Time-dependent morphological alterations and viability of cultured human trabecular cells after exposure to Trypan blue

• Published in: Clinical & experimental ophthalmology Vol. 41; no. 5; pp. 484 - 490
• Main Authors: Tsaousis, Konstantinos T, Kopsachilis, Nikolaos, Tsinopoulos, Ioannis T, Dimitrakos, Stavros A, Kruse, Friedrich E, Welge-Luessen, Ulrich
• Format: Journal Article
• Language: English
• Published: Australia Blackwell Publishing Ltd 01-07-2013; Wiley Subscription Services, Inc
• Subjects: Benzimidazoles; Cell Survival - drug effects; Cells, Cultured; Coloring Agents - toxicity; Fluorescent Dyes; Humans; Microscopy, Phase-Contrast; Middle Aged; Propidium - metabolism; Time Factors; toxicity; Trabecular Meshwork - drug effects; Trabecular Meshwork - pathology; trabecular meshwork cell; Trypan blue; Trypan Blue - toxicity; viability
• ISSN: 1442-6404; 1442-9071
• DOI: 10.1111/ceo.12018

Background
To determine the time‐dependent toxicity of Trypan blue at 0.06% concentration in cultured human trabecular meshwork cells.
Methods
Human trabecular meshwork cells cultured in vitro were exposed to Trypan blue and acute toxicity was evaluated. Cells were exposed for 5, 30, 60 s and for 5, 15, 30, 60 min to a commercially available Trypan blue preparation (Vision Blue; DORC International Zuidland, The Netherlands). Morphology was observed by phase‐contrast microscopy and cell viability was measured using Hoechst 33342 (Intergen; Purchase, NY, USA) and propidium iodide assays to determine the percentage of living and dead cells.
Results
Morphological changes occurred mainly after 5 min of exposure to Trypan blue. Viability was 96.0% ± 3.6%, 94.8% ± 3.5% and 92.5% ± 4.4% after 5, 30 and 60 s of exposure, respectively; a significant toxic effect of Trypan blue was observed after 5, 15, 30 and 60 min of exposure with a viability of 85.0% ± 3.7%, 77.0% ± 6.7%, 70.8% ± 5.9% and 68.3% ± 8.7%, respectively.
Conclusions
Trypan blue is not toxic, in terms of cell viability, over an exposure time of up to 60 s; however, further exposure results in a gradual increase in damage of cultured human trabecular meshwork cells.