Protective Role Of Vitamin D3 In DSS-induced Acute Colitis And AOM-DSS Induced Colitis-Associated Colorectal Cancer In Mice

BackgroundInflammatory bowel disease (IBD) is a group of chronic inflammatory disorders of the gastrointestinal tract, which comprises two major types, ulcerative colitis (UC) and Crohn’s disease (CD). Patients with IBD have increased risk of colorectal cancer (CRC). Vitamin D (VitD) is the precursor to the potent steroid hormone calcitriol, which has widespread actions throughout the body including anti-infection, immunomodulation and cancer prevention. Epidemiologic and clinical researches show that incidence of VitD deficiency is increased among IBD patients and this insufficiency is also a risk factor for CRC. Accumulating results from preclinical and some clinical studies suggest VitD supplementation might suppress the severity of colitis and reduce CRC incidence.Part I Effects of Vitamin D3 on Dextran Sulfate Sodium Induced Colitis in MiceObject To explore the preventive effect of vitamin D3 (VitD3) on dextran sulfate sodium (DSS) induced colitis in mice and its mechanisms.Method C57BL/6 mice were given 2.5% DSS in drinking water for 5 days to induce experimental colitis. We gave VitD3 (32.5 IU/g/week) to mice in the intervention group from 2 weeks before model induction until 10 day after DSS drinking by gastric lavage. Weight change, disease activity index (DAI), macroscopic and histologic colitis scores, myeloperoxidase (MPO) activity and TNF-alpha mRNA level were used to evaluate the severity of colitis. Intestinal epithelial cell (IEC) apoptosis was assessed by immunohistochemical staining of activated caspase 3.Results Weight loss, DAI, macroscopic and histologic colitis scores and MPO activity increased significantly in colitis group (n=10) comparing to control group (n=5) (P<0.01). Serum 25(OH)D level in intervention group (n=10) was significantly higher than control and model group (P<0.05) without serum calcium level elevation. Symptoms of colitis in intervention group improved earlier than model group. Macroscopic and histologic scores, MPO activity and TNF-alpha expression level in intervention group were lower than model group (Histologic score:P=0.06; others:P<0.05). Activated caspase 3 postive cells in intestinal epithelium were significantly reduced in intervention group comparing to model group.Conclusion VitD3 suppresses DSS-induced colitis in mice possibly through inhibiting TNF-alpha expression and IEC apoptosis. than model group (Histologic score:P=0.06; others:P<0.05). Activated caspase 3 postive cells in intestinal epithelium were significantly reduced in intervention group comparing to model group.Part Ⅱ Effects of Vitamin D3 on Azoxymethane and Dextran Sulfate Sodium Induced Colitis-associated Colorectal Cancer in MiceObject To explore the preventive effect of vitamin D3 (VitD3) on azoxymethane (AOM) and DSS induced colitis-associated CRC (CAC) and its mechanisms.Method C57BL/6 mice were given AOM (12.5mg/kg) and 2.5% DSS to induce CAC model. We gave mice in the intervention group VitD3 (32.5IU/g/week) by gastric lavage. Tumor number (TN), tumor load (TL), pathologic evaluation and Ki-67 staining were used to assess CRC formation. Vitamin D receptor (VDR), beta-catenin, forkhead box M1 (FOXM1) and TNF-alpha, IL-6, IL-21 expression levels were measured.Results Mice in both model group (n=10) and intervention group (n=9) were found to have colon cancer macroscopically and pathologically at 12 weeks after model induction. TNs were 5±2 and 3.8±3.99/mouse for model group and intervention group respectively (P=0.08).TLs were 1.28±0.91and 0.83±0.91cm respectively (P=0.08). Percentage of large (D>4mm), medium (2mm<D≤4mm) and small (D≤2mm) tumor were 0,35% and 65% for intervention group, comparing to 10%,54% and 36% for model group. There were no difference in depth of invasion and degree of differentiation pathologically. Ki-67 positive rates were 58.3±11.7% for model group and 43.3±12.1% for intervention group (P-0.054). Mechanistically, we found that VitD3 reduced the expression level of beta-catenin (P<0.05), suppressed its nuclear localization and down-regulated expression of its transcriptional product, Cyclin D1 (P<0.05). FOXM1 expression level was also suppressed in intervention group comparing to model group (P<0.05).TNF-alpha expression was increased significantly both in colitis and CRC (P<0.05).VDR expression level was reduced in colitis (P<0.05) and decreased further in CRC.VitD3 intervention partially prevented reduction of VDR expression.Conclusion VitD3 reduced the size of AOM-DSS induced CRC in mice. Beta-catenin and FOXM1 may play an important role in the anti-oncogenic function of VitD/VDR signaling. VDR and TNF-alpha may participate in the mechanism of colitis-CRC progression.