Department of Animal Sciences, Purdue University, West Lafayette, IN 47906.
Department of Animal Sciences, Purdue University, West Lafayette, IN 47906; Livestock Behavior Research Unit, Agricultural Research Service, United States Department of Agriculture, West Lafayette, IN 47906.
Poult Sci. 2017 Sep 1;96(9):3096-3103. doi: 10.3382/ps/pex160.
Gut mucosa holds a single layer of epithelial cells and the largest mass of lymphoid tissue in the body. Although the epithelial cell culture model is widely used to assess intestinal barrier function, it has limitations for studying cellular interactions, in particular those of the immune system. In this study, a chicken ileal explant culture model was developed for investigating short-term gut inflammatory and secretory responses in an ex vivo environment. Initially, ileal explants from broilers at 21 d of age were cultured ex vivo up to 6 h. Explants cultured for a maximum of 2 h remained over 90% viable, based on lactate dehydrogenase (LDH) release assay. Morphologically, explants cultured for 2 h displayed normal morphology compared to those cultured longer, further confirming that short-term culture for up to 2 h duration is an acceptable model for studying ex vivo regulation of inflammation. Subsequently, lipopolysaccharide (LPS) dose-related responses were determined for explants cultured for 2 h. Results from LDH activity assay showed that the viability of explants was decreased (P ≤ 0.05) at an LPS dose higher than 50 μg/mL. A significant (P ≤ 0.05) nitric oxide release was observed at LPS concentrations of 10 and 20 μg/mL. In addition, the highest inflammatory and secretory responses were detected at 20 μg/mL LPS based on gene expression of TLR-4, IL-1β, IL-8, MUC2, IgA, and pIgR (P ≤ 0.05). However, the gene expression of claudin-1 and claudin-4 were not increased at the determined LPS concentrations (P > 0.05). These results demonstrated the potential usefulness of this intestinal explant culture model for short-term study of biological factors in gut inflammatory and secretory responses, but not a sufficient duration for evaluation of tight junction responsiveness.
肠黏膜由单层上皮细胞和体内最大的淋巴组织组成。尽管上皮细胞培养模型广泛用于评估肠道屏障功能，但它在研究细胞相互作用方面存在局限性，特别是在免疫系统方面。在这项研究中，建立了一种鸡回肠器官培养模型，用于在体外环境中研究短期肠道炎症和分泌反应。最初，将 21 日龄肉鸡的回肠器官离体培养，最长可达 6 小时。基于乳酸脱氢酶（LDH）释放测定，培养 2 小时的器官的活力保持在 90%以上。形态学上，与培养时间更长的器官相比，培养 2 小时的器官显示出正常的形态，进一步证实了短时间培养 2 小时以内是研究体外炎症调节的可接受模型。随后，确定了培养 2 小时的器官对脂多糖（LPS）剂量相关反应。LDH 活性测定结果表明，当 LPS 剂量高于 50μg/mL 时，器官的活力降低（P≤0.05）。在 LPS 浓度为 10 和 20μg/mL 时，观察到显著的（P≤0.05）一氧化氮释放。此外，基于 TLR-4、IL-1β、IL-8、MUC2、IgA 和 pIgR 的基因表达，在 20μg/mL LPS 下检测到最高的炎症和分泌反应（P≤0.05）。然而，在确定的 LPS 浓度下，claudin-1 和 claudin-4 的基因表达没有增加（P>0.05）。这些结果表明，这种肠道器官培养模型在短期研究肠道炎症和分泌反应中的生物学因素方面具有潜在的用途，但不足以评估紧密连接的反应性。
