I. Introduction

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Feldberg, Kellaway, and coworkers (Feldberg and Kellaway, 1938; Feldberg et al., 1938; Kellaway and Trethewie, 1940) observed that perfusion of guinea pig lungs with antigen induced the release of a material named "slow reaction smooth muscle-stimulating substance (SRS¹)" that caused a contraction of the isolated guinea pig ileum bioassay tissue. These observations were confirmed by several workers (Schild et al., 1951; Brocklehurst, 1960) who demonstrated that SRS (renamed slow-reacting substance of anaphylaxis or SRS-A) was also released from the human lung following antigen challenge. Sweatman and Collier (1968) reported that SRS-A constricted human airways and the compound FPL 55712 (Augstein et al., 1973) was shown to inhibit SRS-A-induced contractions in the guinea pig ileum assay. These observations provoked an intense interest in elucidating the biochemical nature of this entity. Initial attempts to characterize this substance revealed that the factor was a low-molecular weight derivative of arachidonic acid (Orange et al., 1973; Bach et al., 1977; Jakschik et al., 1977) containing sulfur (Orange et al., 1973, Parker et al., 1979). SRS-A was identified subsequently to be a family of lipid mediators known as leukotrienes, a name derived from their cell source (leukocytes) and their conjugated double bonds (triene) structure (Borgeat et al., 1976; Borgeat and Samuelsson, 1979a,b,c; Murphy et al., 1979; Corey et al., 1980; Lewis et al., 1980; Morris et al., 1980; Rokach et al., 1980). Leukotriene B₄ (LTB₄) was the first of the leukotrienes to be isolated (Borgeat et al., 1976).

The elucidation of the structures and synthetic pathways for the leukotrienes lead to a considerable amount of research on these arachidonic acid metabolites (Fig. 1). This work involved comprehensive assessments of the biological profiles of both the cysteinyl-leukotrienes (cys-LTs: LTC₄, LTD₄, and LTE₄) as well as dihydroxy-leukotriene (LTB₄) and, more recently, the lipoxins. Lipoxins (LX), an acronym for eicosanoids, which are often generated during the transcellular metabolism of arachidonic acid via the sequential actions of the 15- and 5- or 5- and 12-lipoxygenase enzymatic pathways (Serhan et al., 1984; Samuelsson et al., 1987). When the synthetic ligands were made available many studies documented a myriad of actions for these lipid mediators (Table 1) providing pertinent evidence for their possible patho-physiological roles in inflammatory diseases, in particular asthma. During the last 20 years significant efforts involving diverse chemical strategies have been directed toward the identification and development of receptor antagonists. These compounds have facilitated the identification and characterization of distinct receptors, which are activated by either the dihydroxy- or cysteinyl-leukotrienes.

View larger version (23K): [in this window] [in a new window]   Fig. 1.   Major pathways for leukotriene and lipoxin formation. The leukotrienes and lipoxins are lipid mediators derived from arachidonic acid, which is released from cell membrane phospholipids by the action of phospholipase A2. Leukotriene formation is initiated by 5-lipoxygenase, which catalyzes the dioxygenation of arachidonic acid to 5-HPETE and the subsequent conversion to LTA4. This latter unstable epoxide is transformed either to LTB4 or LTC4, and LTC4 is further catalyzed to LTD4 and LTE4. The tetraene epoxide intermediate can be formed either from LTA4 or 15-HPETE depending upon the interactions of the different lipoxygenases, and this metabolite is enzymatically hydrolyzed to the lipoxins (LXA4 and LXB4).

	II. General Considerations

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The leukotrienes are formed via activation of the 5-lipoxygenase enzyme (5-LO) in collaboration with the "5-lipoxygenase activating protein" (FLAP). A prerequisite for this enzymatic reaction is the hydrolyzation of arachidonic acid from membrane phospholipids by phospholipase A₂. The principal 5-LO products of arachidonic acid metabolism are LTC₄ and LTB₄ as well as 5-hydroxyeicosatetranoic acid (5-HETE). In addition, eicosanoids that are formed by pathways that involve the dual lipoxygenation of arachidonic acid by either 15- and 5-LO or 5- and 12-LO are referred to as lipoxins (Serhan et al., 1984; Samuelsson et al., 1987). The transcellular metabolism of intermediates such as LTA₄ and 15(S)-HETE is associated with LX formation (Serhan, 1994). LX and their carbon 15-epimer-LXs (aspirin-triggered lipoxins; ASA-15-epi-LX) are bioactive and structurally distinct from other eicosanoids in that they carry a conjugated tetraene system and are present in biological matrix in two main forms that are positional isomers, namely, lipoxin A₄ (5S,6R,15S-trihydroxyeicosa-7E,9E,11Z,13E-tetraenoic acid) and lipoxin B₄ (5S,14R,15S-trihydroxyeicosa-6E,8Z,10E,12E-tetraenoic acid; Serhan, 1997; Fig. 1). The aspirin-triggered form carry their C15 alcohol in the R configuration, which is inserted by COX-2 following aspirin treatment (denoted ASA-15-epi-LX). These metabolites are often produced during cell-to-cell interactions, and the principal targets appear to be platelets and leukocytes. During these cellular interactions, platelets convert neutrophil derived LTA₄ to 5,6,-epoxytetraene through the action of platelet 12-LO. However, under these conditions the term 12-LO is a misnomer since this enzymatic activity was originally based on an interaction with arachidonic acid. This enzyme functions as a 15-LO (LX synthase) when the substrate is LTA₄. Thus in an inflammatory condition LTA₄ serves as a pivotal intermediate for both leukotriene and lipoxin formation.

The leukotrienes are formed in different cell types as well as via transcellular metabolism involving multiple cells such as neutrophil and platelets and vascular cells (Feinmark and Cannon, 1986; Maclouf and Murphy, 1988; Sala et al., 1993). Human eosinophils and neutrophils synthesize both LTC₄ and LTB₄, respectively (Bray et al., 1980; Ford-Hutchinson et al., 1980). Monocytes and macrophages also synthesize both LTB₄ and the cys-LTs (Samuelsson, 1983). LTC₄ is metabolized to LTD₄ and LTE₄ by the cells in which this mediator is formed. In addition, the cys-LTs can be transformed into 6-trans-LTB₄ by hypochlorous acid, which is generated during the respiratory burst in leukocytes (Henderson et al., 1982; Lee et al., 1983). LTB₄ is also metabolized in the cells which produce this metabolite, by a unique membrane bound cytochrome P450 enzyme. LTB₄ is metabolized to 20-hydroxy-LTB₄ (Hansson et al., 1981; Shak and Goldstein, 1985; Soberman et al., 1985). There is also evidence for a reductase dehydrogenase in polymorphonuclear leukocytes (PMN) that appears to be specific for LTB₄ (Powell et al., 1989).

The previous IUPHAR publication (Coleman et al., 1995) introduced two main classes of leukotriene receptors. One based on the biological activities of leukotriene B₄ and related hydroxyacids, referred to as BLT receptors, and a second class identified by the cysteinyl-leukotrienes (cys-LTs). The different profiles of biological activity for these two classes of metabolites were the initial basis for these categories and were supported by structure-activity data obtained in studies with a variety of compounds that selectively antagonized the different ligands. Activation of the BLT receptors initially was shown to produce potent chemotactic activities on leukocytes whereas the latter class (CysLT receptors) stimulated smooth muscle as well as other cells. However, the structures of the leukotriene receptors have recently been deduced from the nucleotide sequences of the cDNAs and the encoding proteins are now known for human, mouse, and rat. These data have permitted the IUPHAR committee to establish the nomenclature for the leukotriene receptors, and this is presented in Table 2. The phylogenic tree for the different eicosanoid and bioactive lipid G-protein-coupled receptors (GPCR) is illustrated in Fig. 2 and shows the molecular families with the relationship between leukotrienes and lipoxins as well as other proteins with seven transmembrane helices.

View this table: [in this window] [in a new window]   TABLE 2 Human cloned leukotriene receptorsa Data are from the following references: Fiore et al., 1992, 1994; Perez et al., 1992; Maddox et al., 1997; Takano et al., 1997; Yokomizo et al., 1997; Lynch et al., 1999; Sarau et al., 1999; Heise et al., 2000; Nothacker et al., 2000; Takasaki et al., 2000; Figueroa et al., 2001.

View larger version (24K): [in this window] [in a new window]   Fig. 2.   Dendogram of several GPCRs. The major receptor families for lipid mediators are indicated in this phylogenic tree. There are four known cell surface GPCRs for the leukotrienes, which are classified as either chemoattractants (BLT1 and BLT2) or nucleotide receptors (CysLT1 and CysLT2). The lipoxin receptor (ALX) is also included in the chemoattractant receptor class along with formyl peptide receptors (FPL). This evolutionary tree was constructed using the sequences from the receptors. Construction was performed by using the "All All Program" at the Computational Biochemistry Server at ETHZ (http://cbrg.inf.ethz.ch/ServerBooklet/chapter2-3.html).

The lipoxins, are chemically and functionally different from the leukotrienes (Fig. 1). Although LXA₄ and LXB₄ are similar in structure, these mediators display biological activities that are quite distinct. LXA₄ interactions with neutrophils involves binding sites that are not recognized by LXB₄ (Nigam et al., 1990; Fiore et al., 1992). LXB₄ is a potent agonist for stimulating proliferation and differentiation of granulocyte-monocyte colonies from human mononuclear cells (Popov et al., 1989), increasing the S-phase in the cell cycle and enhancing nuclear protein kinase C activity (Beckman et al., 1992) actions, which have not been reported for LXA₄. However, LXB₄ has also been shown to share actions with LXA₄, such as, both selectively stimulate human peripheral blood monocytes (Maddox and Serhan, 1996) and enhance growth of myeloid progenitor cells (Stenke et al., 1991). Furthermore, LXA₄ does not activate BLT (Fiore et al., 1992) but activates FPRL-1 receptors (Chiang et al., 2000; Resnati et al., 2002; Perretti et al., 2002). These investigators have shown that ALX and FPRL-1 are the same receptor and that LXA₄ is the natural and most potent ligand. In addition, Takano et al. (1997) have identified the amino acid sequence for the receptor associated with the LXA₄ responses. In line with the IUPHAR nomenclature directives, this committee recommends that ALX be used to designate the receptor that has been cloned and is activated by the native ligand LXA₄ (Table 2). LXB₄-induced responses, although different from those of LXA₄, have not to date provided sufficient evidence to specify another receptor. Since this receptor has not been cloned, the LXB₄ response is associated with activation of a putative receptor.

The aim of this review is to present the evidence that led to the leukotriene nomenclature. To this end, information not only from the molecular database but also derived from the properties and significance of leukotriene receptors will be presented. Furthermore, the above nomenclature for the LX receptors is recommended as the framework for this evolving area of receptor research.

	III. Molecular Database for Leukotriene Receptors

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A. Molecular and Structural Aspects of Dihydro-Leukotriene Receptors

1. BLT₁. The cloning and characterization of the BLT₁ receptor was achieved by cDNA subtraction using human leukemic cells HL-60, which were differentiated into granulocyte-like cells (Yokomizo et al., 1997). The BLT₁ receptor was identified as a putative seven transmembrane domain receptor with 352 amino acids. This receptor had been initially misidentified as a purinergic receptor, P2Y7 (Akbar et al., 1996). BLT₁ shares low homology to P2Y receptors and belongs to a family of receptors for chemoattractants including complement receptors and a recently identified novel prostaglandin D₂ receptor, CRTH2 (Hirai et al., 2001). The homology between the BLT₁ receptor for mouse and humans is presented in Fig. 3.

View larger version (69K): [in this window] [in a new window]   Fig. 3.   The sequence alignment of BLT1 and BLT2 from human and mouse receptors. The amino acid sequences were aligned using ClustalW and converted using Boxshade 3.21. The putative transmembrane domains of hBLT1 predicted by Kyte-Doolittle hydrophobicity analysis are overlined and labeled as I-VII. Consensus matches are boxed and shaded with darker shading for identities and light shading for conservative substitutions. The amino acid identity between human and mouse BLT1 was 78.6% whereas BLT2 was 92.7%. The mouse sequence data are available from Swiss-Prot under accession numbers (mBLT1: no entry presently available) and (mBLT2: Q9JJL9).

2. BLT₂. During the analysis of transcriptional regulation of human BLT₁ gene (Kato et al., 2000), a putative ORF for a novel GPCR with structural similarity to BLT₁ was identified (Yokomizo et al., 2000). This novel receptor was also found in a human genome sequence database, reported to act as a low-affinity receptor activated by LTB₄ (Kamohara et al., 2000; Tryselius et al., 2000; Wang et al., 2000) and subsequently referred to as the BLT₂ receptor (Yokomizo et al., 2001b). The gene structure for BLT₂ has also been established (Yokomizo et al., 2001b). Of considerable interest is that the promoter region (Fig. 4) of human BLT₁ overlaps BLT₂ ORF (Kato et al., 2000). This represents the "promoter in ORF", as has been reported in prokaryotes but the biological significance of this rare gene structure is presently not clear. However, there is sufficient evidence that BLT₁ and BLT₂ form a gene cluster both in human (chromosome 14 q11.2-q12;) and mouse (Yokomizo et al., 2000) chromosomes suggesting that these receptors may be generated by gene duplication.

View larger version (12K): [in this window] [in a new window]   Fig. 4.   Structure of the human genomic DNA containing BLT1 and BLT2 located on human chromosome 14q11.2-q12. Chromosome 14 is indicated by the line; the transcribed segments are indicated by open boxes; putative ORFs are shown as filled boxes. Note that the promotor region for BLT1 is located in the ORF of BLT2. This is the first mammalian example of "promotor in ORF" (Yokomizo et al., 2001b).

3. Phenotypes Involving BLT Receptors. Investigations with transgenic mice expressing the human BLT₁ receptor on leukocytes (Chiang et al., 1999) as well as targeted gene disruption of the BLT₁ receptor in knockout mice (BLT/) indicate that an apparent phenotypic difference (Haribabu et al., 2000; Tager et al., 2000) from wild type littermates is not observed unless the animals are subject to experimental disease or injury, which are known to stress the effector immune system (vide infra).

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B. Molecular and Structural Aspects of Cysteinyl-Leukotriene Receptors

1. CysLT₁. The cloning and characterization of the human CysLT₁ receptor (hCysLT₁) was achieved by two groups under the general program of identifying cognate ligands for orphan GPCRs, a process which has been termed "ligand fishing" (Lynch et al., 1999; Sarau et al., 1999). The hCysLT₁ receptor was identified as a 337-amino acid putative seven transmembrane domain receptor, termed either HG55 (Lynch et al., 1999) or HMTMF81 (Sarau et al., 1999) (Fig. 5). The former investigators demonstrated that LTD₄ produced activation of a calcium-activated chloride channel in Xenopus laevis oocytes expressing the cRNA for HG55 but not in control cells or oocytes expressing other GPCRs. This LTD₄-induced stimulation of oocytes was blocked by the selective CysLT₁ receptor antagonist MK-571 (Lynch et al., 1999) (Table 5). Similar results were obtained using the X. laevis melanophore signaling assay and in mammalian monkey kidney COS-7 cells expressing the HG55 (hCysLT₁) receptor (Lynch et al., 1999).

View larger version (39K): [in this window] [in a new window]   Fig. 5.   Comparison of amino acid sequences of the human CysLT1 and CysLT2 receptors. A G-protein-coupled receptor snake diagram depiction of the amino acid sequences of the human CysLT1 and CysLT2 receptors. The amino acid identities between the hCysLT1 and hCysLT2 receptors is 37.3%.

2. CysLT₂. The cloning and characterization of the CysLT₂ receptor (hCysLT₂) was initially reported by Heise et al. (2000) (Fig. 5). This publication confirmed the previous pharmacological characterization of a human CysLT₂ receptor in different tissues, based upon the relative potencies of the cys-LT agonists and the lack of sensitivity of the responses to classical CysLT₁ receptor antagonists, and the antagonist activity of the partial agonist BAY u9773 (Labat et al., 1992; Tudhope et al., 1994; Heise et al., 2000). Subsequent to this initial publication, the Takeda group published an article confirming the identification of the hCysLT₂ (Takasaki et al., 2000), and then a third report by the Nothacker et al. (2000), on the characteristics of the hCysLT₂ was published, which revealed similar distribution and functional data to the previous publications but with more details on the partial agonist activity of BAY u9773 (Nothacker et al., 2000). Recently, the cloned mCysLT₂ has also been reported (Hui et al., 2001).

Of the nonprostanoid eicosanoid GPCRs, the LXA₄ receptor (ALX) was the first recognized at the molecular level (Fiore et al., 1993, 1994). In addition, ALX was initially identified as the only inhibitory or anti-inflammatory receptor that acts via an agonist role as a "stop signal" (Fiore et al., 1994; Serhan, 1994, 1997; Takano et al., 1997). This action appears to be a unique flexibility of GPCR that functions within the immune system. Since LXA₄ shares some structural features with LTC₄ and LTD₄ as well as prostaglandins, LXA₄ competed for CysLT₁ receptors identified on isolated human vascular endothelial cells (Gronert et al., 2001) and mesangial cells (McMahon et al., 2000) and antagonized either LTC₄- or LTD₄-induced bronchoconstriction in humans (Christie et al., 1992) and animals (Badr et al., 1989; Gronert et al., 2001). In addition, lipoxin B₄ has also been reported to activate another receptor. The present nomenclature for the lipoxin receptors is therefore based on the cloned receptor sequence as well as the observation that LXA₄ is the natural and most potent ligand. In contrast, the putative receptor activated by LXB₄ has not been cloned. ALX activation has been reported to generate intracellular stop signals (Serhan et al., 1994; Levy et al., 1997, 1999) and thereby promote resolution of inflammation.

1. Molecular and Structural Aspects of Lipoxin Receptors. Based on the finding that functional ALX are inducible in promyelocytic lineages (HL-60 cells) (Fiore et al., 1993), several putative receptor cDNAs cloned earlier from myeloid lineages and designated orphans were screened for their ability to bind and signal in response to LXA₄ (Fiore et al., 1994). When transfected into CHO cells, one of the orphans (previously denoted as pINF114 or a formyl peptide receptor-like-1 (FPRL-1), displayed both specific [³H]LXA₄ binding with high affinity (K_d of 1.7 nM) and demonstrated ligand selectivity when compared with LXB₄, LTB₄, LTD₄, and prostaglandin E₂ (Fiore et al., 1994). LXB₄ did not act via the ALX receptor and interacted with a specific receptor present on human leukocytes (Maddox and Serhan, 1996). In transfected CHO cells, LXA₄ activated both GTPase and released arachidonic acid from membrane phospholipids, indicating that this cDNA encodes a functional receptor for ALX in myeloid cells. A mouse ALX receptor cDNA was also identified and cloned from a spleen cDNA library. This receptor expressed in CHO cells displayed specific [³H]LXA₄ binding, and LXA₄ initiated GTPase activity (Takano et al., 1997).

Whereas the cascades of cellular events subsequent to GPCR activation have been the subject of many investigations, the exact signal transduction mechanisms for either the leukotrienes or the lipoxins have not been completely elucidated. Generally, agonist interactions with GPCRs involve activation of heterotrimeric G-proteins associated with a group of conventional cellular events. However, effectors for GPCRs that are independent of G-proteins are also known to exist (Hall et al., 1999). G-proteins, composed of -, -, and -subunits each encoded by a different gene, appear often to be cell specific. Upon ligand-receptor activation, the G- and G-subunits stimulate a variety of intracellular molecular systems. Furthermore, G-protein activation leads to increases in intracellular Ca²⁺ and modifications in a number of membrane ion channels.

The cellular responses to ligand activation of GPCRs can also be up-regulated through priming of cells and down-regulated by desensitization. Two types of desensitization have been described, one that results from phosphorylation of the agonist-occupied receptor by G-protein-coupled receptor kinases. These phosphorylated receptors are associated with the arrestin family of proteins. A second type of rapid desensitization (loss of response) following phosphorylation by either second messenger-activated kinases (protein kinase A, protein kinase C) or inhibition of phospholipase C, which are activated by different receptors or signaling processes. Generally, this second type of desensitization does not require agonist-receptor occupancy. In addition, Didsbury et al. (1991) also demonstrated "cross-receptor desensitization", a phenomenon that has been reported for the chemoattractant family receptors. Presently, an exploration of these latter mechanisms associated with the actions of leukotrienes and lipoxins at the molecular level has received little attention.

1. BLT. Investigations involving the intracellular signaling of BLT receptor activation have been performed in peripheral leukocytes specifically granulocytes. One of the problems involved in such studies is that these cells have a limited life span (24 h) making drug and transfection studies difficult. These limitations have caused several investigators to use either CHO cells expressing human BLT receptors (Yokomizo et al., 1997) or to perform reconstitutional studies with the heterotrimeric GTP-binding proteins (Miki et al., 1990; Igarashi et al., 1999). Although high-affinity binding of LTB₄ (BLT₁ receptor) is found essentially in leukocytes and macrophages, the G-proteins associated with the functions in these cells has not been clearly established. Furthermore, the intracellular signaling pathways for BLT may depend on the G-proteins expressed in the different cells. For example, most of the LTB₄-dependent signals in granulocytes appear to be mediated by G_i-like G-proteins, (granulocytes express abundant G_i proteins, mainly G_i2), whereas in the nervous system G_i1 and G_o are mainly present (Simon et al., 1991). In several cell types, LTB₄ signals via G_i-proteins are inhibited by pretreatment of pertussis toxin (PTX). However, LTB₄-induced calcium mobilization in CHO-BLT₁ was not affected by PTX, suggesting the coupling with G_q-like molecules in these latter cells. Chemotaxis and inhibition of adenylyl cyclase by LTB₄ were completely PTX-sensitive in CHO-BLT₁ cells. The coupling of BLT₁ with various G-subunits was examined by cotransfection studies using COS-7 cells, and BLT₁-mediated phospholipase C activation was shown to be mediated by G_i6- and G-subunits released from G_i (Gaudreau et al., 1998). When expressed heterologously in CHO, HeLa, and COS-7 cells, BLT₂ activation led to the inhibition of adenylyl cyclase and an increase in calcium. However, BLT₂ activation was less potent in mobilizing calcium than BLT₁ receptor activation (Yokomizo et al., 2000). BLT₂ was also shown to mediate LTB₄-dependent chemotaxis through G_i-like G-proteins (Kamohara et al., 2000; Yokomizo et al., 2000). Recently, Woo et al. (2002) have suggested that LTB₄ stimulation of the Rac-extracellular signal-regulated kinase cascade associated with the generation of reactive oxygen species-mediated chemotaxis in Rat-2 cells was via activation of the BLT₂ receptor. This suggestion, although not conclusive, was supported by the observations that BLT₁ expression has not been detected in Rat-2 fibroblasts whereas BLT₂ was expressed. Furthermore, the LTB₄ stimulation of reactive oxygen species was observed at high concentrations (0.3-1 µM), which are within the range for BLT₂ activation and are 2 orders of magnitude higher than that observed for activation of BLT₁. In addition, this LTB₄ stimulation was blocked by ZK 158252. In an attempt to understand the mechanisms involved in BLT receptor desensitization, Gaudreau et al. (2002) have reported some initial molecular evidence. These investigators showed that the cytoplasmic tail of BLT₁ receptor was intimately involved in the regulation of desensitization and that the amino acid threonine (Thr³⁰⁸) was implicated in the GPCR-specific kinase phosphorylation associated with this phenomenon. This study therefore provides pertinent leads for understanding those structural elements associated with BLT₁ receptor regulation.

2. CysLT. Unfortunately, most studies concerning the CysLT receptors have involved only LTD₄ activation of CysLT₁ receptors. There is little information available concerning G-protein and Ca²⁺ mobilization when the CysLT₂ receptor is activated. Initial studies (Kuehl et al., 1984; Crooke et al., 1989, 1990; Watanabe et al., 1990) demonstrated that LTD₄ activation of the CysLT₁ receptor lead to G-protein activation and the release of several second intracellular messengers, namely, diacylglycerol, inositol phosphates, and Ca²⁺, events which were followed by activation of protein kinase C (PKC) and accompanied by the mobilization of Ca²⁺ derived from both intracellular and extracellular stores. Clark et al. (1985) demonstrated that LTD₄ activation of CysLT₁ receptors also led to the release of arachidonic acid via stimulation of phospholipase A₂, which was associated with an enhanced transciption of phospholipase A₂ activating protein. Expression of this latter protein was controlled by activation of topoisomerase I, which in turn was regulated by PKC (Mattern et al., 1991).

3. Lipoxins. The cytoplasmic signaling cascade of the ALX receptor is also highly specific and selective for different cell types. In human PMNs, LXA₄ stimulates rapid lipid remodeling and release of arachidonic acid via a PTX-sensitive G-protein (Nigam et al., 1990) and blocked intracellular generation of inositol 1,4,5-trisphosphate (Grandordy et al., 1990) as well as Ca²⁺ mobilization (Lee et al., 1989). In contrast, in human monocytes and THP-1 cells, LXA₄ triggers intracellular calcium release (Romano et al., 1996; Maddox et al., 1997), suggesting a different intracellular signaling pathway than in PMNs despite identical receptor sequences. In addition, distinct signaling in monocyte and PMNs was further supported by different responses to LXA₄ in these cell types. LXA₄ modulates mitogen-activated protein kinase activities in mesangial cells in a PTX-insensitive manner (McMahon et al., 2000), suggesting the presence of an additional ALX receptor subtype and/or signaling pathway for ALX. Since the ALX receptor has been shown to switch recognition and function with certain chemotactic peptides, the G-proteins and intracellular pathways involved may prove to be a difficult but fascinating area to explore. One of the problems presently confronting investigators in this area of research is the availability of the ligands. Studies on G-protein and intracellular messengers are presently limited (Kang et al., 2000), since stable analogs for LXA₄ and LXB₄ have only recently become available.

Within the last few years, a considerable effort at the molecular level has been undertaken to identify the leukotriene receptors. However, data involving chimeric constructs of the leukotriene receptors have only recently been reported (Gaudreau et al., 2002). In contrast, there are several observations that warrant further investigation. For example, the mouse CysLT₁ cloned receptor is activated by all three native ligands and antagonized by MK-571. However, the ligand profile for the mCysLT₁ is quite different from that observed in the human CysLT₁ receptor, since the mouse CysLT₁ receptor exhibited little response to LTC₄. An explanation for this difference is not readily apparent. In addition, MK-571 potentiated Ca²⁺ mobilization in CHO cells transfected with mCysLT₁ long isoform cDNA (Maekawa et al., 2001). The exact reason for this specific effect has not been explored. Recently, Ogasawara et al. (2002) reported different pharmacological properties of the CysLT₂ receptor between human and mouse, an