Utvrđivanje potencijala in vitro modela rožnice za ispitivanje paracelularnog transporta koristeći EDTA kao pospješivač apsorpcije

master's thesis

• Title (english): Identifying the potential of in vitro corneal model for studying paracelular transport using EDTA as an apsorption enhancer
• Author: Ena Ljubas
• Mentor: Jasmina Lovrić (mentor)
• Committee member: Jasmina Lovrić
• Committee member: Anita Hafner
• Committee member: Dubravka Vitali Čepo
• Granter: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of pharmaceutical technology); Zagreb
• Defense date and country: 2015-11-18, Croatia
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Pharmacy

Abstract

Rano predviđanje bioraspoloživosti lijeka u oku kritičan je korak za učinkovit razvoj topikalnih oftalmičkih lijekova. Cilj velikog
projekta, u koji je uključen i ovaj diplomski rad, je standardizirati in vitro/ex vivo modele optimalnog kapaciteta za predviđanje
bioraspoloživosti lijeka, čija bi onda rutinska primjena značajno pridonjela uspješnom razvoju oftalmičkih pripravaka. U ovom
diplomskom radu usredotočili smo se na in vitro model epitela rožnice.Poseban naglasak stavlja se na permeabilnost hidrofilnih
lijekova što, uz mjerenje transepitelnog otpora, predstavlja indikator integriteta modelnog epitela. Time se također utvrđuje
potencijal modela za ispitivanje paracelularnog transporta što je bio cilj ovog diplomskog rada. U tu svrhu bilo je potrebno ispitati
permeabilnost obilježivača velike molekulske mase, fluorescentno obilježenog dekstrana (FD-4), koji ima vrlo nisku permeabilnost te se apsorbira paracelularnim putem. Također, koristeći kelirajući agens, natrijevu sol etilendiamintetraoctene kiseline (EDTA), kao pospješivač apsorpcije, ispitali smo mogućnost povećanja permeabilnosti otvaranjem čvrstih veza. Praćenjem transepitelnog električnog otpora, potvrđen je učinak EDTA na otvaranje čvrstih veza između epitelnih stanica in vitro modela epitela rožnice. Porastom koncentracije EDTA povećava se i prividni koeficijent permeabilnosti obilježivača paracelularnog puta. Ispitivanjem vijabilnosti modela nakon ispitivanja permeabilnosti isključen je citotoksičan učinak EDTA. Time je utvrđen potencijal in vitro modela epitela rožnice za ispitivanje paracelularnog transporta oftalmičkih lijekova.

Abstract (english)

Early prediction of the eye-related bioavailability is critical to efficient development of topical ophthalmic drugs. The aim
of the research project of which this diploma thesis is an integral part is to standardize in vitro/ex vivo models of optimum capacity to predict eye-related bioavailability. The routine use of these models would significantly contribute to the successful development of ophthalmic drugs.In vitro corneal epithelial model was in the focus of this diploma thesis. Special importance is given to the permeability assays with hydrophilic and macromolecular markers of paracellular pathway, which together with transepithelial electric resistance are indicators of barrier integrity. This characterization is important in order to confirm the model applicability in terms of studying paracellular transport, which represents the aim of this diploma thesis. We have determined the permeability of fluorescently labeled dextran (FD-4), a hydrofilic macromolecular marker of paracellular transport. Moreover, by using ethylenediaminetetraacetic acid disodium salt (EDTA), as an enhancer of paracellular absorption, we tested the possibility of permeability enhancement by opening the tight junctions. By measuring the transepithelial electrical resistance, the effect of EDTA on the tight junction opening between the epithelial cells was also confirmed. Furthermore, the apparent permeability coefficient of the paracellular marker increased by increasing the concentration of EDTA. The cytotoxic effect of EDTA was excluded by examining the viability of model after testing permeability. These results indicate the potential of in vitro corneal epithelial model for studying paracellular transport of ophthalmic drugs.

• Keywords: HCE-T; EDTA; FD-4; permeabilnost; vijabilnost; TEER
• Keywords (english): HCE-T; EDTA; FD-4; permeability; viability; TEER
• Language: croatian
• URN:NBN: urn:nbn:hr:163:561840
• Study programme: Title: Pharmacy; Study programme type: university; Study level: integrated undergraduate and graduate; Academic / professional title: magistar/magistra farmacije (magistar/magistra farmacije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Access restricted to students and staff of home institution
• Created on: 2016-03-22 11:06:24