Prostaglandin E2 Signaling Through the EP4 Receptor Regulates the Proliferation of Hematopoietic Stem/Progenitor Cells Under Stress Conditions

Abstract

Abstract 2377Hematopoietic stem cells (HSCs) maintain the balance between self-renewal and differentiation in order to replenish all blood cell lineages throughout the lifespan of the individual. Signaling molecules produced by various niche cells regulate the function and fate of HSCs. Prostaglandin E2 (PGE2) is a known mediator of various pathological states and physiological systems. In the hematopoietic system, PGE2 signaling regulates HSC development and enhances repopulation activity through its interactions with the Wnt/β-catenin pathway. However, it is not been clear which receptor is responsible for PGE2 signaling in adult HSCs.Here we analyzed the function of PGE2 receptors, EP2 and EP4, in the regulation of adult bone marrow (BM) hematopoietic stem/progenitor cells (HSPCs). Quantitative real time PCR (Q-PCR) analysis revealed that EP4 was highly expressed in Lin−Sca-1+c-Kit+ (LSK) fractions compared to Lin− or Lin+ fractions. On the other hand, EP2 mRNA expression was detected at a higher level in LSKCD34+Flt3+ fractions than in other fractions. Digital PCR analysis showed that EP2 transcripts were present in much lower numbers than EP4 transcripts in HSCs.We next analyzed the effects of short-term treatment with an EP2 or EP4 agonist on the colony forming activity of LSK cells. Although the EP2 agonist did not affect the colony formation activity of LSK cells, the EP4 agonist increased the number of colony forming unit in culture (CFU-C) and high proliferative potential-colony forming cells (HPP-CFC) in a dose dependent manner, indicating that PGE2/EP4 signaling affects the proliferation of HSPCs. A knockdown of EP4 reduced the engraftment capacity of LSK cells, suggesting that PGE2/EP4 signaling contributes to the BM repopulation activity of HSPCs.Next, we examined the effect of PGE2/EP4 signaling on the recovery of HSCs after myelosuppression. 5-FU-injected mice were treated with PGE2 and the proportion of LSK cells in the mice at various days after 5-FU injection was examined. The frequency of LSK cells at the early phase of post 5-FU treatment was comparable in control and PGE2-treated mice. In the later phase of post 5-FU treatment, mice treated with PGE2 showed a higher frequency of LSK cells, and the LSK cells derived from the PGE2-treated mice had significantly higher expression of Cdk4, Cdk6, Ctnnb1, and Myc compared to the control mice. These data indicate that PGE2 promoted BM recovery by accelerating the proliferation of HSPCs. The EP4 agonist was as efficient as PGE2 in enhancing BM recovery after myelosuppression. In addition, we found that EP4 agonist treatment induced an increase in the nuclear localization of β-catenin, indicating that PGE2 signaling through EP4 impinges on Wnt/β-catenin signaling in HSCs. Altogether, these data suggest that EP4 plays a crucial role in the PGE2 signaling involved in the proliferation of HSCs under stress conditions. This study was conducted under the collaboration with Dr. Narumiya (Kyoto University Graduate School of Medicine) Disclosures:No relevant conflicts of interest to declare.