Effects of Repeated Aurora-A siRNA Transfection on Cilia Generation and Proliferation of SK-MES-1 or A549 Cells.

Abstract

Suppression of Aurora kinase A (Aurora-A, AURKA) by siRNA of Aurora-A (siAurora-A, siA) has been used in lung tumor treatment. However, the dose and frequency of gene transfection still need to be confirmed further. We imitated multiple administration of solid tumor and attempted to make out the effects of thrice transfection of siAurora-A on cilia generation and apoptosis of SK-MES-1 cells (SK) or A549 cells. The Aurora-A mRNA levels of cells cultured with serum for 6 d or without serum for 2, 4, or 6 d were examined with real-time quantitative PCR; Cells were transfected single or repeatedly with siAurora-A or siControl (siC), their Aurora-A mRNA levels were determined with PCR; Their cilia were examined with immunohistochemistry. Cell viability was measured with the MTT assay. Protein expression was analyzed with western blot. Cell viability showed a downward trend along with the prolongation of starvation time to the second, fourth, and even to the sixth day in both types of cells. But, the expression level of Aurora-A mRNA flipped to rise at the sixth day instead of decreasing at the fourth day. Protein expression trend of total Aurora-A in the two groups was consistent with Aurora-A mRNA expression trend. Compared with siC-3 group (transfected three times with siControl), siAurora-A significantly reduced the Aurora-A mRNA expression in siA-3 group (transfected three times with siAurora-A). Similarly, the cell viability of siA-3 group was lower than that of siC-3 group. The cell viability of siC-3 group was higher than that of serum-free-6d group, but, levels of Aurora-A mRNA expression of siC-3 group had no difference with serum-free-6d group. Finally, among groups transfected once or three times or starved for 6 d, there was no significant difference of ciliated cell proportions in both types of cells respectively. Repeated siAurora-A transfection decreased Aurora-A expression that resulted in effective suppression proliferation of SK-MES-1 or A549 cells, but did not affect cilia generation.