The Specificity of Peptides Bound to Human Histocompatibility Leukocyte Antigen (HLA)-B27 Influences the Prevalence of Arthritis in HLA-B27 Transgenic Rats

Human histocompatibility leukocyte antigen B27 is highly associated with the rheumatic diseases termed spondyloarthropathies, but the mechanism is not known. B27 transgenic rats develop a spontaneous disease resembling the human spondyloarthropathies that includes arthritis and colitis. To investigate whether this disease requires the binding of specific peptides to B27, we made a minigene construct in which a peptide from influenza nucleoprotein, NP383-391 (SRYWAIRTR), which binds B27 with high affinity, is targeted directly to the ER by the signal peptide of the adenovirus E3/gp19 protein. Rats transgenic for this minigene, NP1, were made and bred with B27 rats. The production of the NP383-391 peptide in B27+NP1+ rats was confirmed immunologically and by mass spectrometry. The NP1 product displaced ∼90% of the 3H-Arg-labeled endogenous peptide fraction in B27+NP1+ spleen cells. Male B27+NP1+ rats had a significantly reduced prevalence of arthritis, compared with B27+NP− males or B27+ males with a control construct, NP2, whereas colitis was not significantly affected by the NP1 transgene. These findings support the hypothesis that B27-related arthritis requires binding of a specific peptide or set of peptides to B27, and they demonstrate a method for efficient transgenic targeting of peptides to the ER

Inter-relationship of plasma markers of oxidative stress and thyroid hormones in schizophrenics

<p>Abstract</p> <p>Background</p> <p>The relationship of oxidative stress to thyroid hormones has not been studied in the schizophrenics. The present study determined the status and interrelationship of plasma markers of oxidative stress, nitric oxide and thyroid hormones in thirty (17 males and 13 females) newly diagnosed patients with acute schizophrenia before initiation of chemotherapy. Twenty five (13 males and 12 females) mentally healthy individuals served as controls. Patients and controls with history of hard drugs (including alcohol and cigarette), pre-diagnosis medications (e.g. antiparkinsonian/antipsychotic drugs), chronic infections, liver disease and diabetes mellitus were excluded from the study. Plasma levels of total antioxidant potential (TAP), total plasma peroxides (TPP), nitric oxide (NO), malondialdehyde (MDA), thyroxine (T4), tri-iodothyronine (T3) and thyroid stimulating hormone (TSH) were determined in all participants using spectrophotometric and enzyme linked immunosorbent assay (ELISA) methods respectively. Oxidative stress index (OSI) was calculated as the percent ratio of total plasma peroxides and total antioxidant potential.</p> <p>Findings</p> <p>Significantly higher plasma levels of MDA (p < 0.01), TPP (p < 0.01), OSI (p < 0.01), T3 (p < 0.01) and T4 (p < 0.05) were observed in schizophrenics when compared with the controls. The mean levels of TAP, NO and TSH were significantly lower in schizophrenics (p < 0.01) when compared with the controls. The result shows that T3 values correlate significantly with MDA (p < 0.05) and TPP (p < 0.01) in schizophrenics.</p> <p>Conclusions</p> <p>Higher level of TPP may enhance thyroid hormogenesis in schizophrenics. Adjuvant antioxidant therapy may be a novel approach in the treatment of schizophrenic patients.</p

Intestinal, extra-intestinal and systemic sequelae of Toxoplasma gondii induced acute ileitis in mice harboring a human gut microbiota

Background Within seven days following peroral high dose infection with Toxoplasma gondii susceptible conventionally colonized mice develop acute ileitis due to an underlying T helper cell (Th) -1 type immunopathology. We here addressed whether mice harboring a human intestinal microbiota developed intestinal, extra-intestinal and systemic sequelae upon ileitis induction. Methodology/Principal findings Secondary abiotic mice were generated by broad- spectrum antibiotic treatment and associated with a complex human intestinal microbiota following peroral fecal microbiota transplantation. Within three weeks the human microbiota had stably established in the murine intestinal tract as assessed by quantitative cultural and culture-independent (i.e. molecular 16S rRNA based) methods. At day 7 post infection (p.i.) with 50 cysts of T. gondii strain ME49 by gavage human microbiota associated (hma) mice displayed severe clinical, macroscopic and microscopic sequelae indicating acute ileitis. In diseased hma mice increased numbers of innate and adaptive immune cells within the ileal mucosa and lamina propria and elevated intestinal secretion of pro-inflammatory mediators including IFN-γ, IL-12 and nitric oxide could be observed at day 7 p.i. Ileitis development was accompanied by substantial shifts in intestinal microbiota composition of hma mice characterized by elevated total bacterial loads and increased numbers of intestinal Gram-negative commensals such as enterobacteria and Bacteroides / Prevotella species overgrowing the small and large intestinal lumen. Furthermore, viable bacteria translocated from the inflamed ileum to extra- intestinal including systemic compartments. Notably, pro-inflammatory immune responses were not restricted to the intestinal tract as indicated by increased pro-inflammatory cytokine secretion in extra-intestinal (i.e. liver and kidney) and systemic compartments including spleen and serum. Conclusion/Significance With respect to the intestinal microbiota composition “humanized” mice display acute ileitis following peroral high dose T. gondii infection. Thus, hma mice constitute a suitable model to further dissect the interactions between pathogens, human microbiota and vertebrate host immunity during acute intestinal inflammation

Expression of pendrin in benign and malignant human thyroid tissues

The Pendred syndrome gene (PDS) encodes a transmembrane protein, pendrin, which is expressed in follicular thyroid cells and participates in the apical iodide transport. Pendrin expression has been studied in various thyroid neoplasms by means of immunohistochemistry (IHC), Western blot and RT–quantitative real-time PCR. The expression was related to the functional activity of the thyroid tissue. Follicular cells of normal, nodular goitre and Graves' disease tissues express pendrin at the apical pole of the thyrocytes. In follicular adenomas, pendrin was detected in cell membranes and cytoplasm simultaneously in 10 out of 15 cases. Pendrin protein was detected in 73.3 and 76.7% of the follicular (FTC) and papillary (PTC) thyroid carcinomas, respectively, where pendrin was solely localised inside the cytoplasm. An extensive intracellular immunostaining of pendrin was observed in six out of 11 (54.5%) of positive FTCs and 19 out of 23 (82%) of PTCs. Focal reactivity was detected in one follicular- and three papillary carcinomas, whereas pendrin protein was absent in three of 15 FTC and four of 30 PTC; mRNA of pendrin was detected in 92.4% of thyroid tumours. The relative mRNA expression of pendrin was lower in cancers than in normal thyroid tissues (P<0.001). The pendrin protein level was found to parallel its mRNA expression, which was not, however, related to the tumour size and tumour stage. In conclusion, pendrin is expressed in the majority of differentiated thyroid tumours with high individual variability but its targeting to the apical cell membrane is affected

Evidence for Significant Overlap between Common Risk Variants for Crohn's Disease and Ankylosing Spondylitis

BACKGROUND: A multicenter genome-wide association scan for Crohn's Disease (CD) has recently reported 40 CD susceptibility loci, including 29 novel ones (19 significant and 10 putative). To gain insight into the genetic overlap between CD and ankylosing spondylitis (AS), these markers were tested for association in AS patients. PRINCIPAL FINDINGS: Two previously established associations, namely with the MHC and IL23R loci, were confirmed. In addition, rs2872507, which maps to a locus associated with asthma and influences the expression of the ORMDL3 gene in lymphoblastoid cells, showed a significant association with AS (p = 0.03). In gut biopsies of AS and CD patients, ORMDL3 expression was not significantly different from controls and no correlation was found with the rs2872507 genotype (Spearman's rho: -0.067). The distribution of p-values for the remaining 36 SNPs was significantly skewed towards low p-values unless the top 5 ranked SNPs (ORMDL3, NKX2-3, PTPN2, ICOSLG and MST1) were excluded from the analysis. CONCLUSIONS: Association analysis using risk variants for CD led to the identification of a new risk variant associated with AS (ORMDL3), underscoring a role for ER stress in AS. In addition, two known and five potentially relevant associations were detected, contributing to common susceptibility of CD and AS

Thyroid peroxidase forms thionamide-sensitive homodimers: relevance for immunomodulation of thyroid autoimmunity

Thyroid peroxidase (TPO) is the key enzyme in thyroid hormone production and a universal autoantigen in Graves’ and other autoimmune thyroid diseases. We wished to explore the expression of TPO and whether it was affected by thionamide antithyroid drugs. We studied recombinant TPO, stably expressed by a Chinese hamster ovary cell line (CHO-TPO) and transiently expressed TPO-enhanced green fluorescent protein (eGFP) and -FLAG fusion proteins. Immunoblotting of CHO-TPO cell extracts showed high-molecular weight (HMW) TPO isoforms that were resistant to reduction, as well as 110 kDa monomeric TPO. Co-immunoprecipitation and enzyme-linked-immunosorbent assay (ELISA) binding studies of FLAG- and eGFP-tagged TPO demonstrated TPO dimerisation. CHO-TPO cells cultured in methimazole (MMI) for 10 days showed a significant reduction in HMW-TPO isoforms at MMI concentrations of 1 µM and above (p < 0.01), whereas monomeric TPO expression was unchanged. We observed a similar reduction in HMW-TPO in CHO-TPO cells cultured in propylthiouracil (10 µM and above). Binding of Graves’ disease patient sera and TPO-Fabs to enzymatically active TPO that was captured onto solid phase was not abrogated by MMI. The cellular localisation of TPO in CHO-TPO cells was unchanged by MMI treatment. Our demonstration of homodimeric TPO and the reduction in HMW-TPO isoforms during thionamide treatment of CHO-TPO cells shows, for the first time, an effect of thionamides on TPO structure. This suggests a structural correlate to the effect of thionamides on TPO enzymatic activity and opens up a novel potential mechanism for thionamide immunomodulation of autoimmune thyroid disease

Independent and Opposing Roles For Btk and Lyn in B and Myeloid Signaling Pathways

Transphosphorylation by Src family kinases is required for the activation of Bruton's tyrosine kinase (Btk). Differences in the phenotypes of Btk−/− and lyn−/− mice suggest that these kinases may also have independent or opposing functions. B cell development and function were examined in Btk−/−lyn−/− mice to better understand the functional interaction of Btk and Lyn in vivo. The antigen-independent phase of B lymphopoiesis was normal in Btk−/−lyn−/− mice. However, Btk−/−lyn−/− animals had a more severe immunodeficiency than Btk−/− mice. B cell numbers and response to T cell–dependent antigens were reduced. Btk and Lyn therefore play independent or partially redundant roles in the maintenance and function of peripheral B cells. Autoimmunity, hypersensitivity to B cell receptor (BCR) cross-linking, and splenomegaly caused by myeloerythroid hyperplasia were alleviated by Btk deficiency in lyn−/− mice. A transgene expressing Btk at ∼25% of endogenous levels (Btklo) was crossed onto Btk−/− and Btk−/−lyn−/− backgrounds to demonstrate that Btk is limiting for BCR signaling in the presence but not in the absence of Lyn. These observations indicate that the net outcome of Lyn function in vivo is to inhibit Btk-dependent pathways in B and myeloid cells, and that Btklo mice are a useful sensitized system to identify regulatory components of Btk signaling pathways

A monoclonal antibody against kininogen reduces inflammation in the HLA-B27 transgenic rat

The human leukocyte antigen B27 (HLA-B27) transgenic rat is a model of human inflammatory bowel disease, rheumatoid arthritis and psoriasis. Studies of chronic inflammation in other rat models have demonstrated activation of the kallikrein–kinin system as well as modulation by a plasma kallikrein inhibitor initiated before the onset of clinicopathologic changes or a deficiency in high-molecular-mass kininogen. Here we study the effects of monoclonal antibody C11C1, an antibody against high-molecular-mass kininogen that inhibits the binding of high-molecular-mass kininogen to leukocytes and endothelial cells in the HLA-B27 rat, which was administered after the onset of the inflammatory changes. Thrice-weekly intraperitoneal injections of monoclonal antibody C11C1 or isotype IgG(1 )were given to male 23-week-old rats for 16 days. Stool character as a measure of intestinal inflammation, and the rear limbs for clinical signs of arthritis (tarsal joint swelling and erythema) were scored daily. The animals were killed and the histology sections were assigned a numerical score for colonic inflammation, synovitis, and cartilage damage. Administration of monoclonal C11C1 rapidly decreased the clinical scores of pre-existing inflammatory bowel disease (P < 0.005) and arthritis (P < 0.001). Histological analyses confirmed significant reductions in colonic lesions (P = 0.004) and synovitis (P = 0.009). Decreased concentrations of plasma prekallikrein and high-molecular-mass kininogen were found, providing evidence of activation of the kallikrein–kinin system. The levels of these biomarkers were reversed by monoclonal antibody C11C1, which may have therapeutic potential in human inflammatory bowel disease and arthritis