金黄色葡萄球菌是研究最深入的细菌之一，其在肠道中引起的致病性尚不清楚。最近，我们证明金黄色葡萄球菌感染会引起膜组成的改变，这与肠道功能的伴随损伤有关。在这里，我们使用两种类器官模型，即诱导多能干细胞 (iPSC) 衍生的肠道类器官和结肠肠干细胞衍生的肠道类器官 (colonoid)，来检查甾醇代谢和氧水平如何响应金黄色葡萄球菌感染而变化。 HPLC 定量显示感染细胞和未感染细胞之间的脂质稳态存在差异，其特征是细胞总胆固醇显着降低。由于甾醇代谢的改变通常是由于氧化应激反应引起的，因此我们接下来检查了细胞内和细胞外的氧水平。应用了三种不同的氧测量方法：(1) 细胞穿透纳米粒子，用于量化细胞内氧含量；(2) 传感器板，用于量化培养基中的细胞外氧含量；(3) 用于类器官培养物中氧分布的传感器箔系统。数据显示，肠道类器官模型和 Caco-2 细胞感染后，细胞内和细胞外氧含量均显着下降，甚至在消除细胞外细菌 48 小时后，也没有恢复到感染前的氧水平。总之，我们显示金黄色葡萄球菌感染导致甾醇代谢以及细胞内和细胞外缺氧的改变。这些结果将有助于了解肠上皮持续细菌感染期间的细胞应激反应。© 2023 作者。 FASEB 期刊由 Wiley periodicals LLC 代表美国实验生物学学会联合会出版。

The pathogenicity elicited by Staphylococcus (S.) aureus, one of the best-studied bacteria, in the intestine is not well understood. Recently, we demonstrated that S. aureus infection induces alterations in membrane composition that are associated with concomitant impairment of intestinal function. Here, we used two organoid models, induced pluripotent stem cell (iPSC)-derived intestinal organoids and colonic intestinal stem cell-derived intestinal organoids (colonoids), to examine how sterol metabolism and oxygen levels change in response to S. aureus infection. HPLC quantification showed differences in lipid homeostasis between infected and uninfected cells, characterized by a remarkable decrease in total cellular cholesterol. As the altered sterol metabolism is often due to oxidative stress response, we next examined intracellular and extracellular oxygen levels. Three different approaches to oxygen measurement were applied: (1) cell-penetrating nanoparticles to quantify intracellular oxygen content, (2) sensor plates to quantify extracellular oxygen content in the medium, and (3) a sensor foil system for oxygen distribution in organoid cultures. The data revealed significant intracellular and extracellular oxygen drop after infection in both intestinal organoid models as well as in Caco-2 cells, which even 48 h after elimination of extracellular bacteria, did not return to preinfection oxygen levels. In summary, we show alterations in sterol metabolism and intra- and extracellular hypoxia as a result of S. aureus infection. These results will help understand the cellular stress responses during sustained bacterial infections in the intestinal epithelium.© 2023 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.