Mediators of Inflammation / 2017 / Article / Fig 6

Research Article

Molecular Responses of Human Retinal Cells to Infection with Dengue Virus

Figure 6

Effect of DENV infection on barrier function of human retinal pigment epithelial cells and human retinal endothelial cells: viral strain = Mon601; multiplicity of infection = 1; evaluated up to 60 hours (hr) postinoculation. (a) and (d) Plots of electrical resistance across (a) DENV-infected retinal pigment epithelial monolayers and (d) DENV-infected endothelial monolayers versus mock-infected monolayers, measured at hourly intervals and expressed as transcellular resistance by xCELLigence. Lines indicate mean electrical resistance: blue lines represent DENV-infected monolayers and red lines represent mock-infected monolayers. Error bars indicate standard error of mean. monolayers/condition. Arrowheads mark time of inoculation and 24 and 48 hours postinoculation. Drop in resistance immediately after inoculation with virus is due to removal of plate from xCELLigence for inoculation procedure. (b) DENV- and mock-infected retinal pigment epithelial cells immunolabeled to detect double-stranded RNA (dsRNA) and filamentous- (F-) actin. Alexa Fluor 555 (red) and Alexa Fluor 488 (green) with Hoechst 33342 nuclear counterstain (blue). Images are merged z-stacks. Original magnification: 630x. (c) and (e) Graphs showing relative transcript expression for junctional molecules in (c) DENV-infected retinal pigment epithelial cells and (e) DENV-infected endothelial cells versus mock-infected cells. Reference genes were glyceraldehyde-3-phosphate dehydrogenase and TATA-binding protein. Bars represent mean relative expression, with error bars showing standard deviation. cultures/condition. Data were analyzed by two-tailed Student’s -test. ZO-1 = zonula occludens 1; CLDN5 = claudin 5; OCLN = occludin; JAM-3 = junctional adhesion molecule 3; CTNNB1 = catenin-β1. CLDN5 and VE-cadherin were not detected in epithelial cells.

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