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1.
Methods Mol Biol ; 2212: 169-179, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33733356

RESUMO

In biology, the term "epistasis" indicates the effect of the interaction of a gene with another gene. A gene can interact with an independently sorted gene, located far away on the chromosome or on an entirely different chromosome, and this interaction can have a strong effect on the function of the two genes. These changes then can alter the consequences of the biological processes, influencing the organism's phenotype. Machine learning is an area of computer science that develops statistical methods able to recognize patterns from data. A typical machine learning algorithm consists of a training phase, where the model learns to recognize specific trends in the data, and a test phase, where the trained model applies its learned intelligence to recognize trends in external data. Scientists have applied machine learning to epistasis problems multiple times, especially to identify gene-gene interactions from genome-wide association study (GWAS) data. In this brief survey, we report and describe the main scientific articles published in data mining and epistasis. Our article confirms the effectiveness of machine learning in this genetics subfield.


Assuntos
Biologia Computacional/métodos , Mineração de Dados/métodos , Epistasia Genética , Aprendizado de Máquina , Característica Quantitativa Herdável , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Doença de Crohn/genética , Doença de Crohn/metabolismo , Doença de Crohn/patologia , Genoma Humano , Estudo de Associação Genômica Ampla , Humanos , Padrões de Herança , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Fenótipo , Plantas/genética , Polimorfismo de Nucleotídeo Único
2.
PLoS One ; 15(12): e0244307, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33362238

RESUMO

RATIONALE: Age-related macular degeneration (AMD) is the most prevalent form of irreversible blindness in the developed world. Aging, inflammation and complement dysregulation affecting the retinal pigment epithelium (RPE), are considered significant contributors in its pathogenesis and several evidences have linked tumor necrosis factor alpha (TNF-α) and complement component 3 (C3) with AMD. Acadesine, an analog of AMP and an AMP-activated protein kinase (AMPK) activator, has been shown to have cytoprotective effects in human clinical trials as well as having anti-inflammatory and anti-vascular exudative effects in animals. The purpose of this study was to evaluate if acadesine is able to suppress TNF-α induced C3 in RPE cells. METHODS: ARPE-19 and human primary RPE cells were cultured and allowed to grow to confluence. TNF-α was used for C3 induction in the presence or absence of acadesine. Small molecule inhibitors and siRNA were used to determine if acadesine exerts its effect via the extracellular or intracellular pathway and to evaluate the importance of AMPK for these effects. The expression level of C3 was determined by immunoblot analysis. RESULTS: Acadesine suppresses TNF-α induced C3 in a dose dependent manner. When we utilized the adenosine receptor inhibitor dipyridamole (DPY) along with acadesine, acadesine's effects were abolished, indicating the necessity of acadesine to enter the cell in order to exert it's action. However, pretreatment with 5-iodotubericidin (5-Iodo), an adenosine kinase (AK) inhibitor, didn't prevent acadesine from decreasing TNF-α induced C3 expression suggesting that acadesine does not exert its effect through AMP conversion and subsequent activation of AMPK. Consistent with this, knockdown of AMPK α catalytic subunit did not affect the inhibitory effect of acadesine on TNF-α upregulation of C3. CONCLUSIONS: Our results suggest that acadesine suppresses TNF-α induced C3, likely through an AMPK-independent pathway, and could have potential use in complement over activation diseases.


Assuntos
Aminoimidazol Carboxamida/análogos & derivados , Complemento C3/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Ribonucleosídeos/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Adenosina/metabolismo , Aminoimidazol Carboxamida/metabolismo , Aminoimidazol Carboxamida/farmacologia , Animais , Linhagem Celular , Células Cultivadas , Ativação do Complemento/efeitos dos fármacos , Complemento C3/efeitos dos fármacos , Humanos , Degeneração Macular/metabolismo , Fosforilação , Epitélio Pigmentado da Retina/efeitos dos fármacos , Pigmentos da Retina/metabolismo , Ribonucleosídeos/metabolismo , Ribonucleotídeos/farmacologia , Fator de Necrose Tumoral alfa/metabolismo
3.
Med Sci (Paris) ; 36(10): 886-892, 2020 Oct.
Artigo em Francês | MEDLINE | ID: mdl-33026331

RESUMO

Age-related macular degeneration (AMD) is a complex, highly heritable, multifactorial disease caused by the interplay of age and genetic and environmental risk factors. No treatment has yet been found to treat the slowly progressing atrophic form of AMD. All forms of AMD are invariably associated with an accumulation of mononuclear phagocytes (MP) in the subretinal space, a family of cells that include inflammatory and resident macrophages. We here present an overview of the inflammatory process occurring in AMD and discuss the origin of MPs and the consequences of their accumulation in the subretinal space. Finally, we will review the role played by the established risk factors for AMD to promote the switch from beneficial inflammation in early stage to a deleterious inflammation in the advanced stage of the disease.


Assuntos
Inflamação/complicações , Degeneração Macular/etiologia , Olho/imunologia , Olho/metabolismo , Olho/patologia , Humanos , Privilégio Imunológico/fisiologia , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Mediadores da Inflamação/fisiologia , Degeneração Macular/epidemiologia , Degeneração Macular/imunologia , Degeneração Macular/metabolismo , Fatores de Risco
4.
Med Sci (Paris) ; 36(6-7): 616-625, 2020.
Artigo em Francês | MEDLINE | ID: mdl-32614313

RESUMO

Iron has a fundamental role for cell physiology and especially in retina as a cofactor of many pathways of the visual transduction. A tightly regulated homeostasis avoids the accumulation of prooxidant and proinflammatory free iron. A dysfunction of iron retinal homeostasis is associated with many genetic or age-related degenerative diseases such as age-related macular degeneration (AMD). Here, we describe various mechanisms reported during AMD, enhanced by iron accumulation and its homeostasis dysregulation. We have investigated a local treatment with transferrin, the natural iron carrier, to control these pathological pathways and iron dysfunction, without side effects. Iron has a central role in pathogenesis of AMD and is a target for futures therapies.


Assuntos
Ferro/fisiologia , Degeneração Macular/etiologia , Homeostase/genética , Humanos , Ferro/metabolismo , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/terapia , Redes e Vias Metabólicas/genética , Retina/metabolismo , Retina/patologia , Terapias em Estudo/métodos , Terapias em Estudo/tendências , Transferrina/genética , Transferrina/fisiologia
5.
Proc Natl Acad Sci U S A ; 117(31): 18504-18510, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32699145

RESUMO

The human blood protein vitronectin (Vn) is a major component of the abnormal deposits associated with age-related macular degeneration, Alzheimer's disease, and many other age-related disorders. Its accumulation with lipids and hydroxyapatite (HAP) has been demonstrated, but the precise mechanism for deposit formation remains unknown. Using a combination of solution and solid-state NMR experiments, cosedimentation assays, differential scanning fluorimetry (DSF), and binding energy calculations, we demonstrate that Vn is capable of binding both soluble ionic calcium and crystalline HAP, with high affinity and chemical specificity. Calcium ions bind preferentially at an external site, at the top of the hemopexin-like (HX) domain, with a group of four Asp carboxylate groups. The same external site is also implicated in HAP binding. Moreover, Vn acquires thermal stability upon association with either calcium ions or crystalline HAP. The data point to a mechanism whereby Vn plays an active role in orchestrating calcified deposit formation. They provide a platform for understanding the pathogenesis of macular degeneration and other related degenerative disorders, and the normal functions of Vn, especially those related to bone resorption.


Assuntos
Cálcio/metabolismo , Durapatita/metabolismo , Degeneração Macular/metabolismo , Vitronectina/metabolismo , Sítios de Ligação , Cálcio/química , Durapatita/química , Humanos , Ligação Proteica , Vitronectina/química
6.
Microvasc Res ; 131: 104031, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32531351

RESUMO

The human choroidal vasculature is subject to age-related structural and gene expression changes implicated in age-related macular degeneration (AMD). In this study, we performed both bulk and single-cell RNA sequencing on infant (n = 4 for bulk experiments, n = 2 for single-cell experiments) and adult (n = 13 for bulk experiments, n = 6 for single-cell experiments) human donors to characterize how choroidal gene expression changes with age. Differential expression analysis revealed that aged choroidal samples were enriched in genes encoding pro-inflammatory transcription factors and leukocyte transendothelial cell migration adhesion proteins. Such genes were observed to be differentially expressed specifically within choroidal endothelial cells at the single-cell level. Immunohistochemistry experiments support transcriptional findings that CD34 is elevated in infant choriocapillaris endothelial cells while ICAM-1 is enriched in adults. These results suggest several potential drivers of the pro-inflammatory vascular phenotype observed with advancing age.


Assuntos
Envelhecimento/genética , Corioide/irrigação sanguínea , Células Endoteliais/metabolismo , Mediadores da Inflamação/metabolismo , Inflamação/genética , Degeneração Macular/genética , Análise de Sequência de RNA , Análise de Célula Única , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/metabolismo , Feminino , Regulação da Expressão Gênica , Humanos , Lactente , Recém-Nascido , Inflamação/metabolismo , Degeneração Macular/metabolismo , Masculino , Pessoa de Meia-Idade , Fenótipo
7.
Invest Ophthalmol Vis Sci ; 61(5): 35, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32433758

RESUMO

Purpose: The human retinal pigment epithelium (RPE) accumulates granules significant for autofluorescence imaging. Knowledge of intracellular accumulation and distribution is limited. Using high-resolution microscopy techniques, we determined the total number of granules per cell, intracellular distribution, and changes related to retinal topography and age. Methods: RPE cells from the fovea, perifovea, and near-periphery of 15 human RPE flat mounts were imaged using structured illumination microscopy (SIM) and confocal fluorescence microscopy in young (≤51 years, n = 8) and older (>80 years, n = 7) donors. Using custom FIJI plugins, granules were marked with computer assistance, classified based on morphological and autofluorescence properties, and analyzed with regard to intracellular distribution, total number per cell, and granule density. Results: A total of 193,096 granules in 450 RPE cell bodies were analyzed. Based on autofluorescence properties, size, and composition, the RPE granules exhibited nine different phenotypes (lipofuscin, two; melanolipofuscin, five; melanosomes, two), distinguishable by SIM. Overall, lipofuscin (low at the fovea but increases with eccentricity and age) and melanolipofuscin (equally distributed at all three locations with no age-related changes) were the major granule types. Melanosomes were under-represented due to suboptimal visualization of apical processes in flat mounts. Conclusions: Low lipofuscin and high melanolipofuscin content within foveal RPE cell bodies and abundant lipofuscin at the perifovea suggest a different genesis, plausibly related to the population of overlying photoreceptors (fovea, cones only; perifovea, highest rod density). This systematic analysis provides further insight into RPE cell and granule physiology and links granule load to cell autofluorescence, providing a subcellular basis for the interpretation of clinical fundus autofluorescence.


Assuntos
Lipofuscina/metabolismo , Degeneração Macular/metabolismo , Melanossomas/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Adolescente , Adulto , Idoso de 80 Anos ou mais , Citoplasma/metabolismo , Feminino , Humanos , Imageamento Tridimensional , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Imagem Óptica , Fenótipo , Doadores de Tecidos
8.
Sci Rep ; 10(1): 7656, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32376945

RESUMO

Transplantation of autologous human induced pluripotent stem cell-derived retinal pigment epithelial (hiPSC-RPE) sheets is a promising therapy for age-related macular degeneration (AMD). As melanin content is a representative feature of healthy RPE, we used polarization-sensitive optical coherence tomography (PS-OCT) to estimate the relative melanin content of RPE in diseased and non-diseased area, and in human iPSC-RPE sheets in vitro and in vivo by evaluating the randomness of polarization (entropy). Two aged Japanese women, one with neovascular AMD that underwent transplantation of an autologous hiPSC-RPE cell sheet and another with binocular dry AMD, were selected for this study. Entropy value was minimal in cells containing no melanin, whereas that of human RPE and hiPSC-RPE sheets was high. En face entropy of the cultured hiPSC-RPE sheet was compared with its grey-scale photo and its values were found to be inversely correlated with the extent of absence of pigmentation in vitro. En face entropy maps were compared to colour fundus photographs, fundus autofluorescence images, and fluorescein angiography images from patients. Entropy values of intact and defective RPEs and of iPSC-RPE transplant areas were determined in vivo using PS-OCT B-scan images. PS-OCT was found to be applicable in the estimation of relative melanin content of cultured and transplanted RPEs in regenerative medicine.


Assuntos
Biomarcadores , Células-Tronco Pluripotentes Induzidas/citologia , Melaninas/metabolismo , Epitélio Pigmentado da Retina/diagnóstico por imagem , Epitélio Pigmentado da Retina/metabolismo , Tomografia de Coerência Óptica , Idoso , Idoso de 80 Anos ou mais , Técnicas de Cultura de Células , Diferenciação Celular , Feminino , Angiofluoresceinografia , Células HEK293 , Humanos , Degeneração Macular/diagnóstico por imagem , Degeneração Macular/etiologia , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino , Epitélio Pigmentado da Retina/citologia , Tomografia de Coerência Óptica/métodos
9.
Sci Rep ; 10(1): 7188, 2020 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-32346038

RESUMO

In the present study, we investigated the association between susceptible genetic variants to age-related macular degeneration (AMD) and response to as-needed intravitreal aflibercept injection (IAI) therapy for exudative AMD including both typical neovascular AMD and polypoidal choroidal vasculopathy (PCV) over 12-months. A total of 234 patients with exudative AMD were initially treated with 3 monthly IAI and thereafter as-needed IAI over 12 months. Seven variants of 6 genes including ARMS2 A69S (rs10490924), CFH (I62V:rs800292 and rs1329428), C2-CFB-SKIV2L(rs429608), C3 (rs2241394), CETP (rs3764261) and ADAMTS-9 (rs6795735) were genotyped for all participants using TaqMan technology. After adjusting for age, gender, baseline BCVA and AMD subtype, A (protective) allele of C2-CFB-SKIV2L rs429608 was associated with visual improvement at 12-month (P = 0.003). Retreatment was associated with T(risk) allele of ARMS2 A69S (P = 2.0 × 10-4; hazard ratio: 2.18:95%CI: 1.47-3.24) and C(risk) allele of CFH rs1329428 (P = 2.0 × 10-3; hazard ratio: 1.74:95%CI: 1.16-2.59) after adjusting for the baseline confounders. The need for additional injections was also associated with T allele of ARMS2 A69S (P = 1.0 × 10-5) and C allele of CFH rs1329428 (P = 3.0 × 10-3) after adjusting for the baseline confounders. The variants of ARMS2 and CFH are informative for both physicians and patients to predict recurrence and to quantify the need for additional injections.


Assuntos
Alelos , Neovascularização de Coroide , Frequência do Gene , Genótipo , Degeneração Macular , Receptores de Fatores de Crescimento do Endotélio Vascular/administração & dosagem , Proteínas Recombinantes de Fusão/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Neovascularização de Coroide/tratamento farmacológico , Neovascularização de Coroide/genética , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Feminino , Humanos , Degeneração Macular/tratamento farmacológico , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino , Estudos Retrospectivos
10.
Am J Pathol ; 190(8): 1632-1642, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32339498

RESUMO

Recent studies deciphering the transcriptional profile of choroidal neovascularization (CNV) in body donor eyes with neovascular age-related macular degeneration are limited by the time span from death to preservation and the associated 5'-RNA degradation. This study therefore used CNV and control specimens that were formalin-fixed and paraffin-embedded immediately after surgical extraction and analyzed them by a 3'-RNA sequencing approach. Transcriptome profiles were analyzed to estimate content of immune and stromal cells and to define disease-associated gene signatures by using statistical and bioinformatics methods. This study identified 158 differentially expressed genes (DEGs) that were significantly increased in CNV compared with control tissue. Cell type enrichment analysis revealed a diverse cellular landscape with an enrichment of endothelial cells, macrophages, T cells, and natural killer T cells in the CNV. Gene ontology enrichment analysis found that DEGs contributed to blood vessel development, extracellular structure organization, response to wounding, and several immune-related terms. The S100 calcium-binding proteins A8 (S100A8) and A9 (S100A9) emerged among the top DEGs, as confirmed by immunohistochemistry on CNV tissue and protein analysis of vitreous samples. This study provides a high-resolution RNA-sequencing-based transcriptional signature of human CNV, characterizes its compositional pattern of immune and stromal cells, and reveals S100A8/A9 to be a novel biomarker and promising target for therapeutics and diagnostics directed at age-related macular degeneration.


Assuntos
Neovascularização de Coroide/diagnóstico , Complexo Antígeno L1 Leucocitário/metabolismo , Degeneração Macular/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/metabolismo , Neovascularização de Coroide/metabolismo , Células Endoteliais/metabolismo , Feminino , Humanos , Macrófagos/metabolismo , Degeneração Macular/metabolismo , Masculino , Transcriptoma
11.
J Pathol ; 251(2): 200-212, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32243583

RESUMO

Age-related macular degeneration (AMD) is a complex, multifactorial, progressive disease which represents a leading cause of irreversible visual impairment and blindness in older individuals. Human cytomegalovirus (HCMV), which infects 50-80% of humans, is usually acquired during early life and persists in a latent state for the life of the individual. In view of its previously described pro-angiogenic properties, we hypothesized that cytomegalovirus might be a novel risk factor for progression to an advanced form, neovascular AMD, which is characterized by choroidal neovascularization (CNV). The purpose of this study was to investigate if latent ocular murine cytomegalovirus (MCMV) infection exacerbated the development of CNV in vascular endothelial growth factor (VEGF)-overexpressing VEGF-Ahyper mice. Here we show that neonatal infection with MCMV resulted in dissemination of virus to various organs throughout the body including the eye, where it localized principally to the choroid in both VEGF-overexpressingVEGF-Ahyper and wild-type(WT) 129 mice. By 6 months post-infection, no replicating virus was detected in eyes and extraocular tissues, although virus DNA was still present in all eyes and extraocular tissues of both VEGF-Ahyper and WT mice. Expression of MCMV immediate early (IE) 1 mRNA was detected only in latently infected eyes of VEGF-Ahyper mice, but not in eyes of WT mice. Significantly increased CNV was observed in eyes of MCMV-infected VEGF-Ahyper mice compared to eyes of uninfected VEGF-Ahyper mice, while no CNV lesions were observed in eyes of either infected or uninfected WT mice. Protein levels of several inflammatory/angiogenic factors, particularly VEGF and IL-6, were significantly higher in eyes of MCMV-infected VEGF-Ahyper mice, compared to uninfected controls. Initial studies of ocular tissue from human cadavers revealed that HCMV DNA was present in four choroid/retinal pigment epithelium samples from 24 cadavers. Taken together, our data suggest that ocular HCMV latency could be a significant risk factor for the development of AMD. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Neovascularização de Coroide/virologia , Retinite por Citomegalovirus/virologia , Degeneração Macular/virologia , Muromegalovirus/patogenicidade , Retina/virologia , Latência Viral , Idoso , Idoso de 80 Anos ou mais , Animais , Neovascularização de Coroide/genética , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Retinite por Citomegalovirus/genética , Retinite por Citomegalovirus/metabolismo , Retinite por Citomegalovirus/patologia , Modelos Animais de Doenças , Progressão da Doença , Feminino , Humanos , Proteínas Imediatamente Precoces/metabolismo , Mediadores da Inflamação/metabolismo , Interleucina-6/metabolismo , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino , Camundongos da Linhagem 129 , Camundongos Transgênicos , Pessoa de Meia-Idade , Retina/metabolismo , Retina/ultraestrutura , Fatores de Risco , Transdução de Sinais , Fatores de Tempo , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
12.
Nat Commun ; 11(1): 1609, 2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32231223

RESUMO

In vitro differentiation of human pluripotent stem cells into functional retinal pigment epithelial (RPE) cells provides a potentially unlimited source for cell based reparative therapy of age-related macular degeneration. Although the inherent pigmentation of the RPE cells have been useful to grossly evaluate differentiation efficiency and allowed manual isolation of pigmented structures, accurate quantification and automated isolation has been challenging. To address this issue, here we perform a comprehensive antibody screening and identify cell surface markers for RPE cells. We show that these markers can be used to isolate RPE cells during in vitro differentiation and to track, quantify and improve differentiation efficiency. Finally, these surface markers aided to develop a robust, direct and scalable monolayer differentiation protocol on human recombinant laminin-111 and -521 without the need for manual isolation.


Assuntos
Biomarcadores/metabolismo , Diferenciação Celular/fisiologia , Células Epiteliais/metabolismo , Neurônios/metabolismo , Pigmentos da Retina/metabolismo , Animais , Antígeno CD56 , Células-Tronco Embrionárias , Humanos , Laminina/genética , Degeneração Macular/metabolismo , Coelhos , Epitélio Pigmentado da Retina/metabolismo
13.
Expert Opin Pharmacother ; 21(7): 773-784, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32153203

RESUMO

INTRODUCTION: Age-related macular degeneration (AMD) is the most common cause of blindness among the elderly in the industrialized world. While effective treatment is available for neovascular AMD, no therapy is successful for the non-neovascular form. Herein, the authors report the current knowledge on non-neovascular AMD pathogenesis and the promising research on treatments. AREAS COVERED: In the present review, the authors summarize the most recent advances in the treatment of non-neovascular AMD and provide an update on current treatment strategies. Evidence available from preclinical and clinical studies and from a selective literature search is reported. EXPERT OPINION: When investigating AMD, numerous pathological cascades and alterations of physiological processes have been investigated. It is well-known that AMD is a multifactorial disease, with environmental causes and genetics playing a role. Perturbations in multiple pathogenic pathways have been identified and this led to the development of several molecules directed at specific therapeutic targets. However, despite the huge research effort, the only proven approach so far is oral antioxidant supplementation. We believe that, in addition to successful advancement of promising drugs, further research should be directed at tailoring therapy to specific patient groups, eventually employing a combinational therapy strategy.


Assuntos
Anti-Inflamatórios/uso terapêutico , Antioxidantes/uso terapêutico , Degeneração Macular/tratamento farmacológico , Acuidade Visual/efeitos dos fármacos , Vitaminas/uso terapêutico , Idoso , Anti-Inflamatórios/administração & dosagem , Antioxidantes/administração & dosagem , Ensaios Clínicos como Assunto , Suplementos Nutricionais , Terapia Genética , Humanos , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Transplante de Células-Tronco , Resultado do Tratamento , Vitaminas/administração & dosagem
14.
PLoS One ; 15(3): e0229342, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32155173

RESUMO

We aimed to construct a better model for predicting treatment outcomes of anti-vascular endothelial growth factor therapy for neovascular age-related macular degeneration (nAMD) using the concentrations of aqueous humour proteins at baseline and during treatment. From the data of 48 treatment-naïve nAMD eyes that received intravitreal ranibizumab pro re nata for up to 12 months, we used the aqueous humour concentrations of C-X-C motif chemokine ligand 1 (CXCL1), CXCL12, CXCL13, interferon-γ-induced protein 10, monocyte chemoattractant protein 1 (MCP-1), C-C motif chemokine ligand 11, interleukin 6 (IL-6), IL-10, and matrix metalloproteinase 9 (MMP-9). After stepwise regression, multivariate analysis was performed to identify which predictors were significantly associated with best-corrected visual acuity (BCVA) changes and the number of injections. The results demonstrated that besides male sex (ß coefficient = -0.088, P = 0.040) and central retinal thickness (ß coefficient = 0.00051 per µm, P = 0.027), MCP-1 (ß coefficient = 0.44, P < 0.001) and IL-10 (ß coefficient = -0.16, P = 0.033) were significantly correlated with baseline BCVA. Additionally, high MCP-1 at baseline (ß coefficient = -0.20, P = 0.015) and low CXCL13 at baseline (ß coefficient = 0.10, P = 0.0054) were independently associated with better BCVA change at 12 months. High MMP-9 at the first injection (ß coefficient = 0.56, P = 0.01), CXCL12 at the third injection (ß coefficient = 0.10, P = 0.0002), and IL-10 at the third injection (ß coefficient = 1.3, P = 0.001) were predictor variables associated with the increased number of injections. In conclusion, aqueous humour protein concentrations may have predictive abilities of BCVA change over 12 months and the number of injections in pro re nata treatment of exudative nAMD.


Assuntos
Humor Aquoso/metabolismo , Proteínas do Olho/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Degeneração Macular/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Ranibizumab/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Idoso , Inibidores da Angiogênese/uso terapêutico , Feminino , Humanos , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Masculino , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Estudos Prospectivos , Resultado do Tratamento
15.
Int J Mol Sci ; 21(6)2020 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-32183173

RESUMO

Increased oxidative stress and mitochondrial damage are observed in protein aggregation diseases, such as age-related macular degeneration (AMD). We have recently reported elevated levels of oxidative stress markers, damaged mitochondria, accumulating lysosomal lipofuscin and extracellular drusen-like structures in the retinal pigment epithelial cells (RPE) of the dry AMD-resembling NFE2L2/PGC1α double knockout (dKO) mouse model. Here, we provide evidence of a disturbance in the autolysosomal machinery handling mitochondrial clearance in the RPE cells of one-year-old NFE2L2/PGC1α-deficient mice. Confocal immunohistochemical analysis revealed an upregulation of autophagosome marker microtubule-associated proteins 1A/1B light chain 3B (LC3B) as well as numerous mitophagy markers, such as PTE-induced putative kinase 1 (PINK1) and E3 ubiquitin ligase (PARKIN) together with damaged mitochondria. However, we detected no evidence of increased autolysosome formation in transmission electron micrographs or of colocalization of lysosomal marker LAMP2 (lysosome-associated membrane protein 2) and the mitochondrial marker ATP synthase ß in confocal micrographs. Interestingly, we observed an upregulation of late autolysosomal fusion Ras-related protein (Rab7) in the perinuclear space of RPE cells together with autofluorescence aggregates. Our results reveal that there is at least a relative decrease of mitophagy in the RPE cells of NFE2L2/PGC1α dKO mice. This further supports the hypothesis that mitophagy is a putative therapy target in AMD-like pathology.


Assuntos
Degeneração Macular/metabolismo , Mitofagia , Fator 2 Relacionado a NF-E2/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Epitélio Pigmentado da Retina/metabolismo , Animais , Deleção de Genes , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Lisossomos/metabolismo , Lisossomos/ultraestrutura , Degeneração Macular/genética , Masculino , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Proteínas Quinases/metabolismo , Epitélio Pigmentado da Retina/ultraestrutura , Ubiquitina-Proteína Ligases/metabolismo
16.
J Med Chem ; 63(11): 5697-5722, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32073845

RESUMO

The alternative pathway (AP) of the complement system is a key contributor to the pathogenesis of several human diseases including age-related macular degeneration, paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome (aHUS), and various glomerular diseases. The serine protease factor B (FB) is a key node in the AP and is integral to the formation of C3 and C5 convertase. Despite the prominent role of FB in the AP, selective orally bioavailable inhibitors, beyond our own efforts, have not been reported previously. Herein we describe in more detail our efforts to identify FB inhibitors by high-throughput screening (HTS) and leveraging insights from several X-ray cocrystal structures during optimization efforts. This work culminated in the discovery of LNP023 (41), which is currently being evaluated clinically in several diverse AP mediated indications.


Assuntos
Ácido Benzoico/química , Fator B do Complemento/antagonistas & inibidores , Indóis/química , Síndrome Hemolítico-Urêmica Atípica/metabolismo , Síndrome Hemolítico-Urêmica Atípica/patologia , Ácido Benzoico/metabolismo , Ácido Benzoico/farmacocinética , Sítios de Ligação , Domínio Catalítico , Fator B do Complemento/metabolismo , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Meia-Vida , Humanos , Indóis/metabolismo , Indóis/farmacocinética , Concentração Inibidora 50 , Degeneração Macular/metabolismo , Degeneração Macular/patologia , Simulação de Dinâmica Molecular , Relação Estrutura-Atividade
17.
Sci Rep ; 10(1): 2464, 2020 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-32051464

RESUMO

Age-related macular degeneration is a major cause of vision impairment in the Western world among people of 55 years and older. Recently we have shown that autophagy is dysfunctional in the retinal pigment epithelium (RPE) of the AMD donor eyes (AMD RPE). We also showed increased reactive oxygen (ROS) production, increased cytoplasmic glycogen accumulation, mitochondrial dysfunction and disintegration, and enlarged and annular LAMP-1-positive organelles in AMD RPE. However, the underlying mechanisms inducing these abnormalities remain to be elucidated. Here, by performing a comprehensive study, we show increased PAPR2 expression, deceased NAD+, and SIRT1, increased PGC-1α acetylation (inactive form), lower AMPK activity, and overactive mTOR pathway in AMD RPE as compared to normal RPE. Metabolomics and lipidomics revealed dysregulated metabolites in AMD RPE as compared to normal RPE, including glycerophospholipid metabolism, involved in autophagy, lipid, and protein metabolisms, glutathione, guanosine, and L-glutamic acid, which are implicated in protection against oxidative stress and neurotoxicity, further supporting our observations. Our data show dysregulated metabolic pathways as important contributors to AMD pathophysiology, and facilitate the development of new treatment strategies for this debilitating disease of the visual system.


Assuntos
Degeneração Macular/metabolismo , Redes e Vias Metabólicas , Feminino , Glicogênio/metabolismo , Humanos , Metabolismo dos Lipídeos , Degeneração Macular/genética , Masculino , Metaboloma , Mitocôndrias/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Transcriptoma
18.
Int J Mol Sci ; 21(3)2020 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-32012797

RESUMO

Age-related macular degeneration (AMD) is considered as the main worldwide cause of blindness in elderly adults. Exudative AMD type represents 10 to 15% of macular degeneration cases, but is the main cause of vision loss and blindness. Circadian rhythm changes are associated with aging and could further accelerate it. However, the link between circadian rhythms and exudative AMD is not fully understood. Some evidence suggests that dysregulation of circadian functions could be manifestations of diseases or could be risk factors for the development of disease in elderly adults. Biological rhythms are complex systems interacting with the environment and control several physiological pathways. Recent findings have shown that the dysregulation of circadian rhythms is correlated with exudative AMD. One of the main pathways involved in exudative AMD is the canonical WNT/ß-catenin pathway. Circadian clocks have a main role in some tissues by driving the circadian expression of genes involved in physiological and metabolic functions. In exudative AMD, the increase of the canonical WNT/ß-catenin pathway is enhanced by the dysregulation of circadian rhythms. Exudative AMD progression is associated with major metabolic reprogramming, initiated by aberrant WNT/ß-catenin pathway, of aerobic glycolysis. This review focuses on the interest of circadian rhythm dysregulation in exudative AMD through the aberrant upregulation of the canonical WNT/ß-catenin pathway.


Assuntos
Ritmo Circadiano , Degeneração Macular/etiologia , Degeneração Macular/metabolismo , Via de Sinalização Wnt , Animais , Relógios Circadianos/genética , Glucose/metabolismo , Glicólise , Humanos , Degeneração Macular/patologia , Neovascularização Patológica/metabolismo
19.
Oxid Med Cell Longev ; 2020: 7901270, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32104539

RESUMO

Age-related macular degeneration (AMD) is a common cause of visual impairment in the elderly. There are very limited therapeutic options for AMD with the predominant therapies targeting vascular endothelial growth factor (VEGF) in the retina of patients afflicted with wet AMD. Hence, it is important to remind readers, especially those interested in AMD, about current studies that may help to develop novel therapies for other stages of AMD. This study, therefore, provides a comprehensive review of studies on human specimens as well as rodent models of the disease, to identify and analyze the molecular mechanisms behind AMD development and progression. The evaluation of this information highlights the central role that oxidative damage in the retina plays in contributing to major pathways, including inflammation and angiogenesis, found in the AMD phenotype. Following on the debate of oxidative stress as the earliest injury in the AMD pathogenesis, we demonstrated how the targeting of oxidative stress-associated pathways, such as autophagy and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling, might be the futuristic direction to explore in the search of an effective treatment for AMD, as the dysregulation of these mechanisms is crucial to oxidative injury in the retina. In addition, animal models of AMD have been discussed in great detail, with their strengths and pitfalls included, to assist inform in the selection of suitable models for investigating any of the molecular mechanisms.


Assuntos
Degeneração Macular/metabolismo , Estresse Oxidativo/fisiologia , Animais , Modelos Animais de Doenças , Humanos , Fator 2 Relacionado a NF-E2/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
20.
Exp Cell Res ; 388(1): 111811, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31899207

RESUMO

Age-related macular degeneration is a progressive ocular disease that is the leading cause of vision loss among elderly. AMD usually is divided into two types: wet and dry AMD, which is linked with inflammation. Choroidal Neovascularization (CNV) formation or wet AMD is also associated with oxidative stress. Previously, TSP1 has been shown to have a significant alleviating effect on CNV in TSP1 knockout (TSP1-/-) mice. However, the mechanism by which TSP1 ameliorates CNV remains unclear. Here we report that TSP1 reduces nitric oxide production to prevent cells from forming tubes formation and reduced the levels of vascular endothelial growth factor (VEGF) and lipid peroxides (LPO) during oxidative stress. We measured RF/6A cell viability by CCK-8 assay and apoptosis by flow cytometry. RF/6A cell were transfected with TSP1 and STAT3 overexpression, and then the mRNA and protein levels of TSP1 and also the signal pathways were detected by qRT-PCR and Western blot analysis. Migration assays were performed using a transwell system. Co-Immunoprecipitation was used to analyze the binding relationship between CD47 and SHP-2. The results show that overexpression of TSP1 alleviated the damage of oxidative stress to RF/6A cells including increased cell activity and migration, decreased apoptosis and reduced migration compared to the control group. SHP-2 was activated by TSP1 through its receptor CD47 and STAT3 phosphorylation was reduced by activation of SHP-2, thereby blocking STAT3-iNOS pathway and reducing NO concentration in RF/6A cells ultimately protecting them from oxidative stress. Finally, the CNV mice model confirmed that TSP1 overexpression could protect the mice against CNV in vivo, modified the antioxidants levels and decreased the expression of TNF-α and IL-6 under laser irradiation. These results indicate a potential mechanism of TSP1 to slow down formation of CNV in wet AMD, which may bring hope for new treatment strategies.


Assuntos
Degeneração Macular/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Fator de Transcrição STAT3/metabolismo , Trombospondina 1/metabolismo , Animais , Apoptose , Antígeno CD47/metabolismo , Linhagem Celular , Peroxidação de Lipídeos , Macaca mulatta , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Transdução de Sinais , Trombospondina 1/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
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