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Fig. 5. Involvement of COX-2 in NFAT3-mediated anchorage-independent growth of Cl41 cells induced by TNF-. (A) Cl41-mock vector and Cl41-siCOX-2 cells (2x10⁵) were seeded into each well of a 6-well plate, and cultured in 5% FBS MEM at 37°C. When the cell density reached 70-80%, the cells were exposed to 20 U/ml of TNF- for 24 hours. The induction of COX-2 at the protein level was detected by western blot. (B) The soft agar assay was performed as indicated in Fig. 4A; 2x10⁴ of Cl41-mock vector and Cl41-siCOX-2 cells were exposed to 20 U/ml of TNF- for 3 weeks. (C) Schematic diagram outlining the anchorage-independent cell growth shown in panel B. (D) The soft agar assay was performed as indicated in Fig. 4A, 2x10⁴ of Cl41 cells were exposed to 20 U/ml of TNF- with various concentrations of NS398 for 3 weeks. (E) Schematic diagram outlining the anchorage-independent cell growth shown in panel D. (F) The proposed NFAT pathway involved in TNF--induced anchorage-independent cell growth.

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