Anastomotic failure causes morbidity and mortality even in technically correct anastomoses. Initial leaks must be prevented by mucosal reapproximation across the anastomosis. Healing is a concerted effort between intestinal epithelial cells (IECs), immune cells, and commensal bacteria. IEC TLR4 activation and signaling is required for mucosal healing, leading to inflammatory factor release that recruits immune cells to limit bacteria invasion. TLR4 absence leads to mucosal damage from loss in epithelial proliferation, attenuated inflammatory response, and bacteria translocation. We hypothesize after anastomosis, an imbalance in microbiota will occur due to a decrease in TLR4 expression and will lead to changes in the immune milieu. We isolated fecal content and small intestinal leukocytes from murine, Roux-en-Y and end-to-end anastomoses, to identify microbiome changes and subsequent alterations in the regulatory and pro-inflammatory immune cells 3 days post-operative. TLR4+ IECs were impaired after anastomosis. Microbiome diversity was reduced, with Firmicutes, Bacteroidetes, and Saccharibacteria decreased and Proteobacteria increased. A distinct TCRβhi CD4+ T cells subset after anastomosis was 10–20-fold greater than in control mice. 84% were Th17 IL-17A/F+ IL-22+ and/or TNFα+. iNKT cells were increased and TCRβhi. 75% were iNKT IL-10+ and 13% iNKTh17 IL-22+. Additionally, Treg IL-10+ and IL-22+ cells were increased. A novel dendritic cell subset was identified in anastomotic regions that was CD11bhi CD103mid and was 93% IL-10+. This anastomotic study demonstrated a decrease in IEC TLR4 expression and microbiome diversity which then coincided with increased expansion of regulatory and pro-inflammatory immune cells and cytokines. Defining the anastomotic mucosal environment could help inform innovative therapeutics to target excessive pro-inflammatory invasion and microbiome imbalance.

中文翻译：

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吻合术后微生物组多样性下降，而 Th17、iNKT 和树突状细胞亚群明显扩增

即使在技术上正确的吻合中，吻合失败也会导致发病和死亡。必须通过吻合处的粘膜重新接近来防止最初的渗漏。治愈是肠上皮细胞 (IEC)、免疫细胞和共生细菌之间的共同努力。IEC TLR4 激活和信号传导是粘膜愈合所必需的，导致炎症因子释放，招募免疫细胞以限制细菌入侵。TLR4 缺失会导致上皮增殖丧失、炎症反应减弱和细菌易位造成的粘膜损伤。我们假设吻合后，由于 TLR4 表达下降，会出现微生物群失衡，并导致免疫环境发生变化。我们从小鼠、Roux-en-Y 和端对端吻合中分离了粪便内容物和小肠白细胞，以鉴定术后 3 天微生物组的变化以及调节性和促炎性免疫细胞的后续变化。吻合后 TLR4+ IECs 受损。微生物组多样性降低，厚壁菌门、拟杆菌门和糖杆菌门减少，变形菌门增加。吻合后明显的 TCRβhi CD4+ T 细胞亚群比对照小鼠高 10-20 倍。84% 为 Th17 IL-17A/F+ IL-22+ 和/或 TNFα+。iNKT 细胞增多，TCRβhi 增加。75% 为 iNKT IL-10+，13% 为 iNKTh17 IL-22+。此外，Treg IL-10+ 和 IL-22+ 细胞有所增加。在吻合区域中鉴定出一种新的树突状细胞亚群，即 CD11bhi CD103mid，且 IL-10+ 含量为 93%。这项吻合研究表明，IEC TLR4 表达和微生物组多样性下降，同时调节性和促炎性免疫细胞和细胞因子的扩张增加。定义吻合口粘膜环境有助于为创新疗法提供信息，以针对过度的促炎性侵袭和微生物组失衡。