Figure 5. Effect of TGF-β on TNF-α receptor expression and sensitivity of 4T1 cells to induction of apoptosis by TNF-α.
A. Expression of TNF-α receptor 1 (TNFRSF1a) mRNA and protein in 4T1 cells were treated for 2 days in vitro with 5ng/ml TGF-β or vehicle control. TNFRSF1a mRNA was determined by RTQ-PCR, and the level of TNFR1 protein in 4T1 cell lysates was detected by ELISA assay. Results are the mean +/- SD for 3 determinations. B. RTQ-PCR analysis of TNFRSF1a expression in 4T1 primary tumors following 9 days of treatment in vivo with anti-TGF-β (1D11) or control (CON) antibody. TNFRSF1a expression was normalized to the 28S rRNA in each case. Results are mean +/- SD for 5 tumors/treatment group. C. In vitro apoptosis assay in 4T1 cells after treatment with TGF-β (5ng/ml) or vehicle control for 4 days, with TNF-α (25ng/ml) also present for the final two days. Results are normalized to the no TGF-β, no TNF-α control conditions, and are the mean +/- SD of 3 determinations. * indicates P<0.001, Student's T-test.