miR-126靶向负调控CXCL12招募TAMs抑制非可控性结肠炎癌变的机制

Though the mechanism remains unclear, inflammatory microenvironment is known to play critical role in nonresolving colitis-associated carcinogenesis. We firstly found that miR-126 was down-regulated in the nonresolving colitis-associated colon cancer mouse model previously established, leading to up-regulation of its target gene CXCL12, and activation of signaling pathways such as NF-κB and STAT3. Results also showed that tumor associated macrophages (TAMs) were therefore recruited to intestinal mucosa, releasing IL-6 and IL-1β into circulation. Based on the preliminary data, we hypothesize that miR-126 inhibits nonresolving colitis-associated carcinogenesis by targeting CXCL12 and subsequently impede TAMs recruitment. In this project, we will use cell co-culture technique and blocking agent to study how miR-126 regulates macrophages chemotaxis and secretion by targeting CXCL12, and how macrophages regulate the carcinogenesis of colon cancer cells. Nonresolving colitis-associated colon cancer mice with miR-126 knocked-out in intestinal epithelium will also be used to discuss the role of miR-126 in the "inflammation-cancer" process, as well as TAMs infiltration and signaling pathway activation. Results will be further confirmed by data obtained in tissues from patients with colitis, atypical hyperplasia and colon cancer. The purposes of this study is to elucidate the role of miR-126 in nonresolving colitis-associated colon cancer development, as well as the mechanism of miR-126 inhibits colitis-associated colon cancer by impeding TAMs recruitment via CXCL12, which is a novel project, and by which will discover novel molecular markers for early diagnosis of colitis-associated colon cancer(CAC).

炎症微环境在非可控性结肠炎癌变过程中起着关键作用，其机制不明。我们首次发现：在炎癌小鼠模型的“炎症-不典型增生-癌症”演进过程中miR-126表达持续下调，其靶基因CXCL12上调，NF-κB和STAT3通路活化，肠黏膜炎症微环境内TAMs聚集，释放IL-6、IL-1β等入血。据此，我们提出全新的科学假设：miR-126靶向负调控CXCL12招募TAMs而抑制非可控性结肠炎癌变。本课题采用细胞共培养及阻断策略，研究miR-126 靶向CXCL12调控巨噬细胞的趋化及分泌功能以及巨噬细胞对结肠细胞恶性转化的影响。应用肠上皮特异性miR-126敲除的“炎癌”小鼠模型，研究miR-126对炎癌演进、TAMs募集及炎癌通路活化的影响。并在人结肠炎、不典型增生、癌组织中验证，阐明miR-126靶向CXCL12抑制TAMs招募而抑制结肠炎癌变的机制，是全新的研究，将为结肠炎癌变的早期诊断提供分子靶标。

结肠癌是严重威胁人类健康的恶性肿瘤，炎症微环境在非可控性结肠炎癌变过程中起着关键作用，其机制不明。我们首次发现：在炎癌小鼠模型的“炎-不典型增生-癌”演进过程中miR-126表达持续下调，其靶基因CXCL12上调，NF-κB、STAT3通路活化，肠黏膜炎症微环境内肿瘤相关性巨噬细胞（TAMs）聚集，释放IL-6和IL-1β等入血。据此，我们提出科学假设：miR-126靶向负调控CXCL12招募TAMs抑制非可控性结肠炎癌变。课题采用细胞共培养及阻断策略，研究miR-126 靶向CXCL12调控巨噬细胞的趋化及分泌以及巨噬细胞对结肠细胞恶性转化的影响。应用肠上皮miR-126敲除的“炎癌”小鼠模型，研究miR-126对炎癌演进、TAMs募集及炎癌通路活化的影响。并在人结肠炎、不典型增生、癌组织中验证，以阐明miR-126靶向CXCL12抑制TAMs招募而抑制结肠炎癌变的机制。结果1.构建肠上皮特异性miR-126敲除小鼠，建立AOM-DSS“结肠炎癌”小鼠模型，明确了miR-126在体内具有抑制非可控性结肠炎癌变的作用；2. 研究CXCL12、TAMs及IL-1β、IL-6等炎性因子在结肠炎癌演进中的动态变化，发现miR-126通过靶向负调控CXCL12，招募TAMs分泌IL-6、IL-1β等促炎因子来抑制非可控性结肠炎癌变的发生发展；3. 构建过表达与沉默miR-126结肠癌细胞系，及人巨噬细胞系，在细胞水平明确miR-126对CXCL12的直接靶向作用，分析miR-126对TAMs趋化及炎性因子表达的调控作用，明确了miR-126通过负调控CXCL12作用于来发挥抑癌作用；4. 动物水平应用AMD3100阻断CXCL12/CXCR4轴，分析结肠炎癌演进及TAMs与IL-1β、IL-6在体内的动态变化，明确了CXCL12为miR-126抑制非可控性炎癌变的关键节点；5. 细胞水平研究miR-126通过负调控CXCL12招募TAMs，进而抑制结肠细胞增殖、转移及EMT相关分子表达的作用，深入阐明miR-126通过靶向负调控CXCL12招募TAMs，下游进一步抑制非可控性结肠炎癌变的具体机制。共发表论文8篇，其中SCI 7篇，专利3个。培养博士3人，硕士2人。取得了预期研究成果。根据课题的新发现，开展了肠道菌群相关的前沿研究，为后续结肠炎癌变的诊治提供了丰富的实验基础。