Life science
Medical & healthcare, Drug development

Development of therapeutic strategy for ulcerative colitis bytargeting microbiota-derived metabolite
Institute for Advanced Co-Creation Studies
Associate Professor Hisako Kayama

https://researchmap.jp/0425Kayama_Hisako

Department of Microbiology and Immunology, Graduate School of Medicine
Professor Kiyoshi Takeda

https://researchmap.jp/read0118278

Life science

Abstract
The number of patients with inflammatory bowel diseases (IBD),
such as ulcerative colitis (UC), has been increasing worldwide.
IBD patients are shown to harbor dysbiotic microbial communities,
associated with alterations of intestinal metabolite concentrations.
Extracellular adenosine triphosphate (ATP) released by microbes
contributes to activation of host immune responses in the intestine
through the P2X and P2Y receptors. To avoid inappropriate immune
reactions linking to development of IBD, luminal ATP needs to bestrictly controlled. However, the molecular mechanism underlying
regulation of the luminal ATP concentration in the colon remains
poorly understood. We found that the clearance of microbiota-derived ATP by E-NTPD8 in colonic epithelial cells is essential for
inhibiting the prolonged survival of neutrophils by discouraging theP2X4R-mediated promotion of glycolysis, thereby abrogating innate intestinal pathology. These results reveal the mechanism preventing innate intestinal pathology through modulation of myeloid
cell metabolism and may serve to identify therapeutic targets forIBD.

▍ Background & Results
We previously demonstrated that extracellular ATP released by
microbes contributes to induction of Th17 cells by activating dendritic cells in the intestine. To avoid inadequate immune responses,
extracellular ATP is tightly regulated by epithelial ATP-hydrolyzing
ectoenzymes, such as E-NPPs and E-NTPDs. We have found that
E-NPP3 and E-NTPD7 control luminal ATP concentration in thesmall intestine, which are essential for regulation of innate andadaptive immune responses. However, the molecular mechanism
underlying regulation of the luminal ATP concentration in the large
intestine remains poorly understood.
Among E-NTPD and E-NPP family members, E-NTPD8 washighly expressed in colonic epithelial cells. Intriguingly, patientswith UC exhibited low expression of ENTPD8 mRNA in colonic epithelial cells. Therefore, we generated Entpd8-/- mice to assess theits physiological roles. A higher level of luminal ATP was observedin the colon of Entpd8-/- mice than wild-type mice. Entpd8-/- micedeveloped more severe dextran sodium sulfate-induced colitis
accompanied by increased number of neutrophils in the coloncompared with those in wild-type mice. In this context, the ablationof neutrophils by treatment with anti-Gr-1 antibody mitigated theseverity of colitis in Entpd8-/- mice. In addition, the introduction ofP2rx4 deficiency into Entpd8-/- mice ameliorated DSS-induced colitis with lower levels of neutrophils accumulation. Extracellular ATP
promoted glycolysis in neutrophils through a P2X4 receptor-dependent Ca2+ influx, which is linked to prolonged survival in thesecells. Thus, E-NTPD8 limits intestinal inflammation by controllingmetabolic alteration toward glycolysis via the P2X4 receptor in neutrophils.

▍ Significance of the research and Future perspective
Our study provides evidence for the contribution of excessive
accumulation of microbiota-derived luminal ATP to intestinal inflammation progression via perturbations of host innate immunity
through promoting glycolysis. Modulation of glycolysis in lymphocytes and myeloid cells by small molecule dimethyl fumarate abrogates chemically induced colitis in mice. Thus, the extracellular
ATP, P2X4 receptor, and glycolysis could be a putative therapeutictarget for UC.

Figure 1: E-NTPD8 prevents neutrophil-mediated colitis(A) Expression of ENTPD8 mRNA in colonic epithelial cells. (B and C) Severity of
dextran sodium sulfate-induced colitis. The disease activity index score incorporates bleeding score (0-4) and pasty stool score (0-4).

Figure 2: The role of E-NTPD8-mediated hydrolysis of extracellular ATP in maintenance of the gut homeostasis
E-NTPD8 in colonic epithelial cells maintains the gut homeostasis through hydrolysis of luminal ATP produced by commensal bacteria (left). An increased level ofluminal ATP caused by lack of E-NTPD8 modulates neutrophil physiology, such as
prolonged survival, through P2X4-mediated promotion of glycolysis, which links
to aggravation of colitis (right).

P a t e n tTani, Haruka; Kayama, Hisako; Takeda, Kiyoshi et al. The ATP-hydrolyzing ectoenzyme E-NTPD8 attenuates colitis through modulation of P2X4 receptordependent metabolism in myeloid cells. Proc Natl Acad Sci USA. 2021, 118(39): e2100594118. doi: 10.1073/pnas.2100594118
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L https://www.med.osaka-u.ac.jp/pub/ongene/K e y w o r d ulcerative colitis, microbiota, metabolites, epithelial cells, immune cells
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