Abstract
Abstract: :
Purpose:To study whether Nipradilol, which is used as the ophthalmic solution for the treatment of glaucoma, had the cytoprotective effect, we investigated the effect of Nipradilol on the apoptosis induced by serum withdraw in PC12 cells. Nipradilol is the compound which have ?1, ß blocking and NO releasing property. We also investigate the effect of Timolol, Prazosin and SNAP on PC12 cell death. Methods:Undifferenciated PC12 cells were maintained in RPMI medium containing 10% HS and 5% FBS, which were washed with serum-free RPMI 1640 medium for 3times and replated onto collagen-coated 24 well plates at a density of 2X105 cells per well. Cells were treated with various concentration of Nipradilol, Timolol maleate (ß-blocker), Prazosin hydrochloride (?1 blocker) or S-Nitroso-N-acetyl-DL-penicillamine (SNAP). Cells were stained with Hoechst33258 (10µM) for 30min at 37 ?C observed under the fluorescence microscope. The nuclei shown apoptic and normal feature were counted and indicated as % of survival. Activity of caspase-3 was measured by using fluorogenic substrate peptide, Ac-DEVD-AMC. Cell lysate was incubated with 100µM substrate peptide in 200µl of incubation buffer at 37 ?C for 60 min. The released AMC was measured by the fluoro-micro plate reader Results:C12 cells resulted in apoptosis by serum withdraw and the survival rate was decreased in the time dependent mannar. The addition of Nipradilol into medium showed the cytoprotecive effect on PC12 cell death. Timolol and Prazosin had no protective effect on PC12 cell death. We measured caspase-3 activity to clarify the mechanism of the inhibition of apoptosis under the existence of DTT or not Because the caspase-3 activity could be reactivated by the existence of DTT. The caspase-3 activity was increased by the addition of DTT. Conclusion:These results indicated that Nipradilol has the cytoprotective effect on PC12 cell death. It is one possibility that the protective effect was caused by the caspase-3 inhbition mediated NO related S-nitrosylation.
Keywords: 323 apoptosis/cell death • 491 nitric oxide • 489 neuroprotection