胞外基质蛋白periostin在MSCs调控肠炎及其恶变过程中的功能研究

Recent clinical evidence and studies have revealed that the secreted protein periostin is involved in the development of various tumors including colon cancer. However, the functional mechanisms of periostin responsible for the colitis and its transformation and malignant progression are not clear. In our previous study, we found the tissue and serum periostin are higher in the colitis and its malignant transformation induced by MSCs compared to healthy control. It indicates that periostin plays very important roles in the progression of malignant transformation of colitis under the regulation of MSCs. So this proposal is focused on studying the colitis and colitis-associated cancer by using periostin conditional knockout mice models induced by DSS, AOM/DSS, periostin-/- colon epithelium cells and colon cancer cell line CT26. Firstly, we will confirm the biology functions of periostin responsible for MSCs mediated colitis and its transformation and malignant progression by using tail vein injection and cell co-cultured. And next, we will try to identify the periostin receptor involved in MSCs mediated colitis and its transformation and malignant progression by using Liquid chromatography-mass spectrometry (LC-MS) and co-immunoprecipitation (Co-IP). Furthermore, periostin and its receptors are involved in the main signal path will be confirmed and the function of MSCs in these pathways will be summarized. Simultaneously, we will further analyze some clinical samples of inflammatory bowel disease and colon cancer in order to verify molecular mechanisms and clinical significance. We expect to look for some potential proteins including periostin as a marker of colitis-associated cancer, in order to lay a foundation for the application of clinical medicine.

Periostin是与肿瘤发生发展关系密切的细胞外基质分子，但其在肠炎发生及其恶变中的功能不清。前期研究发现MSCs在促进肠炎恶性转变过程中periostin表达呈现递增趋势。本项目充分利用periostin基因敲除小鼠和DSS、AOM/DSS诱导炎症及其相关性肠癌模型、periostin-/-肠上皮细胞、结肠癌细胞为研究对象，1）采取MSCs尾静脉注射、体外共培养等，明确periostin在MSCs调控肠炎发生和促进肠炎恶变过程中所介导的生物学功能；2）采用Co-IP、cDNA芯片和质谱检测等技术，筛选并鉴定periostin在调控炎症发生发展过程中起作用的受体，明确periostin及其受体共同参与的主要信号通路；3）结合临床肠炎及其肠癌相关标本进行上述分子机制的验证和临床意义分析，寻找包括periostin在内可能的分子作为慢性肠炎恶变的标志物，为其在临床医学上的应用奠定理论基础。

我们前期的大量研究数据不但表明了MSCs在肠炎发生及其恶性转变的过程中都起着重要的调控作用，而且发现细胞外基质分子periostin表达呈现变化。periostin是我们早期在肠癌组织筛选到一个高表达的分泌性蛋白分子，其表达水平与肠癌的恶性程度相关。本项目利用periostin基因敲除小鼠模型和DSS和AOM/DSS诱导急性炎症和慢性肠炎相关大肠癌模型以及相关细胞株等重点研究了periostin在MSCs调控肠炎及其恶性转化过程中的功能及其分子机制。紧紧围绕着项目年度研究计划，首先，进一步证明了MSCs本身对结肠炎相关性肠癌修复作用明显，其机制是periostin通过促进ATM/BID磷酸化激活炎症细胞DNA损伤检控点；但是随着炎症程度加剧，这种修复作用被反转，继而激活Wnt/β-catenin通路更加加重肿瘤的发展。还发现高浓度的periostin或炎症因子促进MSCs异常增殖，异常增殖的MSCs不再具有保护修复作用，而是进一步的促进肠炎的恶化。其次，我们进一步利用periostin基因敲除小鼠和腺病毒介导periostin高表达建立了肠癌肝转移模型，研究发现periostin是促进肠癌细胞肝转移，其分子机制是肠癌细胞转移肝脏后会逐步营造一个肝纤维化的环境，而periostin通过激活TGF-β信号通路加速了这个肝纤维化的过程，同时还发现TGF-β也可通过Smad3促进periostin的表达，从而进一步促进了肠癌肝转移。另外还发现periostin可以促进肝再生，其机制与调控炎症因子和血管新生相关。再者，在执行本项目过程中，我们发现USP39一个全新的分子在调控包括炎症性肠癌在内的恶性肿瘤发生发展过程起更为重要的作用，目前的数据表明其机制可能与调控periostin的RNA剪切或者蛋白表达紧密相关，为下一步深入开展本课题打下了基础。