Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model

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 ARPE19 and use it to test th...

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Veröffentlicht in:Acta ophthalmologica (Oxford, England) England), 2017-09, Vol.95 (S259), p.n/a
Hauptverfasser: Alonso‐Alonso, M.L., Hampton, S.L., Williams, J.L., García‐Gutiérrez, M.T., Fernández‐Bueno, I., Srivastava, G.K., Pastor, J.C., Diebold, Y.
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container_issue S259
container_start_page
container_title Acta ophthalmologica (Oxford, England)
container_volume 95
creator Alonso‐Alonso, M.L.
Hampton, S.L.
Williams, J.L.
García‐Gutiérrez, M.T.
Fernández‐Bueno, I.
Srivastava, G.K.
Pastor, J.C.
Diebold, Y.
description 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 ARPE19 and use it to test the effect of 8 PKI on cell viability and proliferation. Methods ARPE19 cells were grown to a sub‐confluence stage, synchronized, and exposed to different dilutions of glucose oxidase (GOx) 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 nm and 1 μm. Also, ARPE19 cells were exposed to PKI previously and simultaneously to GOx and cell viability and proliferation were assessed (alamarBlue® assays). Results Exposure of ARPE19 to GOx induced dose‐dependent decreased cell viability and increased cell apoptosis. A sub‐lethal GOx dose reducing viability by 30% and increasing apoptosis by 59% was chosen for PKI exposure experiments. Exposure of GOx stressed cells to PKI for 24 hr did not further reduce cell viability after 5 days. No effect on cell proliferation was observed when GOx stressed cells were either pre‐treated or simultaneously exposed to PKI. Conclusions GOx oxidative sub‐lethal stress model in ARPE19 cells may be used for screening new drugs with therapeutic potential for retinal diseases. The PKI did not reduce GOx‐induced cytotoxic effects in RPE cells suggesting plasma kallikrein has no role in this stress mechanism. Support: FP7‐PEOPLE‐2013‐IAPP (612218/3D‐NET) and Regional Junta de Castilla y León Scholarship/European Social Fund Program (Va040‐13).
doi_str_mv 10.1111/j.1755-3768.2017.0F071
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Our aim was to set up an in vitro model of chronic oxidative stress in the retinal pigment epithelium cell line ARPE19 and use it to test the effect of 8 PKI on cell viability and proliferation. Methods ARPE19 cells were grown to a sub‐confluence stage, synchronized, and exposed to different dilutions of glucose oxidase (GOx) 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 nm and 1 μm. Also, ARPE19 cells were exposed to PKI previously and simultaneously to GOx and cell viability and proliferation were assessed (alamarBlue® assays). Results Exposure of ARPE19 to GOx induced dose‐dependent decreased cell viability and increased cell apoptosis. A sub‐lethal GOx dose reducing viability by 30% and increasing apoptosis by 59% was chosen for PKI exposure experiments. Exposure of GOx stressed cells to PKI for 24 hr did not further reduce cell viability after 5 days. No effect on cell proliferation was observed when GOx stressed cells were either pre‐treated or simultaneously exposed to PKI. Conclusions GOx oxidative sub‐lethal stress model in ARPE19 cells may be used for screening new drugs with therapeutic potential for retinal diseases. The PKI did not reduce GOx‐induced cytotoxic effects in RPE cells suggesting plasma kallikrein has no role in this stress mechanism. Support: FP7‐PEOPLE‐2013‐IAPP (612218/3D‐NET) and Regional Junta de Castilla y León Scholarship/European Social Fund Program (Va040‐13).</description><identifier>ISSN: 1755-375X</identifier><identifier>EISSN: 1755-3768</identifier><identifier>DOI: 10.1111/j.1755-3768.2017.0F071</identifier><language>eng</language><publisher>Malden: Wiley Subscription Services, Inc</publisher><subject>Apoptosis ; Aspergillus ; Aspergillus niger ; Benzalkonium chloride ; Caspase ; Caspase-3 ; Cell culture ; Cell proliferation ; Chlorides ; Cytotoxicity ; Diabetes mellitus ; Dilution ; Drug screening ; Edema ; Epithelium ; Exposure ; Glucose oxidase ; Inhibitors ; Kallikrein ; Oxidative stress ; Plasma kallikrein ; Retina ; Retinal pigment epithelium ; Stress ; Viability</subject><ispartof>Acta ophthalmologica (Oxford, England), 2017-09, Vol.95 (S259), p.n/a</ispartof><rights>2017 The Authors Acta Ophthalmologica © 2017 Acta Ophthalmologica Scandinavica Foundation</rights><rights>Copyright © 2017 Acta Ophthalmologica Scandinavica Foundation</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1111%2Fj.1755-3768.2017.0F071$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,1417,1433,27924,27925,45575,46833</link.rule.ids></links><search><creatorcontrib>Alonso‐Alonso, M.L.</creatorcontrib><creatorcontrib>Hampton, S.L.</creatorcontrib><creatorcontrib>Williams, J.L.</creatorcontrib><creatorcontrib>García‐Gutiérrez, M.T.</creatorcontrib><creatorcontrib>Fernández‐Bueno, I.</creatorcontrib><creatorcontrib>Srivastava, G.K.</creatorcontrib><creatorcontrib>Pastor, J.C.</creatorcontrib><creatorcontrib>Diebold, Y.</creatorcontrib><title>Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model</title><title>Acta ophthalmologica (Oxford, England)</title><description>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 ARPE19 and use it to test the effect of 8 PKI on cell viability and proliferation. Methods ARPE19 cells were grown to a sub‐confluence stage, synchronized, and exposed to different dilutions of glucose oxidase (GOx) 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 nm and 1 μm. Also, ARPE19 cells were exposed to PKI previously and simultaneously to GOx and cell viability and proliferation were assessed (alamarBlue® assays). Results Exposure of ARPE19 to GOx induced dose‐dependent decreased cell viability and increased cell apoptosis. A sub‐lethal GOx dose reducing viability by 30% and increasing apoptosis by 59% was chosen for PKI exposure experiments. Exposure of GOx stressed cells to PKI for 24 hr did not further reduce cell viability after 5 days. No effect on cell proliferation was observed when GOx stressed cells were either pre‐treated or simultaneously exposed to PKI. Conclusions GOx oxidative sub‐lethal stress model in ARPE19 cells may be used for screening new drugs with therapeutic potential for retinal diseases. The PKI did not reduce GOx‐induced cytotoxic effects in RPE cells suggesting plasma kallikrein has no role in this stress mechanism. Support: FP7‐PEOPLE‐2013‐IAPP (612218/3D‐NET) and Regional Junta de Castilla y León Scholarship/European Social Fund Program (Va040‐13).</description><subject>Apoptosis</subject><subject>Aspergillus</subject><subject>Aspergillus niger</subject><subject>Benzalkonium chloride</subject><subject>Caspase</subject><subject>Caspase-3</subject><subject>Cell culture</subject><subject>Cell proliferation</subject><subject>Chlorides</subject><subject>Cytotoxicity</subject><subject>Diabetes mellitus</subject><subject>Dilution</subject><subject>Drug screening</subject><subject>Edema</subject><subject>Epithelium</subject><subject>Exposure</subject><subject>Glucose oxidase</subject><subject>Inhibitors</subject><subject>Kallikrein</subject><subject>Oxidative stress</subject><subject>Plasma kallikrein</subject><subject>Retina</subject><subject>Retinal pigment epithelium</subject><subject>Stress</subject><subject>Viability</subject><issn>1755-375X</issn><issn>1755-3768</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><recordid>eNqNkFtLAzEQhYMoWKt_QQI-75rZS7ILvpRSL1CoV_AtJNkEs902NdlW--_ddaXPzsPMMJxzBj6ELoHE0NV1HQPL8yhltIgTAiwmt4TBERodzseHPX8_RWch1IRQoDQboaeZMVq1ATuDN40IK4GXomns0mu7xnb9YaVtnQ_dikV_wDvbeoefH2fYfdtKtHancWi9DgGvXKWbc3RiRBP0xd8co7fb2ev0Ppov7h6mk3mkgFGIpEkYiKzsmpaVkTKBkmkjNIiiKDNpqASRp7pkRWUoqCxRSpCsTCtplFEqHaOrIXfj3edWh5bXbuvX3UsOZUq7uDwtOhUdVMq7ELw2fOPtSvg9B8J7fLzmPRvec-I9Pv6LrzPeDMYv2-j9P118sngZ7D-sRXZF</recordid><startdate>201709</startdate><enddate>201709</enddate><creator>Alonso‐Alonso, M.L.</creator><creator>Hampton, S.L.</creator><creator>Williams, J.L.</creator><creator>García‐Gutiérrez, M.T.</creator><creator>Fernández‐Bueno, I.</creator><creator>Srivastava, G.K.</creator><creator>Pastor, J.C.</creator><creator>Diebold, Y.</creator><general>Wiley Subscription Services, Inc</general><scope>AAYXX</scope><scope>CITATION</scope><scope>7TK</scope></search><sort><creationdate>201709</creationdate><title>Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model</title><author>Alonso‐Alonso, M.L. ; Hampton, S.L. ; Williams, J.L. ; García‐Gutiérrez, M.T. ; Fernández‐Bueno, I. ; Srivastava, G.K. ; Pastor, J.C. ; Diebold, Y.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c1761-bf271a4971aebdfbb2197efae1a8894bf6b1a53e978df61c42cca0493dbfcfcc3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Apoptosis</topic><topic>Aspergillus</topic><topic>Aspergillus niger</topic><topic>Benzalkonium chloride</topic><topic>Caspase</topic><topic>Caspase-3</topic><topic>Cell culture</topic><topic>Cell proliferation</topic><topic>Chlorides</topic><topic>Cytotoxicity</topic><topic>Diabetes mellitus</topic><topic>Dilution</topic><topic>Drug screening</topic><topic>Edema</topic><topic>Epithelium</topic><topic>Exposure</topic><topic>Glucose oxidase</topic><topic>Inhibitors</topic><topic>Kallikrein</topic><topic>Oxidative stress</topic><topic>Plasma kallikrein</topic><topic>Retina</topic><topic>Retinal pigment epithelium</topic><topic>Stress</topic><topic>Viability</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Alonso‐Alonso, M.L.</creatorcontrib><creatorcontrib>Hampton, S.L.</creatorcontrib><creatorcontrib>Williams, J.L.</creatorcontrib><creatorcontrib>García‐Gutiérrez, M.T.</creatorcontrib><creatorcontrib>Fernández‐Bueno, I.</creatorcontrib><creatorcontrib>Srivastava, G.K.</creatorcontrib><creatorcontrib>Pastor, J.C.</creatorcontrib><creatorcontrib>Diebold, Y.</creatorcontrib><collection>CrossRef</collection><collection>Neurosciences Abstracts</collection><jtitle>Acta ophthalmologica (Oxford, England)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Alonso‐Alonso, M.L.</au><au>Hampton, S.L.</au><au>Williams, J.L.</au><au>García‐Gutiérrez, M.T.</au><au>Fernández‐Bueno, I.</au><au>Srivastava, G.K.</au><au>Pastor, J.C.</au><au>Diebold, Y.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model</atitle><jtitle>Acta ophthalmologica (Oxford, England)</jtitle><date>2017-09</date><risdate>2017</risdate><volume>95</volume><issue>S259</issue><epage>n/a</epage><issn>1755-375X</issn><eissn>1755-3768</eissn><abstract>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 ARPE19 and use it to test the effect of 8 PKI on cell viability and proliferation. Methods ARPE19 cells were grown to a sub‐confluence stage, synchronized, and exposed to different dilutions of glucose oxidase (GOx) 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 nm and 1 μm. Also, ARPE19 cells were exposed to PKI previously and simultaneously to GOx and cell viability and proliferation were assessed (alamarBlue® assays). Results Exposure of ARPE19 to GOx induced dose‐dependent decreased cell viability and increased cell apoptosis. A sub‐lethal GOx dose reducing viability by 30% and increasing apoptosis by 59% was chosen for PKI exposure experiments. Exposure of GOx stressed cells to PKI for 24 hr did not further reduce cell viability after 5 days. No effect on cell proliferation was observed when GOx stressed cells were either pre‐treated or simultaneously exposed to PKI. Conclusions GOx oxidative sub‐lethal stress model in ARPE19 cells may be used for screening new drugs with therapeutic potential for retinal diseases. The PKI did not reduce GOx‐induced cytotoxic effects in RPE cells suggesting plasma kallikrein has no role in this stress mechanism. Support: FP7‐PEOPLE‐2013‐IAPP (612218/3D‐NET) and Regional Junta de Castilla y León Scholarship/European Social Fund Program (Va040‐13).</abstract><cop>Malden</cop><pub>Wiley Subscription Services, Inc</pub><doi>10.1111/j.1755-3768.2017.0F071</doi><tpages>1</tpages><oa>free_for_read</oa></addata></record>
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subjects Apoptosis
Aspergillus
Aspergillus niger
Benzalkonium chloride
Caspase
Caspase-3
Cell culture
Cell proliferation
Chlorides
Cytotoxicity
Diabetes mellitus
Dilution
Drug screening
Edema
Epithelium
Exposure
Glucose oxidase
Inhibitors
Kallikrein
Oxidative stress
Plasma kallikrein
Retina
Retinal pigment epithelium
Stress
Viability
title Effects of plasma kallikrein inhibitors in an in vitro RPE oxidative stress model
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