Research on The Effect of Intraepitherial Lymphocytes on Intestinal Mucosal Permeability

上皮内淋巴细胞对肠粘膜通透性影响的研究

• 批准号: 12671278
• 负责人: USUI Noriaki
• 金额: $ 1.86万
• 依托单位: Osaka University (2001-2002)Kinki University (2000)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671278/
• 关键词: Mucosal Permeability; Gut Associated Lymphoid Tissue; Intestinal Mucosal Immunit; Intraepithelial Lymphocytes; Co-culture; Caco-2; TEER; 腸管免疫; CaCo-2; co-culture

Purpose : The aim of this study is to evaluate the influence of gut associated lymphoid tissue (GALT) on intestinal mucosal permeability by using a co-culture system of Caco-2 human enterocytes monolayer with lymphocytes harvested from normal and inflammatory SD ratsMaterials and methods : Human Caco-2 enterocytes were seeded onto the lower aspect of porous membranes of a two-chamber system. "They were cultured for 10 14 days and allowed to form a polarized monolayer. After the measwement of transepithelial electric resistance (TEER) by using an epithelial voltohmmeter, Raji cells (a human lymphoid cell line with predominant B cell features), lymphocytes harvested from rat intestinal Peyer's patches (PPL), lamina propria lymphocytes (LPL) and intraepithelial lymphocytes (IEL) were added to the upper surface of the membranes and allowed to migrate through the pores in between the Caco-2 cell monolayers. After incorporation of Lymphocytes, TEERs were monitored for 3 days. PPL, LPL and IE … More L were also harvested from the bowel of indometacin induced inflammatory SD rats, in order to investigate the influence of bowel inflammation on these lymphocytesResults : TEERs were elevated to 200-300 ohm.cm^2 14 days after seeding 1 x10^6 of Caco-2 cells per well, implying the integrity of tight junction. Co-culture with Raji cells for 3 days caused a decrease in TEERs, in a dose-dependent, manner. Permeability to dextran blue was 0.0t±0.0%, 0.0±0.0%, 0.0±0.0%, 0.6±0.3% and 15.411.5% at the Raji cell numbers of 0, 5x10^5 1x10^6, 2x10^6 and 4x10^6, respectively. Co-culture with 1x10^6 of PPL, 1x10^6 of LPL and 1x10^6 of IEL have not decreased TEERs of Caco-2 cell monolayeis. The intestine of SD rats that have been given Indometacin showed a thinner wall and fraglity, indicating an existence of bowel influnmation. Co-cultwe with PPL, LPL and IEL harvested from Indometacin induced inflammatory bowel also have not decreased TEERs of Caco-2 cell monolayersConclusion : GULT, such as PPL, LPL and IEL, deliverd from normal rats and inflammatory rats indicated no influence on the permeability of polarized human Caco-2 enterocyte monolayers. Further investigation may be required to clarify the mechanism of cell-cellular interaction between GALT and enterocytes Less

目的：本研究的目的是利用Caco-2人单层肠上皮细胞与正常和炎症SD大鼠收获的淋巴细胞共培养系统，评估肠道相关淋巴组织（GALT）对肠粘膜通透性的影响材料和方法：将人 Caco-2 肠细胞接种到两室系统的多孔膜的下部，“将它们培养 10-14 天并使其形成极化。使用上皮电压计测量跨上皮电阻 (TEER) 后，Raji 细胞（一种以 B 细胞为主的人类淋巴细胞系）、从大鼠肠道派尔氏淋巴结 (PPL) 和固有层淋巴细胞 (LPL) 中收集的淋巴细胞。将上皮内淋巴细胞 (IEL) 添加到膜的上表面，并允许其通过 Caco-2 之间的孔迁移淋巴细胞掺入后，还从吲哚美辛诱导的炎症 SD 大鼠的肠道中采集了 TEER 3 天，以研究肠道炎症对这些淋巴细胞的影响。每孔接种 1 x10^6 Caco-2 细胞后 14 天升高至 200-300 ohm.cm^2，这意味着与 Raji 细胞共培养 3 天的紧密连接的完整性导致 TEER 以剂量依赖性方式降低，对葡聚糖蓝的渗透性为 0.0t±0.0%、0.0±0.0%、0.0±0.0%。 Raji 细胞数为 0、5x10^5 时为 0.6±0.3% 和 15.411.5%分别与1x10^6的PPL、1x10^6的LPL和1x10^6的IEL共培养没有降低Caco-2单层细胞的TEER。给予吲哚美辛的大鼠肠壁变薄且脆弱，表明存在肠道感染。与从吲哚美辛诱导的炎症肠中收集的PPL、LPL和IEL共培养也没有降低Caco-2单层细胞的TEER结论：从正常大鼠和炎症大鼠中递送的GULT，例如PPL、LPL和IEL对通透性没有影响可能需要进一步研究以阐明 GALT 和肠上皮细胞之间的细胞-细胞相互作用的机制。