Abstract
Purpose :
Purpose: Benzalkonium chloride (BAK) is a common preservative in topical ophthalmic agents. BAK and other types of preservatives are necessary to maintain a drug’s efficacy and inhibit microbial growth. The effects of BAK on the ocular surface have been well studied and include ocular surface inflammation, cytotoxic damage to corneal epithelial cells, and tear film instability. The purpose of this study is to compare the differences between the human tear proteome in BAK and non BAK users.
Methods :
Methods: Ten volunteers have been recruited into one of two groups: a healthy, non-BAK user control group and a BAK user group. All volunteers signed an informed consent prior to participation in the study. All volunteers were assessed using a slit lamp biomicroscope for signs of ocular surface disease including assessment of corneal staining, tear film break up time, and tear production. Ten µl tear samples were collected using glass microcapillary tubes. Samples for each group were pooled together and total protein concentration was measured using a QuBit 3.0 Fluorometer. 72 µg of each sample was then placed into 2x sample buffer and subject to electrophoresis in a 12% polyacrylamide gel. After separation, each lane was fractionated into six individual sections and sent for mass spectrometry analysis at the UTSW Proteomics Core Facility. Functional analysis of unique and enriched proteins was performed using the WEB-based GEne SeT AnaLysis Toolkit (web-Gestault and the Kyoto Encyclopedia of Genes and Genomes (KEGG). For proteins to be considered enriched, a 3:1 ratio of the spectral index for the enriched protein over the control was required.
Results :
Results: There was a significant difference in conjunctival staining between BAK and non-BAK users (P=0.014). To date, we have identified a total of 579 unique proteins in the tear fluid, with 27 unique to non BAK users and 108 unique to BAK users. An additional 119 unique proteins were found to be enriched in BAK users. Subject recruitment and tear collection is still on-going to validate these findings.
Conclusions :
Conclusions: Enriched and unique proteins found in the BAK group confirm increased cellular toxicity from chronic BAK exposure. These proteins may represent usable biomarkers for studying cellular toxicity at the ocular surface in response to common ocular preservatives and biocides.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.