Abstract: 【Abstract】
Objective:Aim to pick out the siRNA which could most effectively inhibit the expression of caveolin-1 in endothelial cells and to detect the cytotoxicity of the transfection complex. Methods: Four siRNAs were chemically synthesized: siRNA422, siRNA548, siRNA710 were used to inhibit caveolin-1 expression in endothelial cells, FAM-labeled mismatch siRNA was as a negative control. They were all transfected into endothelial cells, respectively. Caveolin-1 mRNA was detected after transfection by Real-Time PCR and its protein expression was measured by Western blot. The cytotoxicity was analyzed with the method of the Sulforhodamine B (SRB). Results ①The transfection rates are all above 85% under the transfection condition of siRNA (20, 30 or 40 nmol•L-1) combined with the same dose of siRNAFect (P<0.01);②The survival rates of transfected cells are all above 80% under the transfection condition of siRNAFect1. 3μL combined with siRNA (10 or 20nmol•L-1)(P<0.05); ③The expression of caveolin-1 mRNA in EA.hy926 cells was significantly decreased after siRNA548 treatment (P<0.01); ④ Compared with siRNA422, siRNA710, vehicle or mismatch siRNA-treated group, the expression of caveolin-1 protein was the least after siRNA548 treatment 48 hours later (P<0.01). Conclusions ①The siRNA548 can inhibit caveolin-1 expression most effectively in endothelial cells. ②siRNA (20 nmol•L-1) combined with siRNAFect (1.3 μL) has lower cytotoxicity and higher transfection efficiency, which is suitable for transfection.
|