The ATP-hydrolyzing ectoenzyme E-NTPD8 attenuates colonic inflammation through regulation of P2X4 receptor-dependent metabolism in myeloid cells [F0536]

Extracellular adenosine triphosphate (ATP) released by mucosal immune cells and by microbiota in the intestinal lumen elicits diverse immune responses that mediate the intestinal homeostasis via P2 purinergic receptors, while overactivation of the ATP signaling leads to disruption of mucosal immune system linked to pathogenesis of intestinal inflammation. In the small intestine, hydrolysis of luminal ATP by E-NTPD7 in epithelial cells is essential for control of the number of Th17 cells. However, the molecular mechanism underlying regulation of microbiota-derived ATP in the colon is poorly understood. Here, we show that E-NTPD8 is highly expressed in large intestinal epithelial cells and hydrolyzes microbiota-derived luminal ATP. Compared to wild-type mice, Entpd8-/- mice develop more severe DSS-induced colitis. In this context, either depletion of neutrophils and monocytes by injecting with anti-Gr-1 antibody or introduction of P2rx4 deficiency into hematopoietic cells ameliorates colitis in Entpd8-/- mice. Increased level of luminal ATP in the colon of Entpd8-/- mice promotes glycolysis in neutrophils and monocytes through P2X4 receptor-dependent Ca2+ influx, which links to prolonged survival and elevated ROS production in these cells. Together, these results indicate that E-NTPD8 limits intestinal inflammation by controlling metabolic alteration toward glycolysis via P2X4 receptor in myeloid cells. We comprehensively analyzed small and large intestinal epithelial cells by performing RNA-seq analysis.肠腔中黏膜免疫细胞与微生物群释放的细胞外三磷酸腺苷（ATP），可通过P2嘌呤能受体（P2 purinergic receptors）触发多样免疫应答，介导肠道稳态；而ATP信号的过度激活则会破坏黏膜免疫系统，与肠道炎症的发病机制密切相关。在小肠中，上皮细胞内的E-NTPD7水解肠腔ATP，这对调控辅助性T细胞17（Th17 cells）的数量至关重要。然而，目前对于结肠中微生物群来源ATP的调控分子机制仍知之甚少。本研究发现，E-NTPD8在大肠上皮细胞中高表达，并可水解微生物群来源的肠腔ATP。与野生型小鼠相比，Entpd8基因敲除小鼠（Entpd8-/- mice）会出现更为严重的葡聚糖硫酸钠（DSS）诱导的结肠炎。在此模型中，通过注射抗Gr-1抗体（anti-Gr-1 antibody）耗竭中性粒细胞与单核细胞，或在造血细胞中引入P2rx4基因缺陷（P2rx4 deficiency），均可缓解Entpd8-/-小鼠的结肠炎症状。Entpd8-/-小鼠结肠肠腔ATP水平升高，可通过依赖P2X4受体（P2X4 receptor）的钙离子内流（Ca2+ influx），促进中性粒细胞与单核细胞的糖酵解过程，进而延长这些细胞的存活时间并增强其活性氧（ROS）生成。综上，上述结果表明，E-NTPD8可通过调控髓系细胞（myeloid cells）中依赖P2X4受体的糖酵解代谢重编程，限制肠道炎症的发生。本研究通过RNA测序分析（RNA-seq analysis），全面解析了小肠与大肠上皮细胞的转录组特征。