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Kang, Hwang, and Kim: Effect of Trypan Blue on the Survival of Cultured Trabecular Meshwork Cells
Original Article

Journal of the Korean Ophthalmological Society 2017; 58(3): 327-332.

Published online: 15 March 2017

DOI: https://doi.org/10.3341/jkos.2017.58.3.327

Effect of Trypan Blue on the Survival of Cultured Trabecular Meshwork Cells

Hye Ri Kang, MD, Seok Jin Hwang, MD, Jae Woo Kim, MD, PhD

Department of Ophthalmology, Catholic University of Daegu School of Medicine, Daegu, Korea.

Address reprint requests to Jae Woo Kim, MD, PhD. Department of Ophthalmology, Daegu Catholic University Medical Center, #33 Duryugongwon-ro 17-gil, Nam-gu, Daegu 42472, Korea. Tel: 82-53-650-4728, Fax: 82-53-627-0133, jwkim@cu.ac.kr

Received 28 July 2016       Revised 17 August 2016       Accepted 31 January 2017

©2017 The Korean Ophthalmological Society

(open-access):

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Purpose

To evaluate the effects of trypan blue (TB) on the survival of cultured human trabecular meshwork cells (HTMCs).

Methods

Primarily cultured HTMCs were exposed to 0.05, 0.10 or 0.50% TB for 1, 5 or 30 min. Cellular survival was assessed using the MTT assay and degree of apoptosis was analyzed with flow cytometry using annexin-V/propidium iodide double staining.

Results

Long-term exposure or high concentration of TB decreased the survival of HTMCs (p < 0.05). In flow cytometric analysis, exposure to 0.50% TB for 30 min increased the degree of apoptosis (p < 0.05). Commercial TB decreased cell survival after exposure for 5 min and increased the degree of apoptosis after exposure for 30 min (p < 0.05).

Conclusions

TB may cause cellular damage of cultured HTMCs and apoptosis could be the underlying mechanism. In TB-assisted cataract surgery, TB should be used for the shortest time possible and removed completely.

Keywords: Apoptosis, Flow cytometry, Survival, Trabecular meshwork cell, Trypan blue

Figures and Tables

Figure 1

Effects of trypan blue on the survival of human trabecular meshwork cells. Exposure to trypan blue for 1 minute decreased cellular survival significantly at the concentration of 0.50%. *p < 0.05.

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Figure 2

Effects of trypan blue on the survival of human trabecular meshwork cells. Exposure to trypan blue for 5 minutes decreased cellular survival significantly at the concentration of 0.50%. *p < 0.05.

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Figure 3

Effects of trypan blue on the survival of human trabecular meshwork cells. Exposure to trypan blue for 30 minutes decreased cellular survival significantly at the concentration of 0.10% and 0.50%. *p < 0.05.

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Figure 4

Effects of 0.06% commercial trypan blue on the survival of human trabecular meshwork cells. Exposure to trypan blue for 5 minutes or 30 minutes decreased cellular survival. *p < 0.05.

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Figure 5

Flow cytometric analysis of apoptosis using annexin-PI double staining. Exposure to 0.50% trypan blue for 30 minutes increased the degree of apoptosis significantly compared to non-exposed control. *p < 0.05.

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Figure 6

Flow cytometric analysis of apoptosis using annexin-propidium iodide double staining. 0.06% commercial trypan blue exposed for 30 minutes increased the degree of apoptosis significantly compared to non-exposed control. *p < 0.05.

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