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. 2009 Feb;11(2):157–166. doi: 10.1593/neo.81074

Figure 2.

Figure 2

ZEB-1 downregulates SEMA3F transcription in H661 cells. (A) H661 cells were transiently transfected with control (CTL; black bars) or ZEB-1 expression vector (ZEB-1; white bars) and were harvested over time to measure ZEB-1, E-Cadherin, and SEMA3F expressions by quantitative real-time RT-PCR. Values are expressed in percentage of GAPDH expression. ZEB-1 protein was checked by Western blot analysis at 48 hours after transfection (top right). α-Tubulin was used as a loading control. The low E-Cadherin mRNA level in H661 cells is in accordance with previous results where H661 cells were selected among NSCLC for their undetectable E-Cadherin [20]. Reasons of the absence of E-Cadherin are the loss of Wnt7a/β-catenin pathway and ZEB-1 expression. (B) H661 cells were transfected with different amounts of ZEB-1 siRNA. ZEB-1, ZEB-2, E-Cadherin, and SEMA3F mRNA were monitored 48 hours after transfection. Values for three independent experiments (except for ZEB-2), done in duplicate, are expressed in percentage of GAPDH expression. Bars, SD. Statistical analysis was performed with Student's t-test: *P < .05, P < .01.