276. RNAi-Based Therapeutics Targeting nNOS for Prevention of Neuroblastoma Cell Apoptosis

摘要: The death of neurons by apoptosis is a key step in natural history of various degenerative diseases. The nNOS enzyme is expressed in brain areas submitted to injury, and its blocking can decrease its neurotoxic effects. This study was aimed to test an RNAi-based strategy targeting nNOS to prevent cellular apoptosis. Firstly, neuroblastoma cells (ATCC® CRL-2266™-SH-SY5Y) were plated in 96-well plates (5X104 cells/well) and incubated for 24 hours with the injuring agent neomycin®(300 µg/ml, Sigma). After that, cells received 0.2 µg of plasmid vectors mixed with Lipofectamine®(Invitrogen) and were incubated for 48h. The two vectors used in our study contained the shRNA 28_hnNOS sequence (pe28_hnNOS) or a scrambled negative control sequence (pScramble) cloned into pSilencerTM 3.1-H1 (Applied Biosystems, Ambion®). Finally, the cell viability was determined by using the colorimetric MTT (3-(4, 5 …