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Fig. 3.
Early TNF release of macrophages depends on the LPS structure of E. coli cells. TNF released by macrophages was detected by L929/NF-κB-luc reporter PXD101 as described in Materials and methods. (A) RAW 264.7 cells released TNF in response to LPS in a concentration dependent manner. (B) In response to MG CL (MG1655) and MG L strains, macrophages released about the same amount of TNF into the supernatant. Introduction of MG OCL bacteria caused higher tumor necrosis factor production, but this difference was not significant. MG CΔL bacteria caused the production of significantly less TNF compared with the TNF produced in response to MG CL. (p cells in response to the MG OCL, the MG CL, and MG L strains, while MG CΔL bacteria caused the production of significantly less TNF ( Fig. 3B). We confirmed the results with ELISA, i.e., the NF-κB activation was indeed due to TNF, and furthermore, the presence of potential contaminating bacterial products did not interfere with the bioassay. The differences in TNF production did not depend on the presence or absence of serum in the medium (data not shown).