Tu1703 Analysis of the Protease MT1-MMP and Its Substrates As Biomarkers and Therapeutic Targets in Inflammatory Bowel Disease

Abstract

Objective: Bone mineral density (BMD) is decreased in patients with inflammatory bowel disease (IBD). Reduced BMD in these patients may be multifactorial. Our recent research demonstrated that serum undercarboxylated osteocalcin (ucOC) levels, which are widely used as a surrogate marker of vitamin K deficiency, were significantly higher in patients with Crohn's disease (CD) than healthy subjects. Vitamin K has been shown to ameliorate liver injury through inhibition of inflammatory cytokine production. We aimed to evaluate the effect of vitamin K on inflammatory responses in the intestine in mice. Methods: CD4, CD11band CD19-positive subpopulations were isolated from splenocytes using the Magnetic Cell Sorting (MACS) separator and these cells were incubated in vitro in the presence or absence of vitamin K2 for 24 h. The mRNA and the culture supernatant of the purified cells were collected. The levels of IL-6 and TNF-α were measured by quantitative RT-PCR and ELISA. Male 8-week-old C57/BL6J mice were fed with the vitamin K deficient diets (K-def) or supplemented diets (K-sup) for 10 days, and then the mice were administered orally with dextran sulfate sodium (DSS) in drinking water for 7 days. Histologic scores of the colon were analyzed 3 days after the cessation of DSS administration. The level of IL6 production from the colonic lamina propria (LP) mononuclear cells was measured by ELISA in the K-def and the K-sup groups. Proportion of the cells costained with CD4 and CD25 was analyzied by flow cytometry. Results: Vitamin K2 reduced the production of IL6 by the splenocytes cultured in vitro, while the expression of TNF-α showed no significant difference. Expression of IL-6 in CD19-positive B cells, but not in CD4and CD11b-positive cells, was significantly decreased in the presence of vitamin K2. The histologic score of DSS colitis was significantly lower in K-sup group than in K-def group. In addition, the expression of IL-6 from LP cells was significantly lower in K-sup group than in the K-def group. The proportion of CD4 and CD25-positive regulatory T cells did not significantly differ between the K-sup and K-def groups. Conclusion: Vitamin K has a suppressive effect on IL-6 production especially in B cells. Vitamin K deficiency deteriorates murine DSS colitis through IL6 production from B cells. Vitamin K may have a potential for the treatment target of IBD.