Angiogenin regulates PKD activation and COX-2 expression induced by TNF-α and bradykinin in the colonic myofibroblast.

INTRODUCTION:The myofibroblast is a gastrointestinal stromal cell that is a target of tumor necrosis factor-alpha (TNF-α), a pro-inflammatory cytokine strongly implicated in colitis-associated cancer. Crosstalk between TNF-α and other pro-inflammatory mediators amplify inflammatory signaling but the mechanism is unknown. Angiogenin (ANG) is a 14-kDa angiogenesis protein that is regulated in patients with inflammatory bowel disease. However, the role of ANG on inflammatory mediator crosstalk in the myofibroblast is unknown. METHODS:The human colonic myofibroblast cell line 18Co, as well as primary mouse and human colonic myofibroblasts, were exposed to TNF-α (10 ng/ml) and bradykinin (BK, 100 nM). ANG was quantified by ELISA. The expression of cyclo-oxygenase-2 (COX-2) and phosphorylation of PKD was assessed by mouse and human colonic myofibroblasts exposed to TNF-α/BK led to enhanced PKD phosphorylation and synergistic COX-2 expression. 18Co cells secrete high levels of ANG (24h, 265 ± 5 pg/ml). The monoclonal antibody 26-2F, which neutralizes ANG, inhibited TNF-α/BK-mediated PKD phosphorylation and synergistic COX-2 expression in primary human myofibroblasts. Likewise, in primary mouse myofibroblasts that do not express ANG (ANG-KO), TNF-α/BK failed to enhance PKD phosphorylation and COX-2 expression. CONCLUSIONS:TNF-α/BK enhance PKD phosphorylation and COX-2 expression in primary mouse and human colonic myofibroblasts. Angiogenin is produced by the myofibroblast, and inhibition of ANG signaling, either by its absence (ANG-KO) or by pharmacologic inhibition, blocks enhanced PKD phosphorylation and synergistic COX-2 expression induced by TNF-α/BK. ANG mediates crosstalk signaling between TNF-α/BK in the regulation of stroma-derived COX-2 and may be a novel therapeutic target for the management of colitis-associated cancer.

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