CCK modulates LPS-induced nitrite production and iNOS expression by peritoneal macrophages. The macrophage culture was pretreated with different CCK concentrations (10−10–10−6 M) or aminoguanidine (200 μM) for 30 min before the addition of LPS (1 μg/mL) to the cell culture medium (time zero). At the indicated time-points, the culture supernatant was collected for nitritequantification by the Griess method (a), while adherent cells were harvested for iNOS expression determination. (b) Total mRNA was extracted and the expression of iNOS mRNA relative to GAPDH mRNA was analyzed by real-time PCR using specific gene primers. (c) Representative immunoblots and densitometry values (iNOS/β-actin ratio) of whole cell lysates. The β-actin levels were used as an internal control. Data is a representative of five independent experiments with similar results. Values are expressed as means ± S.E.M. –10 samples per group. * versus saline and # versus LPS group.
