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- W2392821486 abstract "Objective To investigate the inhibitory effects of RNA interference (RNAi) expression vector on the expression of aquaporin-4(AQP4) gene in astrocyte primary cultures.Methods RNAi expression vector, which specifically suppressed AQP4 gene expression(AQP4 RNAi) and a control vector served as a non-silencing control were constructed, and the neonatal rat astrocytes in primary cultures were nucleofected with the constructed vector. The astrocytes in primary cultures were randomly assigned to non-transfected group, transfected with control vector group, and transfected with AQP4 RNAi expression vector group. The changes of astrocytic morphology were observed under light microscopy and cell growth rate was judged by cell counting at 1~20 day after transfection. The expression levels of AQP4 protein and mRNA were detected by Western blot and real-time PCR respectively; cellular volume was determined using [~ 3 H]-3-O-methyl-D-glucose.Results There was no difference in astrocytic appearance and cell growth rate between AQP4 RNAi expression vector transfected and control vector-treated cells (P0.05). ②Real-time PCR experiment and Western blotting analysis showed that a progressive and parallel reduction of the expression of AQP4 mRNA and protein in cultured astrocytes after transfection with AQP4 RNAi expression vector compared to that of control vector-treated and untransfected wild type astrocytes(P0.05), and reached maximal inhibition, around 85.7% and 77.5% in AQP4 protein and mRNA respectively at 48 hours, and maintained 14 days after transfection. But AQP4 mRNA and protein expression levels were unaffected in control vector-treated and untransfected cells(P0.05). ③After exposure to hypotonic medium(250 mmol·L~ -1 ) for 10,20,30 and 40minutes, the astrocytes showed a time-dependent increase in cellular volume, and the cellular volume was significantly decreased in astrocytes with AQP4 gene silencing than that of control vector-treated cells at the same hypotonic medium(P0.05).Also, there was no difference in cellular volume between control vector-treated and untransfected astrocytes, the results suggested that cellular water permeability in hypotonic medium were decreased significantly in astrocytes with AQP4 gene silencing. Conclusions The RNAi expression vector can effectively inhibit the expression of AQP4 gene in cultured rat astrocytes, and AQP4 may be the major factor responsible for the water transport of cultured astrocytes." @default.
- W2392821486 created "2016-06-24" @default.
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- W2392821486 date "2006-01-01" @default.
- W2392821486 modified "2023-09-23" @default.
- W2392821486 title "RNA interference mediates silencing of aquaporin-4 gene in cultured neonatal rat astrocytes" @default.
- W2392821486 hasPublicationYear "2006" @default.
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