin by LIg was inhibitable nearly as much by the α-galactoside melibiose as by the β-galactoside lactose. Melibiose covalently conjugated to protein and coated on polystyrene wells captured several times more LIg molecules than its lactose analogue. LIg binding to bovine thyroglobulin or rabbit RBC membrane proteins, both bearing TAG was substantially reduced by prior treatment of the proteins with α-galactosidase to remove TAG though enzyme-treated glycans contained newly exposed Lac NAc moieties. Desialylated O-linked oligosaccharides, however, were no ligand for LIg. Unlike LDL, plasma lipoprotein(a) [ Lp(a)] coated on polystyrene well and desialylated by neuraminidase was recognized by LIg through terminal Lac NAc moieties exposed by the enzyme on its apo(a) subunit. Further, same amount of added fluorescence-labelled LIg formed significantly more immune complex with Lp(a) in high Lp(a) plasma than in low Lp(a) plasma. Results suggest (1) possibility of a role for LIg in combating non-primate molecules and cells bearing TAG moiety and (2) a mechanism for Lp(a)-mediated vascular injury as diabetes, infections and inflammations induce greater release of neuraminidase into circulation. [ABSTRACT FROM AUTHOR]

The repeated application of antigens results in the induction of tolerance. Lymph nodes are responsible for this reaction by producing suppressor cells. Using an in vivo transplantation model, we showed recently that stromal cells from different lymph nodes induce different cell populations for suppression, which all produce a tolerogenic phenotype. In this study, we were interested in the role of the spleen in these tolerance reactions. Therefore, tolerance was induced via feeding or injecting ovalbumin several times in control and splenectomized mice. The delayed-type hypersensitivity ( DTH) was measured as well as the cell subset composition of the spleen. The spleen of peripherally tolerized mice showed higher proliferation activity and a specific antibody production compared with orally tolerized mice, where regulatory T cells were predominantly found. Tolerance induction after removal of the spleen resulted in a reduced DTH response in antigen fed animals, whereas skin tolerance induction failed. In conclusion, the results illustrate that lymph nodes from different areas employ their individual pathways for similar immune reactions, and the spleen is part of this reaction initiated at the peripheral site. [ABSTRACT FROM AUTHOR]

It has not been considered so far that antigen-presenting cells (APC) may phagocytose immunogenic material from autologous cancer cells. Assuming the presence of cancer-immunogenic material in APC, we developed a novel autologous priming method that does not require tumour cells or identified peptides. Cancer-immunogenic information came from CD14+ monocytes. When stimulated with CD3-activated T cells, monocytes primed CD3+CD4+ and CD3+CD8+ resting/naïve T cells to become powerful effector cells within 24 h. During priming, depletion of CD14+ monocytes but not CD1a+ CD83+ dendritic cells prevented T cell priming. During cancer cell destruction, dendritic cells, but not monocytes, enhanced cancer cell lysis. The cascade-primed (CAPRI) immune cell quartet comprising monocytes, dendritic cells, CD4+ T and CD8+ T cells induced a significant decrease in the number of suppressive CD25highFoxP3+CD4+ T cells. CAPRI cells induced a marked upregulation of MHC class I and class II expression in cancer cells, which is crucial for autoimmune-like lysis. We show in vivo evidence of the CAPRI cell concept in nude mice. In humans, we present circumstantial clinical evidence showing the efficacy of CAPRI cells in an adjuvant treatment attempt for breast cancer patients with metastasis ( N = 42). Compared to patients at the Munich Tumor Center (no CAPRI treatment N = 428), almost double the expected number of patients survived 5 years ( P = 1.36 × 10−14). The CAPRI method is a safe procedure without negative side effects. High numbers of cancer-specific CAPRI cells can be obtained within a week against different cancer types for efficient adoptive cell therapy. [ABSTRACT FROM AUTHOR]

Diagnostic tests for tuberculosis (TB) using interferon gamma (IFN-γ) responses produced by T lymphocytes after stimulation by early secretory antigen target 6 (ESAT-6), culture filtrate protein 10 (CFP-10) or purified protein derivate (PPD) were carried out using ELISA (enzyme-linked immunosorbent assay) in whole blood culture supernatants from children with suspected TB disease ( n = 21), latent TB infection (LTBI; n = 17) and negative controls (NC; n = 21) from Recife, Pernambuco, Brazil. The results were analysed using the ROC (receiver operating characteristic) curves and the areas under the curve (AUC) generated varied from 0.5 to 1.0 with higher values indicating increased discriminatory ability. Comparisons of AUCs were made using non-parametric assumptions, and the differences were considered significant if P < 0.05. The ROC curve showed a statistical difference ( P = 0.015) between the LTBI and NC groups with an AUC of 0.731, TB disease and NC (AUC = 0.780; P = 0.002) and a group with TB (latent infection + disease, n = 38) and NC (AUC = 0.758; P = 0.001) when the antigen used was ESAT-6. No statistical difference was found between the groups when CFP-10 or PPD was used. In conclusion, the ESAT-6 test may be the most appropriate for diagnosis of childhood TB, both LTBI and TB disease, when associated with epidemiological and clinical data, especially in endemic areas such as Brazil. [ABSTRACT FROM AUTHOR]

It is well established that the passive trans-placental passage of anti-Ro/SSA antibodies from mother to foetus is associated with the risk to develop an uncommon syndrome named neonatal lupus (NLE), where the congenital heart block represents the most severe clinical feature. Recent evidence demonstrated that also adult heart, classically considered invulnerable to the anti-Ro/SSA antibodies, may represent a target of the arrhythmogenicity of these autoantibodies. In particular, the prolongation of the QTc interval appears the most frequent abnormality observed in adults with circulating anti-Ro/SSA antibodies, with some data suggesting an association with an increased risk of ventricular arrhythmias, also life threatening. Moreover, even though the association between anti-Ro/SSA antibodies and conduction disturbances is undoubtedly less evident in adults than in infants, from the accurate dissection of the literature data the possibility arises that sometimes also the adult cardiac conduction tissue may be affected by such antibodies. The exact arrhythmogenic mechanisms involved in foetus/newborns and adults, respectively, have not been completely clarified as yet. However, increasing evidence suggests that anti-Ro/SSA antibodies may trigger rhythm disturbances through an inhibiting cross-reaction with several cardiac ionic channels, particularly the calcium channels (L-type and T-type), but also the potassium channel hERG, whose different expression and involvement in the cardiac electrophysiology during lifespan might account for the occurrence of age-related differences. [ABSTRACT FROM AUTHOR]

MPB70 and MPB83 are among the most studied mycobacterial antigens. They are highly homologous proteins within Mycobacterium tuberculosis complex members, and the orthologs in different members of the complex are virtually identical. They are major antigens highly expressed by Mycobacterium bovis and considerably less abundantly expressed by M. tuberculosis. There are two genes encoding these proteins within an operon of six genes. MPB70 and MPB83 are encoded as precursor proteins with typical signal peptides for export through the general secretory pathway. MPB70 is a soluble secreted protein cleaved by signal peptidase I, while MPB83 is a glycosylated lipoprotein processed by signal peptidase II and located at the surface, possibly with the lipid tail coupled to the N-terminal cystein embedded in the mycobacterial outer membrane. The expression of these genes is controlled by the transcriptional regulator SigK, and a point mutation in SigK explains why some Bacille Calmette-Guérin (BCG) strains express only minute amounts of MPB70 and MPB83. BCG strains that have retained the high expression of MPB70 and MPB83 of wild type M. bovis are linked to a risk of BCG osteitis in <1-year-old children as a complication to BCG vaccination at birth. The proposed mechanism for this is based on homology of these proteins with osteoblast-specific factor II which is a homofilic adhesion protein expressed on osteoblasts in growing bone tissue. T-cell responses and antibody responses to these proteins have been extensively explored for sensitive and specific diagnosis of bovine tuberculosis. [ABSTRACT FROM AUTHOR]