کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
4324865 1613949 2013 9 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Investigation of the performance of PEG–PEI/ROCK-II-siRNA complexes for Alzheimer's disease in vitro
موضوعات مرتبط
علوم زیستی و بیوفناوری علم عصب شناسی علوم اعصاب (عمومی)
پیش نمایش صفحه اول مقاله
Investigation of the performance of PEG–PEI/ROCK-II-siRNA complexes for Alzheimer's disease in vitro
چکیده انگلیسی

Recent studies have showed inhibiting ROCK promoted axonal regeneration and suppressing ROCK-II decreased Aβ formation, suggesting ROCK is a potential target for the treatment of Alzheimer's disease. Because ROCK-II mRNA is abundantly expressed in brain, we targeted ROCK-II mRNA using a siRNA approach. To suppress ROCK-II mRNA expression, we synthesized PEG–PEI/ROCK-II-siRNA complexes and transfected C17.2 neural stem cells in vitro. The characteristics of the complexes were tested using a gel retardation assay. Particle size and zeta potential were examined using dynamic light scattering and the morphology of the complexes were observed by transmission electron microscopy. The toxicity was detected by an MTT assay and transfection efficiency was determined by flow cytometry. Laser confocal microscopy was employed to investigate the cell uptake of the complexes. RT-PCR and western blotting were used to verify the effect of gene silencing. Our results indicated that the characteristics of the complexes depended on the N/P ratios. At a high N/P ratio, PEG–PEI could completely condense the siRNA into small-sized uniform particles. However, high N/P ratios are accompanied with high cytotoxicity. Because of high transfection efficiency and low cytotoxicity, N/P=50 was chosen to transfect C17.2 cells in vitro. Laser confocal microscopy showed that ROCK-II-siRNA with green fluorescence was mainly distributed in the cytoplasm and synapses. Moreover, ROCK-II-siRNA was successfully released from the lysosome. RT-PCR and western blotting demonstrated effective gene silencing. These results indicated that PEG–PEI/ROCK-II-siRNA complexes effectively suppressed ROCK-II mRNA expression, providing the basis for future research in vivo.


► Suppressing the expression of ROCK is a strategy for treating AD.
► The delivery of ROCK-II siRNA is crucial to inhibit ROCK-II mRNA expression in CNS.
► Non-viral vector PEG–PEI was employed for condensed ROCK-II siRNA.
► Complexes at N/P=50 displayed high transfection efficiency and low cytotoxicity.
► PEG–PEI/ROCK-II-siRNA complexes could suppress ROCK-II mRNA expression effectively.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Brain Research - Volume 1490, 15 January 2013, Pages 43–51
نویسندگان
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