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 µg/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 µg/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 µg/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 µg/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 µg/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.

Evaluation of the toxicity of triamcinolone acetonide and dexamethasone sodium phosphate on human lens epithelial cells (HLE B-3).

PURPOSE: The purpose of this study was to compare the in vitro effects of triamcinolone acetonide (TA) and dexamethasone sodium phosphate (DEX) on human lens epithelial cells (HLE B-3). METHODS: HLE B-3 cells were exposed for 24 h to commercially available TA (c-TA) and dimethylsulfoxide-solubilized TA (s-TA). The cells were treated with 1,000 (clinical dose), 750, 500, 200, and 100 µg/mL concentrations of c-TA, s-TA, and supernatant for 24 h. The cells were also treated with DEX at 2, 1, 0.5, 0.2, 0.1 (clinical dose), and 0.05 mg/mL. Cell viability, caspase-3/7 activity, and DNA fragmentation analyses were performed. RESULTS: The mean cell viabilities of HLE B-3 after exposure to c-TA at 1,000, 750, 500, 200, and 100 µg/mL were significantly reduced compared with control untreated cells. The s-TA also significantly reduced cell viability at 1,000, 750, and 500 µg/mL compared with dimethylsulfoxide control. The supernatant did not reduce cell viability. Caspase-3/7 activity significantly increased after treatment with c-TA and s-TA. DNA laddering revealed bands at 200 bp intervals with both c-TA at≥100 µg/mL and s-TA at ≥500 µg/mL. The cell viabilities of HLE B-3 after 24 h exposure to DEX were significantly reduced at 2 and 1 mg/mL but not at lower concentrations tested. Caspase-3/7 activities in HLE B-3 cells were not increased significantly after treatment with DEX at any dose tested. DNA laddering did not reveal any band at any dose tested. CONCLUSION: This study showed that TA at its clinical dose (1,000 µg/mL) in both commercial preparation and solubilized forms decrease HLE B-3 cell viability through an apoptotic pathway. DEX at its clinical dose (0.1 mg/mL) does not decrease cell viability or cause any increase of caspase-3/7 activity. This study suggests that for long-term sustained-release devices, DEX may be less damaging to human lens cells than TA.