RESUMO
Choroidal neovascularization (CNV) is a defining feature of wet age-related macular degeneration. We examined the functional role of CCR3 in the development of CNV in mice and primates. CCR3 was associated with spontaneous CNV lesions in the newly described JR5558 mice, whereas CCR3 ligands localized to CNV-associated macrophages and the retinal pigment epithelium/choroid complex. Intravitreal injection of neutralizing antibodies against vascular endothelial growth factor receptor 2, CCR3, CC chemokine ligand 11/eotaxin-1, and CC chemokine ligand 24/eotaxin-2 all reduced CNV area and lesion number in these mice. Systemic administration of the CCR3 antagonists GW766994X and GW782415X reduced spontaneous CNV in JR5558 mice and laser-induced CNV in mouse and primate models in a dose-dependent fashion. Combination treatment with antivascular endothelial growth factor receptor 2 antibody and GW766994X yielded additive reductions in CNV area and hyperpermeability in mice. Interestingly, topical GW766994X and intravitreal anti-CCR3 antibody yielded strong systemic effects, reducing CNV in the untreated, contralateral eye. Contrarily, ocular administration of GW782415X in primates failed to substantially elevate plasma drug levels or to reduce the development of grade IV CNV lesions. These findings suggest that CCR3 signaling may be an attractive therapeutic target for CNV, utilizing a pathway that is at least partly distinct from that of vascular endothelial growth factor receptor. The findings also demonstrate that systemic exposure to CCR3 antagonists may be crucial for CNV-targeted activity.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Permeabilidade Capilar/efeitos dos fármacos , Neovascularização de Coroide/tratamento farmacológico , Receptores CCR3/antagonistas & inibidores , Degeneração Macular Exsudativa/tratamento farmacológico , Animais , Permeabilidade Capilar/imunologia , Corioide/patologia , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos C57BL , Fator A de Crescimento do Endotélio Vascular/metabolismo , Degeneração Macular Exsudativa/patologiaRESUMO
PURPOSE: We evaluated whether a functional visual acuity (FVA) system can detect subtle changes in central visual acuity that reflect pathological findings associated with age-related macular degeneration (AMD). METHODS: Twenty-eight patients with unilateral AMD and logMAR monocular best corrected VA better than 0 in both eyes, as measured by conventional chart examination, were analyzed between November 2012 and April 2013. After measuring conventional VA, FVA, and contrast VA with best correction, routine eye examinations including spectral domain-optical coherence tomography were performed. Standard Schirmer test was performed, and corneal and lens densities were measured. RESULTS: The FVA score (p < 0.001) and visual maintenance ratio (p < 0.001) measured by the FVA system, contrast VA (p < 0. 01), and conventional VA (p < 0.01) were significantly worse in the AMD-affected eyes than in the fellow eyes. No significant differences were observed in the anterior segment conditions. Forward stepwise regression analysis demonstrated that the length of interdigitation zone disruption, as visualized by optical coherence tomography imaging, correlated with the FVA score (p < 0.01) but not with any other parameters investigated. CONCLUSIONS: The FVA system detects subtle changes in best corrected VA in AMD-affected eyes and reflects interdigitation zone disruption, an anatomical change in the retina recorded by optical coherence tomography. Further studies are required to understand the value of the FVA system in detecting subtle changes in AMD.
Assuntos
Degeneração Macular/fisiopatologia , Acuidade Visual/fisiologia , Idoso , Idoso de 80 Anos ou mais , Feminino , Angiofluoresceinografia , Humanos , Degeneração Macular/diagnóstico , Masculino , Pessoa de Meia-Idade , Retina/fisiopatologia , Tomografia de Coerência Óptica/métodosRESUMO
PURPOSE: To analyze the association between macular pigment optical density (MPOD), which reflects lutein (L), zeaxanthin (Z), and meso-zeaxanthin (MZ) in the macula, and background characteristics. METHODS: Fifty-five healthy adult volunteers were analyzed. Macular pigment optical density was measured using a heterochromatic flicker photometry technique, and serum concentrations of carotenoids and lipoproteins were by high-performance liquid chromatography and enzyme-linked immunosorbent assay, respectively. Dietary intake of nutrient was determined by a validated self-administered questionnaire on ingestion frequency. RESULTS: Macular pigment optical density was positively correlated with serum concentrations of L and Z and dietary L intake and inversely correlated with serum oxidized low-density lipoprotein (LDL). Although MPOD decreased with age (95% confidence interval, -0.011 to -0.002; correlation coefficient, -0.269; P = 0.007), serum L/Z and dietary L intake did not. In contrast, serum oxidized LDL was positively correlated with age (95% confidence interval, 0.69-2.34; correlation coefficient, 0.333; P = 0.0004). After adjusting for age, sex, and oxidized LDL, serum L was positively correlated with MPOD (95% confidence interval, 0.88-1.69; P = 0.000001). After adjusting for age, sex, and serum L, serum oxidized LDL was inversely correlated with MPOD (95% confidence interval, -0.002 to -0.0004; P = 0.006). CONCLUSION: Macular pigment optical density was inversely correlated with serum oxidized LDL. Further study to know the impact of oxidized LDL on MPOD may be warranted.
Assuntos
Lipoproteínas LDL/sangue , Luteína/análise , Macula Lutea/química , Pigmento Macular/análise , Zeaxantinas/análise , Adulto , Cromatografia Líquida de Alta Pressão , Densitometria , Ensaio de Imunoadsorção Enzimática , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Fotometria/métodos , Inquéritos e Questionários , Adulto JovemRESUMO
PURPOSE: Astaxanthin (AST) is a carotenoid found in marine animals and vegetables. The purpose of the present study was to investigate the effect of AST on the development of experimental choroidal neovascularization (CNV) with underlying cellular and molecular mechanisms. METHODS: Laser photocoagulation was used to induce CNV in C57BL/6J mice. Mice were pretreated with intraperitoneal injections of AST daily for 3 days before photocoagulation, and treatments were continued daily until the end of the study. CNV response was analyzed by volumetric measurements 1 week after laser injury. Retinal pigment epithelium-choroid levels of IkappaB-alpha, intercellular adhesion molecule (ICAM)-1, monocyte chemotactic protein (MCP)-1, interleukin (IL)-6, vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR)-1, and VEGFR-2 were examined by Western blotting or ELISA. AST was applied to capillary endothelial (b-End3) cells, macrophages, and RPE cells to analyze the activation of NF-kappaB and the expression of inflammatory molecules. RESULTS: The index of CNV volume was significantly suppressed by treatment with AST compared with that in vehicle-treated animals. AST treatment led to significant inhibition of macrophage infiltration into CNV and of the in vivo and in vitro expression of inflammation-related molecules, including VEGF, IL-6, ICAM-1, MCP-1, VEGFR-1, and VEGFR-2. Importantly, AST suppressed the activation of the NF-kappaB pathway, including IkappaB-alpha degradation and p65 nuclear translocation. CONCLUSIONS: AST treatment, together with inflammatory processes including NF-kappaB activation, subsequent upregulation of inflammatory molecules, and macrophage infiltration, led to significant suppression of CNV development. The present study suggests the possibility of AST supplementation as a therapeutic strategy to suppress CNV associated with AMD.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Anti-Inflamatórios não Esteroides/uso terapêutico , Neovascularização de Coroide/prevenção & controle , Modelos Animais de Doenças , Animais , Western Blotting , Quimiocina CCL2/metabolismo , Corioide/metabolismo , Neovascularização de Coroide/metabolismo , Ensaio de Imunoadsorção Enzimática , Proteínas I-kappa B/metabolismo , Injeções Intraperitoneais , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-6/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Epitélio Pigmentado Ocular/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Xantofilas/uso terapêuticoRESUMO
BACKGROUND: Choroidal neovascularization (CNV) is a critical pathogenesis in age-related macular degeneration, the most common cause of blindness in the developed countries. The aim of the current study was to investigate the effect of lutein supplementation on the development of the murine model of laser-induced CNV together with underlying molecular mechanisms. METHODS AND RESULTS: Mice were orally pretreated with lutein daily from 3 days before laser photocoagulation until the end of the study. The index of CNV volume was significantly suppressed by the treatment with lutein, compared with vehicle-treated animals. Lutein treatment led to significant inhibition of macrophage infiltration into CNV and of the in vivo and in vitro expression of inflammation-related molecules including vascular endothelial growth factor, monocyte chemotactic protein -1, and intercellular adhesion molecule-1. Importantly, lutein suppressed IkappaB-alpha degradation and nuclear translocation of nuclear factor (NF)-kappaB p65 both in vivo and in vitro. Additionally, the development of CNV was significantly suppressed by inhibiting NF-kappaB p65 nuclear translocation, to the levels seen in the lutein treatment. CONCLUSIONS: Lutein treatment led to significant suppression of CNV development together with inflammatory processes including NF-kappaB activation and subsequent upregulation of inflammatory molecules, providing molecular evidence of potential validity of lutein supplementation as a therapeutic strategy to suppress CNV.
Assuntos
Inibidores da Angiogênese/farmacologia , Anti-Inflamatórios/farmacologia , Corioide/efeitos dos fármacos , Neovascularização de Coroide/prevenção & controle , Luteína/farmacologia , Transporte Ativo do Núcleo Celular , Administração Oral , Inibidores da Angiogênese/administração & dosagem , Inibidores da Angiogênese/uso terapêutico , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/uso terapêutico , Quimiocina CCL2/metabolismo , Corioide/metabolismo , Corioide/patologia , Neovascularização de Coroide/etiologia , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Proteínas I-kappa B/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Fotocoagulação a Laser/efeitos adversos , Luteína/administração & dosagem , Luteína/uso terapêutico , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa , Reprodutibilidade dos Testes , Fator de Transcrição RelA/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: Angiotensin-converting enzyme (ACE), also known as kininase II, functions not only to convert angiotensin I to angiotensin II, but also to cleave bradykinin into inactive fragments. Thus, ACE inhibition causes the tissue accumulation of bradykinin, exerting either of two opposite effects: anti- or proangiogenic. The purpose of the present study was to investigate the role of bradykinin in the development of choroidal neovascularization (CNV), with or without ACE inhibition. METHODS: Laser photocoagulation was used to induce CNV in wild-type C57BL/6J mice and angiotensin II type 1 receptor (AT1-R)-deficient mice. Wild-type mice were pretreated with the ACE inhibitor imidapril, with or without the bradykinin B2 receptor (B2-R) antagonist icatibant daily for 6 days before photocoagulation, and the treatment was continued daily until the end of the study. CNV response was analyzed by volumetric measurements using confocal microscopy 1 week after laser injury. The mRNA and protein levels of vascular endothelial growth factor (VEGF), intercellular adhesion molecule (ICAM)-1, and monocyte chemotactic protein (MCP)-1 in the retinal pigment epithelium-choroid complex were examined by RT-PCR and ELISA, respectively. RESULTS: ACE inhibition led to significant suppression of CNV development to the level seen in AT1-R-deficient mice. B2-R blockade together with high-dose but not low-dose ACE inhibition resulted in more potent suppression of CNV than did ACE inhibition alone. B2-R blockade alone exhibited little or no effect on CNV. VEGF, ICAM-1, and MCP-1 levels, elevated by CNV induction, were significantly suppressed by ACE inhibition. VEGF but not ICAM-1 or MCP-1 levels were further attenuated by B2-R blockade with ACE inhibition. CONCLUSIONS: These results suggest a limited contribution of the kallikrein-kinin system to the pathogenesis of CNV, in which the renin-angiotensin system plays more essential roles for facilitating angiogenesis. The present study indicates the possibility of ACE inhibition as a novel therapeutic strategy to inhibit CNV.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Bradicinina/fisiologia , Neovascularização de Coroide/prevenção & controle , Imidazolidinas/farmacologia , Peptidil Dipeptidase A/fisiologia , Animais , Bradicinina/análogos & derivados , Bradicinina/farmacologia , Antagonistas de Receptor B2 da Bradicinina , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Corioide/metabolismo , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Sistema Calicreína-Cinina/fisiologia , Fotocoagulação a Laser , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal , Epitélio Pigmentado Ocular/metabolismo , RNA Mensageiro/metabolismo , Receptor Tipo 1 de Angiotensina/deficiência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: To investigate the role of eicosapentaenoic acid (EPA), the major omega-3 polyunsaturated fatty acid (PUFA), in the development of choroidal neovascularization (CNV), together with underlying molecular mechanisms. METHODS: Six-week-old C57BL/6 mice were fed with laboratory chow with 5% EPA or the omega-6 PUFA linoleic acid (LA) for 4 weeks. Laser photocoagulation was performed to induce CNV, and the volume of CNV tissue was evaluated by volumetric measurements. The expression and production of intercellular adhesion molecule (ICAM)-1, monocyte chemotactic protein (MCP)-1, vascular endothelial growth factor (VEGF) and interleukin (IL)-6 in the retinal pigment epithelium (RPE)-choroid in vivo, and stimulated b-End3 endothelial cells and RAW264.7 macrophages in vitro were evaluated by RT-PCR and ELISA. Fatty acid composition in the serum and the RPE-choroid was analyzed by gas chromatography and high-performance liquid chromatography, respectively. Serum levels of C-reactive protein (CRP), IL-6, VEGF, MCP-1, and soluble ICAM-1 were examined by ELISA. RESULTS: The CNV volume in EPA-fed animals was significantly suppressed compared with that in control mice, whereas the LA-rich diet did not affect CNV. The mRNA expression and protein levels of ICAM-1, MCP-1, VEGF, and IL-6 after CNV induction were significantly reduced in EPA-supplemented mice. In vitro, EPA application led to significant inhibition of mRNA and protein levels of ICAM-1 and MCP-1 in endothelial cells and VEGF and IL-6 in macrophages. EPA-fed mice exhibited significantly higher levels of EPA and lower levels of the omega-6 PUFA arachidonic acid in the serum and the RPE-choroid than control animals. EPA supplementation also led to significant reduction of serum levels of IL-6 and CRP after CNV induction. CONCLUSIONS: The present study demonstrates for the first time that an EPA-rich diet results in significant suppression of CNV and CNV-related inflammatory molecules in vivo and in vitro. These results suggest that frequent consumption of omega-3 PUFAs may prevent CNV and lower the risk of blindness due to age-related macular degeneration.
Assuntos
Anti-Inflamatórios/administração & dosagem , Neovascularização de Coroide/prevenção & controle , Dieta , Ácido Eicosapentaenoico/administração & dosagem , Animais , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Corioide/metabolismo , Corioide/cirurgia , Neovascularização de Coroide/metabolismo , Cromatografia Gasosa , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Ácidos Graxos/sangue , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Fotocoagulação a Laser , Ácido Linoleico/administração & dosagem , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Epitélio Pigmentado Ocular/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: To investigate the involvement of the renin-angiotensin system (RAS) and the nuclear factor (NF)-kappaB pathway with diabetes-induced retinal inflammation. METHODS: Six weeks after induction of diabetes, C57BL/6 mice were treated with the angiotensin II type 1 receptor (AT1-R) blocker (ARB) telmisartan or valsartan, the AT2-R blocker PD123319, or the NF-kappaB inhibitor dehydroxymethylepoxyquinomicin (DHMEQ) daily for 1 week. Retinal mRNA and protein levels of the RAS components were examined by RT-PCR and Western blot, respectively. Leukocyte adhesion to the retinal vasculature was evaluated with a concanavalin A lectin perfusion-labeling technique. Retinal expression levels of intercellular adhesion molecule (ICAM)-1 and vascular endothelial growth factor (VEGF) were examined by RT-PCR and ELISA. ARB or DHMEQ was applied to murine capillary endothelial (b-End3) cells stimulated with a high concentration of glucose to analyze nuclear translocation of NF-kappaB via immunohistochemistry for p65 and mRNA and protein levels of ICAM-1 and monocyte chemotactic protein (MCP)-1. RESULTS: Induction of diabetes led to a significant increase in retinal expression and production of the RAS components including angiotensin II, AT1-R, and AT2-R. Retinal adherent leukocytes were significantly suppressed by AT1-R, but not by AT2-R, blockade. Administration of the ARB, but not of PD123319, inhibited diabetes-induced retinal expression of ICAM-1 and VEGF. DHMEQ also suppressed these cellular and molecular inflammatory parameters in the diabetic retina to the levels obtained with ARB treatment. In vitro, glucose-induced nuclear translocation of NF-kappaB p65 and upregulation of ICAM-1 and MCP-1 were significantly suppressed by application of the ARB. The in vivo treatment with the ARB, as well as DHMEQ, attenuated the diabetes-induced retinal expression of angiotensin II and AT1-R, per se. CONCLUSIONS: The present data revealed significant a contribution of the AT1-R/NF-kappaB pathway to diabetes-induced retinal inflammation, providing a mechanistic reason for targeting AT1-R or NF-kappaB in the treatment of diabetic retinopathy.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Benzamidas/farmacologia , Cicloexanonas/farmacologia , Retinopatia Diabética/prevenção & controle , NF-kappa B/antagonistas & inibidores , Receptor Tipo 1 de Angiotensina/fisiologia , Retinite/prevenção & controle , Bloqueadores do Receptor Tipo 2 de Angiotensina II , Animais , Western Blotting , Adesão Celular , Quimiocina CCL2/metabolismo , Diabetes Mellitus Experimental/complicações , Retinopatia Diabética/etiologia , Retinopatia Diabética/metabolismo , Ensaio de Imunoadsorção Enzimática , Imidazóis/farmacologia , Molécula 1 de Adesão Intercelular/metabolismo , Leucócitos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Piridinas/farmacologia , RNA Mensageiro/metabolismo , Sistema Renina-Angiotensina/fisiologia , Retinite/etiologia , Retinite/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tetrazóis/farmacologia , Tiazolidinas/farmacologia , Valina/análogos & derivados , Valina/farmacologia , Valsartana , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
BACKGROUND: Choroidal neovascularization (CNV) is a critical pathogenesis in age-related macular degeneration, the most common cause of blindness in the developed countries. The aim of the current study was to determine the involvement of the renin-angiotensin system (RAS) with the development of CNV, using human surgical samples and the murine model of laser-induced CNV. METHODS AND RESULTS: In the human and murine CNV tissues, the vascular endothelium expressed angiotensin II type 1 receptor (AT1-R), AT2-R, and angiotensin II. The CNV volume was significantly suppressed by treatment with an AT1-R blocker telmisartan, but not with an AT2-R blocker. AT1-R signaling blockade with telmisartan inhibited various inflammatory mechanisms including macrophage infiltration and upregulation of VEGF, intercellular adhesion molecule-1 (ICAM-1), MCP-1, and IL-6 in the retinal pigment epithelium-choroid complex. A PPAR-gamma antagonist partially but significantly reversed the suppressive effect of telmisartan on in vivo induction of CNV and in vitro upregulation of ICAM-1 and MCP-1 in endothelial cells and IL-6 in macrophages, showing the dual contribution of PPAR-gamma-agonistic and AT1-R-antagonistic actions in the telmisartan treatment. CONCLUSIONS: AT1-R-mediated inflammation plays a pivotal role in the development of CNV, indicating the possibility of AT1-R blockade as a novel therapeutic strategy to inhibit CNV.
Assuntos
Neovascularização de Coroide/etiologia , Mediadores da Inflamação/metabolismo , Inflamação/complicações , Inflamação/etiologia , Receptor Tipo 1 de Angiotensina/metabolismo , Indutores da Angiogênese/antagonistas & inibidores , Angiotensina II/metabolismo , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/fisiopatologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Humanos , Mediadores da Inflamação/antagonistas & inibidores , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , PPAR gama/efeitos dos fármacos , PPAR gama/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Telmisartan , Distribuição TecidualRESUMO
BACKGROUND/AIMS: To study the initial characteristics and response to intravitreal ranibizumab (IVR) treatment of age-related macular degeneration (AMD). METHODS: We reviewed the clinical records of 141 eyes in 141 AMD patients who received monthly IVR for 3â months and thereafter pro re nata (PRN) injections for 9â months as the first treatment for AMD. Patients whose best corrected visual acuity (BCVA) worsened at month 12, and those with increased exudative fundus findings after IVR or an increased central retinal thickness of more than 100 µm at month 12, were considered to be non-responders as judged by BCVA and fundus findings, respectively. Non-responders' initial characteristics were analysed using logistic regression models. RESULTS: 14.9% of eyes were non-responders as judged by BCVA, and 17.0% were non-responders as judged by fundus findings. Initial fibrovascular pigment epithelial detachment (PED) (OR 22.9, 95% CI 2.61 to 201) and serous PED (OR 4.12, 95% CI 1.08 to 15.8) were associated with non-response as judged by BCVA. Initial fibrovascular PED (OR 33.5, 95% CI 2.95 to 381) and type 1 choroidal neovascularization (OR 6.46, 95% CI 1.39 to 30.0) were associated with non-response, as judged by fundus findings. CONCLUSIONS: Although most AMD responded to IVR, non-responders had initial clinical characteristics that might be informative for managing their treatment.
Assuntos
Inibidores da Angiogênese/administração & dosagem , Anticorpos Monoclonais Humanizados/administração & dosagem , Degeneração Macular/tratamento farmacológico , Idoso , Idoso de 80 Anos ou mais , Inibidores da Angiogênese/uso terapêutico , Anticorpos Monoclonais Humanizados/uso terapêutico , Neovascularização de Coroide/complicações , Técnicas de Diagnóstico Oftalmológico , Esquema de Medicação , Feminino , Humanos , Injeções Intravítreas , Degeneração Macular/complicações , Degeneração Macular/fisiopatologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Ranibizumab , Descolamento Retiniano/complicações , Estudos Retrospectivos , Fatores de Risco , Falha de Tratamento , Acuidade Visual/efeitos dos fármacosRESUMO
PURPOSE: Characterization of a mouse model of spontaneous choroidal neovascularization (sCNV) and its effect on retinal architecture and function. METHODS: The sCNV mouse phenotype was characterized by using fundus photography, fluorescein angiography, confocal scanning laser ophthalmoscopy (SLO), optical coherence tomography (OCT), ERG, immunostaining, biochemistry, and electron microscopy. A role for VEGF-A signaling in sCNV was investigated by using neutralizing antibodies and a role for macrophages explored by cell-depletion studies. RESULTS: The sCNV starts between postnatal day 10 and 15 (P10-P15), increasing in number and severity causing RPE disruption and dysfunction. Various morphological methods confirmed the choroidal origin and subretinal position of the angiogenic vessels. At approximately P25, vessels were present in the outer retina with instances of anastomosis of some sCNV lesions with the retinal vasculature. The number of CNV lesions was significantly decreased by systemic blockade of the VEGF-A pathway. Choroidal neovascularization size also was significantly modulated by reducing the number of lesion-associated macrophages. Later stages of sCNV were associated with edema, neuronal loss, and dysfunction. CONCLUSIONS: The sCNV mouse is a new model for the study of both early and late events associated with choroidal neovascularization. Pharmacological reduction in sCNV with VEGF-A antagonists and an anti-inflammatory strategy suggests the model may be useful for investigating novel targets for treating human ocular neovascular disease.
Assuntos
Corioide/irrigação sanguínea , Neovascularização de Coroide/metabolismo , Edema/metabolismo , Epitélio Pigmentado da Retina/ultraestrutura , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Corioide/ultraestrutura , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Edema/patologia , Eletrorretinografia , Ensaio de Imunoadsorção Enzimática , Angiofluoresceinografia , Fundo de Olho , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Microscopia Confocal , Microscopia Eletrônica , Oftalmoscopia , Fenótipo , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/fisiopatologia , Vasos Retinianos/metabolismo , Vasos Retinianos/ultraestrutura , Transdução de Sinais , Tomografia de Coerência ÓpticaAssuntos
Aneurisma/tratamento farmacológico , Edema Macular/tratamento farmacológico , Disco Óptico/irrigação sanguínea , Artéria Retiniana/efeitos dos fármacos , Doenças Retinianas/tratamento farmacológico , Síndrome de Sneddon/complicações , Triancinolona Acetonida/uso terapêutico , Aneurisma/diagnóstico , Aneurisma/etiologia , Angiofluoresceinografia , Glucocorticoides/uso terapêutico , Humanos , Injeções Intraoculares , Edema Macular/diagnóstico , Edema Macular/etiologia , Masculino , Pessoa de Meia-Idade , Artéria Retiniana/patologia , Doenças Retinianas/diagnóstico , Doenças Retinianas/etiologia , Cápsula de Tenon/efeitos dos fármacos , Tomografia de Coerência ÓpticaRESUMO
PURPOSE: Matrix expansion is an early change in age-related maculopathy. The aim of this study was to determine whether connective tissue growth factor (CTGF) regulates the production of extracellular matrix components by retinal pigmented epithelial (RPE) cells. METHODS: ARPE-19 cells were treated with CTGF and analyzed for fibronectin, laminin, and MMP-2 by RT-qPCR, Western blot analysis, or zymography. Cells were also pretreated with an MEK-1/2 inhibitor (PD98059) or a p38 inhibitor (SB203580) and an anti-CTGF antibody to analyze the signaling contributing to fibronectin, laminin, and MMP-2 production. Human maculas were analyzed for mRNA using laser capture microdissected RPE cells and by immunohistochemistry for the topographic distribution of CTGF. RESULTS: CTGF induced fibronectin mRNA (P=0.006) and protein (P=0.006), and laminin mRNA (P=0.006) and protein (P=0.02) by ARPE-19 cells. CTGF also induced MMP-2 mRNA (P=0.002) and protein secretion (P=0.04). Using zymography, CTGF increased the latent and active forms of MMP-2 compared to controls (P=0.02). An anti-CTGF antibody inhibited fibronectin, laminin, and MMP-2 after CTGF stimulation. CTGF increased the phosphorylation of p38 and ERK1/2. Fibronectin and MMP-2 mRNA and protein were suppressed by a MEK-1/2 inhibitor, but not with a p38 inhibitor. Laminin expression was suppressed by both inhibitors. RT-qPCR analysis showed that macular RPE cells from human donors express CTGF. Immunohistochemistry of human maculas showed strong labeling of CTGF in Bruch membrane, including basal deposits and drusen. CONCLUSIONS: CTGF is increased in basal deposits and drusen of AMD specimens, and it induces matrix protein production in ARPE-19 cells through the ERK (p42/p44(mapk)) and p38(mapk) signaling pathways.
Assuntos
Fator de Crescimento do Tecido Conjuntivo/metabolismo , Fator de Crescimento do Tecido Conjuntivo/farmacologia , Proteínas da Matriz Extracelular/biossíntese , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Epitélio Pigmentado da Retina/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Membrana Basal/metabolismo , Western Blotting , Células Cultivadas , Fibronectinas/genética , Fibronectinas/metabolismo , Flavonoides/farmacologia , Humanos , Imidazóis/farmacologia , Técnicas Imunoenzimáticas , Laminina/genética , Laminina/metabolismo , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Piridinas/farmacologia , RNA Mensageiro/metabolismo , Proteínas Recombinantes/farmacologia , Epitélio Pigmentado da Retina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidoresRESUMO
Uveitis is an inflammatory condition that can lead to blindness. It is therefore important to understand the pathophysiology against which to develop targeted therapy. Herein, we tested whether the oxidant-responsive transcription factor Nrf2 is involved in regulating the innate immune response and oxidative damage in the LPS uveitis model. As shown by dihydroethidium staining, intraperitoneally injected LPS increased reactive oxygen species in the retina and iris-ciliary body of Nrf2+/+ and Nrf2-/- mice. After LPS injection, ICAM-1, IL-6, TNF-alpha, COX-2, iNOS, and MCP-1 mRNAs were increased more in the retina and iris-ciliary body of Nrf2-/- than in those of Nrf2+/+ mice. NQO-1 and GCLM, two Nrf2-responsive antioxidant enzymes, had reduced expression in Nrf2+/+ retinas after LPS injection, but no change in expression in Nrf2-/- mice. The number of FITC-Con A-labeled leukocytes adherent to the retinal vascular endothelium increased after LPS treatment in both Nrf2+/+ and Nrf2-/- mice compared to control injections, with more adherent leukocytes in Nrf2-/- than in Nrf2+/+ mice. Pretreatment with the Nrf2 activator 1-(2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl)imidazole increased antioxidant gene expression in the retina, reduced inflammatory mediator expression, and reduced leukocyte adherence to retinal vasculature after LPS treatment in Nrf2+/+ mice, but had no effect on Nrf2-/- mice. Treatment targeting the Nrf2 pathway may be a new therapy for uveitis.
Assuntos
Corpo Ciliar/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Retina/metabolismo , Uveíte/imunologia , Uveíte/metabolismo , Animais , Adesão Celular , Quimiocina CCL2/genética , Quimiocina CCL2/imunologia , Quimiocina CCL2/metabolismo , Corpo Ciliar/imunologia , Corpo Ciliar/patologia , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/imunologia , Ciclo-Oxigenase 2/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica , Glutamato-Cisteína Ligase/genética , Glutamato-Cisteína Ligase/imunologia , Glutamato-Cisteína Ligase/metabolismo , Imunidade Inata , Injeções Intraperitoneais , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/imunologia , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-6/metabolismo , Lipopolissacarídeos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NAD(P)H Desidrogenase (Quinona) , NADPH Desidrogenase/genética , NADPH Desidrogenase/imunologia , NADPH Desidrogenase/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/imunologia , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/imunologia , Óxido Nítrico Sintase Tipo II/metabolismo , Retina/imunologia , Retina/patologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Uveíte/genética , Uveíte/patologiaRESUMO
Interleukin (IL)-6, a potent proinflammatory cytokine, is suggested to be a risk factor for choroidal neovascularization (CNV) because of its increased levels in the serum of patients with age-related macular degeneration; however, the role of IL-6 in CNV has not been defined. The present study reveals the critical contribution of IL-6 signaling and its downstream STAT3 pathway to the murine model of laser-induced CNV. CNV induction by laser treatment stimulated IL-6 expression in the retinal pigment epithelium-choroid complex, and antibody-based blockade of IL-6 receptor or genetic ablation of IL-6 led to significant suppression of CNV. CNV generation was accompanied by STAT3 activation in choroidal endothelial cells and macrophages, and IL-6 receptor blockade resulted in selectively inhibited phosphorylation of STAT3 but not extracellular signal-regulated kinase 1/2. Consistently, pharmacological blockade of STAT3 pathway also suppressed CNV. In addition, IL-6 receptor neutralization led to significant inhibition of the in vivo and in vitro expression of inflammation-related molecules including monocyte chemotactic protein, intercellular adhesion molecule-1, and vascular endothelial growth factor, and of macrophage infiltration into CNV. These results indicate the significant involvement of IL-6 receptor-mediated activation of STAT3 inflammatory pathway in CNV generation, suggesting the possibility of IL-6 receptor blockade as a therapeutic strategy to suppress CNV associated with age-related macular degeneration.