The mechanisms of Ca2+ signaling in human lens epithelial cells upon mechanical stimulationSpasovska, Elena (Avtor) Andjelić, Sofija (Mentor) Gosak, Marko (Komentor) lens epithelial cellscalcium signalingintercellular communicationcataracteye lenscultures of lens epithelial cellsmechanical stimulationapyrasecarbenoxolonegap junctionparacrine mechanismCalcium (Ca2+) is a ubiquitous second messenger that controls diverse cellular functions. Human lens epithelial cells (LECs) are not exception in this respect, because Ca2+ signaling plays a major role in lens homeostasis and its disturbances are also related to cataract formation. The major goal of the present work is analysis, identification and comparison of intercellular Ca2+ signaling in human LECs upon local mechanical stimulation and local mechanical stimulation after applied antagonist’s apyrase (ATP diphosphohydrolase) and carbenoxolone (CBX). The purpose of added antagonists was to determine and better understand paracrine and gap junctional mechanisms in the intercellular aspect of LECs. In our experiments we used fresh postoperative lens capsules (LCs) obtained during cataract surgery and ex vivo explants. We mechanically stimulated each anterior LECs sample with a glass micropipette, which induced an increase in intracellular Ca2+ concentration ([Ca(2+)]i). Changes in [Ca2+]i in the cytosol were monitored using the fluorescent dye Fura-2. We observed the propagation on the Ca2+ signal in form of a wave that propagated radially from the stimulation point. Basically, amplitude of Ca2+ transients was found to decrease with increasing distance from the stimulation point. The Ca2+ signaling analysis in postoperative aLCs with added CBX has shown that the Ca2+ wave propagation is almost completely diminished since only cells nearby the simulation point get activated. In contrast, the application of apyrase showed little effect on the intercellular Ca2+ signaling in fresh postoperative LCs. Furthermore, we additionally studied Ca2+ signaling in ex vivo explant LC culture. It turned out that apyrase has more blocking effect on the spread of Ca2+ signal in ex vivo culture while CBX showed more efficiency in postoperative lenses, which suggests that these pharmacological agents could be a promising tool in regenerative medicine, repair, and lens regeneration.20202020-07-26 07:16:27Magistrsko delo/naloga117796VisID: 176576COBISS_ID: 26682883sl