Figure 3

Knockdown efficiency of COL1A1 and IFITM1 and effect on cell invasion. (a) COL1A1 and IFITM1 mRNA expression was monitored by quantitative reverse transcriptase–PCR in GSC-3 and GSC-9 (n=2). Significant inhibition of 90% of mRNA expression was observed. GADPH was used as internal control. *P<0.05. Each value is expressed as mean±s.e.m. (b) Western blot analysis showed diminution of IFITM1 and COL1A1 proteins expression, β-actin was used as loading control. (c) Densitometry of protein level. The bar graph shows relative IFITM1 and COL1A1 protein expression levels expressed as percentage of IFITM1 and COL1A1 expression in control cells (n=3). We observed >80% inhibition for both genes in GSC-3 and GSC-9 cells, as compared with control cells. (d) Representative fields of Boyden cell invasion assay. After 3 days of invasion, cells that migrated across the membrane were fixed and stained with hematoxylin/eosin and were counted in one hundred different fields. The invaded cells were photographed under the microscope at × 100 magnification. (e) A significant reduction of invasion is observed in both GSC-3 and GSC-9 cells lines with sh-IFITM1 as well as for sh-COL1A1. The results represent a percentage of invaded cells. Data are presented as mean±s.e.m. from three independent experiments. *P<0.05, **P <0.01.