A p < 0.05 was considered significant, whereas not significant (n.s.) difference was associated with a p ≥ 0.05. Statistics were performed RG7420 in comparison with LPS-stimulated PCT-untreated cells (LPS + SF), and the exact significance index

is indicated on the top of the horizontal line encompassing the two statistically compared bars Figure 4 In vitro effect of different concentrations of PCT on S. typhimurium LPS-induced release of IL-10 evaluated by cytokine biochip array. Human PBMC were cultured for 24 h with the following mixtures which had been pre-incubated at 37°C for 30 min: Sterile saline fluid (SF) plus 50 ng/ml PCT (SF + PCT 50); SF plus 500 ng/ml PCT (SF + PCT 500); SF plus 5000 ng/ml PCT (SF + PCT 5000); LPS of S. typhimurium Selleck BI2536 SL1102 (100 ng/ml) plus SF (LPS + SF); LPS (100 ng/ml) plus 50 ng/ml PCT (LPS + PCT 50); LPS (100 ng/ml) plus 500 ng/ml PCT (LPS + PCT 500); LPS (100 ng/ml) plus 5000 ng/ml PCT (LPS + PCT 5000). Results are presented as means ± SEM

of at least four experiments each carried out in duplicate. Statistical significance between groups was assessed by Student’t test. A p < 0.05 was considered significant. Statistics were performed in comparison with LPS-stimulated PCT-untreated cells (LPS + SF), and the exact significance index is indicated on the top of the horizontal line encompassing the two statistically compared bars. The release of IL-4 was not affected by PCT (data not shown). Direct assay (trypan blue test and acridine orange vital staining) of cellular viability always indicated a percentage of more than 95% viable cells in any experimental group, even after 24 h of PBMC incubation, which would indicate that the observed reduction in cytokine release may not be due to cellular toxicity by PCT, LPS or both. Also cell count was carried out at beginning and at the end of each experiment and

these values were not significantly different. Therefore a decrease of cell number should be excluded as a possible cause of reduced cytokine release, during the experiments which involved PCT. Discussion The main and novel findings of the present study are the PCT-induced decrease of bacterial LPS reactivity and the reduction of LPS- induced release of some cytokines/chemokines by PCT in human Megestrol Acetate PBMC. Previous studies from our group [10, 11] and from other investigators [12], demonstrated that antimicrobial peptides (teicoplanin and magainins) and other biological effective molecules presenting a polycationic structure, can neutralize both the LAL reactivity and other effects of LPS including cytokine release [9, 13]. An examination of the PCT primary structure reveals that relevant polycationic motifs (sequence of at least 2–3 bibasic aminoacids within a sequence of four) are present in the whole molecule. Therefore, the whole PCT molecule may account for binding and neutralizing the LPS as well as inhibiting the LPS-stimulated mediators.