Lysophosphatidic acid modulates prostaglandin signalling in bovine steroidogenic luteal cells

Highlights

• LPA increases mPGES1 and cPGES expression and decreases PGFS expression in bSLCs.

• LPA stimulates EP2 and EP4 receptor and PGT expression in bSLCs.

• LPA influences intraluteal PG balance in bSLCs.

• LPA may support CL function and lifespan in cows.

• The results revealed identify linkage between LPA and PG biosynthesis in the bovine CL.

Abstract

We examined whether lysophosphatidic acid affects prostaglandin biosynthesis, transport, and signalling in bovine steroidogenic luteal cells. The aim of the present study was to determine the influence of LPA on PGE2 and PGF2α synthesis and on the expression of enzymes involved in PG biosynthesis (PTGS2, mPGES-1, cPGES, mPGES-2, PGFS and 9-KPR), prostaglandin transporter (PGT), and prostaglandin receptors (EP1, EP2, EP3, EP4 and FP) in bovine steroidogenic luteal cells. We found that LPA inhibited PGF_2α synthesis in steroidogenic luteal cells. Moreover, LPA increased mPGES1 and cPGES and decreased PGFS expression in cultured bovine steroidogenic luteal cells. Additionally, LPA stimulated EP2 and EP4 receptor and PGT expression. This study suggests that LPA activity in the bovine CL directs the physiological intraluteal balance between the two main prostanoids towards luteotropic PGE₂.

Introduction

The corpus luteum (CL) is an endocrine organ established from the remaining cells of the ovulated follicle. The mature CL consists of a heterogeneous cell population. There are at least two steroidogenic cell types: large luteal cells of granulosa cell origin and small luteal cells of theca cell origin [1], [2]. Although progesterone (P4) is the major luteal hormone, the CL also produces other hormones, including prostaglandins (PGs) [3]. Phospholipase activity induces PG biosynthesis, which releases arachidonic acid (AA) from the phospholipid membrane. Thereafter, AA is converted into the PG precursor PG endoperoxide H₂ (PGH₂) by PG endoperoxide synthases (better known as cyclooxygenase) [4]. Prostaglandin E synthases (PGESs) and PGF synthase (PGFS) metabolize prostaglandin H₂ into PGE₂ and PGF_2α, respectively. Additionally, an alternate pathway of PGF_2α synthesis is the conversion of PGE₂ into PGF_2α through PG 9-ketoreductase (9-KPR) activity [5]. Current evidence suggests that three forms of PGES exist: microsomal PGES1 and 2 (mPGES1 and mPGES2) and cytosolic PGES (cPGES) [6], [7], [8], [9].

Newly synthesized PGs are transported through the plasma membrane by prostaglandin transporter (PGT) [10], [11]. This molecule is a broadly expressed, 12-membrane-spanning domain integral membrane protein [12]. A previous study found that luteal PGT expression was lower in the early growing than in late growing, mature, and regressing phases of the oestrous cycle. Therefore, PGT is almost exclusively expressed in large luteal cells (LLCs) [13]. The same study suggested that PGT facilitated both the efflux and influx of available luteal PGE₂ and/or PGF_2α in a competitive manner to affect their autocrine and paracrine actions, as well as their catabolism during the different stages of the CL life span.

Prostaglandin E₂ and PGF_2α primarily exert their effects through G protein-coupled receptors designated EP and FP, respectively [14], [15]. PGE receptor subtypes (EP1, EP2, EP3 (A–D) and EP4) and an FP receptor have been identified. EP2 and EP4 are coupled to adenylate cyclase and generate cAMP, which activates the protein kinase A signalling pathway. EP1 and FP receptors are coupled to phospholipase C to generate two second messengers, inositol triphosphate involved in the liberation of intracellular calcium (Ca²⁺) and diacyl glycerol, an activator of protein kinase C. There are four of EP3 receptor isoforms, A–D. These EP3 receptors exhibit a wide range of action, from the inhibition of cAMP production to the increase in Ca²⁺ and inositol phosphate 3 [14], [16]. Thus, FP and EP are expressed at high levels in the corpus luteum, particularly in the large luteal cells [13], [17].

Lysophosphatidic acid (LPA) is a naturally occurring, small, bioactive phospholipid. This ligand plays several roles in the female reproductive tract [18] by activating its G protein-coupled receptors LPAR1-6 [19]. Previous studies found that LPA stimulated PTGS2 mRNA expression in the porcine endometrium [20] and increased PGE₂ synthesis in the ovine trophectoderm [21] and bovine endometrium [22]. Moreover, infusion of heifers with LPA prevented spontaneous luteolysis, prolonged the functional lifespan of the CL and stimulated luteotropic PGE₂ synthesis in vivo [23]. We have found that LPA is locally produced and released from the bovine CL [24] and that LPA reversed the inhibitory effect of NO donor on the PGE₂/PGF_2α ratio in cultured bovine steroidogenic luteal cells (bSLCs) [25]. The above studies identified a linkage between LPA signalling and PG biosynthesis in bovine CL. However, it is unknown whether LPA can differentially affect PG biosynthesis, transport, and signalling in bSLCs.

Taken together, the evidence suggests that PGF_2α and PGE₂ expression in the bovine CL depends not only on PTGS2 but also on PG synthases, transporter, and specific EP and FP receptors. According to previous results that described LPA as an additional luteosupportive factor in bSLCs [24] and an important modulator of oestrous cycle length in cows [25], we hypothesize that LPA can modulate PG biosynthesis, transport, and signalling in bSLCs. Therefore, the main objective of this study was to evaluate the effect of LPA on PG synthesis via the expression of enzymes responsible for their biosynthesis (PTGS2, mPGES-1, cPGES, mPGES-2, PGFS and 9-KPR), transport (PGT), and receptors (EP1, EP2, EP3, EP4 and FP) in bSLCs.