Sphingosine 1-phosphate (S1P) induces COX-2 expression and PGE 2 formation via S1P receptor 2 in renal mesangial cells Anja Völzke a,1 , Alexander Koch a, ⁎ ,1 , Dagmar Meyer zu Heringdorf a , Andrea Huwiler b , Josef Pfeilschifter a a Pharmazentrum Frankfurt/ZAFES, Klinikum der Johann Wolfgang Goethe-Universität, Theodor-Stern-Kai 7, D-60590 Frankfurt am Main, Germany b Institute of Pharmacology, University of Bern, Friedbühlstrasse 49, CH-3011 Bern, Switzerland abstract article info Article history: Received 18 April 2013 Received in revised form 27 August 2013 Accepted 17 September 2013 Available online 21 September 2013 Keywords: Sphingosine 1-phosphate S1P receptor COX-2 PGE 2 Mesangial cells Understanding the mechanisms of sphingosine 1-phosphate (S1P)-induced cyclooxygenase (COX)-2 expression and prostaglandin E 2 (PGE 2 ) formation in renal mesangial cells may provide potential therapeutic targets to treat inflammatory glomerular diseases. Thus, we evaluated the S1P-dependent signaling mechanisms which are re- sponsible for enhanced COX-2 expression and PGE 2 formation in rat mesangial cells under basal conditions. Fur- thermore, we investigated whether these mechanisms are operative in the presence of angiotensin II (Ang II) and of the pro-inflammatory cytokine interleukin-1β (IL-1β). Treatment of rat and human mesangial cells with S1P led to concentration-dependent enhanced expression of COX-2. Pharmacological and molecular biology ap- proaches revealed that the S1P-dependent increase of COX-2 mRNA and protein expression was mediated via ac- tivation of S1P receptor 2 (S1P 2 ). Further, inhibition of G i and p42/p44 MAPK signaling, both downstream of S1P 2 , abolished the S1P-induced COX-2 expression. In addition, S1P/S1P 2 -dependent upregulation of COX-2 led to sig- nificantly elevated PGE 2 levels, which were further potentiated in the presence of Ang II and IL-1β. A functional consequence downstream of S1P/S1P 2 signaling is mesangial cell migration that is stimulated by S1P. Interestingly, inhibition of COX-2 by celecoxib and SC-236 completely abolished the migratory response. Overall, our results demonstrate that extracellular S1P induces COX-2 expression via activation of S1P 2 and subsequent G i and p42/p44 MAPK-dependent signaling in renal mesangial cells leading to enhanced PGE 2 formation and cell migra- tion that essentially requires COX-2. Thus, targeting S1P/S1P 2 signaling pathways might be a novel strategy to treat renal inflammatory diseases. © 2013 Elsevier B.V. All rights reserved. 1. Introduction In addition to their important role in the regulation of the glomerular filtration rate, renal mesangial cells contribute to the most pathological processes affecting the renal glomerulus [1,2]. Besides enhanced prolif- eration and production of extracellular matrix proteins, secretion of in- flammatory mediators by mesangial cells is a hallmark of many forms of renal diseases. In this context, it was demonstrated that cyclooxygenase (COX)-2 is upregulated in e.g. glomeruli of the anti-Thy-1 glomerulone- phritis model in rats and in the kidney of patients with active lupus nephritis [3,4]. In general, contrary to the constitutively expressed COX-1 [5], COX-2 is inducible by various factors, e.g. cytokines and mi- togens, and is predominantly expressed during acute or chronic inflam- mation (for review, see [6]). Thereby, COX-2 is a key enzyme necessary for the conversion of arachidonic acid into prostaglandin H 2 , which is then immediately converted into several secondary prostaglandins, such as prostaglandin E 2 (PGE 2 ). Previously, several studies investigated the role of sphingosine 1- phosphate (S1P) for the induction of COX-2 and the subsequent forma- tion of pro-inflammatory PGE 2 . S1P is generated intracellularly via the action of two specific enzymes, sphingosine kinase (SK)-1 and SK-2. S1P can act either inside the cell, or extracellularly via activation of five specific G-protein coupled receptors, denoted as S1P 1–5 , which are located at the cell surface and characterized by a cell type-specific ex- pression pattern (for review, see [7,8]). S1P receptors (S1PR) couple to and activate a variety of different G-proteins, namely G i (S1P 1–5 ), G q (S1P 2/3 ), and G 12/13 (S1P 2–5 ), which in turn results in partly diverse functional properties of single S1PR (for review, see [9]). In this context, the role of specific S1PR subtypes for S1P-induced COX-2 expression is still controversial and depends on the cell type and experimental setup used in previous studies. Li et al. [10] suggested that S1P induces COX-2 expression and subsequent PGE 2 formation via S1P 2 in the Biochimica et Biophysica Acta 1841 (2014) 11–21 Abbreviations: AA, arachidonic acid; ActD, actinomycin D; Ang II, angiotensin II; CLX, celecoxib; COX, cyclooxygenase; HuR, human antigen R; IL-1β, interleukin-1β; MAPK, mitogen-activated protein kinase; MEK, MAPK kinase; NAC, N-acetylcystein; PGE 2 , prosta- glandin E 2 ; PLA 2 , group II phospholipase 2a; PTX, pertussis toxin; ROCK, Rho-dependent kinase; S1P, sphingosine 1-phosphat; S1PR, S1P receptors; SK, sphingosine kinase ⁎ Corresponding author. Tel.: +49 69 6301 6352; fax: +49 69 6301 6942. E-mail addresses: voelzke@med.uni-frankfurt.de (A. Völzke), koch@med.uni-frankfurt.de (A. Koch), heringdorf@med.uni-frankfurt.de (D. Meyer zu Heringdorf), huwiler@pki.unibe.ch (A. Huwiler), pfeilschifter@em.uni-frankfurt.de (J. Pfeilschifter). 1 These authors contributed equally to this work. 1388-1981/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.bbalip.2013.09.009 Contents lists available at ScienceDirect Biochimica et Biophysica Acta journal homepage: www.elsevier.com/locate/bbalip