Effects of triamcinolone acetonide on human trabecular meshwork cells in vitro

Aim: To study the effects of triamcinolone acetonide (TA) on cultured human trabecular meshwork (HTM) cells. Materials and Methods: HTM cells were cultured and treated with 125, 250, 500 and 1000 mu g/mL concentration of TA for 24 h. The cells were treated with both crystalline TA (TA-C) (commercial preparation) and solubilized TA (TA-S). Cell viability was measured by a trypan blue dye exclusion test. The activity of caspse-3/7 was measured by a fluorescence caspase kit and DNA laddering was evaluated by electrophoresis on 3% agarose gel. Levels of lactate dehydrogenase (LDH) were assessed with LDH cytotoxicity assay kit-II. Results: Mean cell viabilities of HTM cells after 24 h exposure to TA-C 125, 250, 500, and 1000 mu g/mL were 75.4 +/- 2.45% (P < 0.0001), 49.43 +/- 1.85% (P < 0.0001), 17.07 +/- 2.39% (P < 0.0001), and 3.7 +/- 0.9% (P < 0.0001), respectively, compared with the untreated HTM cells 92.49 +/- 1.21%. The mean cell viabilities with 125, 250, 500, and 1000 mu g/mL of TA-S were 94.47 +/- 1.60% (P > 0.05), 90.13 +/- 0.40% (P < 0.01), 85.57 +/- 0.47% (P < 0.001), and 71.67 +/- 3.30% (P < 0.0001), respectively, compared to DMSO-equivalent cultures. Untreated HTM control had a cell viability of 96.57 +/- 1.98%. DMSO-treated controls of 125, 250, 500, and 1000 mu g/mL had a cell viability of 94.73 +/- 0.57%, 96.97 +/- 1.08%, 93.97 +/- 1.85%, and 97.27 +/- 1.15%, respectively. There was no increase of caspase-3/7 activity in cultures treated with either TA-C or TA-S. DNA laddering showed no bands in the TA-C or TA-S treated cultures. There were significantly higher LDH release rates at all concentrations of TA-C compared to TA-S. Conclusions: Results show that the effect of TA-C and TA-S on HTM cells is due to cell death by necrosis at all concentrations except 125 mu g/mL of TA-S. Elevated levels of LDH confirmed necrotic cell death. Our study also infers the relative safety of TA-S over TA-C.