Comparative Cytotoxicity of Silicone Hydrogel Contact Lenses With Various Contact Lens Care Products Using Human Corneal Epithelial Cells and Flow Cytometry Analysis

Abstract

Abstract: : Purpose:To evaluate cell viability of SV–40 transformed human corneal epithelial cells (HCE–T) exposed to high oxygen permeable silicone hydrogel contact lenses (HiDk SiHy) and contact lens care products (LCP) using flow cytometry. Methods: HCE–T cells were exposed to LCP and to HiDk SiHy treated with LCP and analyzed for cell viability. Cell viability was evaluated using Alamar Blue^TM (AB), a resazurin based dye, and flow cytometry of calcein and ethidium homodimer–1 (CEH) stained cells (Live/Dead Viability/Cytotoxicity Kit from Molecular Probes). Three different cell exposure conditions were evaluated. 1) HiDk SiHy soaked in LCP were seeded with HCE–T, incubated, trypsinized, stained with CEH and analyzed by flow cytometry (BDFACSCailbur^TM 4 color unit cytometer). 2) HCE–T in suspension were exposed 5 and 30 minutes to 50% dilutions of LCP; the cells were stained with CEH and analyzed using flow cytometry. 3) HCE–T were seeded into 96 well plates, exposed to 50% dilutions of three LCP for 5 and 30 minutes, rinsed and evaluated for viability with AB using a CytoFluor® (PerSeptive Biosystems) plate reader. The LCP used were: AQuify^TM MPS (AQ), Clear Care® (CC), and OPTI–FREE® Express® with Aldox^TM (OF). Using flow cytometry, subpopulations of cells were identified and sorted by regions for live (green fluorescent) and dead (red fluorescent) cells. Benzalkonium chloride was used as a positive control for the analysis of dead cells. Results: Alamar Blue cell viability was above 50% for HCE–T exposed to all three LCP for 5 minutes; however, at the 30 minute exposure time cell viability for OF was less than 50%. Flow cytometric analysis demonstrated that CC and AQ are much less cytotoxic to HCE–T in suspension than OF at both exposure times. HCE–T cells grown on HiDk SiHy lenses soaked for one day in CC and AQ were found to be 70% alive and 30% damaged. No live HCE–T cells were detected on HiDk SiHy lenses exposed to OF for one day. Conclusions: HCE–T cells are significantly more viable when grown on HiDk SiHy contact lenses cycled in Clear Care® or Aquify MPS compared to HiDk SiHy contact lenses cycled in OPTI–FREE Express with Aldox.

Keywords: cell death/apoptosis • contact lens