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Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model
Author(s) -
AlonsoAlonso M.L.,
Hampton S.L.,
Williams J.L.,
GarcíaGutiérrez M.T.,
FernándezBueno I.,
Srivastava G.K.,
Pastor J.C.,
Diebold Y.
Publication year - 2017
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2017.0f071
Subject(s) - viability assay , oxidative stress , apoptosis , mtt assay , pharmacology , chemistry , cell growth , biology , biochemistry
Purpose Plasma Kallikrein ( PK ) Inhibitors ( PKI ) are a promising treatment for retinal diseases such as diabetic macular edema, where PK activity is implicated. Our aim was to set up an in vitro model of chronic oxidative stress in the retinal pigment epithelium cell line ARPE 19 and use it to test the effect of 8 PKI on cell viability and proliferation. Methods ARPE 19 cells were grown to a sub‐confluence stage, synchronized, and exposed to different dilutions of glucose oxidase ( GO x) from Aspergillus niger, culture medium (− control) and benzalkonium chloride (+ control) for 24 hr. Cell viability and apoptosis were analyzed ( MTT assay and caspase‐3/7 detection, respectively). The oxidative stressed cells were exposed to 8 different PKI developed by KalVista (Salisbury, UK ) at 100 n m and 1 μ m . Also, ARPE 19 cells were exposed to PKI previously and simultaneously to GO x and cell viability and proliferation were assessed (alamarBlue ® assays). Results Exposure of ARPE 19 to GO x induced dose‐dependent decreased cell viability and increased cell apoptosis. A sub‐lethal GO x dose reducing viability by 30% and increasing apoptosis by 59% was chosen for PKI exposure experiments. Exposure of GO x stressed cells to PKI for 24 hr did not further reduce cell viability after 5 days. No effect on cell proliferation was observed when GO x stressed cells were either pre‐treated or simultaneously exposed to PKI . Conclusions GO x oxidative sub‐lethal stress model in ARPE 19 cells may be used for screening new drugs with therapeutic potential for retinal diseases. The PKI did not reduce GO x‐induced cytotoxic effects in RPE cells suggesting plasma kallikrein has no role in this stress mechanism. Support: FP 7‐ PEOPLE ‐2013‐ IAPP (612218/3D‐ NET ) and Regional Junta de Castilla y León Scholarship/European Social Fund Program (Va040‐13).