Effect of Methotrexate on an In Vitro Patient-Derived Model of Proliferative Vitreoretinopathy
| Autor: | Joseph F. Arboleda-Velasquez, Dhanesh Amarnani, Christina K. Marko, James A. Stefater, Tomasz P. Stryjewski, Lindsay L. Wong, Dean Eliott, Leo A. Kim, Arturo Israel Machuca-Parra |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Adult Male Cell type Proliferative vitreoretinopathy Programmed cell death Cell PVR Cell Culture Techniques Apoptosis Cell Separation Retinal Pigment Epithelium Models Biological Retina methotrexate proliferative vitreoretinopathy 03 medical and health sciences retinal detachment 0302 clinical medicine Cell Movement medicine Humans Fluorescent Antibody Technique Indirect Aged Cell Proliferation Extracellular Matrix Proteins Chemistry Tumor Necrosis Factor-alpha Vitreoretinopathy Proliferative Cell migration Epiretinal Membrane Middle Aged medicine.disease Molecular biology In vitro eye diseases 030104 developmental biology medicine.anatomical_structure Phenotype Cell culture 030221 ophthalmology & optometry Female sense organs Biomarkers Immunosuppressive Agents |
| Zdroj: | Investigative Ophthalmology & Visual Science |
| ISSN: | 1552-5783 0146-0404 |
| Popis: | Purpose The purpose of this study was to develop a method for isolating, culturing, and characterizing cells from patient-derived membranes in proliferative vitreoretinopathy (PVR) to be used for drug testing. Methods PVR membranes were obtained from six patients with grade C PVR. Membrane fragments were analyzed by gross evaluation, fixed for immunohistologic studies to establish cell identity, or digested with collagenase II to obtain single cell suspensions for culture. PVR-derived primary cultures were used to examine the effects of methotrexate (MTX) on proliferation, migration, and cell death. Results Gross analysis of PVR membranes showed presence of pigmented cells, indicative of retinal pigment epithelial cells. Immunohistochemistry identified cells expressing α-smooth muscle actin, glial fibrillary acidic protein, Bestrophin-1, and F4/80, suggesting the presence of multiple cell types in PVR. Robust PVR primary cultures (C-PVR) were successfully obtained from human membranes, and these cells retained the expression of cell identity markers in culture. C-PVR cultures formed membranes and band-like structures in culture reminiscent of the human condition. MTX significantly reduced the proliferation and band formation of C-PVR, whereas it had no significant effect on cell migration. MTX also induced regulated cell death within C-PVR as assessed by increased expression of caspase-3/7. Conclusions PVR cells obtained from human membranes can be successfully isolated, cultured, and profiled in vitro. Using these primary cultures, we identified MTX as capable of significantly reducing growth and inducing cell death of PVR cells in vitro. |
| Databáze: | OpenAIRE |
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