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Peer-reviewed articles


Kao,E. C. Y., Seo,J., McCanna,D. J., Subbaraman,L. N., Jones,L. In vitro assessment of the biocompatibility of chemically treated silicone materials with human lens epithelial cells Nature - Scientific Reports 2022;12(1):Article 4649 [ Show Abstract ]

Cytotoxicity testing is a regulatory requirement for safety testing of new ocular implants. In vitro toxicity tests determine whether toxic chemicals are present on a material surface or leach out of the material matrix. A method of evaluating the cytotoxicity of ocular implants was developed using fluorescent viability dyes. To assess the assay’s sensitivity in detecting toxic substances on biomaterials, zinc diethydithiocarbamate (ZDEC) and benzalkonium chloride (BAK) were deposited on silicone surfaces at different concentrations. Human lens epithelial cells (HLEC) were added to the surface of these treated silicone surfaces and were assessed for viability. The viability of both the adherent and non-adherent cells was determined using confocal microscopy with, annexin V, ethidium homodimer, and calcein. Cell metabolism was also evaluated using resazurin and the release of inflammatory cytokines was quantified using a multiplex Mesoscale Discovery platform. Confocal microscopy was shown to be a sensitive assay for evaluating material toxicity, as significant toxicity (p < 0.05) from ZDEC and BAK-treated surfaces compared to the untreated silicone control was detected. Patterns of cytokine release from cells varied depending on the toxin evaluated and the toxin concentration and did not directly correlate with the reduction in cell metabolic activity measured by alamarBlue.


Chang,J. M. L., Seo,J., Kwan,M. M. Y., Oh,S., McCanna,D. J., Subbaraman,L., Jones,L. Determining the Toxicity of UV Radiation and Chemicals on Primary and Immortalized Human Corneal Epithelial Cells Journal of Visualized Experiments 2021;173(July):e62675 [ Show Abstract ]

This article describes the methods of measuring the toxicity of ultraviolet (UV) radiation and ocular toxins on primary (pHCEC) and immortalized (iHCEC) human corneal epithelial cell cultures. Cells were exposed to UV radiation and toxic doses of benzalkonium chloride (BAK), hydrogen peroxide (H2O2), and sodium dodecyl sulfate (SDS). Metabolic activity was measured using a metabolic assay. The release of inflammatory cytokines was measured using a multi-plex interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor-alpha (TNF-α) assay, and cells were evaluated for viability using fluorescent dyes.

The damaging effects of UV on cell metabolic activity and cytokine release occurred at 5 min of UV exposure for iHCEC and 20 min for pHCEC. Similar percent drops in metabolic activity of the iHCEC and pHCEC occurred after exposure to BAK, H2O2, or SDS, and the most significant changes in cytokine release occurred for IL-6 and IL-8. Microscopy of fluorescently stained iHCEC and pHCEC BAK-exposed cells showed cell death at 0.005% BAK exposure, although the degree of ethidium staining was greater in the iHCECs than pHCECs. Utilizing multiple methods of assessing toxic effects using microscopy, assessments of metabolic activity, and cytokine production, the toxicity of UV radiation and chemical toxins could be determined for both primary and immortalized cell lines.


McCanna,D. J., Oh,S., Seo,J., Coles-Brennan,C., Fadli,Z., Subbaraman,L. N., Jones,L. W. The effect of denatured lysozyme on human corneal epithelial cells Investigative Ophthalmology and Visual Science 2018;59(5):2006-2014 [ Show Abstract ]

PURPOSE. During contact lens wear, the amount of lysozyme deposited on contact lenses varies depending on the lens material. The binding of lysozyme to some contact lens materials may result in a conformational change that denatures the protein to an inactive form. This investigation evaluated the effect that denatured lysozyme has on human corneal epithelial cells (HCECs) by measuring cell viability and the release of inflammatory cytokines. METHODS. HCECs were exposed to lysozyme that was denatured to various activity levels. After 24-hour exposure to the lysozyme (1.9 mg/mL) in growth media, the cells were evaluated for cell viability using confocal microscopy. The metabolic activity of the cells was determined using an alamarBlue assay. Cell supernatants were analyzed for inflammatory cytokines. RESULTS. Using confocal microscopy, there was no detectable change in the viability of the HCECs after exposure to the denatured lysozyme. However, using alamarBlue, a decrease in the metabolic activity of the HCECs exposed to denatured lysozyme was detected. HCECs exposed to lysozyme that was 67%, 47%, and 22% active showed a reduction in metabolic activity when compared with native (100% active) lysozyme and the media controls (P < 0.05). Exposure to the denatured lysozyme also caused an increase in the release of inflammatory cytokines (P < 0.05) from the HCECs. CONCLUSIONS. The results of this study show that denatured lysozyme can have a detrimental effect on HCECs. Both a reduction in metabolic activity and an increase in the release of inflammatory cytokines occurred after HCEC exposure to denatured lysozyme. © 2018 The Authors.


Liu,L. Y., Seo,J., McCanna,D. J., Subbaraman,L. N., Jones,L. W. Assessment of biofilm formation of E. meningoseptica, D. acidovorans, and S. maltophilia in lens cases and their growth on recovery media Contact Lens and Anterior Eye 2016;39(2):117-123 [ Show Abstract ]

Purpose: Bacterial biofilm formation in contact lens cases is a risk factor in the development of both microbial and infiltrative keratitis. This investigation evaluated three emerging pathogens: Stenotrophomonas maltophilia, Elizabethkingia meningoseptica, and Delftia acidovorans for biofilm formation and metabolic activity in lens cases. Also, growth of these bacteria on different media was assessed to optimize recovery conditions. Methods: The three bacteria were incubated in lens cases with different concentrations of tryptic soy broth. Biofilm formation was evaluated by measuring metabolic activity using MTT and enumerating the number of viable bacteria. To determine the optimal recovery media, dilutions of these microorganisms were plated on six different media. The number of colony forming units (CFU) was recorded after 48, 72, and 96 h of incubation at 32 °C and 37 °C for S. maltophilia, and at 37 °C for E. meningoseptica and D. acidovorans. Results: All three microorganisms established biofilms in the lens cases, with significant numbers of CFU recovered. Biofilms of S. maltophilia and E. meningoseptica were metabolically active. Significant reduction in metabolic activity and number of viable S. maltophilia occurred when the incubation temperature was raised from 32 °C to 37 °C (p < 0.05). The metabolic activity of the biofilms increased with greater organic load present. The highest percent recovery for all three organisms was given by Columbia blood agar, followed by chocolate. Conclusion: Based on the results, the presence of the three emerging pathogens present in lens cases and from corneal isolates can be accurately determined if proper growth media and incubation temperatures are utilized.

Scientific Presentations


McCanna D, Oh S, Seo J, Coles_brennan C, Fadli Z, Subbaraman L. In vitro evaluation of the effect of lysozyme coated contact lenses on cell viability and inflammatory response BCLA Clinical Conference and Exhibition, 2015 [ PDF ]

McCanna D, Oh S, Seo J, Subbaraman L, Coles-Brennan C, Fadli Z, Jones L. Effect of Denatured Lysozyme on Human Corneal Epithelial Cells Invest Ophthalmol Vis Sci 2015;56: E-abstract 3511 [ PDF ]


McCanna D,Liu L, Seo J, Subbaraman L, Jones L. Assessment of the growth of Stenotrophomonas maltophilia, Elizabethkingia meningoseptica and Delftia acidovorans in contact lens cases and on recovery media Invest Ophthalmol Vis Sci 2014;55: E-abstract 6051