Xiao Yang1, Harry A Scott1, Soroush Ardekani1, Monique Williams2, Prue Talbot3, Kaustabh Ghosh4. 1. Department of Bioengineering, University of California-Riverside, Riverside, California, United States. 2. Department of Cell Biology and Neuroscience, University of California-Riverside, Riverside, California, United States. 3. Department of Cell Biology and Neuroscience, University of California-Riverside, Riverside, California, United States Stem Cell Center, University of California-Riverside, Riverside, California, United States. 4. Department of Bioengineering, University of California-Riverside, Riverside, California, United States Stem Cell Center, University of California-Riverside, Riverside, California, United States.
Abstract
PURPOSE: Environmental tobacco smoke (ETS) is widely regarded as a major modifiable risk factor for age-related macular degeneration (AMD). Yet, precisely how it exerts its pathologic effects is poorly understood. Since early-stage AMD is characterized by choroidal capillary loss, this study examined the effect of sidestream smoke (SS), the major component of ETS, on the viability of choroidal endothelial cells (EC), with an emphasis on the role of aberrant cell and basement membrane (BM) architecture in mediating SS-induced response. METHODS: Chorioretinal ECs (RF/6A) were treated with SS, and cell viability and architecture were analyzed by colorimetric assay and actin cytoskeletal organization, respectively. The structure of RF/6A EC-secreted BM was examined by immunofluorescence for collagen IV and immunoblotting for lysyl oxidase (LOX), a collagen-crosslinking enzyme. Finally, fresh RF/6A ECs were cultured on decellularized SS-treated BM to evaluate its active role in EC dysfunction. RESULTS: The RF/6A EC viability decreased progressively with increasing SS dose, which correlated strongly with a significant decline in actin cytoskeleton-dependent EC spreading. Sidestream smoke also caused marked disruption of the RF/6A EC-secreted BM that was accompanied by suppression of LOX expression. Further, fresh, non-SS-treated RF/6A ECs exhibited a significant loss in viability and actin cytoskeletal organization when cultured on SS-treated corrupt BM. CONCLUSIONS: These findings indicate that aberrant physical cues in the form of EC and BM architecture likely have an important role in choriocapillaris dysfunction seen in SS-associated early AMD and implicate choroidal BM as a potential target for AMD management strategies. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.
PURPOSE: Environmental tobacco smoke (ETS) is widely regarded as a major modifiable risk factor for age-related macular degeneration (AMD). Yet, precisely how it exerts its pathologic effects is poorly understood. Since early-stage AMD is characterized by choroidal capillary loss, this study examined the effect of sidestream smoke (SS), the major component of ETS, on the viability of choroidal endothelial cells (EC), with an emphasis on the role of aberrant cell and basement membrane (BM) architecture in mediating SS-induced response. METHODS: Chorioretinal ECs (RF/6A) were treated with SS, and cell viability and architecture were analyzed by colorimetric assay and actin cytoskeletal organization, respectively. The structure of RF/6A EC-secreted BM was examined by immunofluorescence for collagen IV and immunoblotting for lysyl oxidase (LOX), a collagen-crosslinking enzyme. Finally, fresh RF/6A ECs were cultured on decellularized SS-treated BM to evaluate its active role in EC dysfunction. RESULTS: The RF/6A EC viability decreased progressively with increasing SS dose, which correlated strongly with a significant decline in actin cytoskeleton-dependent EC spreading. Sidestream smoke also caused marked disruption of the RF/6A EC-secreted BM that was accompanied by suppression of LOX expression. Further, fresh, non-SS-treated RF/6A ECs exhibited a significant loss in viability and actin cytoskeletal organization when cultured on SS-treated corrupt BM. CONCLUSIONS: These findings indicate that aberrant physical cues in the form of EC and BM architecture likely have an important role in choriocapillaris dysfunction seen in SS-associated early AMD and implicate choroidal BM as a potential target for AMD management strategies. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.
Authors: Rohith R Mohan; Andrea P Cabrera; Reed E S Harrison; Ronald D Gorham; Lincoln V Johnson; Kaustabh Ghosh; Dimitrios Morikis Journal: Mol Vis Date: 2016-10-26 Impact factor: 2.367