Apoptosis and morphologic changes in drug-treated trabecular meshwork cells in vitro

Using an in vitro culture system, we investigated whether bovine trabecular meshwork cells undergo apoptosis (programmed cell death) following exposure to anti-glaucoma medications (timolol, pilocarpine and epinephrine) and known inducers of apoptosis (5-fluorouracil, mitomycin-C and dexamethasone). Third to fifth passage bovine trabecular meshwork cells were grown to confluence and incubated for 1-12 days in growth media with timolol (1-1000 microM), pilocarpine (15-15,000 microM), epinephrine (5-5000 microM), 5-fluorouracil (10-100 micrograms ml-1), mitomycin-C (0.01-100 micrograms ml-1) and dexamethasone (0.01-100 microM). The cultures were evaluated for apoptosis by phase-contrast microscopy, transmission electron microscopy and in situ apoptosis labeling. 5-Fluorouracil (10-100 micrograms ml-1), mitomycin-C (0.1-100 micrograms ml-1) and epinephrine (500-5000 microM) induced apoptosis in a dose and time-dependent manner. Timolol, pilocarpine, and dexamethasone-treated specimens did not show evidence of apoptosis at any of the concentrations tested. Trabecular meshwork cells incubated in timolol (100-1000 microM) developed cytoplasmic granules, and specimens treated with pilocarpine (15,000 microM) developed cytoplasmic vacuoles. These granules and vacuoles have the appearance of secondary lysosomes. Dexamethasone-treated cells developed an increased number of mitochondria. This study suggests that the trabecular meshwork may undergo apoptosis following exposure to 5-fluorouracil, mitomycin-C and epinephrine. Timolol, pilocarpine and dexamethasone did not induce apoptosis. However, these drugs can incite characteristic morphologic changes in cultured trabecular meshwork cells.