Xing Chengfeng, Liang Guili, Yu Xin, Zhang Anxing, Luo Xiang, Liu Yu, Tang Zengli, Wu Bian, Song Zhengji, Lan Danfeng
Department of Gastroenterology, The First People's Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, No. 157, Jingbi Road, Kunming 650032, Yunnan, China.
Kunming Medical University, No. 1168, West Chunrong Road, Chenggong District, Kunming 650500, Yunnan, China.
Stem Cells Int. 2023 Mar 7;2023:3328655. doi: 10.1155/2023/3328655. eCollection 2023.
Intestinal epithelial dysfunction is critical in the development of inflammatory bowel disease (IBD). However, most cellular experiments related to epithelial barrier studies in IBD have been based on tumor cell line that lack a variety of intestinal epithelial cell types. Thus, intestinal organoids can present the three-dimensional structure and better simulate the physiological structure and function of the intestinal epithelium . Here, the crypts were isolated from the small intestine of mice; with the participation of major cytokines (EGF, Noggin, and R-Spondin 1 included), the intestinal organoids were established at a density of 100 crypts per well, containing intestinal stem cells (ISC), Paneth cells, goblet cells, and intestinal endocrine cells. We found that tumor necrosis factor-alpha (TNF-) could induce the inflammatory response of intestinal organoids, and a dose of 10 ng/mL could maintain stable passaging of organoids for dynamic observation. After stimulation with TNF-, the intestinal organoid cultures showed lower expression of the cell proliferation-related protein identified by monoclonal antibody Ki 67 (), the ISC marker leucine-rich repeat-containing G protein-coupled receptor 5 (), and the intestinal tight junction proteins occludin () and claudin-1 () while higher expression of the inflammatory cytokine interleukin- (IL-) 15 and the chemokines C-X-C motif ligand 2 () and significantly. In this study, we successfully established an epithelial inflammatory injury model of intestinal organoids, which provides an effective model for studying the pathogenesis and treatment of IBD.
肠上皮功能障碍在炎症性肠病（IBD）的发展中至关重要。然而，大多数与IBD上皮屏障研究相关的细胞实验都是基于缺乏多种肠上皮细胞类型的肿瘤细胞系。因此，肠类器官可以呈现三维结构，更好地模拟肠上皮的生理结构和功能。在这里，从小鼠小肠中分离出隐窝；在主要细胞因子（包括表皮生长因子、头蛋白和R- 斯普隆蛋白1）的参与下，以每孔100个隐窝的密度建立肠类器官，其中包含肠干细胞（ISC）、潘氏细胞、杯状细胞和肠内分泌细胞。我们发现肿瘤坏死因子-α（TNF-）可诱导肠类器官的炎症反应，10 ng/mL的剂量可维持类器官稳定传代以便进行动态观察。用TNF-刺激后，肠类器官培养物中由单克隆抗体Ki 67鉴定的细胞增殖相关蛋白、ISC标志物富含亮氨酸重复序列的G蛋白偶联受体5、肠紧密连接蛋白闭合蛋白和闭合蛋白-1的表达降低，而炎症细胞因子白细胞介素（IL-）15以及趋化因子C-X-C基序配体2和的表达显著升高。在本研究中，我们成功建立了肠类器官的上皮炎症损伤模型，为研究IBD发病机制及治疗提供了有效的模型。
