Kyoko Noguchi

LPA Receptors: Subtypes and Biological Actions

Lysophosphatidic acid (LPA) is a small, ubiquitous phospholipid that acts as an extracellular signaling molecule by binding to and activating at least five known G protein-coupled receptors (GPCRs): LPA(1)-LPA(5). They are encoded by distinct genes named LPAR1-LPAR5 in humans and Lpar1-Lpar5 in mice. The biological roles of LPA are diverse and include developmental, physiological, and pathophysiological effects. This diversity is mediated by broad and overlapping expression patterns and multiple downstream signaling pathways activated by cognate LPA receptors. Studies using cloned receptors and genetic knockout mice have been instrumental in uncovering the significance of this signaling system, notably involving basic cellular processes as well as multiple organ systems such as the nervous system. This has further provided valuable proof-of-concept data to support LPA receptors and LPA metabolic enzymes as targets for the treatment of medically important diseases that include neuropsychiatric disorders, neuropathic pain, infertility, cardiovascular disease, inflammation, fibrosis, and cancer.

FTY720 (fingolimod) efficacy in an animal model of multiple sclerosis requires astrocyte sphingosine 1-phosphate receptor 1 (S1P1) modulation

Sphingosine 1-phosphate (S1P), a lysophospholipid, has gained relevance to multiple sclerosis through the discovery of FTY720 (fingolimod), recently approved as an oral treatment for relapsing forms of multiple sclerosis. Its mechanism of action is thought to be immunological through an active phosphorylated metabolite, FTY720-P, that resembles S1P and alters lymphocyte trafficking through receptor subtype S1P1. However, previously reported expression and in vitro studies of S1P receptors suggested that direct CNS effects of FTY720 might theoretically occur through receptor modulation on neurons and glia. To identify CNS cells functionally contributing to FTY720 activity, genetic approaches were combined with cellular and molecular analyses. These studies relied on the functional assessment, based on clinical score, of conditional null mouse mutants lacking S1P1 in CNS cell lineages and challenged by experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. All conditional null mutants displayed WT lymphocyte trafficking that responded normally to FTY720. In marked contrast, EAE was attenuated and FTY720 efficacy was lost in CNS mutants lacking S1P1 on GFAP-expressing astrocytes but not on neurons. In situ hybridization studies confirmed that astrocyte loss of S1P1 was the key alteration in functionally affected mutants. Reductions in EAE clinical scores were paralleled by reductions in demyelination, axonal loss, and astrogliosis. Receptor rescue and pharmacological experiments supported the loss of S1P1 on astrocytes through functional antagonism by FTY720-P as a primary FTY720 mechanism. These data identify nonimmunological CNS mechanisms of FTY720 efficacy and implicate S1P signaling pathways within the CNS as targets for multiple sclerosis therapies.

Lysophosphatidic acid (LPA) and its receptors.

Lysophosphatidic acid (LPA), a bioactive phospholipid, and its family of cognate G protein-coupled receptors have demonstrated roles in many biological functions in the nervous system. To date, five LPA receptors have been identified, and additional receptors may exist. Most of these receptors have been genetically deleted in mice toward identifying biological and medically relevant roles. In addition, small molecule agonists and antagonists have been reported. Here we review recent data on the nervous system functions of LPA signaling, and summarize data on reported agonists and antagonists of LPA receptors.

LPA4/p2y9/GPR23 mediates rho-dependent morphological changes in a rat neuronal cell line.

Lysophosphatidic acid (LPA) is a potent lipid mediator that evokes a variety of biological responses in many cell types via its specific G protein-coupled receptors. In particular, LPA affects cell morphology, cell survival, and cell cycle progression in neuronal cells. Recently, we identified p2y9/GPR23 as a novel fourth LPA receptor, LPA4 (Noguchi, K., Ishii, S., and Shimizu, T. (2003) J. Biol. Chem. 278, 25600-25606). To assess the functions of LPA4 in neuronal cells, we used rat neuroblastoma B103 cells that lack endogenous responses to LPA. In B103 cells stably expressing LPA4, we observed Gq/11-dependent calcium mobilization, but LPA did not affect adenylyl cyclase activity. In LPA4 transfectants, LPA induced dramatic morphological changes, i.e. neurite retraction, cell aggregation, and cadherin-dependent cell adhesion, which involved Rho-mediated signaling pathways. Thus, our results demonstrated that LPA4 as well as LPA1 couple to Gq/11 and G12/13, whereas LPA4 differs from LPA1 in that it does not couple to Gi/o. Through neurite retraction and cell aggregation, LPA4 may play a role in neuronal development such as neurogenesis and neuronal migration.

LPA4 regulates blood and lymphatic vessel formation during mouse embryogenesis

Lysophosphatidic acid (LPA) is a potent lipid mediator with a wide variety of biological actions mediated through G protein-coupled receptors (LPA(1-6)). LPA(4) has been identified as a G(13) protein-coupled receptor, but its physiological role is unknown. Here we show that a subset of LPA(4)-deficient embryos did not survive gestation and displayed hemorrhages and/or edema in many organs at multiple embryonic stages. The blood vessels of bleeding LPA(4)-deficient embryos were often dilated. The recruitment of mural cells, namely smooth muscle cells and pericytes, was impaired. Consistently, Matrigel plug assays showed decreased mural cell coverage of endothelial cells in the neovessels of LPA(4)-deficient adult mice. In situ hybridization detected Lpa4 mRNA in the endothelium of some vasculatures. Similarly, the lymphatic vessels of edematous embryos were dilated. These results suggest that LPA(4) regulates establishment of the structure and function of blood and lymphatic vessels during mouse embryogenesis. Considering the critical role of autotaxin (an enzyme involved in LPA production) and Gα(13) in vascular development, we suggest that LPA(4) provides a link between these 2 molecules.

Lysophosphatidic Acid Signaling May Initiate Fetal Hydrocephalus

Blockade of lysophosphatidic acid signaling provides a new strategy for treating fetal hydrocephalus. Is the Cause of Hydrocephalus Blood Simple? Hydrocephalus or “water on the brain” is caused by accumulation of cerebrospinal fluid (CSF) in the cerebral ventricles during fetal development and is one of the most common neurological disorders of newborns, occurring in 1 in 1500 live births. One apparent cause of hydrocephalus is bleeding into the cerebral ventricles or brain tissue of the fetus, suggesting that factors or components in blood may trigger development of this severe neurological disorder. The most common treatment is surgical insertion of an intraventricular shunt that drains excess CSF from the cerebral ventricles, but this approach only relieves intracranial pressure and does not solve the root cause of the disorder. Yung et al. set out to investigate which factors in blood trigger hydrocephalus using an in vivo fetal mouse model that they developed. They identify a blood-borne lipid called lysophosphatidic acid (LPA) as a potential cause of hydrocephalus and show that when LPA is prevented from binding to its receptor LPA1 by a receptor antagonist, that hydrocephalus does not develop in fetal mice. The authors injected serum, plasma, or red blood cells into the cerebral ventricles of the brains of fetal mice in utero at 13.5 days of gestation. The animals were then assessed prenatally 1 or 5 days later or postnatally at several different time points. Injection of serum or plasma but not red blood cells induced CSF accumulation and hydrocephalus, with animals displaying enlarged heads, dilated ventricles, and thinning of the cortex. The investigators reasoned that LPA, a blood-borne lipid that is known to be important for the developing cerebral cortex, might be involved in the development of hydrocephalus. When they injected a solution containing LPA into the cerebral ventricles of fetal mice in utero, the mice did indeed develop severe hydrocephalus. The authors wondered how an increase in LPA might affect cortical development and lead to hydrocephalus. They show that injection of LPA resulted in altered adhesion and mislocalization of neural progenitor cells along the surface of the ventricles and that this mislocalization depended on expression of the LPA1 receptor by these cells. When the researchers repeated their experiments with fetal mice lacking the LPA1 receptor, they were unable to induce hydrocephalus. The key finding came with their demonstration that an LPA1 receptor antagonist blocked the ability of LPA to induce hydrocephalus in the fetal mice. These results suggest that LPA and its LPA1 receptor may be new therapeutic targets for developing drugs that could be used in conjunction with surgery to treat this debilitating neurological disease. Fetal hydrocephalus (FH), characterized by the accumulation of cerebrospinal fluid, an enlarged head, and neurological dysfunction, is one of the most common neurological disorders of newborns. Although the etiology of FH remains unclear, it is associated with intracranial hemorrhage. Here, we report that lysophosphatidic acid (LPA), a blood-borne lipid that activates signaling through heterotrimeric guanosine 5′-triphosphate–binding protein (G protein)–coupled receptors, provides a molecular explanation for FH associated with hemorrhage. A mouse model of intracranial hemorrhage in which the brains of mouse embryos were exposed to blood or LPA resulted in development of FH. FH development was dependent on the expression of the LPA1 receptor by neural progenitor cells. Administration of an LPA1 receptor antagonist blocked development of FH. These findings implicate the LPA signaling pathway in the etiology of FH and suggest new potential targets for developing new treatments for FH.

Cysteinyl leukotriene 2 receptor-mediated vascular permeability via transendothelial vesicle transport

Cysteinyl leukotrienes (CysLTs) are potent mediators of inflammation synthesized by the concerted actions of 5‐lipoxygenase (5‐LO), 5‐LO‐activating protein (FLAP), leukotriene C4 synthase, and additional downstream enzymes, starting with arachidonic acid substrate. CysLTs produced by macrophages, eosinophils, mast cells, and other inflammatory cells activate 3 different high‐affinity CysLT receptors: CysLT1R, CysLT2R, and GPR 17. We sought to investigate vascular sites of CysLT2R expression and the role and mechanism of this receptor in mediating vascular permeability events. Vascular expression of CysLT2R was investigated by reporter gene expression in a novel CysLT2R deficient‐LacZ mouse model. CysLT2R was expressed in small, but not large, vessels in mouse brain, bladder, skin, and cremaster muscle. Intravital, in addition to confocal and electron, microscopy investigations using FIT C‐labeled albumin in cremaster postcapillary venule preparations indicated rapid CysLT‐mediated permeability, which was blocked by application of BAY‐u9773, a dual CysLT1R/CysLT2R antagonist or by CysLT2R deficiency. Endothelial human CysLT2R overexpression in mice exacerbated vascular leakage even in the absence of exogenous ligand. The enhanced vascular permeability mediated by CysLT2R takes place via a transendothelial vesicle transport mechanism as opposed to a paracellular route and is controlled via Ca2+ signaling. Our results reveal that CysLT2R can mediate inflammatory reactions in a vascular bed‐specific manner by altering transendothelial vesicle transport‐based vascular permeability.— Moos, M. P. W., Mewburn, J. D., Kan, F. W. K., Ishii, S., Abe, M., Sakimura, K., Noguchi, K., Shimizu, T., Funk, C. D. Cysteinyl leukotriene 2 receptor‐mediated vascular permeability via transendothelial vesicle transport. FASEB J. 22, 4352–4362 (2008)

Stereotyped fetal brain disorganization is induced by hypoxia and requires lysophosphatidic acid receptor 1 (LPA1) signaling

Fetal hypoxia is a common risk factor that has been associated with a range of CNS disorders including epilepsy, schizophrenia, and autism. Cellular and molecular mechanisms through which hypoxia may damage the developing brain are incompletely understood but are likely to involve disruption of the laminar organization of the cerebral cortex. Lysophosphatidic acid (LPA) is a bioactive lipid capable of cortical influences via one or more of six cognate G protein-coupled receptors, LPA1–6, several of which are enriched in fetal neural progenitor cells (NPCs). Here we report that fetal hypoxia induces cortical disruption via increased LPA1 signaling involving stereotyped effects on NPCs: N-cadherin disruption, displacement of mitotic NPCs, and impaired neuronal migration, as assessed both ex vivo and in vivo. Importantly, genetic removal or pharmacological inhibition of LPA1 prevented the occurrence of these hypoxia-induced phenomena. Hypoxia resulted in overactivation of LPA1 through selective inhibition of G protein-coupled receptor kinase 2 expression and activation of downstream pathways including Gαi and Ras-related C3 botulinum toxin substrate 1. These data identify stereotyped and selective hypoxia-induced cerebral cortical disruption requiring LPA1 signaling, inhibition of which can reduce or prevent disease-associated sequelae, and may take us closer to therapeutic treatment of fetal hypoxia-induced CNS disorders and possibly other forms of hypoxic injury.

Roles for lysophospholipid S1P receptors in multiple sclerosis

Sphingosine 1-phosphate (S1P) signaling in the treatment of multiple sclerosis (MS) has been highlighted by the efficacy of FTY720 (fingolimod), which upon phosphorylation can modulate S1P receptor activities. FTY720 has become the first oral treatment for relapsing MS that was approved by the FDA in September 2010. Phosphorylated FTY720 modulates four of the five known S1P receptors (S1P1, S1P3, S1P4, and S1P5) at high affinity. Studies in human MS and its animal model, experimental autoimmune encephalomyelitis (EAE), have revealed that FTY720 exposure alters lymphocyte trafficking via sequestration of auto-aggressive lymphocytes within lymphoid organs, representing the current understanding of its mechanism of action. These effects primarily involve S1P1, which is thought to attenuate inflammatory insults in the central nervous system (CNS). In addition, FTY720’s actions may involve direct effects on S1P receptor-mediated signaling in CNS cells, based upon the known expression of S1P receptors in CNS cell types relevant to MS, access to the CNS through the blood–brain barrier (BBB), and in vitro studies. These data implicate lysophospholipid signaling – via S1P1 and perhaps other lysophospholipid receptors – in therapeutic approaches to MS and potentially other diseases with immunological and/or neurological components.