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1.
Invest Ophthalmol Vis Sci ; 62(7): 21, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34144609

RESUMO

Purpose: For this study we aimed to understand if retinal pigment epithelial (RPE) cells express antimicrobial peptide lysozyme as a mechanism to protect the neuroretina from blood-borne pathogens. Methods: The expression of lysozyme in human and mouse RPE cells was examined by RT-PCR or immune (cyto)histochemistry in cell cultures or retinal sections. RPE cultures were treated with different concentrations of Pam3CSK4, lipopolysaccharides (LPS), staphylococcus aureus-derived peptidoglycan (PGN-SA), Poly(I:C), and Poly(dA:dT). The mRNA expression of lysozyme was examined by qPCR and protein expression by ELISA. Poly(I:C) was injected into the subretinal space of C57BL/6J mice and eyes were collected 24 hours later and processed for the evaluation of lysozyme expression by confocal microscopy. Bactericidal activity was measured in ARPE19 cells following LYZ gene deletion using Crispr/Cas9 technology. Results: The mRNA and protein of lysozyme were detected in mouse and human RPE cells under normal conditions, although the expression levels were lower than mouse microglia BV2 or human monocytes THP-1 cells, respectively. Immunohistochemistry showed punctate lysozyme expression inside RPE cells. Lysozyme was detected by ELISA in normal RPE lysates, and in live bacteria-treated RPE supernatants. Treatment of RPE cells with Pam3CSK4, LPS, PGN-SA, and Poly(I:C) enhanced lysozyme expression. CRISPR/Cas9 deletion of lysozyme impaired bactericidal activity of ARPE19 cells and reduced their response to LPS and Poly(I:C) stimulation. Conclusions: RPE cells constitutively express antimicrobial peptide lysozyme and the expression is modulated by pathogenic challenges. RPE cells may protect the neuroretina from blood-borne pathogens by producing antimicrobial peptides, such as lysozyme.


Assuntos
Lipopeptídeos/fisiologia , Muramidase , Retina , Animais , Antibacterianos/metabolismo , Antibacterianos/farmacologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/metabolismo , Células Cultivadas , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Camundongos , Muramidase/genética , Muramidase/farmacologia , Poli I-C/metabolismo , Poli I-C/farmacologia , Fatores de Proteção , Retina/imunologia , Retina/metabolismo , Epitélio Pigmentado da Retina/fisiologia
2.
Front Immunol ; 12: 688254, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34093593

RESUMO

Several barriers separate the central nervous system (CNS) from the rest of the body. These barriers are essential for regulating the movement of fluid, ions, molecules, and immune cells into and out of the brain parenchyma. Each CNS barrier is unique and highly dynamic. Endothelial cells, epithelial cells, pericytes, astrocytes, and other cellular constituents each have intricate functions that are essential to sustain the brain's health. Along with damaging neurons, a traumatic brain injury (TBI) also directly insults the CNS barrier-forming cells. Disruption to the barriers first occurs by physical damage to the cells, called the primary injury. Subsequently, during the secondary injury cascade, a further array of molecular and biochemical changes occurs at the barriers. These changes are focused on rebuilding and remodeling, as well as movement of immune cells and waste into and out of the brain. Secondary injury cascades further damage the CNS barriers. Inflammation is central to healthy remodeling of CNS barriers. However, inflammation, as a secondary pathology, also plays a role in the chronic disruption of the barriers' functions after TBI. The goal of this paper is to review the different barriers of the brain, including (1) the blood-brain barrier, (2) the blood-cerebrospinal fluid barrier, (3) the meningeal barrier, (4) the blood-retina barrier, and (5) the brain-lesion border. We then detail the changes at these barriers due to both primary and secondary injury following TBI and indicate areas open for future research and discoveries. Finally, we describe the unique function of the pro-inflammatory cytokine interleukin-1 as a central actor in the inflammatory regulation of CNS barrier function and dysfunction after a TBI.


Assuntos
Barreira Hematoencefálica/metabolismo , Barreira Hematorretiniana/metabolismo , Lesões Encefálicas Traumáticas/metabolismo , Mediadores da Inflamação/metabolismo , Inflamação/metabolismo , Interleucina-1/metabolismo , Meninges/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/imunologia , Barreira Hematoencefálica/patologia , Barreira Hematorretiniana/efeitos dos fármacos , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/patologia , Lesões Encefálicas Traumáticas/tratamento farmacológico , Lesões Encefálicas Traumáticas/imunologia , Lesões Encefálicas Traumáticas/patologia , Humanos , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Mediadores da Inflamação/antagonistas & inibidores , Interleucina-1/antagonistas & inibidores , Meninges/efeitos dos fármacos , Meninges/imunologia , Meninges/patologia , Receptores Tipo I de Interleucina-1/metabolismo , Transdução de Sinais
3.
Prog Brain Res ; 256(1): 79-97, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32958216

RESUMO

Over the last decade, new evidence has become increasingly more compelling that commensal microflora profoundly influences the maturation and function of resident immune cells in host physiology. The concept of gut-retina axis is actively being explored. Studies have revealed a critical role of commensal microbes linked with neuronal stress, immune responses, and neurodegeneration in the retina. Microbial dysbiosis changes the blood-retina barrier permeability and modulates T cell-mediated autoimmunity to contribute to the pathogenesis of retinal diseases, such as glaucoma. Heat shock proteins (HSPs), which are evolutionarily conserved, are thought to function both as neuroprotectant and pathogenic antigens of T cells contributing to cell protection and tissue damage, respectively. Activated microglia recruit and interact with T cells during this process. Glaucoma, characterized by the progressive loss of retinal ganglion cells, is the leading cause of irreversible blindness. With nearly 70 million people suffering glaucoma worldwide, which doubles the number of patients with Alzheimer's disease, it represents the most frequent neurodegenerative disease of the central nervous system (CNS). Thus, understanding the mechanism of neurodegeneration in glaucoma and its association with the function of commensal microflora may help unveil the secrets of many neurodegenerative disorders in the CNS and develop novel therapeutic interventions.


Assuntos
Barreira Hematorretiniana , Microbioma Gastrointestinal , Glaucoma , Proteínas de Choque Térmico , Degeneração Neural , Retina , Linfócitos T , Animais , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/metabolismo , Barreira Hematorretiniana/microbiologia , Barreira Hematorretiniana/patologia , Glaucoma/imunologia , Glaucoma/metabolismo , Glaucoma/microbiologia , Glaucoma/patologia , Proteínas de Choque Térmico/imunologia , Proteínas de Choque Térmico/metabolismo , Humanos , Degeneração Neural/imunologia , Degeneração Neural/metabolismo , Degeneração Neural/microbiologia , Degeneração Neural/patologia , Retina/imunologia , Retina/metabolismo , Retina/microbiologia , Retina/patologia , Linfócitos T/imunologia , Linfócitos T/metabolismo
4.
Arterioscler Thromb Vasc Biol ; 38(11): 2691-2705, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30354230

RESUMO

Objective- Blood-CNS (central nervous system) barrier defects are implicated in retinopathies, neurodegenerative diseases, stroke, and epilepsy, yet, the pathological mechanisms downstream of barrier defects remain incompletely understood. Blood-retina barrier (BRB) formation and retinal angiogenesis require ß-catenin signaling induced by the ligand norrin (NDP [Norrie disease protein]), the receptor FZD4 (frizzled 4), coreceptor LRP5 (low-density lipoprotein receptor-like protein 5), and the tetraspanin TSPAN12 (tetraspanin 12). Impaired NDP/FZD4 signaling causes familial exudative vitreoretinopathy, which may lead to blindness. This study seeked to define cell type-specific functions of TSPAN12 in the retina. Approach and Results- A loxP-flanked Tspan12 allele was generated and recombined in endothelial cells using a tamoxifen-inducible Cdh5-CreERT2 driver. Resulting phenotypes were documented using confocal microscopy. RNA-Seq, histopathologic analysis, and electroretinogram were performed on retinas of aged mice. We show that TSPAN12 functions in endothelial cells to promote vascular morphogenesis and BRB formation in developing mice and BRB maintenance in adult mice. Early loss of TSPAN12 in endothelial cells causes lack of intraretinal capillaries and increased VE-cadherin (CDH5 [cadherin5 aka VE-cadherin]) expression, consistent with premature vascular quiescence. Late loss of TSPAN12 strongly impairs BRB maintenance without affecting vascular morphogenesis, pericyte coverage, or perfusion. Long-term BRB defects are associated with immunoglobulin extravasation, complement deposition, cystoid edema, and impaired b-wave in electroretinograms. RNA-sequencing reveals transcriptional responses to the perturbation of the BRB, including genes involved in vascular basement membrane alterations in diabetic retinopathy. Conclusions- This study establishes mice with late endothelial cell-specific loss of Tspan12 as a model to study pathological consequences of BRB impairment in an otherwise intact vasculature.


Assuntos
Barreira Hematorretiniana/metabolismo , Células Endoteliais/metabolismo , Neovascularização Retiniana , Vasos Retinianos/metabolismo , Tetraspaninas/deficiência , Fatores Etários , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Membrana Basal/metabolismo , Membrana Basal/patologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/patologia , Caderinas/genética , Caderinas/metabolismo , Proliferação de Células , Senescência Celular , Proteínas do Sistema Complemento/imunologia , Proteínas do Sistema Complemento/metabolismo , Retinopatia Diabética/genética , Retinopatia Diabética/metabolismo , Retinopatia Diabética/patologia , Células Endoteliais/imunologia , Células Endoteliais/patologia , Oftalmopatias Hereditárias/genética , Oftalmopatias Hereditárias/metabolismo , Oftalmopatias Hereditárias/patologia , Vitreorretinopatias Exsudativas Familiares , Feminino , Genótipo , Imunoglobulinas/imunologia , Imunoglobulinas/metabolismo , Edema Macular/genética , Edema Macular/metabolismo , Edema Macular/patologia , Masculino , Camundongos Knockout , Fenótipo , Doenças Retinianas/genética , Doenças Retinianas/metabolismo , Doenças Retinianas/patologia , Vasos Retinianos/imunologia , Vasos Retinianos/patologia , Transdução de Sinais , Tetraspaninas/genética
5.
Mol Vis ; 24: 379-394, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29853772

RESUMO

Purpose: The purpose of this study was to determine whether the blood-retina barrier is compromised by choroidal murine cytomegalovirus (MCMV) infection, using electron microscopy. Methods: BALB/c mice were immunosuppressed with methylprednisolone and monoclonal antibodies to CD4 and CD8. At several time points post-MCMV intraperitoneal inoculation, the eyes were removed and analyzed with western blotting and immunoelectron microscopy for the presence of MCMV early antigen (EA) and the host protein RIP3. Posterior eyecups from RIP3-/- and RIP3+/+ mice were cultured and inoculated with MCMV. At days 4, 7, and 11 post-infection, cultures were collected and analyzed with plaque assay, immunohistochemical staining, and real-time PCR (RT-PCR). Results: MCMV EA was observed in the nuclei of vascular endothelial cells and pericytes in the choriocapillaris. Disruption of Bruch's membrane was observed, especially at sites adjacent to activated platelets, and a few RPE cells containing some enlarged vesicles were found directly beneath disrupted Bruch's membrane. Some virus particles were also observed in the enlarged vesicles of RPE cells. Levels of the RIP3 protein, which was observed mainly in the RPE cells and the basement membrane of the choriocapillaris, were greatly increased following MCMV infection, while depletion of RIP3 resulted in greatly decreased inflammasome formation, as well as expression of downstream inflammation factors. Conclusions: The results suggest that systemic MCMV spreads to the choroid and replicates in vascular endothelia and pericytes of the choriocapillaris during immunosuppression. Choroidal MCMV infection is associated with in situ inflammation and subsequent disruption of Bruch's membrane and the outer blood-retina barrier.


Assuntos
Corioide/imunologia , Infecções por Citomegalovirus/imunologia , Infecções Oculares Virais/imunologia , Hospedeiro Imunocomprometido , Retina/imunologia , Retinite/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Antígenos Virais/genética , Plaquetas/imunologia , Plaquetas/patologia , Plaquetas/virologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/patologia , Barreira Hematorretiniana/virologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Corioide/irrigação sanguínea , Corioide/patologia , Corioide/virologia , Infecções por Citomegalovirus/patologia , Infecções por Citomegalovirus/virologia , Células Endoteliais , Infecções Oculares Virais/patologia , Infecções Oculares Virais/virologia , Feminino , Proteínas Imediatamente Precoces/genética , Inflamassomos/imunologia , Metilprednisolona/administração & dosagem , Camundongos , Camundongos Endogâmicos BALB C , Muromegalovirus/crescimento & desenvolvimento , Muromegalovirus/patogenicidade , Pericitos/imunologia , Pericitos/patologia , Pericitos/virologia , Proteína Serina-Treonina Quinases de Interação com Receptores/deficiência , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Retina/patologia , Retina/virologia , Epitélio Pigmentado da Retina/imunologia , Epitélio Pigmentado da Retina/patologia , Epitélio Pigmentado da Retina/virologia , Retinite/patologia , Retinite/virologia
6.
Nat Rev Immunol ; 17(12): 761-773, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28869253

RESUMO

Immune privilege is a complex process that protects organs from immune-mediated attack and damage. It is accomplished by a series of cellular barriers that both control immune cell entry and promote the development of tolerogenic immune cells. In this Review, we describe the vascular endothelial and epithelial barriers in organs that are commonly considered to be immune privileged, such as the brain and the eye. We compare these classical barriers with barriers in the intestine, which share features with barriers of immune-privileged organs, such as the capacity to induce tolerance and to protect from external insults. We suggest that when intestinal barriers break down, disruption of other barriers at distant sites can ensue, and this may underlie the development of various neurological, metabolic and intestinal disorders.


Assuntos
Endotélio Vascular/imunologia , Endotélio Vascular/metabolismo , Epitélio/imunologia , Epitélio/metabolismo , Homeostase , Mucosa/imunologia , Mucosa/metabolismo , Animais , Barreira Hematoencefálica/citologia , Barreira Hematoencefálica/imunologia , Barreira Hematoencefálica/metabolismo , Barreira Hematorretiniana/citologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/metabolismo , Suscetibilidade a Doenças , Endotélio Vascular/citologia , Microbioma Gastrointestinal/imunologia , Humanos , Privilégio Imunológico , Imunidade , Imunomodulação , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa/citologia , Especificidade de Órgãos/imunologia
7.
PLoS One ; 12(4): e0175159, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28406972

RESUMO

Ocular toxoplasmosis is mediated by monocytes infected with Toxoplasma gondii that are disseminated to target organs. Although infected monocytes can easily access to outer blood-retinal barrier due to leaky choroidal vasculatures, not much is known about the effect of T. gondii-infected monocytes on outer blood-retinal barrier. We prepared human monocytes, THP-1, infected with T. gondii and human retinal pigment epithelial cells, ARPE-19, grown on transwells as an in vitro model of outer blood-retinal barrier. Exposure to infected monocytes resulted in disruption of tight junction protein, ZO-1, and decrease in transepithelial electrical resistance of retinal pigment epithelium. Supernatants alone separated from infected monocytes also decreased transepithelial electrical resistance and disrupted tight junction protein. Further investigation revealed that the supernatants could activate focal adhesion kinase (FAK) signaling in retinal pigment epithelium and the disruption was attenuated by FAK inhibitor. The disrupted barrier was partly restored by blocking CXCL8, a FAK activating factor secreted by infected monocytes. In this study, we demonstrated that monocytes infected with T. gondii can disrupt outer blood-retinal barrier, which is mediated by paracrinely activated FAK signaling. FAK signaling can be a target of therapeutic approach to prevent negative influence of infected monocytes on outer blood-retinal barrier.


Assuntos
Barreira Hematorretiniana , Quinase 1 de Adesão Focal/imunologia , Monócitos , Comunicação Parácrina/imunologia , Transdução de Sinais/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/parasitologia , Linhagem Celular , Humanos , Interleucina-8/imunologia , Monócitos/imunologia , Monócitos/parasitologia , Epitélio Pigmentado da Retina/imunologia , Epitélio Pigmentado da Retina/parasitologia , Proteína da Zônula de Oclusão-1/imunologia
8.
J Neuroinflammation ; 14(1): 43, 2017 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-28253931

RESUMO

BACKGROUND: Ocular abnormalities present in microcephalic infants with presumed Zika virus (ZIKV) congenital disease includes focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, and lens dislocation. Target cells in the ocular compartment for ZIKV infectivity are unknown. The cellular response of ocular cells to ZIKV infection has not been described. Mechanisms for viral dissemination in the ocular compartment of ZIKV-infected infants and adults have not been reported. Here, we identify target cells for ZIKV infectivity in both the inner and outer blood-retinal barriers (IBRB and OBRB), describe the cytokine expression profile in the IBRB after ZIKV exposure, and propose a mechanism for viral dissemination in the retina. METHODS: We expose primary cellular components of the IBRB including human retinal microvascular endothelial cells, retinal pericytes, and Müller cells as well as retinal pigmented epithelial cells of the OBRB to the PRVABC56 strain of ZIKV. Viral infectivity was analyzed by microscopy, immunofluorescence, and reverse transcription polymerase chain reaction (RT-PCR and qRT-PCR). Angiogenic and proinflammatory cytokines were measured by Luminex assays. RESULTS: We find by immunofluorescent staining using the Flavivirus 4G2 monoclonal antibody that retinal endothelial cells and pericytes of the IBRB and retinal pigmented epithelial cells of the OBRB are fully permissive for ZIKV infection but not Müller cells when compared to mock-infected controls. We confirmed ZIKV infectivity in retinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells by RT-PCR and qRT-PCR using ZIKV-specific oligonucleotide primers. Expression profiles by Luminex assays in retinal endothelial cells infected with ZIKV revealed a marginal increase in levels of beta-2 microglobulin (ß2-m), granulocyte macrophage colony-stimulating factor (GMCSF), intercellular adhesion molecule 1 (ICAM-1), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP1), and vascular cell adhesion molecule 1 (VCAM-1) and higher levels of regulated upon activation, normal T cell expressed and presumably secreted (RANTES) but lower levels of interleukin-4 (IL-4) compared to controls. CONCLUSIONS: Retinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells are fully permissive for ZIKV lytic replication and are primary target cells in the retinal barriers for infection. ZIKV infection of retinal endothelial cells and retinal pericytes induces significantly higher levels of RANTES that likely contributes to ocular inflammation.


Assuntos
Barreira Hematorretiniana/patologia , Células Ependimogliais/patologia , Oftalmopatias/patologia , Infecção por Zika virus/patologia , Zika virus , Adulto , Animais , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/virologia , Células Cultivadas , Chlorocebus aethiops , Células Ependimogliais/imunologia , Células Ependimogliais/virologia , Oftalmopatias/imunologia , Oftalmopatias/virologia , Humanos , Células Vero , Zika virus/imunologia , Zika virus/metabolismo , Infecção por Zika virus/imunologia
9.
Immunity ; 43(2): 343-53, 2015 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-26287682

RESUMO

Activated retina-specific T cells that have acquired the ability to break through the blood-retinal barrier are thought to be causally involved in autoimmune uveitis, a major cause of human blindness. It is unclear where these autoreactive T cells first become activated, given that their cognate antigens are sequestered within the immune-privileged eye. We demonstrate in a novel mouse model of spontaneous uveitis that activation of retina-specific T cells is dependent on gut commensal microbiota. Retina-specific T cell activation involved signaling through the autoreactive T cell receptor (TCR) in response to non-cognate antigen in the intestine and was independent of the endogenous retinal autoantigen. Our findings not only have implications for the etiology of human uveitis, but also raise the possibility that activation of autoreactive TCRs by commensal microbes might be a more common trigger of autoimmune diseases than is currently appreciated.


Assuntos
Intestinos/imunologia , Microbiota/imunologia , Retina/imunologia , Linfócitos T/imunologia , Uveíte/imunologia , Animais , Antígenos de Bactérias/administração & dosagem , Autoantígenos/imunologia , Autoimunidade , Barreira Hematorretiniana/imunologia , Células Cultivadas , Modelos Animais de Doenças , Proteínas do Olho/genética , Proteínas do Olho/imunologia , Proteínas do Olho/metabolismo , Tolerância Imunológica , Intestinos/microbiologia , Ativação Linfocitária , Camundongos Endogâmicos , Camundongos Knockout , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas de Ligação ao Retinol/genética , Proteínas de Ligação ao Retinol/imunologia , Proteínas de Ligação ao Retinol/metabolismo , Uveíte/microbiologia
10.
Neuroscience ; 284: 536-545, 2015 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-25453774

RESUMO

The blood retinal barrier (BRB) can mitigate deleterious immune response. Dysfunction at the BRB can affect disease progression. Under inflammatory conditions Müller glia produce increased pro-inflammatory factors, like nitric oxide (NO). In this study we describe molecular events at the Müller glia during inflammation which could affect inner BRB properties. Griess assay and 4,5-diaminofluorescein diacetate (DAF-2DA) time-lapse fluorescence were used to measure NO production. Western blot was used to analyze the expression of inducible nitric oxide synthase (iNOS) and mitogen-activated protein kinases (MAPK) components. Lucifer Yellow was used to measure permeability. Griess assay and DAF-2DA time-lapse fluorescence images revealed that lipopolysaccharide (LPS) induced inflammation and increased NO production. In parallel, changes were observed in tight junction proteins, zona occludens 1 (ZO-1), connexin 43 (Cx43), and permeability. This was mediated through activation of iNOS and mitogen-activated protein kinase phosphatase-1 (MKP-1), implicated in immune response. Endocannabinoids can exert a protective and anti-inflammatory effect. Exogenous arachidonoyl ethanolamide (AEA) inhibited NO generation and also abolished LPS-induced increase in permeability. Our work suggests that subtle changes in Müller glia function, which act as part of the BRB, could contribute to retinal health. AEA which can reduce inflammatory cytotoxicity has potential as treatment in several ocular manifestations where the integrity of the BRB is crucial.


Assuntos
Ácidos Araquidônicos/farmacologia , Barreira Hematorretiniana/efeitos dos fármacos , Endocanabinoides/farmacologia , Neuroglia/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Óxido Nítrico/metabolismo , Alcamidas Poli-Insaturadas/farmacologia , Animais , Barreira Hematorretiniana/imunologia , Bovinos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Relação Dose-Resposta a Droga , Lipopolissacarídeos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , NF-kappa B/metabolismo , Neuroglia/imunologia , Óxido Nítrico Sintase Tipo II/metabolismo , Retina/efeitos dos fármacos , Retina/imunologia
11.
PLoS One ; 9(10): e108508, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25329075

RESUMO

Inflammation in the diabetic retina is mediated by leukocyte adhesion to the retinal vasculature and alteration of the blood-retinal barrier (BRB). We investigated the role of chemokines in the alteration of the BRB in diabetes. Animals were made diabetic by streptozotocin injection and analyzed for gene expression and monocyte/macrophage infiltration. The expression of CCL2 (chemokine ligand 2) was significantly up-regulated in the retinas of rats with 4 and 8 weeks of diabetes and also in human retinal endothelial cells treated with high glucose and glucose flux. Additionally, diabetes or intraocular injection of recombinant CCL2 resulted in increased expression of the macrophage marker, F4/80. Cell culture impedance sensing studies showed that purified CCL2 was unable to alter the integrity of the human retinal endothelial cell barrier, whereas monocyte conditioned medium resulted in significant reduction in cell resistance, suggesting the relevance of CCL2 in early immune cell recruitment for subsequent barrier alterations. Further, using Cx3cr1-GFP mice, we found that intraocular injection of CCL2 increased retinal GFP+ monocyte/macrophage infiltration. When these mice were made diabetic, increased infiltration of monocytes/macrophages was also present in retinal tissues. Diabetes and CCL2 injection also induced activation of retinal microglia in these animals. Quantification by flow cytometry demonstrated a two-fold increase of CX3CR1+/CD11b+ (monocyte/macrophage and microglia) cells in retinas of wildtype diabetic animals in comparison to control non-diabetic ones. Using CCL2 knockout (Ccl2-/-) mice, we show a significant reduction in retinal vascular leakage and monocyte infiltration following induction of diabetes indicating the importance of this chemokine in alteration of the BRB. Thus, CCL2 may be an important therapeutic target for the treatment of diabetic macular edema.


Assuntos
Barreira Hematorretiniana/citologia , Movimento Celular , Quimiocina CCL2/metabolismo , Retinopatia Diabética/metabolismo , Monócitos/citologia , Animais , Barreira Hematorretiniana/efeitos dos fármacos , Barreira Hematorretiniana/imunologia , Permeabilidade Capilar/efeitos dos fármacos , Contagem de Células , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Quimiocina CCL2/deficiência , Quimiocina CCL2/genética , Retinopatia Diabética/genética , Retinopatia Diabética/imunologia , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Técnicas de Inativação de Genes , Glucose/farmacologia , Humanos , Inflamação/metabolismo , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Ratos , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacos
12.
Biochem Pharmacol ; 86(11): 1603-13, 2013 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-24076420

RESUMO

Diabetic retinopathy is one of the leading causes of blindness and the most common complication of diabetes with no cure available. We investigated the role of phospholipases A2 (PLA2) in diabetic retinopathy using an in vitro blood-retinal barrier model (BRB) and an in vivo streptozotocin (STZ)-induced diabetic model. Mono- and co-cultures of endothelial cells (EC) and pericytes (PC), treated with high or fluctuating concentrations of glucose, to mimic the diabetic condition, were used. PLA2 activity, VEGF and PGE2 levels and cell proliferation were measured, with or without PLA2 inhibition. Diabetes was induced in rats by STZ injection and PLA2 activity along with VEGF, TNFα and ICAM-1 levels were measured in retina. High or fluctuating glucose induced BRB breakdown, and increased PLA2 activity, PGE2 and VEGF in EC/PC co-cultures; inhibition of PLA2 in mono- or co-cultures treated with high or fluctuating glucose dampened PGE2 and VEGF production down to the levels of controls. High or fluctuating glucose increased EC number and reduced PC number in co-cultures; these effects were reversed after transfecting EC with small interfering RNA targeted to PLA2. PLA2 and COX-2 protein expressions were significantly increased in microvessels from retina of diabetic rats. Diabetic rats had also high retinal levels of VEGF, ICAM-1 and TNFα that were reduced by treatment with a cPLA2 inhibitor. In conclusion, the present findings indicate that PLA2 upregulation represents an early step in glucose-induced alteration of BRB, possibly upstream of VEGF; thus, PLA2 may be an interesting target in managing diabetic retinopathy.


Assuntos
Barreira Hematorretiniana/enzimologia , Diabetes Mellitus Experimental/enzimologia , Retinopatia Diabética/enzimologia , Fosfolipases A2/metabolismo , Animais , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/patologia , Bovinos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/imunologia , Diabetes Mellitus Experimental/patologia , Retinopatia Diabética/etiologia , Retinopatia Diabética/imunologia , Retinopatia Diabética/patologia , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/enzimologia , Glucose/farmacologia , Masculino , Microscopia Confocal , Pericitos/citologia , Pericitos/efeitos dos fármacos , Pericitos/enzimologia , Inibidores de Fosfolipase A2/farmacologia , Fosfolipases A2/biossíntese , Ratos , Ratos Sprague-Dawley , Estreptozocina/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo
13.
Nat Rev Immunol ; 13(3): 206-18, 2013 03.
Artigo em Inglês | MEDLINE | ID: mdl-23435332

RESUMO

Complex barriers separate immune-privileged tissues from the circulation. Here, we propose that cell entry to immune-privileged sites through barriers composed of tight junction-interconnected endothelium is associated with destructive inflammation, whereas border structures comprised of fenestrated vasculature enveloped by tightly regulated epithelium serve as active and selective immune-skewing gates in the steady state. Based on emerging knowledge of the central nervous system and information from other immune-privileged sites, we propose that these sites are endowed either with absolute endothelial-based barriers and epithelial gates that enable selective and educative transfer of trafficking leukocytes or with selective epithelial gates only.


Assuntos
Quimiotaxia de Leucócito , Vigilância Imunológica/fisiologia , Modelos Imunológicos , Junções Íntimas/fisiologia , Animais , Barreira Hematoaquosa/imunologia , Barreira Hematoaquosa/fisiologia , Barreira Hematoencefálica/imunologia , Barreira Hematoencefálica/fisiologia , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/fisiologia , Barreira Hematotesticular/imunologia , Barreira Hematotesticular/fisiologia , Fusão Celular , Quimerismo , Células Epiteliais/fisiologia , Células Epiteliais/ultraestrutura , Epitélio/imunologia , Epitélio/fisiologia , Feminino , Humanos , Tolerância Imunológica/imunologia , Vigilância Imunológica/imunologia , Inflamação/imunologia , Inflamação/fisiopatologia , Masculino , Troca Materno-Fetal/imunologia , Infiltração de Neutrófilos , Especificidade de Órgãos , Gravidez , Migração Transendotelial e Transepitelial/fisiologia
14.
Inflamm Res ; 61(7): 759-73, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22487851

RESUMO

OBJECTIVE AND DESIGN: A mathematical analysis of leukocytes accumulating in experimental autoimmune uveitis (EAU), using ordinary differential equations (ODEs) and incorporating a barrier to cell traffic. MATERIALS AND SUBJECTS: Data from an analysis of the kinetics of cell accumulation within the eye during EAU. METHODS: We applied a well-established mathematical approach that uses ODEs to describe the behaviour of cells on both sides of the blood-retinal barrier and compared data from the mathematical model with experimental data from animals with EAU. RESULTS: The presence of the barrier is critical to the ability of the model to qualitatively reproduce the experimental data. However, barrier breakdown is not sufficient to produce a surge of cells into the eye, which depends also on asymmetry in the rates at which cells can penetrate the barrier. Antigen-presenting cell (APC) generation also plays a critical role and we can derive from the model the ratio for APC production under inflammatory conditions relative to production in the resting state, which has a value that agrees closely with that found by experiment. CONCLUSIONS: Asymmetric trafficking and the dynamics of APC production play an important role in the dynamics of cell accumulation in EAU.


Assuntos
Doenças Autoimunes/imunologia , Barreira Hematorretiniana/imunologia , Leucócitos/imunologia , Modelos Biológicos , Uveíte/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Camundongos
15.
Biochem Pharmacol ; 84(1): 88-92, 2012 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-22484312

RESUMO

Diabetic retinopathy is a complex disease that has potential involvement of inflammatory and oxidative stress-related pathways in its pathogenesis. We hypothesized that eriodictyol, one of the most abundant dietary flavonoids, could be effective against diabetic retinopathy, which involves significant oxidative stress and inflammation. The aim of the present study was to investigate the effects of eriodictyol in early retinal and plasma changes of streptozotocin-induced diabetic rats. The effect of eriodictyol treatment (0.1, 1, 10 mg/kg daily for 10 days) was evaluated by TNF-α, ICAM-1, VEGF, and eNOS protein levels measurement in the retina, plasma lipid peroxidation, and blood-retinal barrier (BRB) integrity. Increased amounts of cytokines, adhesion molecule, and nitric oxide synthase were observed in retina from diabetic rats. Eriodictyol treatment significantly lowered retinal TNF-α, ICAM-1, VEGF, and eNOS in a dose-dependent manner. Further, treatment with eriodictyol significantly suppressed diabetes-related lipid peroxidation, as well as the BRB breakdown. These data demonstrated that eriodictyol attenuates the degree of retinal inflammation and plasma lipid peroxidation preserving the BRB in early diabetic rats.


Assuntos
Antioxidantes/uso terapêutico , Barreira Hematorretiniana/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Retinopatia Diabética/prevenção & controle , Flavanonas/uso terapêutico , Peróxidos Lipídicos/sangue , Animais , Antioxidantes/administração & dosagem , Antioxidantes/química , Barreira Hematorretiniana/imunologia , Barreira Hematorretiniana/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/imunologia , Retinopatia Diabética/sangue , Retinopatia Diabética/etiologia , Retinopatia Diabética/imunologia , Relação Dose-Resposta a Droga , Flavanonas/administração & dosagem , Flavanonas/química , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Estrutura Molecular , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Retina/efeitos dos fármacos , Retina/imunologia , Retina/metabolismo
16.
Am J Hematol ; 85(8): 631-3, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20658596

Assuntos
Neoplasias Encefálicas/patologia , Linfoma Difuso de Grandes Células B/patologia , Neoplasias da Retina/patologia , Neoplasias Testiculares/patologia , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Murinos , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Barreira Hematoencefálica/imunologia , Barreira Hematorretiniana/imunologia , Barreira Hematotesticular/imunologia , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/imunologia , Terapia Combinada , Ciclofosfamida/administração & dosagem , Citarabina/administração & dosagem , Dexametasona/administração & dosagem , Doxorrubicina/administração & dosagem , Humanos , Injeções Intraoculares , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/imunologia , Linfoma Difuso de Grandes Células B/radioterapia , Linfoma Difuso de Grandes Células B/cirurgia , Masculino , Metotrexato/administração & dosagem , Pessoa de Meia-Idade , Orquiectomia , Prednisona/administração & dosagem , Procarbazina/administração & dosagem , Indução de Remissão , Neoplasias da Retina/tratamento farmacológico , Neoplasias da Retina/imunologia , Neoplasias da Retina/cirurgia , Rituximab , Neoplasias Testiculares/tratamento farmacológico , Neoplasias Testiculares/imunologia , Neoplasias Testiculares/radioterapia , Neoplasias Testiculares/cirurgia , Vincristina/administração & dosagem , Vitrectomia
17.
MAbs ; 2(2): 176-80, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-21358858

RESUMO

More than a century after the first successful use of serotherapy, antibody-based therapy has been renewed by the availability of recombinant monoclonal antibodies. As in the past, current clinical experience has prompted new pharmacological questions and induced much debate among practitioners, notably in the field of ophthalmology. An examination of the history of antibodies as treatments for ocular disorders reveals interesting parallels to the modern era. The fact that a treatment administered by a systemic route could be efficacious in a local disease was not widely accepted and the "chemical" nature of antibodies was not clearly understood in the late 19(th) century. Clinical studies by Henry Coppez, a Belgian ophthalmologist, established in 1894 that antidiphtheric antitoxins could be used to treat conjunctival diphtheria. Nearly 20 years later, Coppez and Danis described age-related macular degeneration, a disorder which today benefits from ranibizumab therapy. The product, a locally-administered recombinant monoclonal Fab fragment, is directed against vascular endothelial growth factor A. Interestingly, its full-size counterpart, bevacizumab, which is approved for the treatment of solid tumors, has also demonstrated efficacy in age-related macular degeneration when administered either intravenously or locally, which raises new questions about antibody pharmacology and biodistribution.In order to shed some light on this debate, we recount the early history of serotherapy applied to ophthalmology, review the exact molecular differences between ranibizumab and bevacizumab, and discuss what is known about IgG and the blood-retina barrier and the possible role of FcRn, an IgG transporter.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Imunoterapia , Degeneração Macular/tratamento farmacológico , Oftalmologia , Proteínas Recombinantes/uso terapêutico , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais Humanizados , Bevacizumab , Barreira Hematorretiniana/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Degeneração Macular/imunologia , Oftalmologia/tendências , Ranibizumab , Receptores Fc/imunologia , Proteínas Recombinantes/farmacologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores
18.
Vestn Oftalmol ; 125(5): 43-5, 2009.
Artigo em Russo | MEDLINE | ID: mdl-19916335

RESUMO

IgM, IgG, IgA, and IgE in the subretinal fluid (SRF) of the subpopulational composition of peripheral blood T lymphocytes and the parameters of a leukocyte migration inhibition test (LMIT) were comparatively studied in patients with posttraumatic and regmatogenic retinal detachments (RD) complicated by subretinal proliferation (SP). SRF and blood samples taken from 20 patients with SP of various etiologies were studied. SRF immunoglobulins of classes M, G, A, and E were revealed in all the examinees, their higher levels being noted in patients with long-standing (more than 4 months) and severe recurrent RD. Comparative examinations of patients with regmatogenic and posttraumatic RDs revealed a significantly higher content of T lymphocytes (CD3+), T killers (CD8+), and T helpers (CD4+) in the peripheral blood of patients with regmatogenic RD. The patients of both groups were found to have increased LMIT values with phytohemagglutinin, Con-A and retinal antigen, in Group 2, leukocyte migration inhibition with retinal antigens being much more pronounced than that in Group 1.


Assuntos
Barreira Hematorretiniana/imunologia , Imunidade Inata/imunologia , Imunoglobulinas/metabolismo , Linfócitos T/imunologia , Vitreorretinopatia Proliferativa/imunologia , Adolescente , Adulto , Idoso , Humor Aquoso/imunologia , Humor Aquoso/metabolismo , Relação CD4-CD8 , Humanos , Pessoa de Meia-Idade , Adulto Jovem
19.
J Autoimmun ; 33(3-4): 247-54, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19846275

RESUMO

Autoantibody production is associated with a variety of ocular disorders, including autoimmune retinopathy (AIR) and age-related macular degeneration (AMD). A breakdown of immunologic tolerance (ocular immune privilege), including the blood-retinal barrier, anti-immune and anti-inflammatory proteins, and anterior chamber-associated immune deviation may play important roles in these disorders. Although the exact triggers for ocular autoimmunity are unknown, autoimmune targeting of retinal tissue is clearly associated with and may contribute to the pathogenesis of both AIR and AMD. Autoantibody production has long been associated with AIR, a collection of disorders that includes cancer-associated retinopathy, melanoma-associated retinopathy and non-paraneoplastic autoimmune retinopathy. A growing body of evidence indicates that AMD pathogenesis, too, involves ocular inflammation and autoimmunity. Identification and quantification of autoantibodies produced in patients with AIR and AMD may assist with diagnosis, prognosis, and choice of treatments. Animal models that allow investigation of ocular autoimmunity will also be needed to better understand the disease processes and to develop novel therapies. In this review we discuss ocular immune privilege and potential mechanisms of autoimmunity in the eye. We describe how autoimmunity relates to the pathogenesis of AIR and AMD. We explain how the antigen microarray technique is used to detect autoantibodies in patient serum samples, and discuss how current animal models for AMD can be used to investigate autoimmune pathogenesis. Finally, we outline unanswered questions and exciting areas of future study related to autoimmune retinal degeneration.


Assuntos
Autoimunidade , Barreira Hematorretiniana/imunologia , Degeneração Macular/imunologia , Doenças Retinianas/imunologia , Animais , Autoanticorpos/sangue , Barreira Hematorretiniana/metabolismo , Modelos Animais de Doenças , Humanos , Tolerância Imunológica/imunologia , Análise Serial de Proteínas , Retina/imunologia , Retina/patologia
20.
Artigo em Inglês | MEDLINE | ID: mdl-19619127

RESUMO

Leukocyte-endothelial interaction plays an important role in the early phase of the development of diabetic retinopathy. It has been studied extensively linking inflammatory processes to its development conducted to date in rats and mice, and have focused on insulin-deficient models. The molecular and functional changes that are characteristics of inflammation have been detected in retinas from diabetic animals and humans with involvement of multiple pathways that results in the final sequelae of increased permeability of the blood retinal barrier and finally ischemia that drives angiogenesis. Increased expression of Intracellular adhesion molecules heralds the onset of changes that results in attraction of leucocytes such as neutrophils. The consequent release of cytokines and growth factors such as vascular endothelial growth factor (VEGF), tumor necrosis factor-alpha, and interleukin 1-Beta results in increased permeability and retinal edema. Other indirect mediators involved include pathways such as the protein kinase C (PKC), renin-angiotensin system, enzymes such as the poly ADP-ribose polymerase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, nitric oxide synthetase and finally advanced glycation products. Therapy for early diabetic retinopathy may inhibit one or more of these pathways using drugs that can be given systemically, with local ocular applications having a more direct effect as in other eye diseases.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Leucostasia/imunologia , Inibidores da Angiogênese/administração & dosagem , Inibidores da Angiogênese/imunologia , Animais , Barreira Hematorretiniana/efeitos dos fármacos , Barreira Hematorretiniana/imunologia , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/etiologia , Retinopatia Diabética/imunologia , Sistemas de Liberação de Medicamentos/tendências , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/imunologia , Humanos , Leucostasia/complicações , Leucostasia/tratamento farmacológico , Fatores de Tempo , Fator A de Crescimento do Endotélio Vascular/imunologia
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