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Prostanoids: Prostaglandins, Prostacyclins
and Thromboxanes

The prostanoids are part of the oxylipin family of biologically active lipids derived from the action of cyclooxygenases or prostaglandin synthases upon the twenty-carbon essential fatty acids or eicosanoids, mainly arachidonic acid. They can be further subdivided into two main groups, the prostacyclopentanes, comprising the prostaglandins and prostacyclins, and the thromboxanes, each of which is involved in some aspect of the inflammatory response. The prostaglandins were first isolated from semen and named from the prostate gland, thought to be their source, as long ago as the 1930s, but it was the 1960s before the biosynthetic relationship to specific essential fatty acids was described and intensive research into their biological properties began. The Nobel Prize for Medicine for 1982 was awarded to Professors Bengt Samuelsson, John Vane and Sune Bergström for their discoveries in this field (see the review by Samuelsson cited below). In general, prostaglandins occur at very low levels in tissues, of the order of nanomolar concentrations, but they have profound biological activities. While most studies have been concerned with their occurrence and function in mammals, they have also been detected in birds, ray-finned fishes, marine invertebrates, trypanosomes, blood flukes, and some algae and yeasts.

1.  Nomenclature and Structures of Prostanoids

In structure, prostanoids are best considered as derivatives of a C20 saturated fatty acid, prostanoic acid, which does not itself occur in nature. A key feature is a five-membered ring encompassing carbons 8 to 12, as illustrated below. The thromboxanes are similar but have heterocyclic oxane structures. They are all synthesised by specific enzymes, which confer stereospecificity and chirality on every functional group, and are thus distinct from the isoprostanes, which are produced by non-enzymic means.

Structural formulae of prosanoids

In the approved nomenclature, each prostaglandin is named using the prefix 'PG' followed by a letter A to K depending on the nature and position of the substituents on the ring. Thus PGA to PGE and PGJ have a keto group in various positions on the ring, and are further distinguished by the presence or absence of double bonds or hydroxyl groups in various positions in the ring. PGF has two hydroxyl groups while PGK has two keto substituents on the ring. PGG and PGH are bicyclic endoperoxides. An oxygen bridge between carbons 6 and 9 distinguishes prostacyclin (PGI). Thromboxane A (TXA) contains an unstable bicyclic oxygenated ring structure, while thromboxane B (TXB) has a stable oxane ring. In addition, all prostaglandins have a hydroxyl group on carbon 15 and a trans-double bond at carbon 13 of the alkyl substituent (R2).

Further, a numerical subscript (1 to 3) is used to denote the total number of double bonds in the alkyl substituents, and a Greek subscript (α or β) is used with prostaglandins of the PGF series to describe the stereochemistry of the hydroxyl group on carbon 9. This is illustrated for prostaglandins PGE and PGFα of the 1, 2 and 3 series below, as examples.

Prostaglandin precursors and products

The number of double bonds depends on the nature of the fatty acid precursor. Thus, the prostaglandins PGE1, PGE2 and PGE3 are derived from 8c,11c,14c-eicosatrienoic (dihomo-γ-linolenic), 5c,8c,11c,14c-eicosatetraenoic (arachidonic) and 5c,8c,11c,14c,17c-eicosapentaenoic acids, respectively. Of these, PGE2 is the most actively produced, and it is involved in innumerable physiological processes. Dihomo-prostaglandins derived from adrenic acid (22:4(n-6) have also been detected in cell preparations, but no such compounds are produced from docosahexaenoic acid (DHA).

2.  Biosynthesis of Prostaglandins

Cyclooxygenases: Eicosanoids, including the prostanoids, are not stored within cells but are synthesised as required in response to hormonal stimuli. The prostaglandins PGE2 and PGF were first isolated and characterized from human seminal fluid in 1963 by Samuelsson, but prostaglandins and other eicosanoids are now known to be produced in a highly selective manner by most cell types, depending on the activation state and the physiological condition of the tissues in which they occur. The first step in their synthesis is the release of the substrate fatty acid, such as arachidonic acid, from the cellular phospholipids by the action of the enzyme phospholipase A2, and this is discussed in the Introductory document to this series. Next, the free acids are acted upon by one of two related enzymes, cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2), more correctly termed prostaglandin endoperoxide H synthases-1 and -2 (PGHS-1 and PGHS-2). These are key enzymes that catalyse the first committed step in the synthesis of prostanoids from fatty acid precursors; COX-1 is always present in tissues, while COX-2 is induced by appropriate physiological stimuli (cytokines, tumor promoters and growth factors). The two iso-enzymes have approximately 90% sequence identity, and they are very similar in structure. They differ in that COX-2 has a larger pocket at the active site because of an isoleucine to valine substitution. The result is that in comparison with COX-1 it can be more permissive in utilizing fatty acids, such as dihomo-γ-linolenic and eicosapentaenoic acids (and even linoleic and α-linolenic acids) in addition to arachidonic acid and other lipid substrates (see below).

In humans, COX-1 and COX-2 are homodimers of 576 and 581 amino acids, respectively, and each has three mannose-containing oligosaccharides linked to it, one of which facilitates protein folding. Each subunit of the dimer consists of three domains, the epidermal growth factor, the membrane binding domain, and the substantial catalytic domain, which contains two active sites on either side of a heme prosthetic group. A fourth oligosaccharide is found only in COX-2 and regulates its degradation. They are integral membrane proteins of the endoplasmic reticulum and nucleus and are located on one side only of the bilayer (COX-2 localizes to the Golgi in cancer cell lines).

Both enzymes catalyse the same two reactions. Thus, each carries out a cyclooxygenase reaction in which two molecules of oxygen are added to arachidonic acid to form a bicyclic endoperoxide with a further hydroperoxy group in position 15, i.e. to form prostaglandin PGG2. The first reaction occurs at a hydrophobic channel in the centre of the enzyme, before the hydroperoxide intermediate is transferred to the heme-containing site on the surface of the enzyme where it is reduced by a peroxidase to form prostaglandin PGH2. This is highly reactive and is the starting point for the biosynthesis of most other prostanoids.

Biosynthesis of prostaglandins via cyclooxygenases

Although the reactions occur at different sites, they are functionally coupled. The combined reactions are initiated by the oxidation of the heme group involved in the peroxidase reaction by traces of endogenous hydroperoxides with formation of a tyrosyl radical. This abstracts the 13‑pro‑S hydrogen from arachidonic acid and initiates the cyclooxygenase reaction, while during the reduction step the tyrosyl radical is regenerated so that activated COX can carry out multiple turnovers without a need to repeat the activation step. The other precursor polyunsaturated fatty acids interact with the enzymes in similar ways. As the catalytic tyrosyl radical can be transferred to an adjacent tyrosyl residue and become inactive after about 300 turnovers, the enzyme must be re-expressed constantly to generate metabolites.

Scottish thistleThe requirement for two distinct cyclooxygenases is not fully understood. In spite of the structural homology, separate genes encode COX-1 and COX-2 and they are regulated independently by different systems. The enzymes differ in their subcellular localization, substrate specificity and the manner in which they are coupled to upstream and downstream enzymes. In addition, the catalytic domains differ in structure, so that the susceptibilities to some inhibitors are not the same. It is now apparent that the two enzymes have different functional roles. It has been suggested that COX-1 is used for ‘housekeeping’ (homeostatic) purposes, responding rapidly to circulating hormones, which require constant monitoring and regulation. It is a constitutive enzyme that produces prostaglandins in the endoplasmic reticulum, which exit cells and signal through G-protein-linked receptors at the cell surface. However, there are also suggestions that it functions only at relatively high concentrations of arachidonic acid, for example during platelet aggregation, cell injury or acute inflammation. In those tissues where prostaglandins have specialized signalling functions, such as kidney, stomach, vascular endothelium, and especially blood platelets, COX-1 is expressed at higher concentrations, i.e. where the enzyme provides precursors for thromboxane synthesis.

In contrast, COX-2 is an inducible enzyme that is not normally present in tissues other than the kidney and brain (where COX-2 is constitutive in neurons and radial glia, but not other cell types). It is expressed under the control of the pro-inflammatory transcription factor NF‑κB in response to a wide range of extracellular and intracellular stimuli, such as cytokines, growth factors and tumor promoters, and produces prostanoids that are primarily pathophysiological or that function during defined stages of cellular development. It is able to utilize much lower concentrations of arachidonic acid and substrates other than the free acid. COX-2 is especially important in cells that are involved in inflammation, such as macrophages and monocytes, and it is believed to be the form of the enzyme that has the main responsibility for the synthesis of those prostanoids involved in severe inflammatory states, including cancer, rheumatoid arthritis, Alzheimer's disease and respiratory disorders. Some of its products may modulate the transcription of certain genes in the cell nucleus. COX-2 is activated by hydroperoxide concentrations that are approximately tenfold lower than those that activate COX-1, raising the possibility that under limiting concentrations of peroxide, COX-2 may be fully active while COX-1 is not. Induction of COX-2 expression is also regulated by sphingosine-1-phosphate, a further effect of sphingolipids on prostanoid biosynthesis.

Other prostaglandin synthases: PGH2 produced by the COX enzymes is an unstable intermediate from which all other prostanoids are derived by a variety of different enzymic reactions. Some of these are illustrated next for arachidonate as the primary precursor. The nature and proportions of the various enzymes and of the prostanoids produced differ according to cell type. Indeed different forms of some of the enzymes exist in cells that may be functionally similar, but differ in amino acid sequence, structure and co-factor requirements. Thus, PGH2 is converted to PGE2 by prostaglandin E synthases, of which at least three forms exist that are structurally and biologically distinct. The most important of these is a cytosolic enzyme, which is expressed constitutively in many different types of cell and is linked functionally to COX-1 to promote immediate PGE2 production. A second membrane-bound enzyme is induced by inflammatory stimuli and functions in concert with the inducible COX-2. PGD2 is formed in a similar way from PGH2 by the action of prostaglandin D synthases, which exist in two forms that are evolutionarily distinct but convergent in their functions; one is located in the central nervous system and the other in peripheral tissues. In rat peritoneal macrophages, PGD synthase and COX-1 appear to be functionally coupled.

Biosynthesis of prostaglandins - the synthases

Formula of levuglandin E2Levuglandins,  such as LGE2 (also termed ‘isoketals’), are formed from PGH2 by a non-enzymic rearrangement. They have a very short half-life and react more rapidly than most lipid oxidation products with the free primary amine groups of proteins and phosphatidylethanolamine (see below) to form covalent adducts.

The most common stereochemical form of prostaglandin F (PGF) is synthesised by two main routes. For example, it can be produced directly from PGH2 by the action of prostaglandin H-endoperoxide reductase, requiring NADPH. Interestingly, this enzyme can also utilize PGD2 as a substrate for the synthesis of the second of the four stereochemical forms of PGF, i.e. 9α,11β-PGF. As an alternative, PGF is synthesised via PGE2 by the action of an enzyme prostaglandin E 9-ketoreductase.

The cyclopentenone prostaglandins A and J, with reactive α,β-unsaturated keto groups and high biological activity, are produced by spontaneous dehydration reactions from PGE and PGD, respectively, and further modifications can then occur. For example, PGA2 isomerizes to form the highly unstable PGC2, which rapidly undergoes a secondary isomerization to produce PGB2. Similarly, in the presence of human serum albumin in vitro, it has been demonstrated that PGD2 is transformed into three dehydration products, i.e. 15-deoxy-PGD2, Δ12-PGJ2 and 15-deoxy-Δ12,14-PGJ2 (the last two via the intermediate PGJ2).

PGE2 is the major product of prostaglandin biosynthesis pathways following activation by pro-inflammatory substances and with its metabolite PGF, it is involved in a positive feedback loop to regulate COX-2 expression. Before they can function, prostanoids that have been newly synthesised must be transported from the cytosol and cross various membranes by means of active transporter systems.

Endocannabinoid metabolism: There is a significant difference in the substrate requirements of the two iso-enzymes. While both utilize unesterified arachidonic acid as substrate, COX-2 can also metabolize dihomo-γ-linolenic and eicosapentaenoic acids. COX-1 can only utilize free fatty acids, but COX-2 can react with the endocannabinoid 2-arachidonoylglycerol to form esterified 2-prostanoylglycerol derivatives, i.e. hydroxy endoperoxides analogous to PGH2, which can be further metabolized by downstream synthases. Similarly, COX-2 is involved in conversion of anandamide (arachidonoylethanolamine) and arachidonoylglycine to biologically active ‘prostamides’, though with lower efficiency. While these may simply serve as precursors of free prostanoids through hydrolysis, there is increasing evidence that they are new classes of lipid mediators with distinct biological properties of their own. The amide derivatives especially are relatively long-lived in plasma, and amides of PGF are available as drugs to lower ocular pressure and treat glaucoma. There is evidence for effects of 2-prostanoylglycerol on calcium mobilization through distinct and novel receptors as well as activation of the PPARδ receptor. It is subject to hydrolysis by esterases present in blood and some tissues.

Formula of asprin - salicylic acidThe Role of Aspirin: Both COX iso-enzymes and thence prostaglandin synthesis are inhibited by non-steroidal anti-inflammatory drugs ('NSAIDs'), such as aspirin (acetylsalicylic acid) and ibuprofen. Aspirin exerts this inhibition by binding to the cyclooxygenase site and transferring its acetyl group irreversibly to a specific serine residue, which then protrudes into the active site and obstructs the binding of arachidonate. Because of differences in the structures of the binding sites, COX-1 is completely inhibited by this means, but COX-2 is only partially inhibited. In contrast, ibuprofen and all other drugs of this type exert their effects by reversible binding and competition with arachidonic acid for the active sites. The specific inhibition by aspirin is the reason for its well-known analgesic, anti-pyretic and anti-inflammatory effects as a pharmaceutical. Via its effect on COX-1, it inhibits thromboxane synthesis and thence platelet aggregation, and it is now recommended in cardiovascular therapy (the role of COX-2 in atherosclerosis is more complicated).

However, this does not fully explain aspirin's repertoire of anti-inflammatory effects, and it is now known to be intimately involved, through an action with COX-2, in the generation of oxygenated lipid mediators such as the ‘aspirin-triggered’ protectins (resolvins) and the epi-lipoxins, which exert profound anti-inflammatory effects. This may explain some of the clinical benefits of aspirin, especially in neuro-inflammation.

Synthesis of COX-2 is inhibited by steroidal anti-inflammatory drugs at the level of transcription. In addition, as the active site of COX-2 is smaller than that of COX-1, it has proved possible to develop a number of drugs that specifically inhibit the action of COX-2. As well as having analgesic and anti-inflammatory effects, these are used clinically to prevent cancer of the colon. However, some COX-2 selective inhibitors have been associated with an increased risk of cardiovascular disease and have been withdrawn from the market.

Related enzyme activities: Certain pathogenic fungi and yeasts produce 3-hydroxy-eicosanoids from host arachidonic acid and they can hijack the host’s COX-2 enzymes to produce 3-hydroxy-prostaglandins from these that are as active biologically as the normal compounds. In addition, the yeast Candida albicans and other pathogenic fungi produce PGE2 in vitro from exogenous arachidonate by a novel biochemical mechanism, which does not involve the COX enzymes. A prostaglandin H synthase isolated from the red alga Gracilaria vertniculophylla is very different in structure from its animal counterparts, but it appears to function in a similar way, although it is not inhibited by non-steroidal anti-inflammatory drugs.

3. Prostacyclin and Thromboxane Biosynthesis

Prostacyclin (PGI2) and thromboxanes are also synthesised directly from PGH2 as illustrated below. Thus, a prostacyclin synthase converts PGH2 to PGI2 (half-life 42 seconds), while a thromboxane A synthase catalyses the production of TXA2 from PGH2. These enzymes are related to the cytochrome P450 group of proteins and are located on the cytosolic face of the endoplasmic reticulum, so the precursor PGH must cross the membrane. PGI and TXA are the main prostanoids formed in endothelial and smooth muscle cells and in platelets and lung, respectively. In addition, PGI2 and some other prostanoids can be produced by cell-cell interactions by using enzymes in adjacent cells, i.e. PGH2 of platelet origin is converted to PGI2 in the vascular epithelium. Subsequently, PGI2 can be released by endothelial cells to function through a signalling cascade with G-protein coupled receptors on nearby platelets. Similarly, prostacyclin production by erythrocytes is at least in part dependent on PGH2 from lymphocytes. COX-2 is the enzyme that provides PGH2 required for prostacyclin synthesis.

Prostacyclin-thromboxane biosynthesis

While platelets are able to synthesise thromboxane TXA2 from endothelial PGH2 in vitro, this is not believed to be a major pathway in vivo. In rat peritoneal macrophages, thromboxane A synthase and COX-1 appear to be functionally coupled in the endoplasmic reticulum.

Formula of dioxolane A3Dioxolane:  Thrombin-activated human platelets generate an eicosanoid in ng amounts that has been identified as 8-hydroxy-9,10-dioxolane A3 (DXA3), which both stimulates and primes the expression of human neutrophil integrin; it is believed to have a role in innate immunity and acute inflammation. COX1 is the key enzyme involved in its biosynthesis from unesterified arachidonic acid. After synthesis, it is rapidly esterified to position sn-2 of phosphatidylethanolamine in which position sn-1 is occupied by a 16:0, 18:0 or 18:1 vinyl ether or an 18:0 fatty acid; the intact phospholipid remains in the membrane and has similar biological activity to the free eicosanoid. Similar endoperoxides may be formed in tissues via the co-occurrence of LOX and cytochrome P450 or peroxygenase enzymes in tissues.

4.  Prostanoid Catabolism

Prostanoids function close to the site of synthesis, and they are deactivated before they are exported into the circulation as inactive metabolites. Some, such as PGI and TXA, are deactivated spontaneously. However, active enzyme systems also operate, and these function primarily by reaction with the 15(S)-hydroxyl group as discussed in the Introductory web page. For example, prostaglandin PGE2 is oxidized to 15-keto-PGE2, which was long thought to be biologically inactive, as a first step before more extensive oxidation. It is now recognized that inactivation of PGE2 by 15-hydroxyprostaglandin dehydrogenase is a vital step in halting tumor cell proliferation, and that the product 15-keto-PGE2 is an electrophilic molecule that functions in association with the PPARγ (see below) and other proteins to inhibit cell proliferation.

Catabolism of prostanoids and thromboxanes

The vinyl ether moiety in prostacyclin is unstable below pH 8.0, and PGI2 is rapidly deactivated non-enzymatically by a hydrolysis reaction to form 6‑keto-PGF. Similarly, TXA2 contains an unstable ether linkage and is deactivated by non-enzymatic hydrolysis to open the bicyclic oxygenated ring and form inert TXB2. A significant portion of the thromboxanes also undergoes dehydrogenation at C-11 by an 11-dehydroxythromboxane B2 dehydrogenase to form 11-dehydro-TXB2, a metabolite found in human blood plasma and urine, which can be monitored to assess responses to drug treatments.

5.  The Functions of Prostanoids

Prostanoids are ubiquitous lipids in animal tissues that coordinate a multitude of physiological and pathological processes at concentrations down to 10-9g per g of tissue, either within the cells in which they are formed or in closely adjacent cells (they are deactivated too readily to be transported far) in response to specific stimuli. Under normal physiological conditions, they have essential homeostatic functions in the cytoprotection of gastric mucosa, renal physiology, gestation, and parturition, but they are also implicated in a number of pathological conditions, such as inflammation, cardiovascular disease and cancer.

Receptors: Prostanoids and their receptorsProstanoids are sometimes described as local hormones that act in an autocrine fashion close to the site of their synthesis to coordinate the effects of other hormones in the circulation, although some can undergo facilitated transport from the cell via specific transporters to exert paracrine actions. In order to express their activity, they interact with specific G-protein-linked receptors (GPCRs) mainly. These comprise a large protein family with seven trans-membrane domains that sense molecules outside the cell and activate signal transduction pathways inside the cell and thence the cellular responses. When a ligand binds to a GPCR, it causes a conformational change, which allows it to act as a guanine nucleotide exchange factor to activate an associated G protein. Five classes (and several sub-classes) of GPCR have been identified in the mouse and man that interact with prostanoids, and these are specific for PGE2 (designated EP or four subclasses EP1–EP4), PGD2 (DP or two subclasses DP1-DP2), PGF (FP), PGI2 (IP) and TXA2 (TP). The immediate result is an increase or decrease in the rate of generation of cytosolic second messengers (cAMP or Ca2+), a change in membrane potential or activation of a specific protein kinase. The different receptors characterized from diverse cell types tend to have high, but not absolute, specificity for particular prostanoids with characteristic functions in each cell.

Certain of the cyclopentanone prostanoids (PGA and PGJ series) interact with peroxisome proliferator-activated receptors (PPARs) of which there are three, but in this context PPARγ is especially important. This is a nuclear hormone receptor or ligand-activated transcription factor regulating the expression of genes involved in adipogenesis, glucose homeostasis and lipid metabolism. All PPARs heterodimerize with the retinoid X receptor (RXR), which must itself be activated by binding to 9-cis-retinoic acid, and bind to specific regions on the DNA of target genes.

The picture of prostanoid actions is complicated by the fact that a given prostanoid can have a number of different biological functions, sometimes opposing, according to the cell type, the nature of the stimulatory response and the type of receptor. For example, PGE2 can have either pro- or anti-inflammatory effects depending on its interactions with one of four receptors in different cell types. The relative activities of the two iso-enzymes COX-1 and COX-2 are also essential to an understanding of the activity of prostanoids in any given circumstance. However, the complexity of the various interactions can only be hinted at here.

Inflammation and immune responses: Arguably the best known of the functions of prostaglandins and thromboxanes in cells is that they modify the inflammatory response, affecting symptoms, such as pain, fever and swelling. It should be recognized that inflammation is an intrinsically beneficial event that leads to removal of offending molecules and restoration of tissue structure and function. The main cause for concern is when acute inflammation fails to resolve and results in excessive tissue damage. In the early days of prostaglandin research, it was evident that prostaglandins injected into tissues could induce all the symptoms of inflammation. However, it is now recognized that the interactions are complex, and prostanoids can act both in a pro- and anti-inflammatory manner according to the nature of the inflammatory stimulus and the specific prostanoid produced, together with the profile of prostanoid receptors in a given type of cell. For example, EP3 receptors are involved in the development of fever, while EP2 and EP4 function in allergy and bone resorption.

cartoonUnder normal conditions, prostanoid levels in cells are low, but during inflammation both the nature and concentration of prostanoids can change dramatically. For example, macrophages produce both PGE2 and TXA2, but the ratio changes to an excess of PGE2 with an inflammatory stimulus. In these actions, prostanoids are best viewed as part of complex regulatory networks that modulate the actions of immune cells. PGE2 in particular has potent pro-inflammatory effects and is involved in all the processes leading to the classic signs of inflammation, including inducing fever and enhancing pain, On the other hand, it has anti-inflammatory properties also, such as suppressing lymphocyte proliferation and inhibiting the production of certain interleukins and other cytokines. It inhibits the action of 5-lipoxygenase, which is involved in the synthesis of pro-inflammatory leukotrienes, and stimulates the activity of the anti-inflammatory lipoxins. Therefore, PGE2 has a role in initiating the inflammatory response and in its eventual resolution. There is a particular interest in findings that in its pro-inflammatory role PGE2 promotes the growth of colorectal tumors (see below), and it is also involved in the pathology of rheumatoid arthritis and in respiratory diseases; with the latter, it can have both positive and negative effects depending upon circumstances.

Similarly, prostaglandin PGF has an important pro-inflammatory function, especially in patients with chronic inflammatory diseases such as rheumatoid arthritis. PGI2 an important mediator of the oedema and pain that accompany acute inflammation and it is produced rapidly following tissue injury or inflammation. For example, it is the most abundant prostanoid in synovial fluid in human arthritic knee joints.

The high levels of prostanoids found in inflammation are presumed to be due to the recruitment of leukocytes and the induction of the COX-2 enzyme (COX-1 appears to have a minor role only), which then in many tissues produces the pro-inflammatory prostanoids mainly. This explains the interest in COX-2 inhibitors for treating arthritis and other chronic inflammatory diseases. Inhibition of cyclooxygenases also explains the role of non-steroidal drugs, such as aspirin, in reducing the symptoms of fever. In the brain, COX-2 is present in neurons and has been implicated in the progression of Alzheimer's disease.

Immune responses are initiated and coordinated by T lymphocytes. Prostanoids are known to interact with T cells in a variety of ways, and appear to modify their development and maturation. Thus, PGE2 inhibits lymphocyte activation and proliferation, while TXA2 has opposing effects. Again, the actions of COX-2 (and COX-1) may be the key to triggering antigen-specific inflammation. However, this view may be too simplistic, and there is evidence that COX-2 is pro-inflammatory in the early stages of inflammation, but is beneficial at later stages by generating anti-inflammatory prostanoids. COX-1 derived prostanoids may sustain the inflammatory response.

Structural formula for 15-deoxy-PGJ2Although PGD2 has pro-inflammatory properties in allergic responses and in brain in the perception of pain, it is also recognized to be a key anti-inflammatory prostanoid that may be involved in the resolution of inflammation. Appreciable amounts are found only in the brain and in mast cells. Similarly, PGJ2, Δ12-PGJ2 and the short-lived 15-deoxy-Δ12,14-PGJ2, the J-series of prostaglandins produced by non-enzymatic dehydration of PGD2, are now well established as anti-inflammatory regulators, which function mainly if not solely via an interaction with PPARγ as discussed briefly above. They may also be involved in the immune response as they are produced in antigen-presenting cells such as activated T lymphocytes. 15-Deoxy-PGJ2, for example, functions in the resolution of the inflammatory response by inducing apoptotic cell death of activated macrophages. As it contains an electrophilic α,β-unsaturated ketone moiety in its cyclopentenone ring, it can act as an endogenous electrophile, which can undergo Michael addition with key cellular nucleophiles such as the free cysteine residues of proteins; covalent modifications of this type may be one mechanism by which it induces many of its biological responses. Its effects on cancer as an inhibitor of tumorigenesis are discussed below.

Polyunsaturated fatty acids of the omega-3 family are known to have anti-inflammatory properties. One explanation is that they inhibit the release of arachidonate from membrane phospholipids for eicosanoid production, or they may compete with arachidonate for the same enzymes of eicosanoid biosynthesis. Another reason may be that the 3-series prostanoids derived from eicosapentaenoic acid (EPA) have different biological activities from those of the 2-series. The protectins, resolvins and maresins must also be considered in this context. Similarly, prostaglandins derived from dihomo-γ-linolenic acid (20:3(n-6)), i.e. 1-series prostanoids, have properties distinct from those of the 2-series, and for example, PGE1 has been shown to suppress inflammation and promote vasodilation.

Opposing effects of thromboxane and prostacyclin.Cardiovascular effects: Two prostanoids are especially important and have essential but opposing functions in the maintenance of vascular homeostasis, i.e. thromboxane TXA2 and prostacyclin PGI2 (PGE2 and PGD2 are also relevant). TXA2 is synthesised mainly in platelets (which express only COX-1), production being enhanced during platelet activation, and it promotes platelet aggregation, vasoconstriction, and smooth muscle proliferation, even though it has a half-life of only 20-30 seconds. This is part of an essential repair mechanism for damaged vessel walls, but when the damage is too great blood clots can result with the potential to cause strokes or heart attacks. In contrast, PGI2 is the main product of macro-vascular endothelial cells. It is produced as required and exerts its effects a potent vasodilator locally through a specific IP receptor; it also inhibits platelet aggregation and smooth muscle cell proliferation. Thus, it contributes substantially to myocardial protection. Both TXA2 and PGI2 are therefore important mediators of pathological vascular events including thrombosis and atherogenesis, and it is evident that the correct balance between the two prostanoids is essential to good cardiovascular health. The ratio of TXA2:PGI2 seems to be more important than the absolute amounts of these mediators that are produced in vivo. In platelets and certain other cells, PGI2 is believed to function by elevating cAMP concentrations and activating adenyl cyclase, while TXA2 has the opposite effect. Further relevant factors are increased expression and activation of the TP receptor (for TXA2) in atherosclerotic lesions, which can directly accelerate atherogenesis and plaque growth.

The cardio-protective effect of aspirin, established by clinical trials, is exerted by the irreversible long-term inhibition of platelet COX-1 and thence of TXA2 biosynthesis for the lifetime of a platelet in the circulation (aspirin has little effect on PGI synthesis). Indeed, aspirin appears to be the only COX inhibitor with proven cardio-protective activity. In contrast, there is some concern that specific COX-2 inhibitors may have pro-thrombotic effects by inhibiting prostacyclin synthesis relative to that of thromboxanes. In clinical practice, such potential adverse effects of these drugs have to be balanced against positive effects in other tissues since only 1-2% of patients are believed to be at risk. Once more, polyunsaturated fatty acids of the omega-3 family are believed to have beneficial effects via the action of specialized pro-resolving mediators.

Lung: PGE2 can have anti-inflammatory and anti-asthmatic effects by activating the EP3 receptor. The role of PGD2 is more complex, but it may be pro-inflammatory.

Gastrointestinal system: COX-1 is always present throughout the human gastrointestinal tract, and produces PGI2 and PGE2, which have protective effects on the gastrointestinal mucosa. Both of these prostanoids reduce acid secretion from parietal cells, while increasing blood flow and stimulating the secretion of mucus. In this instance, the non-steroidal anti-inflammatory drugs, such as aspirin, have negative effects, while the COX-2 inhibitors can be beneficial. On the other hand, these findings are challenged by studies showing that COX-2 is expressed in the intestinal mucosa, and is induced in ulceration, for example, when large amounts of prostaglandins are produced that assist in healing.

Kidney function: Prostaglandins generated by both COX-1 and COX-2, especially PGE2, assist in the regulation of kidney function by maintaining vascular tone, blood flow, and salt and water excretion. PGE2 is required for the regulation of sodium re-absorption, while PGI2 (and possibly PGE2) increases potassium secretion. In addition, PGI2 with its well-known vasodilatory properties increases renal blood flow and the flow of fluids through the kidney. These actions are again mediated via specific receptors.

Reproductive system: Prostaglandins produced both by COX-1 and COX-2 are involved in many aspects of reproduction in females, from ovulation and fertilization through to labour. They are produced in the fetus and in the placenta as well as in other reproductive tissues. In particular, the synthesis of PGE2 and PGF is increased appreciably during labour, and these prostaglandins are in fact used as drugs to induce labour. PGF is used to induce ovulation in dairy cows and to induce abortions in women in midtrimester.

Scottish thistleCancer: COX-2 is over-expressed in many cancers, including those of the breast, colon and prostate. In particular, PGE2 produced by the enzyme occurs at much higher concentrations in tumor than in normal tissues. It promotes survival of tumor cells by inhibiting apoptosis and inducing proliferation, and by increasing cell motility and migration. In addition, via its effect on the immune system and inflammation, it has adverse effects in relation to the destruction of tumors. In consequence both the non-steroidal anti-inflammatory drugs, such as aspirin, and the COX-2 inhibitors have been found to have beneficial effects towards some types of cancer. Also, it is established that EP1 receptors are involved in chemically induced colon cancer. In contrast, both pro- and anti-tumorigenic activities have been demonstrated for PGD2 depending on the experimental model. Similarly, PGE3, derived from the n-3 eicosapentaenoic acid, has anti-proliferative activity in various cancers, possibly by interfering with PGE2 activity.

Thromboxane TXA4 is a pro-carcinogenic mediator that affects a number of tumor cell survival pathways, including cell proliferation, apoptosis and metastasis, and its activity is again balanced by that of prostacyclin.

15-Deoxy-Δ12,14-PGJ2 (15d-PGJ2), a potent anti-inflammatory regulator that functions via its interaction with PPARγ, also regulates adipogenesis and tumorigenesis and is produced by a variety of cells. An active transport system may carry it to the cells where it is required, and thence it is transported into the nucleus, where it affects gene transcription. Unlike PGE2, 15d-PGJ2 is a potent anti-tumor agent, inhibiting tumor growth both in vitro and in vivo in many tissues. It appears to act in a number of ways, for example directly by inhibiting proliferation and stimulating apoptosis. Also, it can interact indirectly to inhibit migration of tumor cells, and it can affect surrounding cells to reduce the expression of key receptors. However, some experimental conditions have been identified in which it exerts contrary effects. In general, PGE2 and 15d-PGJ2 have profound but opposing effects on tumorigenesis. It is evident that the prostaglandin synthases that are responsible for their biosynthesis are likely to be key targets for the development of anticancer drugs.

Protein metabolism: γ-Keto aldehydes such as the levuglandins (see above) and isolevuglandins, the latter produced in an analogous manner to the isoprostanes, have a remarkable reactivity towards proteins, forming adducts with greatly modified biological functions. Thus, these di-aldehydes react with lysyl residues on proteins to form first Schiff base adducts and thence pyrrole derivatives, which are able to form intra- and intermolecular protein-protein cross-links. Pyrrole adducts are in turn sensitive to oxygen and are further oxidized in vivo to stable lactam and hydroxylactam products. Protein adducts of this type are not at all easy to analyse, but those in brain have been correlated with the severity of Alzheimer’s disease, for example. Indeed, levuglandins and isolevuglandins are believed to be among the most potent neurotoxic products of lipid oxidation.

Levuglandins also react with phosphatidylethanolamine to form hydroxy-lactam derivatives, which may be better markers of oxidative injury from a practical standpoint, as they are more easily analysed.

Parasitic infections: It has been established that a number of parasitic organisms produce prostaglandins in the same way as their mammalian hosts, and by similar enzymic mechanisms. They may play a part in the pathogenesis of parasitic diseases.

6.   Some Exotic Prostanoids

Marine invertebrates, including sponges, corals, and molluscs, contain a wide range of prostaglandins, many of which are of the conventional type such as PGE2, PGF2 and so forth. They are presumed to perform similar functions as in mammals, and are also involved in the regulation of oogenesis and spermatogenesis, ion transport and defence. One species of coral (Plexaura homomalla) contains up to 8% of its dry mass as prostanoid esters, and for many years this was a primary source of material for experimental work. In many marine invertebrates, the prostaglandins exist largely in esterified form rather than the free state. It is perhaps more surprising that some red algae such as Gracilaria species contain prostaglandins (PGE2 and PGF) and are known to have a cyclooxygenase gene, but the function of these oxylipins in the organisms is not known.

Some exotic marine prostanoids

In addition, a number of novel prostanoids have been discovered, some examples of which are illustrated above, which differ in stereochemistry from the typical prostanoids, or contain acetyl groups, or are substituted with halogen atoms, such as chlorine or bromine. Little is known of the biochemistry or function of the "clavulones, bromovulones or punaglandins" in marine organisms, but there is increasing interest in them because of reported anti-tumor activities.

7.  Prostanoid Analysis

Analysis of prostanoids is not a simple task because they occur at such low levels in tissues and because of their high reactivity. Extraction must be carried out under mild conditions as rapidly as possible, and solid-phase extraction methods are now available that set the standard. Subsequent analysis usually involves HPLC linked to mass spectrometry. Immunoassays are available that may be suitable for some clinical applications, but they are not sensitive to minor differences in prostanoid structure.

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Lipid listings Credits/disclaimer Updated: May 4th, 2017 Author: William W. Christie LipidWeb icon