Chitosan Oligosaccharides Suppress Nuclear Factor-Kappa B Activation and Ameliorate Experimental Autoimmune Uveoretinitis in Mice.

We investigated the therapeutic potential and mechanism of chitosan oligosaccharides (COS) for experimental autoimmune uveoretinitis (EAU) in mice. EAU was induced in C57/BL6 mice by injection of human interphotoreceptor retinoid-binding protein (IRBP) peptides. At the same time, a high or low dose (20 or 10 mg/kg) of COS or phosphate-buffered saline (PBS) was given to mice daily after EAU induction. We found that mouse EAU is ameliorated by the high-dose COS treatment when compared with PBS treatment. In the retinas of high-dose COS-treated mice, the nuclear translocation of NF-κB subunit (p65) was suppressed, and the expression of several key EAU inflammatory mediators, IFN-γ, TNF-α, IL-1α, IL-4, IL-5, IL-6, IL-10, IL-17 and MCP-1 was lowered. These results suggest that COS may be a potential treatment for posterior uveitis.

IFN-ß inhibits the increased expression of IL-9 during experimental autoimmune uveoretinitis.

PURPOSE: It has been shown that IL-9 plays a proinflammatory role in the pathogenesis of certain autoimmune diseases. This study was designed to investigate the possible role of IL-9 in the development of experimental autoimmune uveoretinitis (EAU) and the effect of IFN-ß on its expression. METHODS: EAU was induced in B10RIII mice by immunization with interphotoreceptor retinoid-binding protein peptide 161-180 (IRBP(161-180)). IFN-ß was administered subcutaneously to IRBP(161-180) immunized mice every other day from day one before immunization to the end of the study. Splenocytes and draining lymph node (DLN) cells from EAU mice or control mice or EAU mice treated with IFN-ß or PBS were stimulated with anti-CD3/CD28 or IRBP(161-180) for 3 days. Naïve T cells cultured under Th1 or Th17 polarizing conditions were incubated in the presence or absence of IFN-ß for 4 days. Effector/memory T cells were activated by anti-CD3/CD28 in the presence or absence of IFN-ß for 3 days. IFN-ß-treated monocytes were cocultured with naïve T cells or effector/memory T cells for 3 days. Culture supernatants were collected and IL-9 was detected by ELISA. RESULTS: IL-9 expression in splenocytes and DLN cells was increased in EAU mice during the inflammatory phase and returned back to lower levels during the recovery phase. IFN-ß in vivo treatment significantly inhibited EAU activity in association with a down-regulated expression of IL-9. In vitro polarized Th1 and Th17 cells both secreted IL-9 and the addition of IFN-ß suppressed production of IL-9 by both Th subsets. Beside its effect on polarized Th cells, IFN-ß also suppressed the secretion of IL-9 by effector/memory T cells. However, IFN-ß-treated monocytes had no effect on the production of IL-9 when cocultured with naïve or effector/memory T cells. CONCLUSION: IL-9 expression is increased during EAU which could be suppressed by IFN-ß.

Posttranslational modification of differentially expressed mitochondrial proteins in the retina during early experimental autoimmune uveitis.

PURPOSE: Posttranslational modification of proteins plays an important role in cellular functions and is a key event in signal transduction pathways leading to oxidative stress and DNA damage. In this study, we used matrix-assisted laser desorption/ionization- time of flight (MALDI-TOF) to investigate the posttranslational modifications of the differentially expressed proteins in the retinal mitochondria during early experimental autoimmune uveitis (EAU). METHODS: EAU was induced in 18 B10RIII mice with 25 µg of inter-photoreceptor retinoid-binding protein (IRBP) emulsified with complete Freund's adjuvant (CFA); 18 mice treated with CFA without IRBP served as controls. Retinas were removed from the experimental and control groups on day 7 post immunization; mitochondrial fractions were extracted and subjected to 2 dimentional-difference in gel electrophoresis (2D-DIGE); and the protein spots indicating differential expression were subjected to MALDI-TOF for protein identification and indication of any posttranslational modifications. RESULTS: Of the 13 proteins found to be differentially expressed by 2D-DIGE (including upregulated aconitase, mitochondrial heat shock protein (mtHsp) 70, lamin-1, syntaxin-binding protein, αA crystallin, ßB2 crystallin, along with downregulated guanine nucleotide-binding protein and ATP synthase) nine were found to undergo posttranslational modification. Oxidation was a common modification found to occur on aconitase, mtHsp 70, ATP synthase, lamin-1, ßB2-crystallin, guanine nucleotide-binding protein, and manganese superoxide dismutase (MnSOD). In addition, aconitase hydratase, mtHsp 70, guanine nucleotide-binding protein, ATP synthase, syntaxin-binding protein, ßB2-crystallin, and lamin-1 were also modified by carbamidomethylation. αA-crystallin had a pyro-glu modification. CONCLUSIONS: Several proteins present in the retinal mitochondria are posttranslationally modified during early EAU, indicating the presence of oxidative stress and mitochondrial DNA damage. The most common modifications are oxidation and carbamidomethylation. A better understanding of the proteins susceptible to posttranslational modifications in the mitochondria at the early stage of the disease may serve to advance therapeutic interventions to attenuate disease progression.

Coordinated gene expression of Th17- and Treg-associated molecules correlated with resolution of the monophasic experimental autoimmune uveitis.

PURPOSE: To investigate the role of T-cell-mediated immune response in a monophasic experimental autoimmune uveitis (EAU). METHODS: A monophasic EAU was induced in Lewis rats by immunization with interphotoreceptor retinoid-binding protein peptide. Optimized quantitative real-time RT-PCR was used for consecutive measurement of the relative expression of Th17-associated molecules, including interleukin 6 (IL-6), transforming growth factor-ß (TGF-ß), interleukin 23p19 (IL-23p19), interleukin 23p40 (IL-23p40), CD4, CD8, major histocompatibility complex I (MHC I), major histocompatibility complex II (MHC II), interleukin 17 (IL-17), interleukin 17F (IL-17F), interleukin 17 receptor A (IL-17RA), retinoic acid-related orphan receptor γt (RORγt) and Chemokine receptor 6 (CCR6), in addition to Treg-related forkhead box P3 (Foxp3), C-X-C chemokine receptor type 5 (CXCR5), and cluster of differentiation 25 (CD25) at the initiation, effector, and resolution phases of EAU and compared with those at 14 days post-immunization of control animals. Immunohistochemisty was used to examine IL-17 expression in retinas. Glial fibrillary acidic protein retinal astrocytes, Neuronal class III ß-Tubulin(Tuj1(+))retinal ganglion cells, and infiltrating CD11b(+) microglia were analyzed by fluorescent microscopy in a kinetic manner. RESULTS: Our results indicated well organized T-cell activity, measured by relative expression of multiple T-cell-related factors at the mRNA level, synchronized with the initiation of autoimmune inflammation, and thereafter resolution of the monophasic EAU. Immune balance was achieved several times through coordinated expression of Th17- and Treg-related factors. The expression pattern of these factors and results from immunochemistry with an IL-17 antibody indicated that there may be intensive crosstalk between infiltrating immune cells and the resident neural cells, which were significantly activated during the course of disease. CONCLUSIONS: T-cell-mediated immune response played a positive role in resolution of the monophasic EAU.

Xenopus IRBP, a phylogenetically remote protein, is uveitogenic in Lewis rats.

Mammalian interphotoreceptor retinoid-binding proteins (IRBPs) are highly uveitogenic in Lewis rats. Xenopus laevis IRBP resembles mammalian IRBP in its four-fold structure, and has approximately 70% amino acid sequence identity with the bovine protein. This study investigated the uveitogenicity of recombinant Xenopus IRBP and two of its derived peptides in Lewis rats. Rats immunized with Xenopus IRBP developed uveoretinitis as well as pineal inflammation. The Xenopus molecule was, however, less immunopathogenic than the bovine IRBP. Of the two Xenopus IRBP peptides tested, 1180-1191 was remarkably uveitogenic, whereas sequence 521-540 exhibited low activity. It is assumed, therefore, that as with bovine IRBP, peptide 1180-1191 is the major uveitogenic sequence in Xenopus IRBP. The role individual residues of these peptides play in the immunopathogenic process is discussed. Our data thus demonstrate that despite its being phylogenetically remote, Xenopus IRBP is uveitogenic in Lewis rats

Envolvimento renal na infecçäo do trato urinário recorrente: importância da prote1na transportadore de retinol como marcador de disfunçäo tubular / Renal envolvemen in urinary tract infection recurring: retinol binding protein importance how marker of tubular disfuncion

Com o objetivo de pesquisar o envolvimento renal na infecçäo do trato urinário (ITU), realizaram-se a ultra-sonografia e a cintilografia renal em 66 crianças (média de idade = 30 anos). Em 20 crianças e em 23 mulheres comrecorrência da ITU (ITUr), realizou-se, também, a dosagem da proteína transportadora de retinol (RBP) na urina. Como controles, foram estudadas41 crianças (média de idade =7,2 anos) e 24 mulheres (média de idade = 32 anos). Havia cicatriz pielonefrítica em 25 porcento das crianças e em 26 porcento das mulheres com ITUr. A excreçäo urinária de RBP foi significantemente maior nas crianças e nas mulheres com ITU que nas normais. Nossos resultados demonstrtam disfunçäo tubular renal na infecçäo do trato urinário recorrente.(au)