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. 2000 Apr 15;105(8):1147–1156. doi: 10.1172/JCI7545

Figure 4.

Figure 4

The ERK and p38 pathways regulate toxin A–induced IL-8 gene expression. (a) THP-1 cells were transiently transfected with a luciferase reporter construct carrying the IL-8 gene promoter region. Then, cells were preincubated with PD98059 (20 μM) for 30 minutes and stimulated with C. difficile toxin A (100 nM) or LPS (1 μg/mL) for 4 hours. PD98059 prevented IL-8 gene expression by each stimulus. Means and SE are shown (n = 3; AP < 0.05). (b) Transfected cells were preincubated with SB203580 (10 μM) for 30 minutes and then stimulated with C. difficile toxin A (100 nM) or LPS (1 μg/mL) for 4 hours. SB203580 blocked IL-8 gene expression induced by toxin A, but not by LPS (n = 3; AP = 0.003). (c) The involvement of the p38 pathway in toxin A–induced IL-8 gene expression was confirmed by transient expression of dominant-negative mutants of MKK3 and MKK6 (MKK3/6 dominant negative [dn]). Total DNA was kept constant at 7 μg using control vector and included 1 μg reporter plasmid DNA. Whereas MKK3 dn or MMK6 dn did not significantly affect IL-8 reporter gene expression, transfection of both mutants abrogated luciferase activity induced by toxin A and by LPS. Means and SE of 2 experiments, each done in triplicate, are shown (AP < 0.01).