Prostaglandin F2 and EP2 Agonists Exert Different Effects on 3D 3T3-L1 Spheroids during Their Culture Phase.

Yosuke Ida,Fumihito Hikage,Megumi Watanabe,Masato Furuhashi,Araya Umetsu,Hiroshi Ohguro

doi:10.3390/biomedicines9121821

Abstract

To elucidate the effects of switching a PGF2α agonist, bimatoprost acid (BIM-A), to an EP2 agonist (Omidenepag—OMD; butaprost—Buta) or reversing the switching on adipose tissue, two-dimensional (2D) and three-dimensional (3D) cultures of 3T3-L1 cells were analyzed by lipid staining and according to the mRNA expression of adipogenesis-related genes (Pparγ, Ap2, and Leptin), components of the extracellular matrix (ECM; collagen1 (Col1), Col4, Col6, and fibronectin (Fn)), and the sizes and stiffness of the 3D spheroids. Switching from BIM-A to EP2 agonists caused (1) suppression of lipid staining and downregulation of most adipogenesis-related genes, (2) smaller and stiffer 3D spheroids, and (3) upregulation of Col1 and Fn, downregulation of Col4 (2D), or up-regulation of all ECM genes (3D, BIM-A to OMD), as well as downregulation of Col6 (3D, BIM-A to Buta). In contrast, reversing the switching resulted in (1) an enhancement in lipid staining (2D) and a significant upregulation of adipogenesis-related genes (2D, 3D Buta to BIM-A), (2) larger and slightly stiffer 3D spheroids, and (3) upregulation of Col1 and Fn (2D). These collective findings indicate that the switching orders of BIM-A and EP2 agonists have a significant effect on lipid metabolism, ECM expression, and the physical stiffness of 3T3-L1 cells.

Highlights

Glaucomatous optic neuropathy (GON), a condition that is associated with elevated intraocular pressure (IOP), is a leading cause of irreversible blindness throughout the world [1,2], and the only effective evidenced-based therapies for GON are hypotensive therapies with medications and/or surgery [3]
In our previous studies of the effects of the mono-treatment of PGF2α-ags and EP2 agonists using 2D- and 3D-cultured 3T3-L1 cells, we reported that both receptors are coexpressed within the 3T3-L1 cells, and that PGF2α-ags and EP2 agonists exert different effects on adipogenesis and the physical properties of the 2D- and 3D-cultured 3T3-L1 cells [8,15]
In terms of adipogenesis and the physical stiffness of the 3D spheroids, since we showed that these changes were more evident in the later (Days 5 to 7) rather than the earlier period during the course of the 3D cultures [8,16], the unexpected results for Pparγ in the expression of Ap2 observed in cases of switching from OMD or Buta to bimatoprost acid (BIM-A), which was assumed to be due of 15 to the effects of a mono-treatment, may have been caused by some unknown9 cross-linkages between EP2 and PGF2α-ags receptors that are related to signaling (Table 1)

Summary

Introduction

Glaucomatous optic neuropathy (GON), a condition that is associated with elevated intraocular pressure (IOP), is a leading cause of irreversible blindness throughout the world [1,2], and the only effective evidenced-based therapies for GON are hypotensive therapies with medications and/or surgery [3]. To establish a more suitable in vivo model to replicate an adipocyte-spreading environment within a 3D conic space, a three-dimensional (3D) drop culture technique was used This 3D in vivo model relatively accurately replicated a significant PGF2α-ags-induced

Methods

Results

Conclusion