Lysophosphatidic acid (LPA) receptors of the EDG family are differentially activated by LPA species. Structure-activity relationship of cloned LPA receptors

We examined the structure-activity relationship of cloned lysophosphatidic acid (LPA) receptors (endothelial cell differentiation gene (EDG) 2, EDG4, and EDG7) by measuring [Ca(2+)](i) in Sf9 insect cells expressing each receptor using LPA with various acyl chains bound at either the sn-1 or the sn-2 position of the glycerol backbone. For EDG7 the highest reactivity was observed with LPA with Delta9-unsaturated fatty acid (oleic (18:1), linoleic (18:2), and linolenic (18:3)) at sn-2 followed by 2-palmitoleoyl (16:1) and 2-arachidonoyl (20:4) LPA. In contrast, EDG2 and EDG4 showed broad ligand specificities, although EDG2 and EDG4 discriminated between 14:0 (myristoyl) and 16:0 (palmitoyl), and 12:0 (lauroyl) and 14:0 LPAs, respectively. EDG7 recognizes the cis double bond at the Delta9 position of octadecanoyl residues, since 2-elaidoyl (18:1, trans) and 2-petroselinoyl (18:1, cis-Delta12) LPA were poor ligands for EDG7. In conclusion, the present study demonstrates that each LPA receptor can be activated differentially by the LPA species.     

Mechanisms of the lysophosphatidic acid-induced increase in [Ca(2+)](i) in skeletal muscle cells.

Although lysophosphatidic acid (LPA) is known to increase intracellularfree calcium concentration ([Ca(2+)](i)) in different cell types, the effect of LPA on the skeletal muscle cells is not known. The present study was therefore undertaken to examine the effect of LPA on the [Ca(2+)](i) in C2C12 cells. LPA induced a concentration and time dependent increase in [Ca(2+)](i), which was inhibited by VPC12249, VPC 32183 and dioctanoyl glycerol pyrophosphate, LPA1/3 receptor antagonists. Pertussis toxin, a G(i) protein inhibitor, also inhibited the LPA-induced increase in [Ca(2+)](i). Inhibition of tyrosine kinase activities with tyrphostin A9 and genistein also prevented the increase in [Ca(2+)](i) due to LPA. Likewise, wortmannin and LY 294002, phosphatidylinositol 3-kinase (PI3-K) inhibitors, inhibited [Ca(2+)](i) response to LPA. The LPA effect was also attenuated by ethylene glycolbis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), an extracellular Ca(2+) chelator, Ni(2+) and KB-R7943, inhibitors of the Na(+)-Ca(2+) exchanger; the receptor operated Ca(2+) channel (ROC) blockers, 2-aminoethoxydiphenyl borate and SK&F 96365. However, the L-type Ca(2+) channel blockers, verapamil and diltiazem; the store operated Ca(2+) channel blockers, La(3+) and Gd(3+); a sarcoplasmic reticulum calcium pump inhibitor, thapsigargin; an inositol trisphosphate receptor antagonist, xestospongin and a phospholipase C inhibitor, U73122, did not prevent the increase [Ca(2+)](i) due to LPA. Our data suggest that the LPA-induced increase in [Ca(2+)](i) might occur through G(i)-protein coupled LPA(1/3) receptors that may be linked to tyrosine kinase and PI3-K, and may also involve the Na(+)-Ca(2+) exchanger as well as the ROC. In addition, LPA stimulated C2C12 cell proliferation via PI3-K. Thus, LPA may be an important phospholipid in the regulation of [Ca(2+)](i) and growth of skeletal muscle cells.     

Lysophosphatidic acid (LPA) receptors are activated differentially by biological fluids: possible role of LPA-binding proteins in activation of LPA receptors

Lysophosphatidic acid (LPA) exerts multiple biological functions through G protein-coupled receptors (EDG2/LPA(1), EDG4/LPA(2), and EDG7/LPA(3)) and is present in serum where it is associated with albumin. In this study we examined LPA activity in various biological fluids by measuring the LPA-induced increase in the intracellular concentration of calcium ion in three types of Sf9 insect cells, each expressing one of the LPA receptors. Using this system, we found that EDG2 and EDG4, but not EDG7, were activated strongly by addition of incubated plasma. By contrast, LPA detected in seminal plasma, which contains a low concentration of albumin, selectively activated EDG7. After LPA in these samples was extracted and reconstituted, it activated all three receptors. We also found that serum albumin readily inhibits the activation of EDG7 but not the activation of EDG2 or EDG4. In addition, plasma from Nagase analbuminemic rats but not plasma from control Sprague-Dawley rats was found to strongly activate EDG7, although the plasma of these two types of rats contained equal amounts of LPA and activated both EDG2 and EDG4. The present study shows that serum albumin can negatively regulate EDG7 but not EDG2 or EDG4, and suggests that protein factors are present in seminal plasma and deliver LPA efficiently to EDG7 but not to EDG2 or EDG4.     

Dioleoyl phosphatidic acid increases intracellular Ca2+ through endogenous LPA receptors in C6 glioma and L2071 fibroblasts

Phosphatidic acid (PA) increased intracellular Ca(2+) concentration ([Ca(2+)](i)) in C6 rat glioma and L2071 mouse fibroblast cells. Dioleoyl PA (PA, 18:1) was the most efficacious, followed by dipalmitoyl PA (16:0 PA) and dimyristoyl PA (14:0 PA). Lysophosphatidic acid (LPA) also increased the [Ca(2+)](i) in the both cells. PA desensitized LPA-induced Ca(2+) response completely in C6 cells, but partly in L2071 cells. Treatment of pertussis toxin (PTX), a specific inhibitor of G(i/o)-type G proteins, completely ameliorated LPA- and PA-induced Ca(2+) response in C6 cells. However, in L2071 cells, PTX inhibited PA-induced Ca(2+) increase by 80% and LPA-induced one by 20%. Ki16425, a specific inhibitor of LPA(1)/LPA(3) receptors, completely inhibited both LPA- and PA-induced Ca(2+) responses in C6 cells. On the other hand, in L2071 cells, Ki16425 completely inhibited PA-induced Ca(2+) response, but partly LPA-induced one. VPC32183, another specific inhibitor of LPA(1)/LPA(3) receptors, completely inhibited LPA- and PA-induced Ca(2+) responses in both C6 and L2071 cells. Therefore, PA and LPA appear to increase [Ca(2+)](i) through Ki16425/VPC32183-sensitive LPA receptor coupled to PTX-sensitive G proteins in C6 cells. In L2071 cells, however, LPA increases [Ca(2+)](i) through Ki16425-insensitive LPA receptor coupled to PTX-insensitive G proteins and Ki16425-sensitive LPA receptor coupled to PTX-sensitive G protein, whereas PA utilized only the latter pathway. Our results suggest that PA acts as a partial agonist on endogenous LPA receptors, which are sensitive to Ki16425 and coupled to PTX-sensitive G protein, but not on LPA receptors, which are not sensitive to Ki16425 and coupled to PTX-insensitive G protein.     

Identification of an EDG7 variant, HOFNH30, a G-protein-coupled receptor for lysophosphatidic acid

We have identified a cDNA, designated HOFNH30, which encodes a 354 amino acid G-protein-coupled receptor (GPCR). This receptor has 96% amino acid identity to the Jurkat-T cell-derived EDG7 and could be a splice variant. RT-PCR analysis demonstrated that HOFNH30 mRNA is expressed in placenta whereas EDG7 mRNA shows highest expression in prostate. The HOFNH30 gene is localized to human chromosome 1p22. 3-1p31.1. When HOFNH30 was expressed in RBL-2H3 cells, LPA and phosphatidic acid (PA) induced a calcium mobilization response with EC(50) values of 13 nM and 3 microM, respectively. LPA also induced phosphorylation of mitogen-activated protein kinase (p42(MAPK) and p44(MAPK)) in HOFNH30-transfected but not vector-transfected RBL-2H3 cells. In the present study, we have identified a novel variant from the EDG receptor family, a GPCR for which LPA is a high-affinity endogenous ligand.     

Lysophospholipid receptor-dependent and -independent calcium signaling

Changes in cellular Ca(2+) concentrations form a ubiquitous signal regulating numerous processes such as fertilization, differentiation, proliferation, contraction, and secretion. The Ca(2+) signal, highly organized in space and time, is generated by the cellular Ca(2+) signaling toolkit. Lysophospholipids, such as sphingosine-1-phosphate (S1P), sphingosylphosphorylcholine (SPC), or lysophosphatidic acid (LPA) use this toolkit in a specific manner to initiate their cellular responses. Acting as agonists at G protein-coupled receptors, S1P, SPC, and LPA increase the intracellular free Ca(2+) concentration ([Ca(2+)](i)) by using the classical, phospholipase C (PLC)-dependent pathway as well as PLC-independent pathways such as sphingosine kinase (SphK)/S1P. The S1P(1) receptor, via protein kinase C, inhibits the [Ca(2+)](i) transients caused by other receptors. Both S1P and SPC also act intracellularly to regulate [Ca(2+)](i). Intracellular S1P mobilizes Ca(2+) in intact cells independently of G protein-coupled S1P receptors, and Ca(2+) signaling by many agonists requires SphK-mediated S1P production. As shown for the FcepsilonRI receptor, PLC and SphK may contribute specific components to the overall [Ca(2+)](i) transient. Of the many open questions, identification of the intracellular S1P target site(s) appears to be of particular importance.     

Edg-6 as a putative sphingosine 1-phosphate receptor coupling to Ca(2+) signaling pathway

The endothelial differentiation gene-6 (Edg-6) was recently identified as an orphan G-protein-coupled receptor. Its predicted amino acid sequence is very close to Edg family of receptor proteins whose ligand is supposed to be lysophosphatidic acid (LPA) or lysosphingolipid such as sphingosine 1-phosphate (S1P) and sphingosylphosphorylcholine (SPC). Transfection of the Edg-6 into Chinese hamster ovary (CHO) cells and K562 cells resulted in the appearance of high-affinity [(3)H]S1P binding activity. Among lipids employed, S1P and, even though less potent, SPC, displaced the [(3)H]S1P binding, but LPA was inactive. In Edg-6-transfected CHO cells, an increase in cytosolic Ca(2+) concentration in response to S1P or SPC was clearly enhanced without change in the LPA-induced action as compared with the vector-transfected cells. The enhancement of the Ca(2+) response was associated with a significant accumulation of inositol phosphate, reflecting activation of phospholipase C. Similar enhancement of Ca(2+) response to S1P or SPC was also observed in Edg-6-expressing K562 cells. These lipid-induced actions in CHO cells and K562 cells expressing Edg-6 were markedly suppressed by pertussis toxin treatment. We conclude that Edg-6 is one of S1P or lysosphingolipid receptors that couple to phospholipase C-Ca(2+) system through pertussis toxin-sensitive G-proteins.     

Gintonin, newly identified compounds from ginseng, is novel lysophosphatidic acids-protein complexes and activates G protein-coupled lysophosphatidic acid receptors with high affinity

Recently, we isolated a subset of glycolipoproteins from Panax ginseng, that we designated gintonin, and demonstrated that it induced [Ca2+]i transients in cells via G protein-coupled receptor (GPCR) signaling pathway(s). However, active components responsible for Ca2+ mobilization and the corresponding receptor(s) were unknown. Active component(s) for [Ca2+]i transients of gintonin were analyzed by liquid chromatography-electrospray ionization-tandem mass spectrometry and ion-mobility mass spectrometry, respectively. The corresponding receptor(s)were investigated through gene expression assays. We found that gintonin contains LPA C18:2 and other LPAs. Proteomic analysis showed that ginseng major latex-like protein and ribonuclease-like storage proteins are protein components of gintonin. Gintonin induced [Ca2+]i transients in B103 rat neuroblastoma cells transfected with human LPA receptors with high affinity in order of LPA2 >LPA5 > LPA1 > LPA3 > LPA4. The LPA1/LPA3 receptor antagonist Ki16425 blocked gintonin action in cells expressing LPA1 or LPA3. Mutations of binding sites in the LPA3 receptor attenuated gintonin action. Gintonin acted via pertussis toxin (PTX)-sensitive and -insensitive G protein-phospholipase C (PLC)-inositol 1,4,5-trisphosphate (IP3)-Ca2+ pathways. However, gintonin had no effects on other receptors examined. In human umbilical vein endothelial cells (HUVECs) gintonin stimulated cell proliferation and migration. Gintonin stimulated ERK1/2 phosphorylation. PTX blocked gintonin-mediated migration and ERK1/2 phosphorylation. In PC12 cells gintonin induced morphological changes, which were blocked by Rho kinase inhibitorY-27632. Gintonin contains GPCR ligand LPAs in complexes with ginseng proteins and could be useful in the development of drugs targeting LPA receptors.     

Lysophosphatidic acid-induced Ca2+ mobilization requires intracellular sphingosine 1-phosphate production. Potential involvement of endogenous EDG-4 receptors

Lysophosphatidic acid (LPA)-mediated Ca(2+) mobilization in human SH-SY5Y neuroblastoma cells does not involve either inositol 1,4, 5-trisphosphate (Ins(1,4,5)P(3))- or ryanodine-receptor pathways, but is sensitive to inhibitors of sphingosine kinase. This present study identifies Edg-4 as the receptor subtype involved and investigates the presence of a Ca(2+) signaling cascade based upon the lipid second messenger molecule, sphingosine 1-phosphate. Both LPA and direct G-protein activation increase [(3)H]sphingosine 1-phosphate levels in SH-SY5Y cells. Measurements of (45)Ca(2+) release in premeabilized SH-SY5Y cells indicates that sphingosine 1-phosphate, sphingosine, and sphingosylphosphorylcholine, but not N-acetylsphingosine are capable of mobilizing intracellular Ca(2+). Furthermore, the effect of sphingosine was attenuated by the sphingosine kinase inhibitor dimethylsphingosine, or removal of ATP. Confocal microscopy demonstrated that LPA stimulated intracellular Ca(2+) "puffs," which resulted from an interaction between the sphingolipid Ca(2+) release pathway and Ins(1,4,5)P(3) receptors. Down-regulation of Ins(1,4,5)P(3) receptors uncovered a Ca(2+) response to LPA, which was manifest as a progressive increase in global cellular Ca(2+) with no discernible foci. We suggest that activation of an LPA-sensitive Edg-4 receptor solely utilizes the production of intracellular sphingosine 1-phosphate to stimulate Ca(2+) mobilization in SH-SY5Y cells. Unlike traditional Ca(2+) release processes, this novel pathway does not require the progressive recruitment of elementary Ca(2+) events.     

The influence of lysophosphatidic acid on the functions of human dendritic cells

Lysophosphatidic acid (LPA) is a bioactive lipid mediator which is generated by secretory phospholipase A(2). In this study, we studied the biological activity of LPA on human dendritic cells (DCs), which are specialized APCs characterized by their ability to migrate into target sites and secondary lymphoid organs to process Ags and activate naive T cells. We show that immature and mature DCs express the mRNA for different LPA receptors such as endothelial differentiation gene (EDG)-2, EDG-4, and EDG-7. In immature DCs, LPA stimulated pertussis toxin-sensitive Ca(2+) increase, actin polymerization, and chemotaxis. During the maturation process, DCs lost their ability to respond toward LPA with Ca(2+) transients, actin polymerization, and chemotaxis. However, LPA inhibited in a pertussis toxin-insensitive manner the secretion of IL-12 and TNFalpha as well as enhanced secretion of IL-10 from mature DCs. Moreover, LPA did not affect the endocytic or phagocytic capacities and the surface phenotype of DCs, although it increased the allostimulatory function of mature DC and inhibited their capacity to induce Th1 differentiation. In summary, our study implicates that LPA might regulate the trafficking, cytokine production, and T cell-activating functions of DCs.     

The essential role of H2O2 in the regulation of intracellular Ca2+ by epidermal growth factor in rat-2 fibroblasts

We have investigated a new mechanism by which epidermal growth factor (EGF) increases intracellular Ca(2+) ([Ca(2+)](i)) in Rat-2 fibroblasts. EGF induced a transient increase of [Ca(2+)](i), and sustained Ca(2+) increase disappeared in the absence of extracellular Ca(2+). However, EGF had no effect on the formation of inositol phosphates. Expression of N17Rac or scrape-loading of C3 transferase blocked the elevation of [Ca(2+)](i) by EGF, but not by lysophosphatidic acid (LPA). EGF increased intracellular H(2)O(2), with a maximal increase at 5 min, which was blocked by catalase, scrape-loading of C3 transferase, or expression of N17Rac. H(2)O(2) scavengers, catalase and N-acetyl-L-cysteine, also blocked the Ca(2+) response to EGF, but not to LPA. In the presence of EGTA, preincubation with EGF completely inhibited subsequent Ca(2+) response to extracellular H(2)O(2) and vice versa. Incubation with EGF or phosphatidic acid abolished subsequent elevation of [Ca(2+)](i) by phosphatidic acid or EGF, respectively. Furthermore, preincubation with LPA inhibited the subsequent Ca(2+) response to EGF, but not vice versa. These results suggested that intracellular H(2)O(2) regulated by Rac and RhoA, but not inositol phosphates, was responsible for the EGF-stimulated elevation of [Ca(2+)](i). It was also suggested that EGF cross talked with LPA in the regulation of [Ca(2+)](i) by producing intracellular H(2)O(2).     

A novel phosphatidic acid-selective phospholipase A1 that produces lysophosphatidic acid

Lysophosphatidic acid (LPA) is a lipid mediator with diverse biological properties, although its synthetic pathways have not been completely solved. We report the cloning and characterization of a novel phosphatidic acid (PA)-selective phospholipase A(1) (PLA(1)) that produces 2-acyl-LPA. The PLA(1) was identified in the GenBank(TM) data base as a close homologue of phosphatidylserine (PS)-specific PLA(1) (PS-PLA(1)). When expressed in insect Sf9 cells, this enzyme was recovered from the Triton X-100-insoluble fraction and did not show any catalytic activity toward exogenously added phospholipid substrates. However, culture medium obtained from Sf9 cells expressing the enzyme was found to activate EDG7/LPA(3), a cellular receptor for 2-acyl-LPA. The activation of EDG7 was further enhanced when the cells were treated with phorbol ester or a bacterial phospholipase D, suggesting involvement of phospholipase D in the process. In the latter condition, an increased level of LPA, but not other lysophospholipids, was confirmed by mass spectrometry analyses. Expression of the enzyme is observed in several human tissues such as prostate, testis, ovary, pancreas, and especially platelets. These data show that the enzyme is a membrane-associated PA-selective PLA(1) and suggest that it has a role in LPA production.     

Alkyl lysophosphatidic acid and fluoromethylene phosphonate analogs as metabolically-stabilized agonists for LPA receptors

We describe an efficient method for the synthesis of alkyl lysophosphatidic acid (LPA) analogs as well as alkyl LPA mono- and difluoromethylene phosphonate analogs. Each alkyl LPA analog was evaluated for subtype-specific LPA receptor agonist activity using a cell migration assay for LPA(1) activation in cancer cells and an intracellular calcium mobilization assay for LPA(2) and LPA(3) activation. Alkyl LPAs induced pronounced cell migration activity with equivalent or higher potency than sn-1-oleoyl LPA, while the alkyl LPA fluoromethylene phosphonates proved to be less potent agonists in this assay. However, each alkyl LPA analog activated Ca(2+) release by activation of LPA(2) and LPA(3) receptors. Interestingly, the absolute configuration of the sn-2 hydroxyl group of the alkyl LPA analogs was not recognized by any of the three LPA receptors. The use of alkyl LPA analogs further expands the scope of structure-activity studies, which will better define LPA-LPA receptor interactions.     

Ca(2+) signalling is not required for chemotaxis in Dictyostelium

Dictyostelium cells can move rapidly towards a source of cyclic-AMP (cAMP). This chemoattractant is detected by G-protein-linked receptors, which trigger a signalling cascade including a rapid influx of Ca(2+). We have disrupted an inositol 1,4,5-trisphosphate (InsP(3)) receptor-like gene, iplA, to produce null cells in which Ca(2+) entry in response to chemoattractants is abolished, as is the normal increase in free cytosolic Ca(2+) ([Ca(2+)](c)) that follows chemotactic stimulation. However, the resting [Ca(2+)](c) is similar to wild type. This mutant provides a test for the role of Ca(2+) influx in both chemotaxis and the signalling cascade that controls it. The production of cyclic-GMP and cAMP, and the activation of the MAP kinase, DdERK2, triggered from the cAMP receptor, are little perturbed in the mutant; mobilization of actin into the cytoskeleton also follows similar kinetics to wild type. Mutant cells chemotax efficiently towards cAMP or folic acid and their sensitivity to cAMP is similar to wild type. Finally, they move at similar speeds to wild-type cells, with or without chemoattractant. We conclude that Ca(2+) signalling is not necessary for chemotaxis to cAMP.     

EDG3 is a functional receptor specific for sphingosine 1-phosphate and sphingosylphosphorylcholine with signaling characteristics distinct from EDG1 and AGR16.

AGR16/H218/EDG5 and EDG1 are functional receptors for lysosphingolipids, whereas EDG2 and EGD4 are receptors for lysophosphatidic acid (LPA). The present study demonstrates that EDG3, the yet poorly defined member of the EDG family G protein-coupled receptors, shows identical agonist specificity, but distinct signaling characteristics, compared to AGR16 and EDG1. Overexpression of EDG3 conferred a specific [32P]S1P binding, which was displaced by S1P and sphingosylphosphorylcholine (SPC), but not by LPA or other related lipids. In cells overexpressing EDG3, S1P induced inositol phosphate production and [Ca2+]i increase in a manner only partially sensitive to pertussis toxin (PTX), which was similar to the case of AGR16, but quite different from the case of EDG1, in which the S1P-induced responses were totally abolished by PTX. EDG3 also mediated activation of mitogen-activated protein kinase (MAPK) in PTX-sensitive and Ras-dependent manners, as in the cases of EDG1 and AGR16, although EDG3 and EDG1 were more effectively coupled to activation of MAPK, compared to AGR16. Additionally, EDG3 mediated a decrease in cellular cyclic AMP content, like EDG1, but contrasting with AGR16 which mediated an increase in cyclic AMP. These and previous results establish that EDG1, AGR16 and EDG3 comprise the lysosphingolipid receptor subfamily, each showing distinct signaling characteristics.     

Evidence that IP3 and ryanodine-sensitive intra-cellular Ca2+ stores are not involved in acute hyposmotically-induced prolactin release in Tilapia.

Prolactin (PRL) cells from the euryhaline tilapia, Oreochromis mossambicus, behave like osmoreceptors by responding directly to reductions in medium osmolality with increased secretion of the osmoregulatory hormone PRL. Extracellular Ca(2+) is essential for the transduction of a hyposmotic stimulus into PRL release. In the current study, the presence and possible role of intracellular Ca(2+) stores during hyposmotic stimulation was investigated using pharmacological approaches. Changes in intracellular Ca(2+) concentration were measured with fura-2 in isolated PRL cells. Intracellular Ca(2+) stores were depleted in dispersed PRL cells with thapsigargin (1 microM) or cyclopiazonic acid (CPA, 10 microM). Pre-incubation with thapsigargin prevented the rise in [Ca(2+)](i) induced by lysophosphatidic acid (LPA, 1 microM), an activator of the IP(3) signalling cascade, but did not prevent the hyposmotically-induced rise in [Ca(2+)](i) in medium with normal [Ca(2+)] (2mM). Pre-treatment with CPA produced similar results. Prolactin release from dispersed cells followed a pattern that paralleled observed changes in [Ca(2+)](i). CPA inhibited LPA-induced prolactin release but not hyposmotically-induced release. Xestospongin C (1microM), an inhibitor of IP(3) receptors, had no effect on hyposmotically-induced PRL release. Pre-exposure to caffeine (10mM) or ryanodine (1microM) did not prevent a hyposmotically-induced rise in [Ca(2+)](i). Taken together these results indicate the presence of IP(3) and ryanodine-sensitive Ca(2+) stores in tilapia PRL cells. However, the rapid rise in intracellular [Ca(2+)] needed for acute PRL release in response to hyposmotic medium can occur independently of these intracellular Ca(2+) stores.     

EDG receptors and hepatic pathophysiology of LPA and S1P: EDG-ology of liver injury

The biological roles of phospholipid growth factors lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) have been broadly investigated. The cellular effects of LPA and S1P are mediated predominantly via endothelial differentiation gene (EDG) receptors. Yet, the biological significance of LPA, S1P and their EDG receptors in cells of the liver remains unclear. Recent data demonstrate the presence of EDG2 and EDG4 mRNA for LPA receptor in a murine hepatocyte cell line transformed with human TGF-alpha, and in primary mouse hepatocytes. EDG2 receptor protein is expressed in mouse liver, where it appears to be located in nonparenchymal cells. Moreover, we have obtained data suggesting that proliferation of small hepatocyte-progenitors and stem (oval) cells during liver injury is associated with the expression of EDG2 and EDG4 receptors. LPA, and possibly S1P, appear to be essential factors that control proliferation and motility of hepatic stellate cells (HSC) and hepatoma cells. It is proposed that LPA, S1P and their respective EDG receptors play important roles in pathophysiology of chronic liver injury and fibrogenesis. The underlying mechanisms recruited by LPA and S1P in pathogenesis of liver injury remain to be investigated.