The ATP-hydrolyzing ectoenzyme E-NTPD8 attenuates colonic inflammation through regulation of P2X4 receptor-dependent metabolism in myeloid cells [F1347]

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 analyzed colonic myeloid cells with 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嘌呤能受体介导多样免疫应答，进而维持肠道稳态；而ATP信号通路的过度激活则会引发黏膜免疫系统紊乱，与肠道炎症的发病机制相关。在小肠中，上皮细胞内的E-NTPD7水解肠腔ATP，这对调控Th17细胞的数量至关重要。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-/-小鼠的DSS诱导性结肠炎症状更为严重。在此背景下，通过注射抗Gr-1抗体耗竭中性粒细胞与单核细胞，或是在造血细胞中引入P2rx4缺陷，均可缓解Entpd8-/-小鼠的结肠炎症状。Entpd8-/-小鼠结肠内的肠腔ATP水平升高，可通过P2X4受体依赖的钙离子内流，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提升其活性氧（ROS）生成量。综上，上述结果表明，E-NTPD8可通过调控髓系细胞中P2X4受体介导的糖酵解代谢重编程，限制肠道炎症的发生。我们采用RNA测序（RNA-seq）技术对结肠髓系细胞进行了分析。