Congenital Heart Defects in Patients with Classic Bladder Exstrophy: A Hitherto Neglected Association?

INTRODUCTION: Classic bladder exstrophy (BE) is regarded as an isolated malformation without any further anomalies, but some studies have indicated a higher incidence of cardiac anomalies. This cross-sectional study is planned to evaluate the prevalence of congenital heart defects (CHDs) and the clinical relevance for patients with BE admitted for primary closure. MATERIALS AND METHODS: Patients were prospectively recruited between March 2012 and January 2019. Patients' profiles including demographic data, results of transthoracic echocardiography (TTE), as well as essential peri- and postoperative data were assessed. RESULTS: Thirty-nine (25 boys and 14 girls) patients with BE (median age 61 days) underwent delayed primary bladder closure. Thirty-seven (24 boys and 13 girls) patients had received TTE 1 day before surgery. CHD was detected in 7 (18.9%) out of the 39 patients, but no clinical differences between patients with and without CHD were observed peri- or postoperatively. DISCUSSION AND CONCLUSION: This prospective systematic evaluation shows an even higher rate of CHD in patients with BE than assumed previously. Although peri- and postoperative outcome did not differ between patients with and without CHD, we consider TTE an important additional method for ensuring a safe peri- and postoperative courses and a short- and long-term care for patients with CHD.

No association of IL-12p40 pro1.1 polymorphism with juvenile idiopathic arthritis.

BACKGROUND: IL-12p40 plays an important role in the activation of the T-cell lines like Th17 and Th1-cells. Theses cells are crucial in the pathogenesis of juvenile idiopathic arthritis. A polymorphism in its promoter region and the genotype IL12p40 pro1.1 leads to a higher production of IL-12p40. We studied whether there is a difference in the distribution of the genotype in patients with JIA and the healthy population. METHODS: In 883 patients and 321 healthy controls the IL-12p40 promoter genotype was identified by ARMS-PCR. RESULTS: There is no association of IL-12p40 pro polymorphism neither in patients with JIA compared to controls nor in subtypes of JIA compared to oligoarthritis. We found a non-significant tendency of a higher prevalence of the genotype pro1.1 in systemic arthritis (32.4%) and in rheumatoid factor negative polyarthritis (30.5%) and a lower pro1.1 genotype in persistent oligoarthritis (20.7%) and in enthesitis-related arthritis (17%). Likelihood of the occurrence of genotype IL12-p40 pro1.1 in patients with systemic arthritis (OR 1.722, CI 95% 1.344-2.615, p 0.0129) and RF-negative polyarthritis (OR 1.576, CI 95% 1.046-2.376, p 0.0367) compared to persistent oligoarthritis was significantly higher. This was also true for comparison of their homozygous genotypes IL-12p40 pro 1.1 and 2.2 in systemic arthritis (OR 1.779, CI 95 % 1.045-3.029, p 0.0338). However, in Bonferroni correction for multiple hypothesis this was not significant. CONCLUSION: A tendency of a higher prevalence of the genotype IL-12p40 pro1.1 in systemic arthritis and in rheumatoid factor negative polyarthritis was observed but not significant. Further investigations should be done to clarify the role IL-12p40 in the different subtypes of JIA.

Impaired suppression of synovial fluid CD4+CD25- T cells from patients with juvenile idiopathic arthritis by CD4+CD25+ Treg cells.

OBJECTIVE: Natural CD4+CD25+FoxP3+ Treg cells play a crucial role in maintaining immune homeostasis and controlling autoimmunity. In patients with juvenile idiopathic arthritis (JIA), inflammation occurs despite the increased total numbers of Treg cells in the synovial fluid (SF) compared to the peripheral blood (PB). This study was undertaken to investigate the phenotype of CD4+ T cells in PB and SF from JIA patients, the function of synovial Treg cells, and the sensitivity of PB and SF CD4+CD25- effector T cells to the immunoregulatory properties of Treg cells, and to study the suppression of cytokine secretion from SF effector T cells by Treg cells. METHODS: The phenotypes of effector T cells and Treg cells of PB and SF from JIA patients and healthy donors were determined by flow cytometry. The functionality of isolated Treg cells and effector T cells was quantified in (3) H-thymidine proliferation assays. Cytokine levels were analyzed using Bio-Plex Pro assay. RESULTS: Compared to PB, SF showed significantly elevated numbers of activated and differentiated CD4+CD45RO+ T cells. Sensitivity of SF effector T cells to the suppressive effects of Treg cells from both PB and SF was impaired, correlating inversely with the expression of CD69 and HLA-DR. However, SF effector T cell cytokine secretion was partly suppressed by SF Treg cells. CONCLUSION: Our findings indicate that regulation is impaired in the SF of patients with JIA, as shown by the resistance of effector T cells to immunoregulation by functional Treg cells. This resistance of the SF effector T cells might be due to their activated phenotype.

Bench-to-bedside review: adenosine receptors--promising targets in acute lung injury?

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are life-threatening disorders that have substantial adverse effects on outcomes in critically ill patients. ALI/ARDS develops in response to pulmonary or extrapulmonary injury and is characterized by increased leakage from the pulmonary microvasculature and excessive infiltration of polymorphonuclear cells into the lung. Currently, no therapeutic strategies are available to control these fundamental pathophysiological processes in human ALI/ARDS. In a variety of animal models and experimental settings, the purine nucleoside adenosine has been demonstrated to regulate both endothelial barrier integrity and polymorphonuclear cell trafficking in the lung. Adenosine exerts its effects through four G-protein-coupled receptors (A1, A2A, A2B, and A3) that are expressed on leukocytes and nonhematopoietic cells, including endothelial and epithelial cells. Each type of adenosine receptor (AR) is characterized by a unique pharmacological and physiological profile. The development of selective AR agonists and antagonists, as well as the generation of gene-deficient mice, has contributed to a growing understanding of the cellular and molecular processes that are critically involved in the development of ALI/ARDS. Adenosine-dependent pathways are involved in both protective and proinflammatory effects, highlighting the need for a detailed characterization of the distinct pathways. This review summarizes current experimental observations on the role of adenosine signaling in the development of acute lung injury and illustrates that adenosine and ARs are promising targets that may be exploited in the development of innovative therapeutic strategies.

70-kDa heat shock proteins: specific interactions with HLA-DR molecules and their peptide fragments.

Heat shock protein 70 (HSP70):peptide complexes are involved in MHC class I and class II-restricted antigen presentation enabling enhanced activation of antigen-specific T cells. Here, we investigated the potential of bacterial and mammalian HSP70 molecules to interact with peptide fragments from HLA-DR and the corresponding complete HLA-DR molecules. Peptide fragments were found to interact with DnaK, the HSP70 homologue from E. coli, but less with stress-inducible human Hsp70. Only a peptide sequence exclusively found in rheumatoid arthritis-protective HLA-DR molecules did not interact with DnaK. Subsequently, we investigated the interaction of complete HLA-DR molecules with HSP70 and detected a specific HSP70:HLA-DR interaction, with highest affinity for human stress-inducible Hsp70. In contrast to the peptide fragments, no allele-specific differences in Hsp70 affinity were detected with complete HLA-DR molecules. Interaction with HLA-DR molecules was increased at lowered pH values, whereas HSP70-chaperoned peptides were released at acidic pH, thus HSP70 could serve as scanner and carrier for antigenic peptides of self or foreign origin and transfer chaperoned peptides onto MHC class II molecules in acidic late endosomal compartments. Our findings indicate that direct interaction between mammalian HSP70 and HLA-DR molecules could be involved in the HSP70-mediated enhancement of MHC class II-restricted peptide presentation and CD4(+) T cell activation.

Large-scale mapping of mutations affecting zebrafish development.

BACKGROUND: Large-scale mutagenesis screens in the zebrafish employing the mutagen ENU have isolated several hundred mutant loci that represent putative developmental control genes. In order to realize the potential of such screens, systematic genetic mapping of the mutations is necessary. Here we report on a large-scale effort to map the mutations generated in mutagenesis screening at the Max Planck Institute for Developmental Biology by genome scanning with microsatellite markers. RESULTS: We have selected a set of microsatellite markers and developed methods and scoring criteria suitable for efficient, high-throughput genome scanning. We have used these methods to successfully obtain a rough map position for 319 mutant loci from the Tübingen I mutagenesis screen and subsequent screening of the mutant collection. For 277 of these the corresponding gene is not yet identified. Mapping was successful for 80 % of the tested loci. By comparing 21 mutation and gene positions of cloned mutations we have validated the correctness of our linkage group assignments and estimated the standard error of our map positions to be approximately 6 cM. CONCLUSION: By obtaining rough map positions for over 300 zebrafish loci with developmental phenotypes, we have generated a dataset that will be useful not only for cloning of the affected genes, but also to suggest allelism of mutations with similar phenotypes that will be identified in future screens. Furthermore this work validates the usefulness of our methodology for rapid, systematic and inexpensive microsatellite mapping of zebrafish mutations.

The heat shock protein Hsp70 enhances antigen-specific proliferation of human CD4+ memory T cells.

Heat shock proteins (HSP) can interact with a wide variety of peptides and the resulting HSP:peptide complexes are known to be highly immunogenic. The ability of HSP:peptide complexes to elicit CD8+ T cell responses by cross-presentation of exogenous antigen via MHC class I is well known. In contrast, their role in the activation of CD4+ T cells is less clearly defined, although several recent studies in mice and T cell lines suggest an involvement of HSP in the presentation of antigenic peptides via MHC class II. In this study we have investigated the potential of antigenic peptides from tetanus toxin and influenza hemagglutinin complexed to the human stress-inducible Hsp70 to enhance activation and proliferation of human memory CD4+ T cells. Hsp70:peptide complexes were found to amplify the proliferation of antigen-specific CD4+ T cells as confirmed by HLA-DR tetramer staining. Complex formation of the antigenic peptide with Hsp70 was absolutely required to elicit an antigen-specific amplification. This effect was most pronounced at low doses of antigen and decreasing APC/CD4+ T cell ratios. Taken together, we show the potential of Hsp70 to enhance antigen-specific CD4+ T cell proliferation and to increase the immunogenicity of presented peptides in human CD4+ T cells.

Autoantibodies in juvenile idiopathic arthritis: glucose-6-phosphate isomerase is not a specific target.

OBJECTIVE: Antibodies recognizing the ubiquitous cytosolic enzyme glucose-6-phosphate isomerase (GPI) cause arthritis in the K/BxN mouse model. Studies have shown that these antibodies are not specific for rheumatoid arthritis (RA) in humans. We evaluated GPI as a target of autoantibodies in juvenile idiopathic arthritis (JIA). METHODS: We studied 324 serum and 48 synovial fluid (SF) samples from 103 patients with JIA, 36 with RA, and 8 with arthralgia and 11 controls. Anti-GPI antibodies were assessed by densitometrically evaluating immunoblots and ELISA using native and recombinant GPI. We determined the GPI activity of the soluble antigen in serum and SF. RESULTS: Although several samples contained anti-GPI-IgG antibodies, this was not specific for JIA or its subgroups, or for RA. Other proteins in the GPI preparation were also frequently recognized by antibodies. Additionally, we observed increased GPI activity in patients with the systemic manifestation of JIA, but not in other patients. Neither anti-GPI concentrations nor GPI activity were associated with disease activity. CONCLUSION: In addition to the findings in RA, our results indicate that GPI is not a general target of autoantibodies in JIA.

Determination of the peptide binding motif and high-affinity ligands for HLA-DQ4 using synthetic peptide libraries.

Juvenile idiopathic arthritis (JIA) is considered to be an autoimmune disease. Various human leukocyte antigen (HLA) associations for different subgroups of this heterogeneous disease have been found. For early-onset pauciarticular arthritis (now oligoarthritic JIA), a strong association with the HLA class II haplotype DQA1*0401/DQB1*0402 (DQ4) has been described. We determined the peptide-binding specificities of this HLA-DQ molecule by screening a synthetic acetylated nonapeptide amide library with one defined and eight random sequence positions. A characteristic binding motif could be deduced. By use of these data, we designed defined specific nonapeptides and identified high-affinity ligands binding to HLA-DQ4. The peptide binding motif of HLA-DQ4 is very similar to the motif of HLA-DQ7, also associated with oligoarthritic JIA. It is, however, different from binding motifs of neutral or protective HLA-DQ molecules. Our results further support the idea of differential peptide presentation in the pathogenesis of oligoarthritic JIA.

Retinal defects in the zebrafish bleached mutant.

The recessive zebrafish mutant bleached has, apart from its defects in pigmentation, a heritable defect leading to larval blindness. Here, we analyze the retina of homozygous bleached larvae, employing morphological and electrophysiological methods. Electroretinography revealed a complete lack of electrical signals in response to light. Histological analysis of mutant retinae showed a severely affected outer retina with a hypopigmented pigment epithelium and a disorganized outer nuclear layer containing few or no intact photoreceptors. Using the TUNEL assay for cell death detection, we noticed a strong increase of apoptotic cells in all retinal cell layers, starting in young larvae even before retinal support of visual function. At later stages cell death is most pronounced at the marginal zone, where new cells are constantly added to the retina. At early stages increased apoptosis is mainly confined to the retina, while at later stages elevated cell death is al so apparent in extra-retinal tissues, particularly in the brain. Hence, the lack of visual responses in homozygous bleached larvae can be attributed to a severe defect of the outer retina, preceded by increased levels of apoptotic cell death in all retinal cell layers.