miR-155 expression in T cells is dependent on the H. pylori-mediated induction of Foxp3.

(A) In vivo analysis of Foxp3 expression in human gastric biopsies. Total RNA extracted from the antrum of four patients before and after 80 days of infection (patient 13, 29, 71 and 77) was used for the measurement of Foxp3 expression by qPCR. (B) Total RNA from CCRF-CEM cells was assayed for Foxp3 mRNA expression after H. pylori infection or incubation with H. pylori LPS and GGT, and compared to non-infected cells. qPCR data show an induction of Foxp3 after challenge with live bacteria, LPS and purified GGT. Data are mean ±SD of two independent experiments. (C) Western blot analysis of Foxp3 protein induction in CCRF-CEM cells infected with H. pylori (P12, 3 h). Actin serves as a loading control. Foxp3 band intensities shown. Blots are representative of two independent experiments. (D) Human PBMCs were assayed by qPCR for Foxp3 expression after H. pylori infection or GGT treatment over time (1 to 6 h). The results obtained from 5 individual donors show an increase of Foxp3 mRNA after 6 h. (E) miR-155 expression was measured by qPCR in the Foxp3 KD CCRF-CEM cells at 3 h p.i. Results are expressed as a ratio between infected and non-infected samples. Error bars show SD of two independent sets of experiments. Foxp3 (#Foxp3) KD was achieved using siRNA technology and confirmed by Western blot 48 hours post-transfection.