Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products
The utilization of in vitro tests with a tiered testing strategy for detection of mild ocular irritants can reduce the use of animals for testing, provide mechanistic data on toxic effects, and reduce the uncertainty associated with dose selection for clinical trials. The first section of this thesi...
| Main Author: | |
|---|---|
| Language: | en |
| Published: |
2009
|
| Subjects: | |
| Online Access: | http://hdl.handle.net/10012/4338 |
| id |
ndltd-WATERLOO-oai-uwspace.uwaterloo.ca-10012-4338 |
|---|---|
| record_format |
oai_dc |
| collection |
NDLTD |
| language |
en |
| sources |
NDLTD |
| topic |
in vitro Alternative Methods alamarBlue rhodamine tight junctions bovine lens viability toxicity mitochondria mitochondria integrity benzalkonium chloride sodium dodecyl sulphate sodium lauryl sulfate scanning electron microscopy Draize Draize maximal average scores zonula occludens cosmetic directive PETA humane society animal league people for the ethical treatment of animals interagency coordinating committee on the validation of alternative methods ICCVAM ECVAM JaCVAM BCOP bovine cornea ocular irritants ocular irritation ocular toxicity human corneal epithelial cells vitro confocal confocal microscopy metabolic activity optical quality reactive oxygen species contact lens contact lens care solutions barrier function microbial keratitis mulitipurpose solutions tight junction cell damage claudin ZO-1 occludin MDCK Madin Madin-Darby canine kidney cells cornea human cornea human corneal epithelium epithelial cell line human corneal epithelial cell line sodium fluorescein sodium fluorescein permeability fluorescein permeability ocular surface cell monolayer Araki-Sasaki cell physiology risk assessment safety assessment ScanTox scanning laser lens epithelium corneal cells in vitro model ophthalmic formulations multiple instillation back vertex animal testing rabbit testing alternatives to animal testing cultured bovine lens toxicity of chemicals irritation irritants laser scanner organ culture delayed toxicity toxins hydrogen peroxide cornea toxicity cornea toxicity models prediction of human toxicity no observable adverse effect level lowest observable adverse effect level NOAEL LOAEL toxicity thresholds safety factors cornea uncertainty factors preservatives disinfectants ophthalmic products preclinical preclinical testing epithelial barrier drug penetration clinical confocal microscopy animal rights rabbit cornea human cornea human clinical effects toxic animal rights activist sensitive measures toxic effect toxicity threshold agar overlay agar diffusion agar overlay method agar diffusion method cytochrome cytochrome c apoptosis necrotic apoptotic necrosis caspase rhodamine 123 resazuran resorufin cell death cell viability metabolic dye microsomal microsomal enzymes cytotoxicity cytotoxic cytotoxic effect MTT XTT WST-1 plasma membrane mitochondrial mitochondrial morphology ocular toxicity potential ocular toxicity human corneal epithelial cell line confocal analysis corneal epithelium cell fluorescence alamarBlue assay rhodamine dye animal welfare toxic injury degraded mitochondria epithelial monolayer disinfectants membrane integrity eye toxicity eye viability dye toxicity in humans human toxicity effects on the mitochondria mitochondrial toxicity in vitro battery in vitro test battery ophthalmic eye drop direct contact product safety cytotoxicity potential molecular molecular biology refine reduce replace sensitivity and relevance sensitivity relevance rabbit ocular irritation test product development cytotoxicity models cytotoxicity alternative methods replacements for animal testing three r's beagle tiered testing tiered testing strategy replacements animal testing mechanistic toxicity cornea mitochondria dose response threshold Vision Science and Biology |
| spellingShingle |
in vitro Alternative Methods alamarBlue rhodamine tight junctions bovine lens viability toxicity mitochondria mitochondria integrity benzalkonium chloride sodium dodecyl sulphate sodium lauryl sulfate scanning electron microscopy Draize Draize maximal average scores zonula occludens cosmetic directive PETA humane society animal league people for the ethical treatment of animals interagency coordinating committee on the validation of alternative methods ICCVAM ECVAM JaCVAM BCOP bovine cornea ocular irritants ocular irritation ocular toxicity human corneal epithelial cells vitro confocal confocal microscopy metabolic activity optical quality reactive oxygen species contact lens contact lens care solutions barrier function microbial keratitis mulitipurpose solutions tight junction cell damage claudin ZO-1 occludin MDCK Madin Madin-Darby canine kidney cells cornea human cornea human corneal epithelium epithelial cell line human corneal epithelial cell line sodium fluorescein sodium fluorescein permeability fluorescein permeability ocular surface cell monolayer Araki-Sasaki cell physiology risk assessment safety assessment ScanTox scanning laser lens epithelium corneal cells in vitro model ophthalmic formulations multiple instillation back vertex animal testing rabbit testing alternatives to animal testing cultured bovine lens toxicity of chemicals irritation irritants laser scanner organ culture delayed toxicity toxins hydrogen peroxide cornea toxicity cornea toxicity models prediction of human toxicity no observable adverse effect level lowest observable adverse effect level NOAEL LOAEL toxicity thresholds safety factors cornea uncertainty factors preservatives disinfectants ophthalmic products preclinical preclinical testing epithelial barrier drug penetration clinical confocal microscopy animal rights rabbit cornea human cornea human clinical effects toxic animal rights activist sensitive measures toxic effect toxicity threshold agar overlay agar diffusion agar overlay method agar diffusion method cytochrome cytochrome c apoptosis necrotic apoptotic necrosis caspase rhodamine 123 resazuran resorufin cell death cell viability metabolic dye microsomal microsomal enzymes cytotoxicity cytotoxic cytotoxic effect MTT XTT WST-1 plasma membrane mitochondrial mitochondrial morphology ocular toxicity potential ocular toxicity human corneal epithelial cell line confocal analysis corneal epithelium cell fluorescence alamarBlue assay rhodamine dye animal welfare toxic injury degraded mitochondria epithelial monolayer disinfectants membrane integrity eye toxicity eye viability dye toxicity in humans human toxicity effects on the mitochondria mitochondrial toxicity in vitro battery in vitro test battery ophthalmic eye drop direct contact product safety cytotoxicity potential molecular molecular biology refine reduce replace sensitivity and relevance sensitivity relevance rabbit ocular irritation test product development cytotoxicity models cytotoxicity alternative methods replacements for animal testing three r's beagle tiered testing tiered testing strategy replacements animal testing mechanistic toxicity cornea mitochondria dose response threshold Vision Science and Biology McCanna, David Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| description |
The utilization of in vitro tests with a tiered testing strategy for detection of mild ocular irritants can reduce the use of animals for testing, provide mechanistic data on toxic effects, and reduce the uncertainty associated with dose selection for clinical trials. The first section of this thesis describes how in vitro methods can be used to improve the prediction of the toxicity of chemicals and ophthalmic products. The proper utilization of in vitro methods can accurately predict toxic threshold levels and reduce animal use in product development. Sections two, three and four describe the development of new sensitive in vitro methods for predicting ocular toxicity. Maintaining the barrier function of the cornea is critical for the prevention of the penetration of infections microorganisms and irritating chemicals into the eye. Chapter 2 describes the development of a method for assessing the effects of chemicals on tight junctions using a human corneal epithelial and canine kidney epithelial cell line. In Chapter 3 a method that uses a primary organ culture for assessing single instillation and multiple instillation toxic effects is described. The ScanTox system was shown to be an ideal system to monitor the toxic effects over time as multiple readings can be taken of treated bovine lenses using the nondestructive method of assessing for the lens optical quality. Confirmations of toxic effects were made with the utilization of the viability dye alamarBlue. Chapter 4 describes the development of sensitive in vitro assays for detecting ocular toxicity by measuring the effects of chemicals on the mitochondrial integrity of bovine cornea, bovine lens epithelium and corneal epithelial cells, using fluorescent dyes.
The goal of this research was to develop an in vitro test battery that can be used to accurately predict the ocular toxicity of new chemicals and ophthalmic formulations. By comparing the toxicity seen in vivo animals and humans with the toxicity response in these new in vitro methods, it was demonstrated that these in vitro methods can be utilized in a tiered testing strategy in the development of new chemicals and ophthalmic formulations. |
| author |
McCanna, David |
| author_facet |
McCanna, David |
| author_sort |
McCanna, David |
| title |
Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| title_short |
Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| title_full |
Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| title_fullStr |
Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| title_full_unstemmed |
Development of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic Products |
| title_sort |
development of sensitive in vitro assays to assess the ocular toxicity potential of chemicals and ophthalmic products |
| publishDate |
2009 |
| url |
http://hdl.handle.net/10012/4338 |
| work_keys_str_mv |
AT mccannadavid developmentofsensitiveinvitroassaystoassesstheoculartoxicitypotentialofchemicalsandophthalmicproducts |
| _version_ |
1716573302722396160 |
| spelling |
ndltd-WATERLOO-oai-uwspace.uwaterloo.ca-10012-43382013-01-08T18:52:09ZMcCanna, David2009-04-27T14:06:30Z2009-04-27T14:06:30Z2009-04-27T14:06:30Z2009http://hdl.handle.net/10012/4338The utilization of in vitro tests with a tiered testing strategy for detection of mild ocular irritants can reduce the use of animals for testing, provide mechanistic data on toxic effects, and reduce the uncertainty associated with dose selection for clinical trials. The first section of this thesis describes how in vitro methods can be used to improve the prediction of the toxicity of chemicals and ophthalmic products. The proper utilization of in vitro methods can accurately predict toxic threshold levels and reduce animal use in product development. Sections two, three and four describe the development of new sensitive in vitro methods for predicting ocular toxicity. Maintaining the barrier function of the cornea is critical for the prevention of the penetration of infections microorganisms and irritating chemicals into the eye. Chapter 2 describes the development of a method for assessing the effects of chemicals on tight junctions using a human corneal epithelial and canine kidney epithelial cell line. In Chapter 3 a method that uses a primary organ culture for assessing single instillation and multiple instillation toxic effects is described. The ScanTox system was shown to be an ideal system to monitor the toxic effects over time as multiple readings can be taken of treated bovine lenses using the nondestructive method of assessing for the lens optical quality. Confirmations of toxic effects were made with the utilization of the viability dye alamarBlue. Chapter 4 describes the development of sensitive in vitro assays for detecting ocular toxicity by measuring the effects of chemicals on the mitochondrial integrity of bovine cornea, bovine lens epithelium and corneal epithelial cells, using fluorescent dyes. The goal of this research was to develop an in vitro test battery that can be used to accurately predict the ocular toxicity of new chemicals and ophthalmic formulations. By comparing the toxicity seen in vivo animals and humans with the toxicity response in these new in vitro methods, it was demonstrated that these in vitro methods can be utilized in a tiered testing strategy in the development of new chemicals and ophthalmic formulations.enin vitroAlternative MethodsalamarBluerhodaminetight junctionsbovine lensviabilitytoxicitymitochondriamitochondria integritybenzalkonium chloridesodium dodecyl sulphatesodium lauryl sulfatescanning electron microscopyDraizeDraize maximal average scoreszonula occludenscosmetic directivePETAhumane societyanimal leaguepeople for the ethical treatment of animalsinteragency coordinating committee on the validation of alternative methodsICCVAMECVAMJaCVAMBCOPbovine corneaocular irritantsocular irritationocular toxicityhuman corneal epithelial cellsvitroconfocalconfocal microscopymetabolic activityoptical qualityreactive oxygen speciescontact lenscontact lens care solutionsbarrier functionmicrobial keratitismulitipurpose solutionstight junctioncell damageclaudinZO-1occludinMDCKMadinMadin-Darbycanine kidney cellscorneahuman corneahuman corneal epitheliumepithelial cell linehuman corneal epithelial cell linesodium fluoresceinsodium fluorescein permeabilityfluorescein permeabilityocular surfacecell monolayerAraki-Sasakicell physiologyrisk assessmentsafety assessmentScanToxscanning laserlens epitheliumcorneal cellsin vitro modelophthalmic formulationsmultiple instillationback vertexanimal testingrabbit testingalternatives to animal testingcultured bovine lenstoxicity of chemicalsirritationirritantslaser scannerorgan culturedelayed toxicitytoxinshydrogen peroxidecornea toxicitycornea toxicity modelsprediction of human toxicityno observable adverse effect levellowest observable adverse effect levelNOAELLOAELtoxicity thresholdssafety factorscorneauncertainty factorspreservativesdisinfectantsophthalmic productspreclinicalpreclinical testingepithelial barrierdrug penetrationclinical confocal microscopyanimal rightsrabbit corneahuman corneahuman clinical effectstoxicanimal rights activistsensitive measurestoxic effecttoxicity thresholdagar overlayagar diffusionagar overlay methodagar diffusion methodcytochromecytochrome capoptosisnecroticapoptoticnecrosiscaspaserhodamine 123resazuranresorufincell deathcell viabilitymetabolic dyemicrosomalmicrosomal enzymescytotoxicitycytotoxiccytotoxic effectMTTXTTWST-1plasma membranemitochondrialmitochondrial morphologyocular toxicity potentialocular toxicityhuman corneal epithelialcell lineconfocal analysiscorneal epitheliumcell fluorescencealamarBlue assayrhodamine dyeanimal welfaretoxic injurydegraded mitochondriaepithelial monolayerdisinfectantsmembrane integrityeye toxicityeyeviability dyetoxicity in humanshuman toxicityeffects on the mitochondriamitochondrial toxicityin vitro batteryin vitro test batteryophthalmic eye dropdirect contactproduct safetycytotoxicity potentialmolecularmolecular biologyrefine reduce replacesensitivity and relevancesensitivityrelevancerabbit ocular irritation testproduct developmentcytotoxicity modelscytotoxicity alternative methodsreplacements for animal testingthree r'sbeagletiered testingtiered testing strategyreplacements animal testingmechanistic toxicitycornea mitochondriadose responsethresholdDevelopment of Sensitive In Vitro Assays to Assess the Ocular Toxicity Potential of Chemicals and Ophthalmic ProductsThesis or DissertationSchool of OptometryDoctor of PhilosophyVision Science and Biology |