Figure 4: VIP/SP and ACh/nicotine modulate inflammatory mediators in a cumulative manner. (a) RAW peritoneal macrophages were incubated with a dose range of VIP (0–10 μM) for 30 min prior to 3 hrs LPS stimulation (100 ng/mL). TNF production was measured with ELISA. (b) cells were pretreated with 10 μM ACh/nicotine for 30 min, then stimulated with 10 μM VIP or SP, and followed by 100 ng/mL LPS challenge. TNF levels were measured after 3 hrs and depicted as % of vehicle-treated cells. Data are mean s.e.m. of three independent experiments done in triplicate. * ; ** . E.P. van der Zanden et al., Tytgat Institute AMC, unpublished 2012.