The effect of TGFβ on LPS-induced chemokine mRNA and protein levels. Adherent TG-elicited macrophages (1 × 10⁷ cells/100-mm Petri dish) were stimulated with LPS (10 ng/mL) in the presence or absence of TGFβ (20 ng/mL) for the indicated times. (A) Total RNA was prepared and levels of KC, MIP-2, IP-10, and MCP-1 mRNA were analyzed by Northern hybridization using 20 μg total RNA in each lane. Blots were hybridized with the indicated radiolabeled cDNA probes. Similar results were obtained in 3 separate experiments. (B) The blot in panel A was quantified by phosphorimage analysis. Values presented are in arbitrary units where each chemokine level is normalized to the level of GAPDH mRNA in the same sample. (C) The levels of chemokine protein in the culture medium were analyzed by ELISA. Values are means ± SEM of 3 separate experiments. □ indicates no treatment (UT); ▨, LPS only; and ▪, TGF + LPS.