IL-1 beta/IL-6共刺激通过JAK/STAT和p38信号事件协同介导DUOX2/DUOXA2复合物上调

Chronic inflammation increases the susceptibility of the colon to neoplasia and cancer through establishment of an inflammatory microenvironment and persistent release of reactive oxygen species (ROS), inducing genome damage. Interleukin -1beta (IL-1beta), a pro-inflammatory cytokine, plays a pivotal role in the pathogenesis of acute and chronic intestinal inflammation, regulating both innate and adaptive immune responses. IL-1beta has been shown to promote accumulation of IL-17A through lymphoid and Th17 infiltration in the colon tumor microenvironment, supporting increased inflammation and correlating directly with poor patient prognosis. New insights into the signaling pathways supported by IL-1beta in the pathogenesis of colon cancer, including angiogenesis and metastasis, may provide downstream targets for treatments that minimally perturb intestinal immunity. Prior studies from our group have demonstrated significant up-regulation of IL-1, dual oxidase 2 (DUOX2) and its partner maturation factor, DUOXA2, in surgically resected colon cancer specimens compared with adjacent normal colonic epithelium. To elucidate a direct link between IL-1 family cytokines, DUOX2/DUOXA2 derived ROS, and colorectal cancer, we examined human colon cancer cells (HT29, Ls513, T84 and Colo205) stimulated with IL-1beta, in cooperation with IL-6. Co-stimulation resulted in a dramatic, synergistic up-regulation of a hydrogen peroxide producing (Amplex Red oxidizing) DUOX2 enzyme complex and was directly associated with enhanced histone H2AX phosphorylation (gammaH2AX), a marker of DNA double strand breaks. Interestingly, this concentration- and time-dependent induction of expression and oxidative response was not mediated by other IL-1 family members (IL-18, IL-33 or IL-37). Investigations with the interleukin-1 receptor antagonist anakinra established that signaling for IL-1beta/IL-6 co-treatments proceeded through the IL-1 receptor for all cell lines, though minimal receptor expression is present in T84 and Ls513 cells. Similarly, exposure to tocilizumab, an IL-6R antagonist, demonstrated that the IL-1beta/IL-6 synergistic increase in DUOX2/DUOXA2 complex expression is IL-6 receptor dependent. Perturbation of the IL-1 and IL-6 signaling pathway elements MYD88, IRAK1, STAT1, and STAT3 by siRNA knockdown demonstrated their significant contribution to DUOX2/DUOXA2 up-regulation, while dependence on RELA was absent. Knockdown facilitated by MAPK14 siRNA and SB203580 p38 alpha/beta inhibition also demonstrated a role for p38alpha in the regulation of DUOX2/DUOXA2 expression by the IL-1beta/IL-6 signaling axis. Current studies are focused on elucidating the DUOX2 promoter transcription factor binding site(s) responsible for DUOX2 up-regulation through chromatin immunoprecipitation.