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1.
Invest Ophthalmol Vis Sci ; 61(1): 1, 2020 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-31995153

RESUMO

Purpose: Vacuolar protein sorting 35 (Vps35) mutations and protein dysfunction have been linked to the hyperphosphorylation and accumulation of tau protein in a number of central neurodegenerative disorders. The aims of the present study were to investigate the mechanism underlying the tau hyperphosphorylation caused by Vps35 deficiency. Methods: The cells used in this study were primary retinal ganglion cells (RGCs). The rat retinal glutamate excitotoxicity model was used in vivo. Fresh retinal tissues or eyeballs were collected at different time points. The expression and interactions of Vps35, Cdk5/p35, tau hyperphosphorylation, LAMP1, EEA1 and UBE1 in RGCs were studied by immunofluorescence staining, Western blotting, and immunoprecipitation. Results: The downregulation and overexpression of Vps35 increased and decreased the expression of p35 and tau hyperphosphorylation, respectively. More important, roscovitine, a Cdk5 inhibitor, could effectively decrease the hyperphosphorylated tau level induced by Vps35 deficiency. Furthermore, this study confirmed that the inhibition of Vps35 could increase the activity of Cdk5/p35 by affecting the lysosomal degradation of p35 and lead to the degeneration of RGCs. Conclusions: These findings demonstrate the possibility that Cdk5/p35 acts as a "cargo" of Vps35 and provide new insights into the pathogenesis of RGC degeneration caused by hyperphosphorylated tau protein. Vps35 is a potential target for basic research and clinical treatment of RGC degeneration in many ocular diseases such as glaucoma.


Assuntos
Quinase 5 Dependente de Ciclina/metabolismo , Fosfotransferases/metabolismo , Células Ganglionares da Retina/metabolismo , Proteínas de Transporte Vesicular/deficiência , Proteínas tau/metabolismo , Animais , Western Blotting , Células Cultivadas , Quinase 5 Dependente de Ciclina/antagonistas & inibidores , Regulação para Baixo , Técnica Indireta de Fluorescência para Anticorpo , Ácido Glutâmico/toxicidade , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Masculino , Camundongos , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Degeneração Retiniana/induzido quimicamente , Degeneração Retiniana/metabolismo , Roscovitina/farmacologia , Transfecção , Enzimas Ativadoras de Ubiquitina/metabolismo , Proteínas de Transporte Vesicular/metabolismo
2.
Life Sci ; 242: 117229, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31887298

RESUMO

AIMS: Neutrophil elastase (NE) is a critical proteolytic enzyme that is involved in cancer. We previously reported high NE expression in peripheral blood neutrophils from acute promyelocytic leukemia (APL) patients. The present study aimed to elucidate the specific role and mechanisms of NE in APL development. MATERIALS AND METHODS: NE expression was detected in APL bone marrow samples and analyzed in the BloodSpot database. CCK-8 assay and flow cytometry were used to assess cell proliferation and cell cycle distribution, respectively. The expression levels of proliferation and differentiation markers were measured by Western blotting and quantitative real-time PCR. The co-expression and interaction of NE and p200 cut-like homeobox 1 (CUX1) were evaluated by indirect immunofluorescence, co-immunoprecipitation, and in situ proximity ligation assay. KEY FINDINGS: NE was highly expressed in APL bone marrow and blood neutrophils. NE overexpression promoted the proliferation and inhibited the differentiation of NB4 cells, whereas NE downregulation achieved the opposite results in U937 cells. Mechanistically, NE interacted with and effectively hydrolyzed the tumor suppressor p200 CUX1. Rescue experiments revealed that p200 CUX1 upregulation reversed the functional influence of NE on APL cells. SIGNIFICANCE: NE-mediated proteolysis of the tumor suppressor p200 CUX1 promotes APL progression. NE/p200 CUX1 axis is a novel and promising therapeutic target for APL treatment.


Assuntos
Proteínas de Homeodomínio/metabolismo , Leucemia Promielocítica Aguda/enzimologia , Elastase de Leucócito/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Adolescente , Adulto , Western Blotting , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Feminino , Citometria de Fluxo , Técnica Indireta de Fluorescência para Anticorpo , Células HL-60 , Proteínas de Homeodomínio/fisiologia , Humanos , Imunoprecipitação , Leucemia Promielocítica Aguda/metabolismo , Elastase de Leucócito/fisiologia , Masculino , Proteólise , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Repressoras/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sincalida/metabolismo , Fatores de Transcrição/fisiologia , Células U937
3.
Rev Chilena Infectol ; 36(5): 650-655, 2019 Oct.
Artigo em Espanhol | MEDLINE | ID: mdl-31859807

RESUMO

Ehrlichiosis is a disease transmitted by tick's bite that affect dogs and humans caused by the species Ehrlichia canis and E. chaffeensis, respectively. These bacteria are obligated intracellular gram negatives, with a cocoid to pleomorph aspect and can infect monocytes and trigger symptoms such as high fever, anorexia, thrombocytopenia, hemorrhages, anemia, and some serious problems such as splenomegaly, hepatomegaly and meningitis. There are several diagnostic tests for ehrlichiosis such as the hematological ones that evaluate the morphology of the monocytes in search of morulae; serological tests that includes the search of anti-Ehrlichia antibodies, although they might be limited due to cross reaction with other species. In other hand, the culture of Ehrlichia species is an effective method to obtain antigens and even develop indirect immunofluorescence assays (IFA). The polymerase chain reaction offers a definitive diagnosis associated to the use of genus-specific and species-specific primers, as well as its increased sensibility and specificity, compared to the others methods. Thus, in this review, we will discuss various methods applied to the diagnosis of this disease, as well as the advantages and disadvantages that these present.


Assuntos
Ehrlichiose/diagnóstico , Animais , Cães , Ehrlichia canis/isolamento & purificação , Ehrlichia chaffeensis/isolamento & purificação , Ensaio de Imunoadsorção Enzimática/métodos , Técnica Indireta de Fluorescência para Anticorpo/métodos , Humanos , Reação em Cadeia da Polimerase/métodos , Testes Sorológicos/métodos
4.
An Bras Dermatol ; 94(5): 578-581, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31777359

RESUMO

The authors have successfully treated and monitored a case of paraneoplastic pemphigus in association with follicular dendritic cell sarcoma aggravated by hyaline-vascular Castleman's disease. The patient was a 56-year-old female who presented with recalcitrant erosive lichen planus of the oral cavity, tongue, and genital mucosa, along with polymorphous eruptions throughout her body. Histological examination of the cutaneous lesions, indirect immunofluorescence on rat bladder epithelium, and western blot of human keratinocyte proteins identified anti-epidermal antibodies in the patient's serum. Positron emission tomography and computed tomography scans found a mass in her retroperitoneal region. Pathology and immunohistochemistry investigation further corroborated the diagnosis of follicular dendritic cell sarcoma originated from hyaline-vascular Castleman's disease. Complete remission was achieved and the patient has been monitored for four years.


Assuntos
Hiperplasia do Linfonodo Gigante/complicações , Hiperplasia do Linfonodo Gigante/patologia , Sarcoma de Células Dendríticas Foliculares/etiologia , Sarcoma de Células Dendríticas Foliculares/patologia , Pênfigo/etiologia , Pênfigo/patologia , Biópsia , Western Blotting , Sarcoma de Células Dendríticas Foliculares/cirurgia , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Hialina , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons , Tomografia Computadorizada por Raios X , Resultado do Tratamento
5.
Invest Ophthalmol Vis Sci ; 60(14): 4904-4914, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31770435

RESUMO

Purpose: Uveal melanoma is a common primary intraocular malignancy accompanied by high mortality. Previous evidence has highlighted the implication of microRNAs (miRNAs) and long noncoding RNAs (lncRNAs) in uveal melanoma. Accordingly, we further uncovered the possible role of lncRNA plasmacytoma variant translocation 1 gene (PVT1) and microRNA-17-3p (miR-17-3p) in uveal melanoma. Methods: A series of experiments were performed to examine the relationship among lncRNA PVT1, miR-17-3p, and murine double minute clone 2 oncoprotein (MDM2). Afterward, gain- and loss-of-function approaches were used with uveal melanoma cells to verify the role of lncRNA PVT1, miR-17-3p, and MDM2 in the tumorigenesis and development of uveal melanoma. Results: Highly expressed lncRNA PVT1 and MDM2, yet lowly expressed miR-17-3p, were identified in ocular uveal melanoma tissues versus normal adjacent tissues. Then, dual luciferase reporter gene assay, RNA binding protein immunoprecipitation, and RNA pull-down assays showed that lncRNA PVT1 specifically bound to miR-17-3p, and that MDM2 was a target gene of miR-17-3p. Gain- and loss-of-function studies elucidated that silencing of lncRNA PVT1 or overexpression of miR-17-3p resulted in decreased MDM2 expression and increased transcriptional activity of p53, in addition to inhibiting uveal melanoma cell proliferation, migration, and invasion, yet promoted cell apoptosis in vitro. In addition, lncRNA PVT1 silencing or miR-17-3p overexpression was noted to inhibit tumor growth in vivo. Conclusions: Downregulation of lncRNA PVT1 could potentially promote miR-17-3p expression to suppress tumorigenesis and development of uveal melanoma by activating the p53 signaling pathway through binding to MDM2.


Assuntos
Inativação Gênica/fisiologia , Melanoma/prevenção & controle , MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , RNA Longo não Codificante/genética , Neoplasias Uveais/prevenção & controle , Animais , Western Blotting , Movimento Celular/fisiologia , Sobrevivência Celular , Primers do DNA/química , Técnica Indireta de Fluorescência para Anticorpo , Genes Reporter , Hibridização in Situ Fluorescente , Masculino , Melanoma/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Ativação Transcricional , Transplante Heterólogo , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima , Neoplasias Uveais/metabolismo
6.
Invest Ophthalmol Vis Sci ; 60(14): 4596-4605, 2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31675076

RESUMO

Purpose: Choroidal neovascularization (CNV) is the principal pathological factor contributing to blindness in neovascular age-related macular degeneration (nAMD). Infiltration of M2 macrophage is thought to contribute to CNV progress, although the way that regulates its differentiation remains unclear. Here, we investigate the role of CHI3L1 in M2 differentiation and angiogenesis in CNV. Methods: Serums from nAMD patients were tested for CHI3L1 expression. Mice were subjected to laser injury to induce CNV, and lesion expansion were tracked using fundus fluorescence angiography (FFA) and immunofluorescence analysis. Several strategies were taken to verify the contribution of M2 macrophage and CHI3L1: macrophage depletion by clodrosome, local CHI3L1 inhibition using intravitreally injection neutralize antibody (mAY), and depletion of CHI3L1 receptor (IL13-Ra2) by small-interfering RNA (siRNA). Tuber analysis was used to further determine angiogenetic effect of CHI3L1. Anti-VEGFA was used as positive control for mAY. Results: Serum levels of CHI3L1 were highly elevated in nAMD patients. CHI3L1 was expressed by infiltrating M2 macrophages and was elevated as CNV progress in a mice model. System macrophage depletion and local suppression of CHI3L1 alleviated CNV formation while enhancing anti-VEGFA therapeutic effect. Stimulation of macrophage with recombinant CHI3L1 activated MAPK signaling cascade and induced transition to M2, while siRNA knockdown of IL13-Ra2 abolished it. In an in vitro coculture system, supernatants from CHI3L1-stimulated M2 macrophages and promoted tube vascularization. Conclusions: These results unveil novel angiogenic regulation of CHI3L1 and M2 polarized macrophages in CNV development. These mechanistic insights may point to CHI3L1 as a new therapeutic target for treatment for nAMD.


Assuntos
Diferenciação Celular/fisiologia , Proteína 1 Semelhante à Quitinase-3/fisiologia , Neovascularização de Coroide/fisiopatologia , Macrófagos/fisiologia , Degeneração Macular Exsudativa/fisiopatologia , Idoso , Animais , Anticorpos Neutralizantes/farmacologia , Western Blotting , Neovascularização de Coroide/sangue , Neovascularização de Coroide/prevenção & controle , Modelos Animais de Doenças , Angiofluoresceinografia , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Separação Imunomagnética , Injeções Intravítreas , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Degeneração Macular Exsudativa/sangue , Degeneração Macular Exsudativa/prevenção & controle
7.
Invest Ophthalmol Vis Sci ; 60(13): 4378-4387, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31634395

RESUMO

Purpose: Iron supplementation therapy is used for iron-deficiency anemia but has been associated with macular degeneration in a 43-year-old patient. Iron entry into the neurosensory retina (NSR) can be toxic. It is important to determine conditions under which serum iron might cross the blood retinal barrier (BRB) into the NSR. Herein, an established mouse model of systemic iron overload using high-dose intraperitoneal iron dextran (IP FeDex) was studied. In addition, because the NSR expresses the iron regulatory hormone hepcidin, which could limit iron influx into the NSR, we gave retina-specific hepcidin knockout (RS-HepcKO) mice IP FeDex to test this possibility. Methods: Wild-type (WT) and RS-HepcKO mice were given IP FeDex. In vivo retina imaging was performed. Blood and tissues were analyzed for iron levels. Quantitative PCR was used to measure levels of mRNAs encoding iron regulatory and photoreceptor-specific genes. Ferritin and albumin were localized in the retina by immunofluorescence. Results: IP FeDex in both WT and RS-HepcKO mice induced high levels of iron in the liver, serum, retinal vascular endothelial cells (rVECs), and RPE, but not the NSR. The BRB remained intact. Retinal degeneration did not occur. Conclusions: Following injection of high-dose IP FeDex, iron accumulated in the BRB, but not the NSR. Thus, the BRB can shield the NSR from iron delivered in this manner. This ability is not dependent on NSR hepcidin production.


Assuntos
Barreira Hematorretiniana/metabolismo , Células Endoteliais/metabolismo , Sobrecarga de Ferro/metabolismo , Complexo Ferro-Dextran/administração & dosagem , Ferro/metabolismo , Vasos Retinianos/metabolismo , Albuminas/metabolismo , Animais , Modelos Animais de Doenças , Ferritinas/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Hepcidinas/farmacologia , Injeções Intraperitoneais , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Transferrina/metabolismo
8.
Mem Inst Oswaldo Cruz ; 114: e190253, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31664313

RESUMO

BACKGROUND: Timely diagnosis is recommended by the Brazilian Visceral Leishmaniasis (VL) Surveillance and Control Program to reduce case fatality. Attempts at assessing this topic in Brazil are scarce. OBJECTIVE: This study aimed to describe where, when, and how the diagnosis of VL has been performed in a Brazilian endemic setting. METHODS: Data of all autochthonous cases confirmed between 2011 and 2016 (N = 81) were recorded. The care-seeking itinerary until the confirmation of VL diagnosis was assessed among 57 patients. FINDINGS: The majority of VL cases (79.1%) were reported by referral hospitals. The patients mainly sought primary health care centres at the onset of symptoms. However, they had to visit seven health services on average to achieve a confirmed diagnosis. The time from the onset of symptoms to the diagnosis of VL (TD) ranged from 1-212 (median, 25) days. The TD was longer among adult patients. There was a direct correlation between the patient's age and TD (r = 0.22; p = 0.047) and a higher occurrence of deaths due to the disease among older patients (p = 0.002). Almost all the patients (98.9%) underwent laboratory investigation, and the VL diagnosis was mainly confirmed based on clinical-laboratory criteria (92.6%). Positive results for the indirect fluorescence antibody test (22.7%) and parasitological examination plus rk39-based immunochromatographic tests (21.3%) were commonly employed. MAIN CONCLUSIONS: VL diagnosis was predominantly conducted in hospitals with a long TD and wide application of serology. These findings may support measures focused on early diagnosis, including a greater involvement of the primary health care system.


Assuntos
Assistência à Saúde/estatística & dados numéricos , Leishmaniose Visceral/diagnóstico , Adolescente , Adulto , Brasil/epidemiologia , Criança , Pré-Escolar , Cromatografia de Afinidade , Assistência à Saúde/classificação , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Lactente , Recém-Nascido , Leishmaniose Visceral/epidemiologia , Masculino , Estudos Retrospectivos
9.
Klin Lab Diagn ; 64(9): 546-552, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31610107

RESUMO

The aim of the study was to characterize the dynamics of immunoglobulin IgG and IgM level in syphilis patients serum at different stages of the disease before and after the therapy towards 12 diagnostic antigens of T. pallidum in an microarray assay and to evaluate these data as possible prognostic markers. The dynamics of immunoglobulin IgG and IgM level was measured in the reaction of indirect immunofluorescence using microarray and compared to the results of non-treponemal RPR test and treponemal tests as EIA and reaction of passive hemagglutination. In microarray assay diagnostically high level of IgM in patients with primary, secondary and early latent and late latent syphilis decreased dramatically to zero after the successful therapy. Continuously high level of IgM after the therapy proposes the persistence of infection agents in the organism and points out the need of additional antimicrobial treatment. In most of the cases anti-treponemal IgG level also declined after the successful therapy and this confirms the appropriate treatment. The results of microarray assay coincide with the results of other mentioned laboratory tests for syphilis diagnostics. Microarray assay with the recombinant T. pallidum antigens gives the perspective for creating methods with wider spectrum of diagnostic and therapy control options using the IgM immunoglobulin level as a marker for successful syphilis treatment.


Assuntos
Anticorpos Antibacterianos/sangue , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Sífilis/diagnóstico , Sífilis/tratamento farmacológico , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Sífilis/sangue , Sorodiagnóstico da Sífilis , Treponema pallidum
10.
Rev Bras Parasitol Vet ; 28(4): 816-820, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31618304

RESUMO

Toxoplasma gondii and Neospora caninum are Apicomplexan intracellular protozoan parasites that affect numerous animal species, thus leading to severe diseases and economic losses, depending on the vertebrate species involved. The role of the avian species in maintaining and transmission of these coccidia has been studied for several years as they tend to serve as a potential source of infection for mammals and humans. The present study aimed to assess the serological exposure of Orinoco goose (Neochen jubata) to T. gondii and N. caninum. Between 2010 and 2013, 41 free-ranging Orinoco geese were captured in the Araguaia River, Brazil. The presence and titration of IgY antibodies to both coccidia were assayed via indirect immunofluorescent antibody test (IFAT). While IgY antibodies for N. caninum were present in 5 animals, with titers of 20, the antibodies for T. gondii were found in 35 animals, with titers ranging from 20 to 640. Considering that the Orinoco goose's meat is consumed by the local population in the studied area, it may represent an important source of T. gondii infection for humans. Due to its migratory behavior, this goose may play a pivotal role in the natural dispersion of both parasites. Furthermore, molecular studies are required for genotyping the isolates of T. gondii that occurs in this avian species.


Assuntos
Anticorpos Antiprotozoários/sangue , Doenças das Aves/diagnóstico , Coccidiose/veterinária , Gansos/parasitologia , Neospora/imunologia , Toxoplasma/imunologia , Toxoplasmose Animal/diagnóstico , Animais , Doenças das Aves/epidemiologia , Doenças das Aves/parasitologia , Brasil/epidemiologia , Coccidiose/diagnóstico , Coccidiose/epidemiologia , Coccidiose/parasitologia , Técnica Indireta de Fluorescência para Anticorpo , Toxoplasmose Animal/epidemiologia , Toxoplasmose Animal/parasitologia
11.
Invest Ophthalmol Vis Sci ; 60(13): 4416-4424, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31639828

RESUMO

Purpose: Lenses have an intracellular hydrostatic pressure gradient to drive fluid from central fiber cells to surface epithelial cells. Pressure is regulated by a feedback control system that relies on transient receptor potential vanilloid (TRPV)1 and TRPV4 channels. The ciliary muscle transmits tension to the lens through the zonules of Zinn. Here, we have examined if ciliary muscle tension influenced the lens intracellular hydrostatic pressure gradient. Methods: We measured the ciliary body position and intracellular hydrostatic pressures in mouse lenses while pharmacologically causing relaxation or contraction of the ciliary muscle. We also used inhibitors of TRPV1 and TRPV4, in addition to phosphoinositide 3-kinase (PI3K) p110α knockout mice and immunostaining of phosphorylated protein kinase B (Akt), to determine how changes in ciliary muscle tension resulted in altered hydrostatic pressure. Results: Ciliary muscle relaxation increased the distance between the ciliary body and the lens and caused a decrease in intracellular hydrostatic pressure that was dependent on intact zonules and could be blocked by inhibition of TRPV4. Ciliary contraction moved the ciliary body toward the lens and caused an increase in intracellular hydrostatic pressure and Akt phosphorylation that required intact zonules and was blocked by either inhibition of TRPV1 or genetic deletion of the p110α catalytic subunit of PI3K. Conclusions: These results show that the hydrostatic pressure gradient within the lens was influenced by the tension exerted on the lens by the ciliary muscle through the zonules of Zinn. Modulation of the gradient of intracellular hydrostatic pressure in the lens could alter the water content, and the gradient of refractive index.


Assuntos
Corpo Ciliar/metabolismo , Cristalino/metabolismo , Ligamentos/metabolismo , Músculo Liso/metabolismo , Canais de Cátion TRPV/metabolismo , Animais , Técnica Indireta de Fluorescência para Anticorpo , Pressão Hidrostática , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Midriáticos/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Pilocarpina/farmacologia , Canais de Cátion TRPV/antagonistas & inibidores , Tropicamida/farmacologia
12.
Actas dermo-sifiliogr. (Ed. impr.) ; 110(8): 673-680, oct. 2019. tab, ilus
Artigo em Espanhol | IBECS | ID: ibc-185506

RESUMO

La dermatosis ampollar IgA lineal es una enfermedad vesicoampollar subepidérmica, adquirida, mediada por inmunoglobulinas. Presentamos nuestra serie con el objetivo de describir las características clínicas, evolución y tratamientos instaurados. Se realizó un estudio descriptivo, observacional retrospectivo. Se incluyeron 17 pacientes. Como antecedentes 2 niños recibieron vacunas 2 semanas antes del inicio de los síntomas; en 2 casos la enfermedad estuvo precedida por cuadros respiratorios broncoobstructivos. Un paciente recibió antibioticoterapia endovenosa antes del inicio del cuadro. Hallamos asociación con hepatitis autoinmune en un caso y con alopecia areata en otro. Un niño padecía asociación VACTERL. El diagnóstico se confirmó con histopatología e inmunofluorescencia directa. Como tratamiento 16 pacientes recibieron dapsona, 8 de ellos asociaron corticoides orales y 2 esteroides tópicos. Destacamos la presencia de rebrotes con compromiso perioral ante cuadros infecciosos e inmunizaciones, la asociación con síndrome de VACTERL y con hepatitis autoinmune


Linear IgA bullous dermatosis is an acquired subepidermal immunoglobulin-mediated vesiculobullous disease. In this retrospective, observational, descriptive study, we describe the clinical characteristics, treatments, and outcomes of 17 patients with linear IgA bullous dermatosis. Two children had been vaccinated 2 weeks before the onset of symptoms, 2 had had bronco-obstructive respiratory symptoms, and 1 had received intravenous antibiotic therapy. We also observed an association with autoimmune hepatitis in one patient and alopecia areata in another. One boy had VACTERL association. Diagnosis was confirmed by histopathology and direct immunofluorescence. Sixteen patients were treated with dapsone, which was combined with oral corticosteroids in 8 cases and topical corticosteroids in two. Of note in this series was the occurrence of relapses in the perioral area coinciding with infections and vaccination, and the association between linear IgA bullous dermatosis and autoimmune hepatitis and VACTERL association


Assuntos
Humanos , Masculino , Feminino , Lactente , Pré-Escolar , Criança , Dermatose Linear Bolhosa por IgA/diagnóstico , Dermatose Linear Bolhosa por IgA/tratamento farmacológico , Dermatoses Faciais/tratamento farmacológico , Dermatoses do Pé/tratamento farmacológico , Dermatoses da Perna/tratamento farmacológico , Epidemiologia Descritiva , Estudos Retrospectivos , Dermatose Linear Bolhosa por IgA/patologia , Dapsona/uso terapêutico , Corticosteroides/uso terapêutico , Esteroides/uso terapêutico , Administração Tópica , Técnica Indireta de Fluorescência para Anticorpo/métodos , Diagnóstico Diferencial
14.
Invest Ophthalmol Vis Sci ; 60(12): 3842-3853, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31529081

RESUMO

Purpose: Outer blood retinal barrier breakdown is a neglected feature of diabetic retinopathy (DR). We demonstrated that the agonism of the δ opioid receptor (DOR) by epicatechin preserves the tight junction proteins in ARPE-19 cells under diabetic conditions. Presently, we aimed to evaluate the possible role of the DOR on the maintenance of tight junction of RPE layer and on the early markers of experimental DR. Methods: DR markers and external retinal tight junction proteins were evaluated in CL57B diabetic mice submitted to intravitreous injection of short hairpin RNA (shRNA)-DOR (108 transducing units [TU]/mL) treated or not with DOR agonist (0.05 g/animal/d of epicatechin in drinking water) for 16 weeks. The presence of DOR in human retina from postmortem eyes from diabetic and nondiabetic donors were also performed. Results: DOR is present in RPE layer and in neuro retina. The treatment with DOR agonist prevented the upregulation of the early markers of retinopathy (glial fibrillary acidic protein, VEGF) and the downregulation of pigment epithelium-derived factor, occludin, claudin-1, and zonula occludens-1 tight junction expressions. The silencing of DOR in retina of diabetic mice partially abolished the protective effects of epicatechin. In human retina specimens, DOR is present throughout the retina, similarly in nondiabetic and diabetic donors. Conclusions: This set of experiments strongly indicates that the DOR agonism preserves RPE tight junctions and reduces the early markers of retinopathy in model of diabetes. These novel findings designate DOR as a potential therapeutic tool to treat DR with preservation of the RPE tight junction proteins.


Assuntos
Catequina/farmacologia , Diabetes Mellitus Experimental/prevenção & controle , Retinopatia Diabética/prevenção & controle , Receptores Opioides delta/agonistas , Epitélio Pigmentado da Retina/metabolismo , Junções Íntimas/metabolismo , Idoso , Animais , Glicemia/metabolismo , Western Blotting , Claudina-1/metabolismo , Diabetes Mellitus Experimental/metabolismo , Retinopatia Diabética/metabolismo , Retinopatia Diabética/patologia , Impedância Elétrica , Proteínas do Olho/metabolismo , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Fatores de Crescimento Neural/metabolismo , Ocludina/metabolismo , RNA Interferente Pequeno , Reação em Cadeia da Polimerase em Tempo Real , Receptores Opioides delta/metabolismo , Serpinas/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
15.
Invest Ophthalmol Vis Sci ; 60(12): 3718-3726, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31479112

RESUMO

Purpose: To evaluate the effects of human amniotic membrane-derived fibroblast (AMF) cell supernatant (AMF-sup) on corneal epithelium. Methods: The phenotype of AMF cells was analyzed by flow cytometry using cell-surface markers. AMF cells were also induced to form osteoblasts and neural cells, and cell phenotypes were observed by staining and RT-PCR. Cultivated human corneal limbal epithelial sheets generated using AMF-sup were analyzed using immunohistochemistry and colony-forming efficiency, and the wound healing of epithelial defects was observed using a tissue-punch method. The effects of instillation of each supernatant in a rabbit corneal epithelial wound healing model were compared. Results: Mesenchymal stem cell (CD29, CD44, CD73, and CD90) and neural crest (CD49d and CD56) markers were expressed on the AMF cell surface. Following induction of differentiation, isolated AMF cells showed characteristics of osteoblasts and neural cells. Application of AMF-sup resulted in maintenance of the limbal epithelial phenotype and immature state, and significantly promoted wound healing in cultivated human corneal limbal epithelial sheets (P < 0.05) and rabbit corneal epithelium (P < 0.05) compared with the control. Conclusions: These data suggest that AMF cells have multi-differentiation potential, and that AMF-sup is effective in maintaining the limbal epithelial phenotype and promoting corneal epithelial wound healing, which may be of value in ocular surface reconstruction.


Assuntos
Âmnio/citologia , Fatores Biológicos/farmacologia , Lesões da Córnea/tratamento farmacológico , Epitélio Anterior/efeitos dos fármacos , Fibroblastos/fisiologia , Animais , Biomarcadores/metabolismo , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Lesões da Córnea/metabolismo , Lesões da Córnea/patologia , Epitélio Anterior/metabolismo , Epitélio Anterior/patologia , Feminino , Citometria de Fluxo , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Limbo da Córnea/citologia , Camundongos , Células NIH 3T3 , Fenótipo , Coelhos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Cicatrização/fisiologia
16.
Invest Ophthalmol Vis Sci ; 60(12): 3696-3707, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31487370

RESUMO

Purpose: We assess if α retinal ganglion cells (αRGCs) and intrinsically photosensitive retinal ganglion cells (ipRGCs) survive in mouse models of glaucoma. Methods: Two microliters of N-methyl-D-aspartate (NMDA; 1 mM) or PBS were injected intraocularly 7 days before sacrifice. Immunohistochemical analyses of the retina were performed using antibodies against RNA-binding protein with multiple splicing (RBPMS), osteopontin, and melanopsin. Immunohistochemical analyses also were performed in adult mice with glutamate/aspartate transporter (GLAST) deletion (GLAST knockout [KO] mice), a mouse model of normal tension glaucoma. Results: NMDA-induced loss of RBPMS-positive total RGCs was 58.4% ± 0.4% compared to PBS-treated controls, whereas the loss of osteopontin-positive αRGCs was 5.0% ± 0.6% and that of melanopsin-positive ipRGCs was 7.6% ± 1.6%. In GLAST KO mice, the loss of total RGCs was 48.4% ± 0.9% compared to wild-type mice, whereas the loss of αRGCs and ipRGCs was 3.9% ± 0.4% and 9.3% ± 0.5%, respectively. The distribution of survived total RGCs, αRGCs, and ipRGCs was similar regardless of the location of the retina. Conclusions: These results suggest that αRGC and ipRGC are highly tolerant to NMDA-induced neurotoxicity and NTG-like neurodegeneration in GLAST KO mice.


Assuntos
Modelos Animais de Doenças , Agonistas de Aminoácidos Excitatórios/toxicidade , Glaucoma de Baixa Tensão/prevenção & controle , N-Metilaspartato/toxicidade , Disco Óptico/efeitos dos fármacos , Células Ganglionares da Retina/efeitos dos fármacos , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Animais , Sobrevivência Celular/fisiologia , Técnica Indireta de Fluorescência para Anticorpo , Injeções Intravítreas , Luz , Glaucoma de Baixa Tensão/metabolismo , Glaucoma de Baixa Tensão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia de Fluorescência , Disco Óptico/metabolismo , Disco Óptico/efeitos da radiação , Osteopontina/metabolismo , Proteínas de Ligação a RNA/metabolismo , Degeneração Retiniana/metabolismo , Degeneração Retiniana/patologia , Degeneração Retiniana/prevenção & controle , Células Ganglionares da Retina/classificação , Células Ganglionares da Retina/metabolismo , Células Ganglionares da Retina/efeitos da radiação
17.
Invest Ophthalmol Vis Sci ; 60(12): 3776-3785, 2019 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-31503282

RESUMO

Purpose: To investigate the therapeutic effects of targeting signal transducer and activator of transcription-3 (STAT3) activation on the ocular surface damage of dry eye in mice. Methods: Adult Balb/C and C57BL/6 mice with benzalkonium chloride (BAC) treatment, lacrimal gland excision, and meibomian gland dysfunction were used as dry eye models. The levels of phosphorylated STAT3 (p-STAT3) were detected with immunofluorescence staining and Western blotting. STAT3 inhibition was performed by topical application of STAT3 inhibitor S3I-201. Corneal epithelial barrier function, tear production, and conjunctival goblet cell density were quantified with fluorescein sodium staining, phenol red cotton test, and histochemical staining. The expressions of matrix metalloproteinase (MMP)-3/9, TUNEL, and inflammation cytokines were assessed with immunofluorescence staining, qPCR, and ELISA assays. The therapeutic effect of S3I-201 was further compared with the Janus kinase inhibitor tofacitinib and ruxolitinib. Results: Elevated levels of nuclear p-STAT3 were detected in the corneal and conjunctival epithelium of three dry eye models. Topical application of S3I-201 improved corneal epithelial barrier function, increased tear production and conjunctival goblet cell density in BAC-induced dry eye mice. Moreover, S3I-201 decreased the expression of MMP-3/9, suppressed the apoptosis of corneal and conjunctival epithelial cells, and reduced the levels of IL-1ß, IL-6, IL-17A, and IFN-γ. Compared with tofacitinib and ruxolitinib, the STAT3 inhibitor S3I-201 showed superior improvement of tear production and inflammatory cytokine expression in lacrimal gland. Conclusions: Elevated STAT3 activation is involved in the pathogenesis of dry eye, while targeting STAT3 effectively alleviates BAC-induced ocular surface damage.


Assuntos
Modelos Animais de Doenças , Síndromes do Olho Seco/tratamento farmacológico , Proteínas Inibidoras de STAT Ativados/uso terapêutico , Fator de Transcrição STAT3/antagonistas & inibidores , Administração Oftálmica , Animais , Western Blotting , Túnica Conjuntiva/metabolismo , Citocinas/metabolismo , Síndromes do Olho Seco/metabolismo , Síndromes do Olho Seco/patologia , Ensaio de Imunoadsorção Enzimática , Epitélio/metabolismo , Epitélio Anterior/metabolismo , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Células Caliciformes/patologia , Marcação In Situ das Extremidades Cortadas , Metaloproteinase 3 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Soluções Oftálmicas , Fosforilação , Reação em Cadeia da Polimerase em Tempo Real , Fator de Transcrição STAT3/metabolismo , Lágrimas/fisiologia
18.
Ann Clin Lab Sci ; 49(4): 496-502, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31471339

RESUMO

GOALS: The existence of anti-dense fine speckled (DFS) 70 autoantibodies are considered an exclusionary biomarker for systemic autoimmune rheumatic diseases (SARDs). Several tests to confirm the presence of anti-DFS70 autoantibodies have been introduced, and the use of them in specimens with a DFS pattern in indirect immunofluorescence-antinuclear antibody (IIF-ANA) testing has been suggested. However, these additional tests have only been evaluated in a small number of samples; their clinical usefulness therefore requires further evaluation. METHODS: A total of 213 serum specimens showing DFS (n=155) or homogeneous (H; n=58) patterns were included. All specimens were tested by western blotting (WB) and enzyme immunoassay (EIA). Clinical information regarding SARDs was analyzed. RESULTS: The detection rates for WB and EIA for anti-DFS70 autoantibodies in specimens with a DFS pattern were 86.5% and 73.5%, respectively. Detection rates in specimens with a low IIF-ANA titer were significantly lower than those in specimens with a high titer. The detection rate of anti-DFS70 autoantibodies in 58 specimens with an H pattern was 10.3% (6/58). Among 155 subjects with a DFS pattern in IIF-ANA staining, only five were diagnosed with SARD. CONCLUSIONS: There is little need to confirm the presence of anti-DFS70 autoantibodies using other methods. When a DFS pattern is observed in IIF-ANA staining, it is more important to confirm the presence of other autoantibodies related to SARDs than to identify anti-DFS70 autoantibodies. Finally, more careful interpretation of IIF-ANA to specimens with a low IIF-ANA titer is needed.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/imunologia , Anticorpos Antinucleares/sangue , Autoanticorpos/sangue , Programas de Rastreamento , Fatores de Transcrição/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Doenças Autoimunes/sangue , Doenças Autoimunes/diagnóstico , Doenças Autoimunes/imunologia , Criança , Pré-Escolar , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem
19.
Invest Ophthalmol Vis Sci ; 60(10): 3538-3546, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31415077

RESUMO

Purpose: To determine if trigeminal innervations of the corneal epithelium maintains its integrity and homeostasis through controlling the nicotinamide adenine dinucleotide (NAD) content of this tissue. Methods: Corneal denervation of C57BL/6 mice was induced by squeezing the nerve bundles that derive from the trigeminal ganglion and was confirmed by whole-mount corneal nerve staining and the sensation test. The apoptosis of the corneal epithelium was examined by TUNEL assay and annexin V/propidium iodide staining. NAD biosynthesis-related enzymes were analyzed by quantitative PCR, immunofluorescence staining, and Western blotting. FK866, an inhibitor of nicotinamide phosphoribosyltransferase (NAMPT), exogenous nicotinamide mononucleotide (NMN), and NAD+ were used to evaluate the effect of NAD+ on the apoptosis of cultured corneal epithelial cells and epithelial detachment in denervated mice. Protein expression that related to apoptosis and phosphorylation were analyzed by Western blotting. Results: The denervated mice showed spontaneous corneal epithelial detachment and cell apoptosis accompanied with impaired epithelial NAD+ contents due to low levels of NAMPT. Similarly, inhibition of NAMPT recapitulated epithelial detachment as in denervated mice and induced apoptosis in cultured corneal epithelial cells. The replenishment of NMN or NAD+ partially slowed down corneal nerve fiber degeneration, reduced the epithelial defect in denervated mice, and improved apoptosis induction in FK866-treated cells by restoring the activation levels of SIRT1, AKT, and CREB. Conclusions: Corneal denervation lowered epithelial NAD+ contents through reducing the expression of NAMPT and caused cell apoptosis and epithelial defects, suggesting that corneal innervations contribute to epithelial homeostasis by regulating NAD+ biosynthesis.


Assuntos
Apoptose , Córnea/inervação , Denervação , Epitélio Anterior/patologia , NAD/metabolismo , Nervo Trigêmeo/fisiologia , Animais , Anexina A5/metabolismo , Western Blotting , Proteína de Ligação a CREB/metabolismo , Córnea/metabolismo , Doenças da Córnea/diagnóstico , Doenças da Córnea/metabolismo , Epitélio Anterior/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Marcação In Situ das Extremidades Cortadas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Sirtuína 1/metabolismo , Doenças do Nervo Trigêmeo/diagnóstico , Doenças do Nervo Trigêmeo/metabolismo , Proteína X Associada a bcl-2/metabolismo
20.
J Fish Dis ; 42(10): 1447-1455, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31418903

RESUMO

Vibrio ordalii is an extracellular, Gram-negative bacterium that produces vibriosis in salmonids. While pathogenesis is not fully understood, this bacterium has numerous likely genes for adhesion, colonization, invasion factors and, as recently suggested, intracellular behaviour. Therefore, this study aimed to clarify possible intracellular behaviour for V. ordalii Vo-LM-18 and ATCC 33509T in the fish-cell lines SHK-1 and CHSE-214. Confocal microscopy revealed Vo-LM-18 and ATCC 33509T inside cytoplasm in both fish-cell lines at 4 hr post-inoculation (hpi). At 8 and 16 hpi, the proportion of fish cells invaded by both strains increased. Moreover, intracellular V. ordalii were observed after 8 hpi inside mouse embryonic fibroblasts (MEF), demonstrating that entry was not due to a cellular phagocytosis process. Flow cytometry confirmed immunocytochemistry results, with both V. ordalii evidencing statistically significant differences in the number of infected cells between 8 and 16 hpi. Interestingly, V. ordalii infection did not significantly damage fish cells, as determined by LDH liberation. Viable counts at 8 hpi detected, on average for both lines, 176 ± 47 CFU/ml of culturable intracellular Vo-LM-18 and ATCC 33509T cells. These in vitro findings support the facultative intracellular behaviour of V. ordalii and may be of importance for understanding pathogenicity and survival in aquatic environments.


Assuntos
Doenças dos Peixes/microbiologia , Salmão , Vibrioses/veterinária , Vibrio/fisiologia , Animais , Linhagem Celular , Citometria de Fluxo/veterinária , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Microscopia Confocal/veterinária , Vibrioses/microbiologia
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