Inhibiting neutrophils activation by long-term administration of 5-amino salicylic acid reduces colitis-associated colorectal tumor burden in ApcMin/+ mice. Mus musculus

Objective The risk of ulcerative colitis (UC)-associated colorectal cancer (CRC) increases with the duration of UC. Oral 5-aminosalicylic acid (5-ASA) formulations are the first-line treatment for mild-to-moderate UC; nevertheless, preventive effect of oral 5-aminosalicylic acid (5-ASA) formulations on UC-associated CRC remains unsolved. We investigated the impact of 5-ASA to colitis-associated neoplasia in C57BL/6J-ApcMin heterozygous (ApcMin/+) mice. Design ApcMin/+ mice exposed to 1.5 w/v% dextran sulfate sodium (DSS) ad libitum for 3 days followed by 25 days of tap water to develop colitis-associated neoplasia. Mice were intracolorectally administrated with 5-ASA to evaluate the effects on the number of tumors. The mechanism of action was presumed by microarray analysis using in vivo samples and was confirmed by in vitro culture of HT-29, a human colorectal adenocarcinoma cell line with supernatant of human primary neutrophils. Results A remission between acute and recurrent episodes of diarrhea, which were ameliorated by 5-ASA treatment were observed in this model. The number of tumors was reduced by continuous 5-ASA administration in this model. This reduction was abolished by withdrawal of 5-ASA in remission period compared to continuous 5-ASA administration. Microarray analysis revealed that neutrophils were involved in the protective mechanism of 5-ASA against proliferation of tumors. Additionally, calprotectin production from human primary neutrophil as activation marker was strongly correlated with proliferation of HT-29. Overall design: ApcMin/+ mice were given drinking water supplemented with 1.5 w/v% dextran sulphate sodium (DSS) for 3 days to induce colitis, and were intracolorectally administrated with 5-aminosalicylic acid (5-ASA) or Saline (Control) for 28 days. ApcMin/+ mice (Sham) were given drinking water alone for 28 day. Mice of each groups (Sham, 5-ASA and Control) were sacrified and were collected macroscopically distinguished non-tumor regions in colon on day 28.

研究目标：溃疡性结肠炎（ulcerative colitis, UC）相关结直肠癌（colorectal cancer, CRC）的发病风险随UC病程延长而升高。口服5-氨基水杨酸（5-aminosalicylic acid, 5-ASA）制剂是轻中度UC的一线治疗药物，但口服5-ASA制剂对UC相关CRC的预防作用仍未明确。本研究探讨了5-ASA对C57BL/6J-ApcMin杂合子（ApcMin/+）小鼠结肠炎相关肿瘤发生的影响。实验设计：将ApcMin/+小鼠自由饮用1.5w/v%葡聚糖硫酸钠（dextran sulfate sodium, DSS）溶液3天，随后更换为普通自来水饲养25天，以诱导结肠炎相关肿瘤发生。对小鼠经结肠内给予5-ASA，以评估其对肿瘤数量的影响。通过体内样本的基因芯片分析推测其作用机制，并利用人结肠腺癌细胞系HT-29与人类原代中性粒细胞上清共培养的体外实验验证该机制。实验结果：本模型中，急性与复发性腹泻症状可经5-ASA治疗得到缓解。持续给予5-ASA可降低本模型的肿瘤数量；与持续给药组相比，缓解期停用5-ASA的小鼠，其肿瘤数量减少的效应被抵消。基因芯片分析显示，中性粒细胞参与了5-ASA抑制肿瘤增殖的保护机制。此外，作为中性粒细胞活化标志物的人类原代中性粒细胞钙卫蛋白（calprotectin）的产生水平与HT-29细胞增殖显著相关。整体实验设计：将ApcMin/+小鼠饮用添加1.5w/v%葡聚糖硫酸钠（DSS）的饮水3天以诱导结肠炎，随后经结肠内给予5-氨基水杨酸（5-ASA）或生理盐水（对照组），共给药28天。另有ApcMin/+小鼠（假手术组）仅饮用普通自来水28天。于第28天处死各组小鼠（假手术组、5-ASA组与对照组），收集结肠肉眼可见的非肿瘤区域样本。