Essential role of prostaglandin E2 and the EP3 receptor in lymphatic vessel development during zebrafish embryogenesis

Ryo Iwasaki,Kyoshiro Tsuge,Koichiro Kishimoto,Atsuo Kawahara,Hirofumi Hohjoh,Tomoaki Inazumi,Yuta Hayashi,Takuya Iwaana,Soken Tsuchiya,Yukihiko Sugimoto

doi:10.1038/s41598-019-44095-5

Abstract

Lymphatic endothelial cells arise from the venous endothelial cells in embryonic lymphatic development. However, the molecular mechanisms remain to be elucidated. We here report that prostaglandin (PG) E2 plays essential roles in the embryonic lymphatic development through the EP3 receptor, one of the PGE2 receptors. Knockdown of the EP3 receptor or inhibition of cyclooxygenases (COX; rate-limiting enzymes for PG synthesis) impaired lymphatic development by perturbing lymphatic specification during zebrafish development. These impairments by COX inhibition were recovered by treatment with sulprostone (EP1/3 agonist). Knockdown of the EP3 receptor further demonstrated its requirement in the expression of sex determining region Y-box 18 (sox18) and nuclear receptor subfamily 2, group F, member 2 (nr2f2), essential factors of the lymphatic specification. The EP3 receptor was expressed in the posterior cardinal vein (region of embryonic lymphatic development) and the adjacent intermediate cell mass (ICM) during the lymphatic specification. COX1 was expressed in the region more upstream of the posterior cardinal vein relative to the EP3 receptor, and the COX1-selective inhibitor impaired the lymphatic specification. On the other hand, two COX2 subtypes did not show distinct sites of expression around the region of expression of the EP3 receptor. Finally, we generated EP3-deficient zebrafish, which also showed defect in lymphatic specification and development. Thus, we demonstrated that COX1-derived PGE2-EP3 pathway is required for embryonic lymphatic development by upregulating the expression of key factors for the lymphatic specification.

Highlights

The lymphatic system is a major component of the vertebrate vasculature and plays pivotal roles in the collection of interstitial fluid, absorbance of dietary lipids, and trafficking of immune cells[1]
parachordal lymphangioblast (PL) formation was severely impaired in morphants injected with EP3 MO1 or MO2 (Fig. 1A), and the ratio of PL-positive segments was significantly reduced in these morphants (Fig. 1B)
We investigated the formation of thoracic duct (TD), which is located immediately ventral to the dorsal aorta in the trunk, at 5 days post fertilization (Fig. 1E,F)

Summary

Introduction

The lymphatic system is a major component of the vertebrate vasculature and plays pivotal roles in the collection of interstitial fluid, absorbance of dietary lipids, and trafficking of immune cells[1]. The formation of lymphatic vessels is known to occur during embryogenesis and the progression of various cancers[22], and COX2-derived PGE2 was found to facilitate lymphangiogenesis during tumor development through the EP3 receptor in mice[23,24]. PGE2 was shown to accelerate the formation of lymphatic vessels through the EP3 receptor in granulation tissues[25,26]. These data suggest the possibility that the PGE2-EP3 pathway plays important roles in the formation of lymphatic vessels. We report novel functions of the PGE2-EP3 pathway in the formation of lymphatic vessels during early development

Methods

Results

Conclusion