Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
1.
Toxicol Pathol ; 47(4): 528-541, 2019 06.
Artículo en Inglés | MEDLINE | ID: mdl-31064296

RESUMEN

In a juvenile toxicology program, an unexpected finding of vacuolation of inner nuclear, ganglion cell, and nerve fiber layers of the retina was observed microscopically in routine Davidson's fixed and hematoxylin and eosin-stained tissue sections of eyes in beagle dogs at approximately 5 weeks of age. There was no necrosis or degeneration of the affected cells and no associated inflammation. Fluorescein angiography revealed no vascular leakage. Optical coherence tomography (OCT) indicated swollen cells in the same layers of the retina as observed at light microscopic examination. Transmission electron microscopy revealed that the retinal vacuolation likely was consistent with intracellular swelling of amacrine, horizontal, and/or bipolar cells of the inner nuclear layer as affected cells had an expanded cytoplasm but contained normal nucleus and organelles. As assessed by animal behavior and full-field electroretinography, the retinal vacuolation appeared to have no impact on visual function. Retinal vacuolation was seen in approximately 40% of dogs at 5 weeks of age using OCT and/or light microscopic examination. Because the change was transient and age related, did not result in degenerative retinal changes, and was not present in dogs older than 5 weeks of age, it was considered a background developmental observation in beagle dogs.


Asunto(s)
Retina/crecimiento & desarrollo , Toxicología/métodos , Vacuolas/fisiología , Factores de Edad , Animales , Animales Recién Nacidos , Perros , Angiografía con Fluoresceína , Microscopía , Microscopía Electrónica de Transmisión , Retina/diagnóstico por imagen , Tomografía de Coherencia Óptica
2.
Apoptosis ; 20(12): 1613-22, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26404525

RESUMEN

Retinoblastoma (Rb) is an aggressive childhood cancer of the developing retina. This disease is associated with epigenetic deregulation of several cancer pathways including upregulation of the proto-oncogene spleen tyrosine kinase (SYK). We have previously demonstrated that lymphocyte-derived microparticles (LMPs) possess strong cytotoxic effect on cancer cells. This report demonstrated that LMPs have potent pro-apoptotic properties on human Rb cells, which was associated with a strong reduction of SYK expression. Perturbing SYK activity in Rb cells induced cell apoptosis and upregulated expression of p53 and p21. Interestingly, inhibition of p53 or knockdown of p21, abolished LMP-induced caspase-3 activity and cell death. Blocking oxidized phospholipid-rich LMPs with a specific antibody significantly prevented LMP-induced Rb apoptosis and reversed the expression patterns of SYK, p53, p21. In summary, our results suggest that LMPs are important pro-apoptotic regulators for Rb cells through reduction of SYK expression and upregulation of the p53-p21 pathway which ultimately activates caspase-3. These data may open unexpected avenues for the development of LMPs as a novel therapeutic strategy that would be particularly useful and relevant for the treatment of Rb.


Asunto(s)
Apoptosis/genética , Micropartículas Derivadas de Células/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Linfocitos/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Retinoblastoma/metabolismo , Caspasa 3/metabolismo , Línea Celular , Línea Celular Tumoral , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Epigénesis Genética/genética , Humanos , Proto-Oncogenes Mas , Quinasa Syk , Proteína p53 Supresora de Tumor/metabolismo , Regulación hacia Arriba/genética
3.
PLoS One ; 15(8): e0237858, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32822382

RESUMEN

The main objective of the study was to analyze deviations in retinal nerve fiber layer (RNFL) thickness measurements caused by the displacement of circular optic disc optical coherence tomography scans. High-density radial scans of the optic nerve heads of cynomolgus monkeys were acquired. The retinal nerve fiber layer was manually segmented, and a surface plot of the discrete coordinates was generated. From this plot, the RNFL thicknesses were calculated and compared between accurately centered and intentionally displaced circle scans. Circle scan displacement caused circumpapillary retinal nerve fiber layer thickness deviations of increasing magnitude with increasing center offset. As opposed to the human eye, horizontal displacement resulted in larger RNFL thickness deviations than vertical displacement in cynomolgus monkeys. Acquisition of high-density radial scans allowed for the mathematical reconstruction and modelling of the nerve fiber layer and extrapolation of its thickness. Accurate and strictly repeatable circle scan placement is critical to obtain reproducible values, which is essential for longitudinal studies.


Asunto(s)
Disco Óptico/diagnóstico por imagen , Tomografía de Coherencia Óptica/métodos , Animales , Femenino , Macaca fascicularis , Masculino , Modelos Biológicos , Fibras Nerviosas/ultraestructura , Disco Óptico/anatomía & histología , Retina
4.
Mol Ther Methods Clin Dev ; 13: 1-13, 2019 Jun 14.
Artículo en Inglés | MEDLINE | ID: mdl-30581889

RESUMEN

Hereditary hearing loss often results from mutation of genes expressed by cochlear hair cells. Gene addition using AAV vectors has shown some efficacy in mouse models, but clinical application requires two additional advances. First, new AAV capsids must mediate efficient transgene expression in both inner and outer hair cells of the cochlea. Second, to have the best chance of clinical translation, these new vectors must also transduce hair cells in non-human primates. Here, we show that an AAV9 capsid variant, PHP.B, produces efficient transgene expression of a GFP reporter in both inner and outer hair cells of neonatal mice. We show also that AAV9-PHP.B mediates almost complete transduction of inner and outer HCs in a non-human primate. In a mouse model of Usher syndrome type 3A deafness (gene CLRN1), we use AAV9-PHP.B encoding Clrn1 to partially rescue hearing. Thus, we have identified a vector with promise for clinical treatment of hereditary hearing disorders, and we demonstrate, for the first time, viral transduction of the inner ear of a primate with an AAV vector.

5.
Sci Rep ; 6: 37391, 2016 11 22.
Artículo en Inglés | MEDLINE | ID: mdl-27874077

RESUMEN

Pathological choroidal neovascularization (CNV) is the common cause of vision loss in patients with age-related macular degeneration (AMD). Macrophages possess potential angiogenic function in CNV. We have demonstrated that human T lymphocyte-derived microparticles (LMPs) exert a potent antiangiogenic effect in several pathological neovascularization models. In this study, we investigated the alteration of proangiogenic properties of macrophages by LMPs treatment in vitro and in vivo models. LMPs regulated the expression of several angiogenesis-related factors in macrophages and consequently stimulated their antiangiogenic effects evidenced by the suppression of the proliferation of human retinal endothelial cells in co-culture experiments. The involvement of CD36 receptor in LMPs uptake by macrophages was demonstrated by in vitro assays and by immunostaining of choroidal flat mounts. In addition, ex vivo experiments showed that CD36 mediates the antiangiogenic effect of LMPs in murine and human choroidal explants. Furthermore, intravitreal injection of LMPs in the mouse model of laser-induced CNV significantly suppressed CNV in CD36 dependent manner. The results of this study suggested an ability of LMPs to alter the gene expression pattern of angiogenesis-related factors in macrophages, which provide important information for a new therapeutic approach for efficiently interfering with both vascular and extravascular components of CNV.


Asunto(s)
Micropartículas Derivadas de Células/metabolismo , Neovascularización Coroidal/patología , Linfocitos/metabolismo , Macrófagos/metabolismo , Neovascularización Fisiológica , Animales , Biomarcadores/metabolismo , Antígenos CD36/metabolismo , Polaridad Celular , Proliferación Celular , Regulación de la Expresión Génica , Humanos , Rayos Láser , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células RAW 264.7
6.
Invest Ophthalmol Vis Sci ; 54(6): 4061-71, 2013 Jun 12.
Artículo en Inglés | MEDLINE | ID: mdl-23620430

RESUMEN

PURPOSE: StarGen is an equine infectious anemia virus (EIAV)-based lentiviral vector that expresses the photoreceptor-specific adenosine triphosphate (ATP)-binding cassette transporter (ABCA4) protein that is mutated in Stargardt disease (STGD1), a juvenile macular dystrophy. EIAV vectors are able to efficiently transduce rod and cone photoreceptors in addition to retinal pigment epithelium in the adult macaque and rabbit retina following subretinal delivery. The safety and biodistribution of StarGen following subretinal delivery in macaques and rabbits was assessed. METHODS: Regular ophthalmic examinations, IOP measurements, ERG responses, and histopathology were carried out in both species to compare control and vector-treated eyes. Tissue and fluid samples were obtained to evaluate the persistence, biodistribution, and shedding of the vector following subretinal delivery. RESULTS: Ophthalmic examinations revealed a slightly higher level of inflammation in StarGen compared with control treated eyes in both species. However, inflammation was transient and no overt toxicity was observed in StarGen treated eyes and there were no abnormal clinical findings. There was no StarGen-associated rise in IOP or abnormal ERG response in either rabbits or macaques. Histopathologic examination of the eyes did not reveal any detrimental changes resulting from subretinal administration of StarGen. Although antibodies to StarGen vector components were detected in rabbit but not macaque serum, this immunologic response did not result in any long-term toxicity. Biodistribution analysis demonstrated that the StarGen vector was restricted to the ocular compartment. CONCLUSIONS: In summary, these studies demonstrate StarGen to be well tolerated and localized following subretinal administration.


Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Vectores Genéticos , Virus de la Anemia Infecciosa Equina/genética , Degeneración Macular/congénito , Células Fotorreceptoras de Vertebrados/metabolismo , Transducción Genética , Animales , Western Blotting , Líquidos Corporales/metabolismo , Citomegalovirus/genética , Electrorretinografía , Ensayo de Inmunoadsorción Enzimática , Femenino , Expresión Génica , Terapia Genética , Proteínas Fluorescentes Verdes/genética , Presión Intraocular , Macaca mulatta , Degeneración Macular/genética , Degeneración Macular/metabolismo , Degeneración Macular/fisiopatología , Masculino , Reacción en Cadena de la Polimerasa , Conejos , Enfermedad de Stargardt , Distribución Tisular , Transfección
7.
Invest Ophthalmol Vis Sci ; 52(9): 6949-58, 2011 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-21791590

RESUMEN

PURPOSE: The pharmacokinetics of ophthalmic biotherapeutics are difficult to determine in human vitreous humor. Because of the high transparency of living tissue to near-infrared (NIR) light, the temporal changes in vitreous concentrations of a biomolecule labeled with an NIR fluorescent probe can be monitored in situ with a scanning laser ophthalmoscope (SLO). METHODS: A humanized IgG was labeled with the NIR probe IRDye800CW (CVX-4164). Rabbits were given CVX-4164 intravitreally, and NIR fluorescence intensity was measured in the central plane of the vitreous humor with an SLO. Fluorescence intensities were converted to concentrations by using standard curves. RESULTS: Little background fluorescence was detected, and the minimum detectable concentration of CVX-4164 was <10 nM. Vitreal concentrations of CVX-4164 determined in situ declined with time, with C(max) ≈ 1 µM and t½ = 145 hours (112-µg dose). The t½ of CVX-4164 was approximately three times greater than that of the IRDye800CW alone, whereas the vitreal clearance (CL) and volume of distribution (V(ss)) of the native dye were approximately 2000- and 550-fold greater than that of the conjugate. CVX-4164 concentrations determined in situ were 2.6 to 4.4 times higher than those determined by ex vivo NIR fluorescence or ELISA in homogenized vitreous humor, reflecting the greater spatial resolution of in situ imaging. Moreover, vitreal concentrations determined in situ were >3 orders of magnitude greater than plasma concentrations of CVX-4164, as determined by ELISA, and had a different kinetic profile. CONCLUSIONS: This study demonstrates the feasibility of determining the pharmacokinetics of intraocular biotherapeutics labeled with NIR fluorescent probes by in situ monitoring.


Asunto(s)
Colorantes Fluorescentes/farmacocinética , Inmunoglobulina G/metabolismo , Indoles/farmacocinética , Oftalmoscopía/métodos , Espectroscopía Infrarroja Corta/métodos , Cuerpo Vítreo/metabolismo , Animales , Ensayo de Inmunoadsorción Enzimática , Semivida , Rayos Láser , Masculino , Conejos , Espectrometría de Fluorescencia , Distribución Tisular
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA