New tregitopes inducing adaptive regulatory T cells in mice.

Epitopes of regulatory T cells (tregitopes) represent linear sequences of amino acids that induce CD4+CD25+Foxp3+ T lymphocytes expansion both in vitro and in vivo. The tregitopes' effectiveness was confirmed in autoimmune disease mouse models and in murine transplant models. Therefore, tregitopes together with regulatory T cells (Tregs) could play a major role in maintaining immune tolerance. The purpose of the presented study was a selection of potential tregitopes and assessment of their impact on Tregs expansion. Eight peptides were selected based on the previously published in silico model and their immunotolerogenic functions. To verify, if selected peptides are potential TCR ligands, the affinity of selected peptides to overrepresented in patients with autoimmune diseases, HLA-DRB1*04:01 allele, was measured by surface plasmon resonance. In order to evaluate the impact of potential tregitopes on the induction of Tregs in in vitro conditions, C57BL6Foxp3GFP mouse antigen presenting cells were co-cultured with naive syngeneic T cells under stimulation of selected peptides. CD4+CD25+Foxp3+ and CD4+CD25+Foxp3+IL-10+ cells frequency was analyzed using flow cytometry. Based on Tregs induction, two tregitopes derived from yeast and adenovirus protein were identified. In summary, the performed studies allowed an identification of novel putative tregitopes, which application potential includes their use as immunomodulators in mice.

Proteome of spleen CD4 lymphocytes in mouse preimplantation pregnancy.

Pregnancy exerts profound impact on female immune system. The first signs of pregnancy recognition by immune system are observed even before implantation. The most visible effects are present in the local compartment, i.e. in uterine draining lymph nodes and the decidua, while peripheral changes are less obvious. In our recent paper we indicated that costimulation phenotype of APCs in spleens of female mice during the preimplantation period of pregnancy differs from mice in pseudopregnancy. However, the effect of differential costimulation in the context of the T lymphocyte function at periphery in early pregnancy is still unknown. For that reason, we decided to investigate global protein expression in splenic CD4(+) lymphocytes in order to identify and validate the most important biomarkers characteristic for the preimplantation period of pregnancy at periphery. Two-dimensional electrophoresis (2-DE) and mass spectrometry (MS) were utilized to analyze the protein expression pattern of magnetically sorted CD4(+) lymphocytes from spleens of pregnant and pseudopregnant females at 3.5 days after mating. The first goal of this study was to create a 2-DE map of the splenic CD4(+) T cells of pregnant mice. As a result, 106 protein spots from 373 were identified using MS. The comparison of lymphocyte protein patterns between pregnant and pseudopregnant mice depicted differential expression of 11 identified proteins belonging to the group of proteins involved in cytoskeletal structure, cell motility and metabolism. Profoundly diminished expression of cofilin-1, F-actin capping protein subunit alpha and malate dehydrogenase proteins in lymphocytes of pregnant mice indicates that preimplantation pregnancy could change the activation state of peripheral CD4(+) lymphocytes.

SPIn: model selection for phylogenetic mixtures via linear invariants.

In phylogenetic inference, an evolutionary model describes the substitution processes along each edge of a phylogenetic tree. Misspecification of the model has important implications for the analysis of phylogenetic data. Conventionally, however, the selection of a suitable evolutionary model is based on heuristics or relies on the choice of an approximate input tree. We introduce a method for model Selection in Phylogenetics based on linear INvariants (SPIn), which uses recent insights on linear invariants to characterize a model of nucleotide evolution for phylogenetic mixtures on any number of components. Linear invariants are constraints among the joint probabilities of the bases in the operational taxonomic units that hold irrespective of the tree topologies appearing in the mixtures. SPIn therefore requires no input tree and is designed to deal with nonhomogeneous phylogenetic data consisting of multiple sequence alignments showing different patterns of evolution, for example, concatenated genes, exons, and/or introns. Here, we report on the results of the proposed method evaluated on multiple sequence alignments simulated under a variety of single-tree and mixture settings for both continuous- and discrete-time models. In the simulations, SPIn successfully recovers the underlying evolutionary model and is shown to perform better than existing approaches.

Susceptibility testing and resistance phenotype detection in Staphylococcus aureus strains isolated from patients with atopic dermatitis, with apparent and recurrent skin colonization.

BACKGROUND: Staphylococcus aureus colonization is accepted to be an important triggering factor in patients with atopic dermatitis (AD) and antibiotic resistance has been recognized to be a serious problem as a consequence and for the management of AD treatment. OBJECTIVES: To investigate the antibiotic resistance pattern of S. aureus strains isolated from patients with AD with apparent (lesional and nonlesional skin areas) and recurrent skin colonization and strains obtained from healthy nasal carriers. METHODS: Eighty-seven patients (age 23+/-11.5 years) with mild to severe AD (SCORAD 46.9+/-16.6), 21 patients (age 19.8+/-6.7 years) before antistaphylococcal treatment and 177 healthy nasal carriers (age 27.5+/-8.4 years) were microbiologically assessed for carriage of S. aureus. Colonization of lesional and nonlesional skin areas was quantified by counting the number of colony forming units on the skin surface (log(10) CFU cm(-2)). Antimicrobial susceptibility and resistance phenotypes of 179 S. aureus strains were assessed with the agar disc-diffusion method. RESULTS: Staphylococcus aureus was isolated from 87% of lesional and 44% of nonlesional skin samples from patients with AD. The colonization density of S. aureus was markedly higher in lesional than in nonlesional skin (P<0.001), and was positively correlated with AD severity (P<0.001) and total serum IgE (P<0.05). Patients with AD had a significantly higher prevalence of chloramphenicol-resistant S. aureus than nasal carriers (P<0.01). Similar rates of resistance were expressed to tetracycline, erythromycin, mupirocin, clindamycin and penicillin. Nearly 35% of S. aureus strains from the lesional skin demonstrated different antimicrobial sensitivity pattern compared with strains from nonlesional skin of the same patients with AD. The trend of increasing resistance to chloramphenicol, erythromycin and fusidic acid was observed among S. aureus strains recovered from patients after approximately 75 days of antibiotic treatment. Methicillin-resistant S. aureus isolates were cultured from two patients, one during exacerbation and the other after subsequent bacterial recolonization. CONCLUSIONS: Discrepancies in antibiotic sensitivity pattern were observed among S. aureus strains colonizing different sites of AD skin (lesional and nonlesional areas), and also in AD patients with prior antibiotic treatment. Therefore, clinicians should consider repeat microbial susceptibility testing on different body sites of patients with AD when clinically indicated.

Current status of fungal cell wall components in the immunodiagnostics of invasive fungal infections in humans: galactomannan, mannan and (1-->3)-beta-D-glucan antigens.

Early diagnosis of fungal infections and the implementation of appropriate treatment represent major issues for clinicians, nowadays. Histopathological demonstration of microorganisms in tissue specimens or growth of fungal agents in culture media is still considered the "gold standard", but obtaining such specimens may be difficult. Several groups have investigated serological assays for cell wall elements unique to fungal organisms in serum or other body fluids to improve diagnostics in patients with haematological malignancies or undergoing haematopoietic stem-cell transplantation. In this review we have concentrated on the currently available assays allowing for detection of highly immunogenic components of fungal cell wall: galactomannan, mannan, and also (1-->3)-beta-D-glucan. Rapid serological tests appear to be useful for screening high-risk haematological patients, since they allow for the early diagnosis of invasive fungal infections, including infections with the most common pathogens such as Aspergillus and Candida. Based on current literature, factors increasing the probability of obtaining false-positive or false-negative results detected by each test were also analysed and tabulated.

[Histocompatibility HLA system of man. Consideration in the light of current concepts. I. Organization and polymorphism]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. I. Organizacja i polimorfizm.

The HLA system, which represents the major histocompatibility complex (MHC) of man, encompasses approximately one thousandth of the human genome and is localised on the short arm of chromosome 6 (band 6p 21.3). The HLA gene complex shows an extreme polymorphism which can be demonstrated by molecular genetic methods. The genes so far recognised in the HLA can be subdivided into three major classes: 1) the HLA class I (HLA-ABC) and class II genes (HLA-D); 2) immune function related genes (C2, C4A, C4B, TNFA and TNFB, transporter and proteasome genes); 3) other genes apparently not related to immune functions (CYP 21, valyl-tRNA synthetase). The loci HLA-A, -B, and -C represent the classical HLA class I loci with gene products expressed on nearly all nucleated cells; HLA-E, -F and -G the non-classical HLA class I loci, code for products with a limited tissue distribution and a restricted polymorphism. Classical class I and II MHC antigens are integral membrane proteins composed of two pairs of structurally similar extracellular domains. The X-ray studies indicated the presence of peptides bound to HLA molecules within the groove. The groove has on its floor several amino acids where peptides, derived from the antigen being presented, are bound. Although the principle of allele specific motives ruling the peptide binding seems to be become more established, the further biological impact of the allelic variation remains subject for future studies.

[Histocompatibility HLA system of man. Considerations in the light of current concepts. VII. Nonclassical HLA- E, F, and H loci]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. VII. Nieklasyczne uklady HLA-E, F i H.

Current opinions connected with HLA-E and HLA-F genes determining "nonclassical" (HLA-Ib) class I antigens of the Main Histocompatibility Complex MHC, and formed in the consequence of mutation or partial deletion of HLA-H pseudogene loci were presented. The expression of protein products of HLA-E and -F genes on some cells and tissues, their polymorphism, and also their biological functions in organisms were qualified by the use of molecular technics. The kind and frequency of occurrence of mutations 845 A (C282Y) and 187 G (H63D) in gene HLA-H were analysed, and in this context some genetic aspects of hereditary hemochromatozy (HH) were discussed.

[Histocompatibility HLA system of man. Considerations in the light of current concepts. VI. HLA-G antigen polymorphism]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. VI. Antygenowy polimorfizm ukladu HLA-G.

The results of hitherto researches over gene products of HLA-G locus, i.e. membrane bound forms (M*HLA-G) and soluble forms (S*HLA-G) were verified. Current opinions connected with synthesis and molecular organisation of five HLA-G isoformic molecules (HLA-G1, -G2, -G3, -G4, -G5) were presented. The polymorphism of HLA-G gene and its expression were qualified. Special attention has been turned on presumably significant function of HLA-G in maintenance of maternal-placenta immunological tolerance, binding of monomeric peptides, and antigens presentation, taking into consideration cytomegaloviruses (CMV). In spite of significant progress in the understanding of mutual relationship between mother and placenta many phenomena with participation of HLA-G gene products, especially in pathological conditions, are still unknown.

[Human histocompatibility HLA system. Considerations in the light of current concepts. VIII. Polymorphism of HLA-related MIC system]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. VIII. Polimorfizm ukladu MIC HLA-pokrewnego.

The recent advances about a new distinct family of polymorphic genes MIC (PERB11) "mapped" in the region of the major histocompatibility complex of antigens MHC were presented. Some aspects connected with their molecular organisation, degree of polymorphism, expression, and immunogenetical function were discussed. Special attention sacrificed to results of investigations over existence of particular MICA allele association with some diseases. Many relating problems, especially functional MICA and MICB genes are still in interests both clinicians, and immunogenetists.

[Histocompatibility HLA system in man. Considerations in the light of current concept. Part III. The relationship between HLA antigens and disease susceptibility]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. III. Wspólzaleznosc miedzy antygenami HLA a wystepowaniem okreslonych chorób.

A large number of diverse diseases are associated with genetic markers of the MHC complex. The analysis of such disease associations is complicated by the diversity of genetic information located within the HLA complex. Recent advances in the structural analysis of HLA genes and gene products contribute a new perspective to questions of HLA-associated disease. HLA-associated disease are caused by an interplay of many different genes and environmental factors, where HLA complex genes most often confer the strongest genetic predisposition. With this knowledge at hand, identification of individuals at high risk to develop autoimmune and other potentially disabling diseases should be used in order to effectively prevent or halt disease development. This may be a major part of the clinical HLA field in the near future. In this article, we present and interpret some such recent information, especially a possible molecular mechanism of the HLA associations.

[Human histocompatibility HLA system. Considerations in the light of current concepts. II. Immunogenetic function]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. II. Funkcja immunogenetyczna.

Investigation of self-nonself recognition in the immune system has focused on classical class I and class II MHC antigens. Their extensive polymorphism and high level expression rendered them amenable to experimentation with the traditional immunologic tools, serology and transplantation. Much has been learned about their functions as a peptide--presenting receptors important for immune responses against pathogens. The central unit for regulation of the specific immune system is a trimolecular complex made up of the T cell antigen receptor (TCR), the MHC molecule and the MHC ligand. The third component is a peptide derived as a degradation product from a protein. During recent years there has been some progress in understanding the interaction between MHC molecules and their peptide ligands: MHC molecules are peptide receptors of peculiar specificity, being able to accommodate millions of different peptides provided they share some common features. In this review is to discuss the relationship between MHC molecules and their natural peptide ligands with special emphasis on MHC class I molecules, because more is known about these than class II molecules.

[Immuno-genetic significance in sensorineural hearing loss (SNHL) in the light of our own investigations]. / Immunogenetyczne uwarunkowania w gluchocie odbiorczej (SNHL) w swietle wyników badan wlasnych.

The immunogenetic analysis in 152 patients with sensorineural hearing loss (SNHL) was studied. HLA class I antigens and haplotypes were calculated on the basis of typical linkage between HLA-A, -B, and C loci. Soluble form of HLA antigens (sHLA-I) in blood serum in a semiquantitative microabsorption test according to McLean et al. were also determined. It was found that the frequency of HLA-A2, -A25(10), -B39(16), -Cw5, and -Cw7 were different in patients with SNHL in compared to the control group (n = 1554). This deference's were highly significant, especially for HLA-Cw7 antigens (chi 2 = 88.88; p < 0.0000...1). The frequency of haplotype HLA-A25-B18-Cw7 in patients was also different (f = 362 x 10(4) vs 51 x 10(4)), and highly significant compared to the healthy individuals (n = 1944) (chi 2 = 27.37; p < 0.0000...1). Among 23 typed HLA class I antigens, significantly higher mean level was observed for 14 sHLA antigens then in control group (n = 248). The levels of sHLA-I antigens in patients sera may suggests participation of sHLA molecules in etiopathogenesis of SNHL. This hypothesis may indicate the existence of autoimmunogenetic background of SNHL within high concentrations of certain sHLA-I antigens.

[Histocompatibility HLA system of man. Considerations in the light of current concepts. IV. Soluble HLA class I antigens (sHLA-I)]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. IV. Rozpuszczalne antygeny ukladu HLA klasy I (sHLA-I).

Soluble class I human leukocyte antigens (sHLA) have been detected in serum, sweat, lymphatic fluid, urine and cerebrospinal fluid. The levels vary among different individuals and are significantly affected by inflammatory diseases and organ rejection. This article discusses the clinical significance of levels of serum HLA class I antigens, both in patients with viral diseases and following organ transplantation, as well as the potential involvement of such antigens in the immune response. The potential use of sHLA in clinical medicine is far-reaching. sHLA-peptide complexes may find wide application, particularly for the treatment recipients. Although further in vivo studies are required, available data show the efficacy of sHLA to regulate T-cell function.

[Histocompatibility HLA system of man. Considerations in the light of current concepts. V. sHLA-I in patient's body fluids]. / Uklad zgodnosci tkankowej czlowieka--HLA. Rozwazania w swietle aktualnych pogladów. V. sHLA-I w plynach ustrojowych chorych.

The retrospective estimation of concentrations of soluble HLA class I antigens (sHLA-I) in blood serum and in other body fluids of patients was executed. The physiological concentration of sHLA-I is significantly upregulated in various diseases and during inflammation, and following organ transplantation. This suggested that sHLA-I might serve as a marker of pathological changes. Observed differences in sHLA-I levels among different diseases could reflect variability in genetic factors, pathophysiology or disease activity. The authors show on usefulness of obtained results to comparative analysis of particular HLA class I allospecificities concentrations characteristic for some diseases, for example HLA-Cw7 in patients with SNHL, or HLA-B27 in zzsk. Estimation of sHLA-I concentrations in blood sera of patients could represents a good diagnostic and prognostic marker in monitoring of course disease and its activity.

Soluble HLA class I antigens in serum of healthy individuals--population study.

Variability of concentrations of s-HLA-I depending on allelic specificity substantiated realisation of research in population of 248 healthy, unrelated individuals. Defined phenotypes from tissue typing of polymorphic HLA complex enabled concentrations measurements of 1553 serum samples for HLA--A, B, and C loci determined antigens. Semi-quantitative technique of inhibition microcytotoxic reaction according to Tait et al. (1981) and Mclean et al. (1983) with usage of policlonal sera anti-HLA was applied. For most of numbers of the allelic specificity the concentration of antigen material in soluble form (s-HLA-I) in blood serum were nominally very high and high. For certain numbers of specificity e.g. HLA--A26, A29, B39, B52, B56, Cw5, Cw6 the percentage of sera, where the s-HLA concentrations were decreased was observed. The results were presented as mean values of inhibition microcytotoxic reaction--according to NIH classification. Authors point on usefulness of results for s-HLA comparative analysis of particular HLA allelic specificity, specific for certain diseases e.g. Cw7 for SNHL patients and B27 for ankylosing spongilitis.

HLA class I haplotypes in families of children with coeliac disease.

The purpose of the study was the analysis of HLA-AB haplotypes frequency in the families of children with coeliac disease. Haplotypes present in 46 probands' families including 69 affected children, 49 healthy siblings and 91 parents were verified. HLA antigens were typed by Terasaki and McClelland's routine two-step-microcytotoxic assay in NIH modification. Among 138 haplotypes, the following were significantly more frequent in affected children: HLA-A1-B8 (3116 x 10(4)), HLA-A3-B8 (290 x 10(4)) and HLA-A2-B8 (217 x 10(4)). Total frequency of haplotypes including HLA-B8 antigen in comparison to control population equalled 3986 x 10(4) vs. 763 x 10(4). HLA-A1-B8 haplotype frequency was twice lower in probands' healthy siblings and parents, equalling 1837 x 10(4) and 1868 x 10(4), respectively. Highly significantly more frequent HLA-A1-B8 haplotype found in probands' families may indicate the correlation between inherited gene products and increased risk of coeliac disease incidence.

HLA class I antigens in families with coeliac disease.

The aim of the study was the analysis of the frequency of HLA class I antigens in the Polish population of children suffering from coeliac disease and their families, as well as the estimation of the relative risk of incidence, etiologic fraction (EF) and preventive fraction (PF) indexes. Forty-six probands' families were included in the typing: 69 children with coeliac disease confirmed clinically and histologically, 49 healthy siblings and 91 parents. The HLA antigens were typed with routine Terasaki an McClelland's two-stage microcytotoxic assay in NIH modification. The following antigens occurred significantly more frequently (p < 0.0000...1) in phenotypes of children with coeliac disease: HLA-A1 (chi 2 = 35.90; RR = 4.3; EF = 0.44), -B8 (chi 2 = 88.20; RR = 8.8; EF = 0.58) and Cw7 (chi 2 = 55.24; RR = 7.5; EF = 0.69). The positive correlation for the specificity of HLA-A1, -B8 was proved also in siblings (chi 2 = 16.03; chi 2 = 18.10) and parents (chi 2 = 15.67; chi 2 = 32.67). The presence of antigens HLA-A1, -B8 in the phenotype may be the risk factor predisposing for the manifestation of hypersensitivity to gluten.

[Selected parameters of immune reactivity in sensorineural hearing loss patients. II. CD3+, CD4+ and CD8+ subsets of T lymphocytes]. / Wybrane parametry reaktywnosci immunologicznej chorych z czuciowo-nerwowym uposledzeniem sluchu. II. Subpopulacje CD3+, CD4+ i CD8+ limfocytów T.

Subpopulations CD3+, CD4+ and CD8+ of T lymphocytes in 67 healthy individuals and in 36 sensorineural hearing loss (SNHL) patients with unknown etiology were determined. The patients were studied in two groups, i.e. 29 patients with bilateral and progressive hearing loss and 7 patients with sudden deafness. The ratio of CD4+ and CD8+ was also calculated. The results were presented as the mean value for percentage and absolute numbers of the cells together with statistical analysis. The acquired results of two group patients with SNHL were completed before and at the beginning of ubiquitine therapy. The highly significant differences (p < 0.001) for CD8+ subset in comparison with the results of control group was found. The restitution effect of ubiquitine treatment for CD3+ and CD4+ T lymphocytes subpopulation in SNHL patients was observed. Additionally in patients with sudden hearing loss the CD8+ subset was also normalized. The ratio of CD4+:CD8+ in patients with bilateral and progressive hearing loss was lower (1.27 +/- 0.21) that in control group (1.66 +/- 0.25) and significantly different (t = 7.31; p < 0.001). It was shown that the distribution of examined CD system markers which depended on CD4+:CD8+ ratio was normal only in 27.6% of patients with SNHL. It was a group of 9 patients in whom hearing loss appeared after bacterial or virus infections. No changes in the CD4+:CD8+ subpopulation ratio at the beginning of ubiquitine therapy was observed. The distribution of normal results for T cell subpopulations in patients before and during treatment was presented in table III. Our results indicate that the observations may have an important diagnostic value. The authors suggest that the CD system antigens determination in monitoring the clinical status of SNHL patients and also in patients treated with drugs influencing the immune system is needed.

[Selected parameters of immunologic reactivity in patients with sensorineural hearing loss. I. Receptor activity of T lymphocytes]. / Wybrane zagadnienia reaktywnosci immunologicznej chorych z czuciowo-nerwowym uposledzeniem sluchu. I. Receptorowa aktywnosc limfocytów T.

Receptors activity of peripheral T cells for sheep erythrocytes was analyzed in 67 healthy individuals and in the 29 patients with progressive, bilateral hearing loss and in 7 patients with sudden deafness on unknown etiology. The ability of T lymphocytes to form non-immune complete and incomplete rosettes in TEt and TEa tests was evaluated before and at the beginning of ubiquitine therapy. The mean values of rosette forming cells (RFC) in percentage and absolute numbers and the statistical analysis were presented. It was found that the receptor activity of T lymphocytes in TEt test was not significant different from the results of healthy control group. On the other hand highly significant differences were observed (p < 0.001) in subset of "active" T-cells determined in TEa rosette test. The relative coefficient of receptor activity of lymphocytes (CRAL) calculated for complete TEa rosettes was high different (p < 0.0008). Normalization of the results of TE rosette tests after first period of ubiquitin biotherapy was observed.

[Soluble class HLA antigens in serum of patients with brain tumors]. / Rozpuszczalne antygeny HLA klasy I w surowicach chorych na guzy mózgu.

The levels of soluble class I HLA antigens (s-HLA-I) in blood sera of patients with brain gliomas were studied before and after operation. It was found that sHLA material in sera was markedly decreased. The authors suggest the use of these examinations as one of the parameters of immune state of the oncologic patients before operation and in postoperative period.