The cytotoxic and stress responses of human trabecular meshwork cells treated with triamcinolone acetonide.
OBJECTIVE
To evaluate the cytotoxic effect of triamcinolone acetonide (TA) on cultured human trabecular meshwork (TM) cells.


METHODS
TA (0.1 mg/ml, 1 mg/ml) or the vehicle (benzyl alcohol, 0.0025%, 0.025%) was added to human TM cell cultures on day 0 and collected subsequently on day 1, 3, or 5. The amount of cell proliferations with or without TA treatment was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylterazolium bromide (MTT) assay. All samples were read in triplicate (n=4 in all cases). By using real-time quantitative polymerase chain reaction (PCR), gene expression levels of c-fos, c-jun, caspase-3, c-myc, and p53 were determined after TA treatments at 0 min, 10 min, 20 min, 30 min, 50 min, 80 min, 2 h, 12 h, 24 h, and 48 h. Unpaired t-test was used to test the drug and concentration effects of TA, ANOVA was used to test the time effects of TA, and the Bonferroni test was used to correct multiple comparisons. Apoptosis of TM cells as a result of TA treatment were assessed by the terminal uridyl nick end labeling (TUNEL) assay.


RESULTS
Both concentrations of TA caused a significant reduction in the number of human TM cells as early as day 1 and across five days of the treatment period. Significantly increased expressions of c-jun, c-fos, c-myc, p53, and caspase 3 were observed at different time points after both 0.1 mg/ml and 1 mg/ml TA treatment. Significantly increased apoptotic cells were observed after TA treatment for three days.


CONCLUSIONS
Our results showed that TA was cytotoxic to human TM cells in culture and the presence of TA caused apoptotic cell death. It gave evidence that the underlying mechanism of TA caused ocular hypertension and may be associated with necrosis and apoptosis of the TM cells.
Wang
Dan Yi
DY
Department of Ophthalmology & Visual Sciences, the Chinese University of Hong Kong, Hong Kong, China.
Fan
Bao Jian
BJ
Yam
Gary Y H
GY
Lam
Dennis S C
DS
Pang
Chi Pui
CP
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002452:Cell Count; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression / Q000187:drug effects; D005938:Glucocorticoids / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014129:Trabecular Meshwork / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D014222:Triamcinolone Acetonide / Q000008:administration & dosage / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D005938:Glucocorticoids; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; D053148:Caspase 3; D014222:Triamcinolone Acetonide