Fig. 3.

pLTA inhibited rhTNF-α-mediated iNOS and NO production. (A) HT-29 cells were treated with the indicated dose of pLTA for 20 h and then restimulated with cytokine cocktail (100 ng/ml of rhTNF-α, 300 U/ml of IFN-γ, and 10 ng/ml of IL-1α) for 48 h. Total RNA was extracted, cDNA was synthesized, and PCR was performed with specific primers for iNOS and GAPDH (upper panel). The iNOS mRNA quantity was normalized with GAPDH after density analysis using Image J software (lower panel). Experiments were replicated at least three times. One representative data point is displayed. (B) HT-29 cells were preincubated with pLTA for 20 h and retreated with cytokine cocktail for 24 h. NO production was analyzed with a Nitric Oxide Colorimetric Assay Kit (Abcam) according to the manufacturer’s instruction. Data are given as means ± standard deviations for triplicate samples in a single experiment. *, p < 0.05. UT indicates untreated cells.