RESUMEN

Roughly one-half of mice with partial defects in two immune tolerance pathways (AireGW/+Lyn-/- mice) spontaneously develop severe damage to their retinas due to T cell reactivity to Aire-regulated interphotoreceptor retinoid-binding protein (IRBP). Single-cell T cell receptor (TCR) sequencing of CD4+ T cells specific for a predominate epitope of IRBP showed a remarkable diversity of autoantigen-specific TCRs with greater clonal expansions in mice with disease. TCR transgenic mice made with an expanded IRBP-specific TCR (P2.U2) of intermediate affinity exhibited strong but incomplete negative selection of thymocytes. This negative selection was absent in IRBP-/- mice and greatly defective in AireGW/+ mice. Most P2.U2+/- mice and all P2.U.2+/-AireGW/+ mice rapidly developed inflammation of the retina and adjacent uvea (uveitis). Aire-dependent IRBP expression in the thymus also promoted Treg differentiation, but the niche for this fate determination was small, suggesting differences in antigen presentation leading to negative selection vs. thymic Treg differentiation and a stronger role for negative selection in preventing autoimmune disease in the retina.

Asunto(s)

Presentación de Antígeno , Receptores de Antígenos de Linfocitos T , Animales , Ratones , Autoantígenos , Modelos Animales de Enfermedad , Ratones Endogámicos , Ratones Transgénicos

RESUMEN

Interphotoreceptor retinoid-binding protein (IRBP) is an abundant glycoprotein in the subretinal space bound by the photoreceptor (PR) outer segments and the processes of the retinal pigmented epithelium (RPE). IRBP binds retinoids, including 11-cis-retinal and all-trans-retinol. In this study, visual function for demanding visual tasks was assessed in IRBP knock-out (KO) mice. Surprisingly, IRBP KO mice showed no differences in scotopic critical flicker frequency (CFF) compared to wildtype (WT). However, they did have lower photopic CFF than WT. IRBP KO mice had reduced scotopic and photopic acuity and contrast sensitivity compared to WT. IRBP KO mice had a significant reduction in outer nuclear layer (ONL) thickness, PR outer and inner segment, and full retinal thickness (FRT) compared to WT. There were fewer cones in IRBP KO mice. Overall, these results confirm substantial loss of rods and significant loss of cones within 30 days. Absence of IRBP resulted in cone circuit damage, reducing photopic flicker, contrast sensitivity, and spatial frequency sensitivity. The c-wave was reduced and accelerated in response to bright steps of light. This result also suggests altered retinal pigment epithelium activity. There appears to be a compensatory mechanism such as higher synaptic gain between PRs and bipolar cells since the loss of the b-wave did not linearly follow the loss of rods, or the a-wave. Scotopic CFF is normal despite thinning of ONL and reduced scotopic electroretinogram (ERG) in IRBP KO mice, suggesting either a redundancy or plasticity in circuits detecting (encoding) scotopic flicker at threshold even with substantial rod loss.

Asunto(s)

Proteínas del Ojo , Visión Nocturna , Retina , Proteínas de Unión al Retinol , Retina/fisiología , Retina/ultraestructura , Estimulación Luminosa , Proteínas del Ojo/genética , Proteínas del Ojo/fisiología , Proteínas de Unión al Retinol/genética , Proteínas de Unión al Retinol/fisiología , Ratones Noqueados , Animales , Ratones , Fusión de Flicker/genética , Fusión de Flicker/fisiología , Visión de Colores/genética , Visión de Colores/fisiología , Agudeza Visual/genética , Agudeza Visual/fisiología , Visión Nocturna/genética , Visión Nocturna/fisiología , Tomografía de Coherencia Óptica , Masculino , Femenino

RESUMEN

We report the first molecular detection of Leishmania infection (subgenus Viannia) in the yellow-faced parrot (Alipiopsitta xanthops), at a wildlife rehabilitation center located in the city of Campo Grande, Brazil, an endemic area for leishmaniasis. PCRs targeting kinetoplast DNA (kDNA) and the small subunit of ribosomal RNA of Leishmania spp. were performed, both positive, followed by the sequencing of the amplified region of the SSU rDNA gene, which confirmed the identity of the parasite. This is the first report of success obtained in the use of PCR targeting the IRBP (Interphotoreceptor retinoid-binding protein) gene as an internal control in the molecular diagnosis of pathogens in bird species.

Asunto(s)

Loros , Leishmaniasis/diagnóstico

RESUMEN

Experimental autoimmune uveoretinitis (EAU) in mice provides a useful platform to study the pathogenesis and experimental therapeutics of human uveitis. One often used EAU model employs C57BL/6 (B6) mice sensitized with a peptide residue having 1 to 20 amino acids of human interphotoreceptor retinoid binding protein (hIRBP1-20). The model using the B6 background has permitted a liberal use of genetically engineered strains and has provided insights for understanding uveoretinitis. However, this is usually acute/monophasic and does not represent human uveoretinitis that is characterized as a chronic/recurrent disease. Several chronic/recurrent EAU models have been developed; of these, we employed administration of staphylococcal enterotoxin B (SEB) for relapse in the present study, and found that recurrence was induced at day 24 after primary immunization, which is thought to be the convalescent phase. We reported the activation of invariant natural killer T (iNKT)-cells upon primary immunization of the EAU model mice with the ligand RCAI-56, which was found to mitigate the disease in our previous study. Here, we first attempted to ameliorate EAU in the relapse model using a preventive regimen by activating iNKT cells at the same time relapse induction (day 24) or in a regimen after 3 days of relapse induction (day 27). The preventive as well as post-inductive regimens were successful in reducing histopathological scores by inhibiting the Ag-specific Th17-biased response. Collectively, activation of iNKT cells may be useful to mitigate the relapse response of EAU induced with SEB.

Asunto(s)

Enfermedades Autoinmunes/prevención & control , Modelos Animales de Enfermedad , Células T Asesinas Naturales/fisiología , Retinitis/prevención & control , Uveítis/prevención & control , Animales , Enfermedades Autoinmunes/inmunología , Proliferación Celular , Proteínas del Ojo/toxicidad , Femenino , Citometría de Flujo , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Recurrencia , Retinitis/inmunología , Proteínas de Unión al Retinol/toxicidad , Células TH1/inmunología , Células Th17/inmunología , Células Th2/inmunología , Uveítis/inmunología

RESUMEN

Interphotoreceptor retinoid-binding protein (IRBP), also known as retinol binding protein 3 (RBP3), is a lipophilic glycoprotein specifically secreted by photoreceptors. Enriched in the interphotoreceptor matrix (IPM) and recycled by the retinal pigment epithelium (RPE), IRBP is essential for the vision of all vertebrates as it facilitates the transfer of retinoids in the visual cycle. It also helps to transport lipids between the RPE and photoreceptors. The thiol-dependent antioxidant activity of IRBP maintains the delicate redox balance in the normal retina. Thus, its dysfunction is suspected to play a role in many retinal diseases. We have reviewed here the latest research on IRBP in both retinal health and disease, including the function and regulation of IRBP under retinal stress in both animal models and the human retina. We have also explored the therapeutic potential of targeting IRBP in retinal diseases. Although some technical barriers remain, it is possible that manipulating the expression of IRBP in the retina will rescue or prevent photoreceptor degeneration in many retinal diseases.

RESUMEN

Since the publication of our previous paper, Visual cycle proteins: Structure, function, and roles in human retinal disease (Tsin, et.al, JBC 293:13016, 2018) there has been significant progress on multiple topics discussed in this paper. In the present communication, we further explore research advances on two visual cycle proteins: DES1 and IRBP. In addition, we emphasize the progress of clinical translation of other visual cycle protein research, including the breakthrough of FDA-approved gene therapy for Leber's congenital amaurosis, and additional gene therapies at different stages of clinical trials for various retinal diseases such as retinitis pigmentosa, diabetic retinopathy, and Stargardt's disease.

RESUMEN

IL-22 has opposing effects in different tissues, from pro-inflammatory (skin, joints) to protective (liver, intestine) but little is known about its effects on neuroinflammation. We examined the effect of IL-22 on retinal tissue by using the model of experimental autoimmune uveitis (EAU) in IL-22-/- mice, as well as by intraocular injections of recombinant IL-22 or anti-IL-22 antibodies in wild type animals. During EAU, IL-22 was produced in the eye by CD4+ eye-infiltrating T cells. EAU-challenged IL-22-/- mice, as well as WT mice treated systemically or intraocularly with anti-IL-22 antibodies during the expression phase of disease, developed exacerbated retinal damage. Furthermore, IL-22-/- mice were more susceptible than WT controls to glutamate-induced neurotoxicity, whereas local IL-22 supplementation was protective, suggesting direct or indirect neuroprotective effects. Mechanistic studies revealed that retinal glial Müller cells express IL-22rα1 in vivo, and in vitro IL-22 enhanced their ability to suppress proliferation of effector T cells. Finally, IL-22 injected into the eye concurrently with IL-1, inhibited the (IL-1-induced) expression of multiple proinflammatory and proapoptotic genes in retinal tissue. These findings suggest that IL-22 can function locally within the retina to reduce inflammatory damage and provide neuroprotection by affecting multiple molecular and cellular pathways.

Asunto(s)

Autoinmunidad , Sistema Nervioso Central/inmunología , Sistema Nervioso Central/metabolismo , Susceptibilidad a Enfermedades , Interleucinas/metabolismo , Animales , Enfermedades Autoinmunes/etiología , Enfermedades Autoinmunes/metabolismo , Enfermedades Autoinmunes/patología , Autoinmunidad/genética , Sistema Nervioso Central/patología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/etiología , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/patología , Células Ependimogliales/inmunología , Células Ependimogliales/metabolismo , Células Ependimogliales/patología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Interleucinas/genética , Interleucinas/farmacología , Activación de Linfocitos/inmunología , Ratones , Ratones Noqueados , Enfermedades del Sistema Nervioso/etiología , Enfermedades del Sistema Nervioso/metabolismo , Enfermedades del Sistema Nervioso/patología , Neuroprotección/genética , Índice de Severidad de la Enfermedad , Linfocitos T/inmunología , Linfocitos T/metabolismo , Uveítis/etiología , Uveítis/metabolismo , Uveítis/patología , Interleucina-22

RESUMEN

Non-infections uveitis in humans is an autoimmune disease of the retina and uvea that can be blinding if untreated. Its laboratory equivalent is experimental autoimmune uveitis (EAU) induced in susceptible rodents by immunization with retinal antigens and described elsewhere in this series (Agarwal et al., Methods Mol Biol, 900:443-469, 2012). Evaluation and quantitation of the disease is usually performed by fundus examination and/or histopathology, which provide limited information on structural and no information on functional changes as disease progresses. Here, we describe methods for systematic evaluation of disease using noninvasive clinical assessments by fundus examination and photography, optical coherence tomography, and functional evaluation by electroretinography, which are then compared to histopathology. Using these methodologies, we demonstrate that clinical variants of disease can be accurately evaluated both clinically and functionally, facilitating longitudinal follow-up and providing information that cannot be obtained by fundoscopy and histology alone. These methodologies can be useful to obtain additional information and to evaluate effects of therapeutic modalities under investigation.

Asunto(s)

Enfermedades Autoinmunes/inmunología , Inflamación/inmunología , Tomografía de Coherencia Óptica/métodos , Uveítis/inmunología , Animales , Antígenos/inmunología , Enfermedades Autoinmunes/patología , Modelos Animales de Enfermedad , Fondo de Ojo , Humanos , Inflamación/patología , Retina/inmunología , Retina/patología

RESUMEN

OBJECTIVE: Experimental autoimmune uveitis (EAU) represents autoimmune uveitis in humans, among which B10.RIII and C57BL/6 are the frequently used strains in mice, but to date, no study has been reported to compare EAU disease between the two strains. Here we compared the differences in morphology, pathology, visual function of the retinal inflammation and Th1/Th17 immune responses in the EAU models induced by the interphotoreceptor retinoid binding protein (IRBP) between the B10.RIII and C57BL/6 strains, using fundus and histological examinations, optical coherence tomography, electroretinography and immunoassays. METHOD: EAU induced in B10.RIII mice exhibited a shortterm severe inflammation with massive ocular infiltrates of inflammatory cells and extensive destruction of the retina that culminated in rapid degeneration of the retina and permanent loss of visual function. In contrast, C57BL/6 mice developed chronic inflammation with recurring and persistent retinitis for several months, highlighting moderate scores of disease severity and visual signal in comparison with those in B10.RIII mice. Consistent with the clinical manifestations, increased Th1/Th17 effector responses were detected in the uveitic eyes of B10.RIII strain than those in C57BL/6 strain. These data demonstrate distinguishing features of retinal inflammation and T-cell immune responses involved in IRBP-induced EAU between B10.RIII and C57BL/6 strains. CONCLUSION: Our findings suggest that the persistent-recurring EAU model induced in C57BL/6 mice may serve as a better tool to represent distinct aspects of human uveitis.

Asunto(s)

Enfermedades Autoinmunes/inmunología , Proteínas del Ojo/inmunología , Proteínas de Unión al Retinol/inmunología , Células TH1/inmunología , Células Th17/inmunología , Uveítis/inmunología , Animales , Enfermedades Autoinmunes/patología , Enfermedad Crónica , Modelos Animales de Enfermedad , Humanos , Inflamación/inmunología , Inflamación/patología , Ratones , Especificidad de la Especie , Células TH1/patología , Células Th17/patología , Uveítis/patología

RESUMEN

Recent discoveries on the role of commensal microbiota have significantly changed our understanding of human physiology. The host-microbiota interplay is now an important aspect to take into account to understand immune responses and immunological diseases. Autoimmune uveitis is a sight-threatening disease that arises without a known infectious etiology. It is unknown where and how autoreactive T cells become primed to trigger disease in the eye, which is an immune privileged site. We recently reported data supporting the notion that retina-specific T cells receive a signal in the gut from commensal microbiota-derived cross-reactive antigen(s) and trigger autoimmune uveitis in the R161H mouse model. Here we discuss our published findings, as well as our recent attempts to identify the responsible microbe(s) by using different antibiotic treatments, 16S rDNA sequencing and homology searches for candidate antigenic mimic(s) of the retinal antigen.

Asunto(s)

Antígenos/inmunología , Enfermedades Autoinmunes/microbiología , Microbioma Gastrointestinal , Uveítis/inmunología , Uveítis/microbiología , Animales , Enfermedades Autoinmunes/inmunología , Autoinmunidad , Humanos , Retina/inmunología , Linfocitos T/inmunología

RESUMEN

In Drosophila, P-element transposition causes mutagenesis and genome instability during hybrid dysgenesis. The P-element 31-bp terminal inverted repeats (TIRs) contain sequences essential for transposase cleavage and have been implicated in DNA repair via protein-DNA interactions with cellular proteins. The identity and function of these cellular proteins were unknown. Biochemical characterization of proteins that bind the TIRs identified a heterodimeric basic leucine zipper (bZIP) complex between an uncharacterized protein that we termed "Inverted Repeat Binding Protein (IRBP) 18" and its partner Xrp1. The reconstituted IRBP18/Xrp1 heterodimer binds sequence-specifically to its dsDNA-binding site within the P-element TIRs. Genetic analyses implicate both proteins as critical for repair of DNA breaks following transposase cleavage in vivo. These results identify a cellular protein complex that binds an active mobile element and plays a more general role in maintaining genome stability.

Asunto(s)

Elementos Transponibles de ADN , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Animales , ADN/metabolismo , Daño del ADN , Reparación del ADN , Drosophila/genética , Drosophila/metabolismo , Mutación , Multimerización de Proteína

RESUMEN

Adhesion molecules play a central role in leukocyte adhesion to the blood-retinal barrier (BRB) during uveitis. VCAM-1 expression on the BRB has been already described but although structurally similar, ICAM-1 has shown in various autoimmunity models to have distinct role and expression. Here, we induced uveitis in C57Bl/6 mice by adoptive transfer of semi-purified T cells from IRBP1-20-immunized mice. Using Flow cytometry analysis on transferred cells and immunofluorescence staining on retina we have studied the comparative ocular expression of both ICAM-1 and VCAM-1 and their ligands LFA-1 and VLA-4 at the surface of uveitogenic cells. Our results showed that LFA-1 and VLA-4 are expressed on both T and non T cells, VLA-4 sparsely and LFA-1 ubiquitously. Considering retinal expression, ICAM-1 is faintly present and VCAM-1 is absent in naive eyes. Only ICAM-1 is present on infiltrating cells in the retina and vitreous, while only VCAM-1 extends to perivascular glial cells and all along the internal limiting membrane. Finally, ICAM-1 is strongly expressed on the RPE, where VCAM-1 expression is much weaker. VCAM-1 seems most strongly expressed on the internal BRB while ICAM-1 predominates on the external BRB. Those major differences in the expression pattern could represent differential entry pathways for inflammatory cells to penetrate the eye.

Asunto(s)

Enfermedades Autoinmunes/metabolismo , Barrera Hematorretinal , Molécula 1 de Adhesión Intercelular/biosíntesis , Linfocitos T/inmunología , Uveítis/metabolismo , Molécula 1 de Adhesión Celular Vascular/biosíntesis , Traslado Adoptivo , Animales , Enfermedades Autoinmunes/inmunología , Adhesión Celular , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Uveítis/inmunología , Uveítis/patología

RESUMEN

The surface plasmon resonance (SPR) biosensor method is a highly sensitive, label-free technique to study the non-covalent interactions of biomolecules, especially protein-protein and protein-small molecule interactions. We have explored this robust biosensor platform to study the interactions of carotenoid-binding proteins and their carotenoid ligands to assess the specificity of interaction, kinetics, affinity, and stoichiometry. These characterizations are important to further study uptake and transport of carotenoids to targeted tissues such as the macula of the human eye. In this review, we present an overview of the SPR method and optimization of assay conditions, and we discuss the particular challenges in studying carotenoid-protein interactions using SPR.

Asunto(s)

Carotenoides/metabolismo , Proteínas Portadoras/metabolismo , Resonancia por Plasmón de Superficie/métodos , Transporte Biológico , Humanos , Mácula Lútea/metabolismo , Unión Proteica

RESUMEN

PURPOSE: Interphotoreceptor retinoid-binding protein's (IRBP) role in facilitating the exchange of retinoids between rod and cone photoreceptors, RPE, and Müller cells in the visual cycle remains a mystery. Interphotoreceptor retinoid-binding protein's ability to bind the pericellular matrix of the cone outer segment and Müller cell villi suggests a function in all-trans and 11-cis retinol targeted trafficking in the cone visual cycle. We hypothesize that IRBP facilitates delivery and uptake of all-trans retinol to and release of 11-cis retinol from rat Müller cells (rMC-1). METHODS: Rat Müller cells were incubated with all-trans retinol and BSA or bovine IRBP (bIRBP). Retinoids in the cell homogenates and conditioned media were analyzed by high performance liquid chromatography (HPLC). RESULTS: Cells incubated with 10 µM retinol and BSA had 2100 pmol of all-trans retinol per milligram homogenate protein compared with 3450 pmol when retinol was delivered by bIRBP; these cells also had 450 pmol all-trans retinyl ester per milligram when retinol was delivered by BSA compared with 270 pmol when retinol was delivered by bIRBP. Conditioned media from cells incubated with retinol delivered by BSA did not contain11-cis retinol. However, cells with retinol delivered by bIRBP released 130 pmol/mL of 11-cis retinol into the cell media. Incubation with 5.0 mM deferoxamine (an iron chelator) reduced IRBP-dependent 11-cis retinol retrieval by 60%. CONCLUSIONS: Promoting Müller cell uptake of all-trans retinol and release of 11-cis retinol is a previously unrecognized function of IRBP that may be critical to cone function and integrity.

Asunto(s)

Células Ependimogliales/metabolismo , Proteínas del Ojo/metabolismo , Células Fotorreceptoras Retinianas Conos/metabolismo , Proteínas de Unión al Retinol/metabolismo , Visión Ocular/fisiología , Vitamina A/farmacocinética , Animales , Bovinos , Recuento de Células , Células Cultivadas , Cromatografía Líquida de Alta Presión , Medio de Cultivo Libre de Suero , Células Ependimogliales/citología , Células Ependimogliales/efectos de los fármacos , Ratas , Células Fotorreceptoras Retinianas Conos/citología , Células Fotorreceptoras Retinianas Conos/efectos de los fármacos

RESUMEN

We report here identification of novel mimicry epitopes for interphotoreceptor retinoid-binding protein (IRBP) 201-216, a candidate ocular antigen that causes experimental autoimmune uveoretinitis (EAU) in A/J mice. One mimicry epitope from Ehrlichia canis (EHC), designated EHC 44-59, induced cross-reactive T cells for IRBP 201-216 capable of producing T helper (Th)1 and Th17 cytokines, but failed to induce EAU in A/J mice. In addition, animals first primed with suboptimal doses of IRBP 201-216 and subsequently immunized with EHC 44-59 did not develop EAU; rather, the mimicry epitope prevented the disease induced by IRBP 201-216. However, alteration in the composition of EHC 44-59 by substituting alanine with valine at position 49, similar to the composition of IRBP 201-216, enabled the mimicry epitope to acquire uveitogenicity. The data provide new insights as to how microbes containing mimicry sequences for retinal antigens can prevent ocular inflammation by acting as naturally occurring altered peptide ligands.

Asunto(s)

Enfermedades Autoinmunes del Sistema Nervioso/prevención & control , Ehrlichia canis/inmunología , Ehrlichiosis/prevención & control , Imitación Molecular/inmunología , Retinitis/prevención & control , Uveítis/prevención & control , Secuencia de Aminoácidos , Animales , Enfermedades Autoinmunes del Sistema Nervioso/inmunología , Enfermedades Autoinmunes del Sistema Nervioso/microbiología , Bovinos , Ehrlichia canis/genética , Ehrlichiosis/inmunología , Ehrlichiosis/microbiología , Proteínas del Ojo/administración & dosificación , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Femenino , Ligandos , Ratones , Ratones Endogámicos A , Datos de Secuencia Molecular , Retinitis/inmunología , Retinitis/microbiología , Proteínas de Unión al Retinol/administración & dosificación , Proteínas de Unión al Retinol/genética , Proteínas de Unión al Retinol/metabolismo , Uveítis/inmunología , Uveítis/microbiología

RESUMEN

Despite presence of circulating retina-specific T cells in healthy individuals, ocular immune privilege usually averts development of autoimmune uveitis. To study the breakdown of immune privilege and development of disease, we generated transgenic (Tg) mice that express a T cell receptor (TCR) specific for interphotoreceptor retinoid-binding protein (IRBP), which serves as an autoimmune target in uveitis induced by immunization. Three lines of TCR Tg mice, with different levels of expression of the transgenic R161 TCR and different proportions of IRBP-specific CD4⁺ T cells in their peripheral repertoire, were successfully established. Importantly, two of the lines rapidly developed spontaneous uveitis, reaching 100% incidence by 2 and 3 months of age, respectively, whereas the third appeared "poised" and only developed appreciable disease upon immune perturbation. Susceptibility roughly paralleled expression of the R161 TCR. In all three lines, peripheral CD4⁺ T cells displayed a naïve phenotype, but proliferated in vitro in response to IRBP and elicited uveitis upon adoptive transfer. In contrast, CD4⁺ T cells infiltrating uveitic eyes mostly showed an effector/memory phenotype, and included Th1, Th17 as well as T regulatory cells that appeared to have been peripherally converted from conventional CD4⁺ T cells rather than thymically derived. Thus, R161 mice provide a new and valuable model of spontaneous autoimmune disease that circumvents the limitations of active immunization and adjuvants, and allows to study basic mechanisms involved in maintenance and breakdown of immune homeostasis affecting immunologically privileged sites such as the eye.

Asunto(s)

Autoantígenos/inmunología , Autoinmunidad/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Retina/inmunología , Animales , Enfermedades Autoinmunes/inmunología , Linfocitos T CD4-Positivos/inmunología , Citocinas/inmunología , Proteínas del Ojo/inmunología , Humanos , Memoria Inmunológica/inmunología , Ratones , Ratones Transgénicos , Receptores de Antígenos de Linfocitos T/biosíntesis , Proteínas de Unión al Retinol/inmunología , Linfocitos T Reguladores/inmunología , Células TH1/inmunología , Células Th17/inmunología , Uveítis/inmunología

RESUMEN

The recently described taxon Drymoreomys albimaculatus is endemic to the Brazilian Atlantic Forest and its biology and genetics are still poorly known. Herein, we present, for the first time, the karyotype of the species using classical and molecular cytogenetics, which showed 2n=62, FN=62, and interstitial telomeric signals at the sex chromosomes. Nuclear and mitochondrial DNA sequences from the two karyotyped individuals verify the taxonomic identity as the recently described Drymoreomys albimaculatus and confirm the relationship of the species with other Oryzomyini. Additionally, external morphological information is provided.

RESUMEN

Most human autoimmune diseases have a relapsing-remitting or a chronic progressive course, while animal models are usually acute and monophasic. In our experimental animal model the disease can be either monophasic or remitting, depending on the autoantigen used for induction, and it appears to lie in the effector phenotype of the elicited T helper cell response. Since both, monophasic and relapsing courses of disease are induced by immunization as well as by adoptive transfer of peptide-specific, CD4(+) T cells, we were able to directly compare the transcriptomes of pathogenic T cell lines by gene array analysis and qPCR as well as protein expression. Upregulated genes were only determined in T cells inducing relapsing uveitis and belong to certain pathways of antigen presentation, activation, inflammation, migration and survival, comprising WNT, Hedgehog, MAP-kinase and JAK/STAT-pathways. These pathways are partially interacting with each other, and the central molecule upregulated in T cells causing relapsing disease was found to be IFN-γ. Here the course of the autoimmune diseases strictly depends on the characteristics of the autoreactive T cells, which are already determined at their early stage of antigen-specific activation. Our rat models of experimental autoimmune uveitis could help elucidating the immune mechanisms behind relapsing autoimmunity in order to develop better therapeutic strategies.

Asunto(s)

Enfermedades Autoinmunes/etiología , Enfermedades Autoinmunes/patología , Animales , Presentación de Antígeno/inmunología , Autoantígenos/efectos adversos , Autoantígenos/inmunología , Enfermedades Autoinmunes/inmunología , Modelos Animales de Enfermedad , Humanos , Inflamación/etiología , Inflamación/inmunología , Inflamación/patología , Proteína Glutamina Gamma Glutamiltransferasa 2 , Ratas , Ratas Endogámicas Lew , Recurrencia , Uveítis/etiología , Uveítis/inmunología , Uveítis/patología

RESUMEN

The close packing of vertebrate photoreceptors presents a challenge to the exchange of molecules between the outer segments, retinal pigmented epithelium (RPE), and Müller glia. An extracellular hyaluronan scaffold separates these cells while soluble interphotoreceptor matrix (IPM) proteins traffic visual cycle retinoids, fatty acids, and other molecules between them. In the IPM, retinoids and fatty acids are carried by interphotoreceptor retinoid-binding protein (IRBP). The fact that much of the retina's IRBP can be extracted by saline wash has led to the notion that IRBP does not bind to the retina, but freely distributes itself within the subretinal space. In this study, we challenge this idea by asking if there are specialized IPM domains that bind IRBP, perhaps facilitating its ability to target delivery/uptake of its ligands. Xenopus is an ideal animal model to study the role of the IPM in RPE-photoreceptor interactions. Here, we took advantage of the large size of its photoreceptors, ability to detach the retina in light, sustainability of the retina in short term organ culture, and the availability of recombinant full-length Xenopus IRBP and antisera directed against Xenopus IRBP. We compared the distribution of wash resistant native IRBP, and that of IRBP-Alexa 647 binding in Xenopus retina. IRBP and cone opsin were localized using anti-Xenopus IRBP serum, and monoclonal COS-1 respectively. Cone matrix sheath proteoglycans were localized with wheat germ agglutinin (WGA), and diffuse IPM proteoglycans with peanut agglutinin (PNA). Wholemounts and frozen sections were compared by immunofluorescence from retinas detached under Ringer's followed by additional washes, or detached directly under 4% paraformaldehyde without Ringer's wash. Undetached Lowicryl embedded retinas were subjected to IRBP immunogold electron microscopy (EM). Immunogold labeled a diffuse network of filamentous structures, and a separate distinct flocculant material directly coating the outer segments, filling the rod periciliary ridge, and associated with Müller microvilli. By immunofluorescence, Ringer's wash removed most of the diffuse IRBP, but not that coating the outer segments. IRBP-Alexa 647 bound to the cone outer segments and Müller villi region, and comparably less to rod outer segments. Co-incubation with unlabeled IRBP markedly reduced this binding; ovalbumin-Alexa 647 and Alexa 647 dye alone showed no binding. Our data suggest that the pericellular matrix of the cone outer segments and Müller microvilli provide specialized domains that facilitate IRBP's functions.

Asunto(s)

Proteínas del Ojo/metabolismo , Neuroglía/metabolismo , Células Fotorreceptoras Retinianas Conos/metabolismo , Segmento Externo de las Células Fotorreceptoras Retinianas/metabolismo , Proteínas de Unión al Retinol/metabolismo , Animales , Carbocianinas/metabolismo , Opsinas de los Conos/metabolismo , Opsinas de los Conos/ultraestructura , Matriz Extracelular/metabolismo , Técnica del Anticuerpo Fluorescente Indirecta , Colorantes Fluorescentes/metabolismo , Inmunohistoquímica , Microscopía Electrónica , Microvellosidades/metabolismo , Neuroglía/ultraestructura , Técnicas de Cultivo de Órganos , Aglutinina de Mani/metabolismo , Células Fotorreceptoras Retinianas Conos/ultraestructura , Segmento Externo de las Células Fotorreceptoras Retinianas/ultraestructura , Aglutininas del Germen de Trigo/metabolismo , Xenopus laevis

RESUMEN

Eales' disease is an idiopathic retinal vasculitis of the eye. The disease is predominantly characterized by recurrent vitreous hemorrhage. Interphotoreceptor retinol-binding protein 3 plays a significant role in the etiopathogenesis of this condition. It transports retinoids between the retinal pigment epithelium and the photoreceptors; hence, this protein is a potential target for docking studies. In silico data reveal that herbal molecules interact with regulatory domains of interphotoreceptor retinol-binding protein 3 (IRBP-3), resulting into significant docking score and also forms H-bond and several hydrophobic interactions between active residues of IRBP-3. These interactions between the active residues may lead to significant conformational change in that particular portion of the protein. This efficacy and suitability of ligand was determined on the basis of binding energy calculations. Ginkgolide showed minimum binding energy calculations among selected 10 other natural ligands. This fact of virtual screening for potential ligand can give new insights toward the therapeutic intonations and alterations toward the advances in treatment for Eales' disease.