Platelet Activating Factor

The term platelet-activating factor was introduced to define the activity of a then unknown metabolite, which induced the aggregation of blood platelets released from basophils stimulated with immunoglobulin E. In 1979 independently in the laboratories of D.J. Hanahan, J. Benveniste and F. Snyder, a phospholipid identified as 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine, an ether analogue of phosphatidylcholine, was shown to be responsible for this activity and for the activity of a compound that had been termed ‘antihypertensive polar renal lipid’. In the light of what is now known of the manifold biological activities of this lipid, platelet-activating factor or PAF is not an especially appropriate name, but it has stuck. It can be considered a special case of the more abundant ether lipids.

Formula of platelet-activating factorIt is an unusual lipid in many ways, as lipids with alkyl groups only in position sn-1 are not common in animals, as is acetic acid esterified directly to glycerol. In general, the alkyl groups tend to be mainly saturated and C16 or C18 in chain length, although vinyl ether (plasmalogen) forms have also been detected. Short-chain fatty acids other than acetate (e.g. propionyl, butyryl) in position sn-2 are only occasionally found.

PAF was the first intact phospholipid known to have messenger functions, i.e. in which the signalling results from the molecule binding to specific receptors rather than from physicochemical effects on the plasma membrane or other membranes of the cell. There is a strict structural requirement for binding to its unique single G-protein-coupled receptor, which is expressed by numerous cells including all those of the innate immune system, and for recognition as a substrate by enzymes. Thus, there is a many-hundred-fold specificity for the ether bond in position sn-1 of PAF in comparison to the 1-acyl analogue, together with considerable specificity for a short acyl chain in position sn-2 and for the phosphocholine head group.

While both PAF and lyso-PAF were reported to be present in plants in one study, e.g. nettles and strawberries, others have not been able to detect ether lipids.

 

Biosynthesis

PAF is synthesised by a variety of cells, but especially those involved in host defence, such as platelets, endothelial cells, neutrophils, monocytes, and macrophages. In the main pathway, a distinct membrane-bound acetyltransferase (LPCAT2) catalyses the transfer of an acetyl residue from acetyl-CoA to 1-alkyl-sn-glycero-3-phosphocholine (lyso-PAF), generated by the action of phospholipase A2 on phosphatidylcholine.

Biosynthesis of platelet-activating factor

PAF is synthesised continuously by cells but at low levels, controlled by the activity of PAF acetyl hydrolases (see below). However, it is produced in much greater quantities by inflammatory cells when required in response to cell-specific stimuli. Studies with the purified acetyltransferase have shown that with cells in the resting state, the enzyme can utilize arachidonoyl-CoA to produce the membrane-bound PAF precursor 1-alkyl-2-arachidonoylglycerophosphocholine with even greater facility than the generation of PAF per se. Only when the cells are subjected to acute inflammatory stimulation does the activated enzyme produce PAF in appreciable amounts, probably after phosphorylation by a protein kinase, while simultaneously arachidonate is released for eicosanoid production.

A second lyso-PAF acetyltransferase (LPCAT1) is expressed in the lungs mainly, where it produces PAF and dipalmitoyl-phosphatidylcholine essential for respiration under noninflammatory conditions. This is a constitutively expressed enzyme, while LPCAT2 is inducible.

Alternatively, PAF can be produced by acetylation of 1-alkyl-sn-glycero-3-phosphate, which is subsequently converted to 1-alkyl-2-acetylglycerol and thence to PAF, i.e. by a mechanism analogous to the biosynthesis of phosphatidylcholine. This ‘de novo’ pathway is also believed to be noninflammatory.

PAF-like molecules with some biological activity can also be produced in tissues by nonenzymatic oxidation of polyunsaturated fatty acids in phosphatidylcholine, resulting in cleavage at the first double bond leaving a short-chain acid with a terminal aldehyde group in position 2 (a ‘core aldehyde’). Such compounds are formed in lipoproteins and are present in human atherosclerotic lesions. They bring about platelet aggregation at nanomolar concentrations and may be involved in thrombosis and acute coronary events. Such oxidatively truncated phospholipids (and PAF) are also pro-apoptotic by a mechanism that is independent of the PAF receptor, and they have a substantial influence on regulated cell death.

 

Formula of a core aldehyde

 

Biochemical Function

Initially, PAF was found to effect aggregation of platelets at concentrations as low as 10-11 M, and it induced a hypertensive response at very low levels also. More generally, it is now recognised that its primary role is to mediate intercellular interactions. For example, by binding to its specific receptor, PAF activates the cytoplasmic phospholipase A2 and phospholipase C. The result of the latter is an increase in intracellular Ca2+ downstream of the cell and activation of protein kinase C. It is now known to exert effects on many different types of noninflammatory biological events and functions, including glycogen degradation, reproduction, brain function and blood circulation.

Much recent work has been concerned with the function of PAF as a mediator of inflammation, and in the mechanism of the immune response. While it is presumed to have evolved as part of a protective mechanism in the innate host defence system, there is particular interest in its involvement in uncontrolled pathological conditions. For example, it has a number of pro-inflammatory properties, and in excess it has been implicated in the pathogenesis of a number of disease states, ranging from allergic reactions to stroke, sepsis, myocardial infarction, colitis and multiple sclerosis. PAF can activate human inflammatory cells at concentrations as low as 10-14 M. In relation to asthma, PAF is able to act directly as a chemotactic factor and indirectly by stimulating the release of other inflammatory agents. Administration of PAF can produce many of the symptoms observed in asthma, probably via the formation of leukotrienes as secondary mediators. However, the amount of PAF produced by cellular stimuli of various kinds is dependent on the nature of the cell and specific agonists. In essence it is a hormone that acts locally, as it is found only on the surface of activated cells so restricting the inflammatory response.

Recently, PAF has been shown to be an anti-obesity factor, functioning through stimulation of its receptor in brown but not white adipose tissue. Reduction in this activity may be responsible for increasing adiposity with age.

The nature of the alkyl group in position sn-1 may be important to such processes. For example, it has been established that C16-PAF and C18-PAF cause death to cerebellar granule neurons, but that they signal through different pathways. In addition, PAF receptor signalling can be either pro- or antiapoptotic, depending upon the nature of the sn-1 alkyl moiety, probably because of differential binding of each isomer to the receptor. A synthetic analogue of PAF, 1-O-octadecyl-2-O-methylglycero-3-phosphocholine (‘edelfosine’) is a potent anticancer agent in animal models, but appears too toxic for use with humans.

Alkylacetylglycerols, analogues of 1,2-diacyl-sn-glycerols, have biological activity also, some of which is independent of subsequent conversion to PAF. Phosphatidylethanolamine analogues of PAF have also been studied, but are much less potent biologically. A further comparable signalling molecule, N-acetylsphingosine, is produced by a CoA-independent transacetylase, which transfers the acetyl group of PAF to sphingosine (see also our web page on ceramides).

Catabolism: Control of PAF concentration and activity is regulated partly by tight control of its synthesis, and partly by the action of specific acetylhydrolases, two small subfamilies of which exist (both Ca2+-independent) and are classified as part of the large phospholipase A2 family of enzymes. These are not active against conventional phospholipids, and their main function is to remove the acetyl group from PAF thus eliminating its biological activity. One isoform of the subfamily of group VIII PAF-acetylhydrolases is enriched in brain and erythrocytes, it is cytosolic and it is completely specific for PAF. A second isoform is present mainly in liver and kidney, where it is located in both the cytosol and membranes, and has a broader specificity in that it will also hydrolyse truncated acyl moieties from oxidized phospholipids.

Plasma PAF-acetylhydrolase is associated with both circulating LDL and HDL particles and functions on the lipid-aqueous interface, where it is sometimes termed the ‘lipoprotein-associated phospholipase A2’ (group VII family). This is secreted by macrophages and is a 45 kDa protein, which circulates in plasma in its active form. These enzymes also hydrolyse unmodified fatty acyl residues up to 5 or 6 carbon atoms long in the sn-2 position, although even this restriction is relaxed when the terminal end of the fatty acyl moiety is oxidized (i.e. aldehydic or carboxylic), such as the oxidatively truncated phospholipids described above. Indeed, the enzyme will hydrolyse phospholipids containing hydroperoxyoctadecadienoyl and F2-isoprostane residues. The effect is to remove any oxidized phospholipids from lipoproteins and from atherosclerotic plaques that might otherwise contribute to their inflammatory properties. Thus, while oxysterols accumulate as atherosclerotic lesions mature, formation and destruction of oxidized phosphatidylcholines is a continuous process in both early and advanced lesions. Similarly, by removing intracellular truncated phospholipids, PAF acetylhydrolase protects cells from apoptosis. Expression of these enzymes is up-regulated at the transcriptional level by mediators of inflammation in response to inflammatory stimuli, but they are susceptible to oxidative inactivation.

The other products of PAF hydrolases are lysophospholipids, but at concentrations orders of magnitude lower than normal circulating levels, so they are not expected to make a significant inflammatory contribution. The ether linkage in the lysophospholipid can be cleaved oxidatively by the microsomal alkylglycerol monooxygenase to yield a fatty aldehyde, which is then further oxidized to the corresponding acid as described in our web page on ether lipids.

PAF-acetylhydrolase has transacetylase activity also, and is able to transfer short-chain fatty acids from PAF to ether/ester-linked lysophospholipids.

 

Suggested Reading

  • Christie, W.W. and Han, X. Lipid Analysis - Isolation, Separation, Identification and Lipidomic Analysis (4th edition), 446 pages (Oily Press, Woodhead Publishing and now Elsevier) (2010).
  • Farooqui, A.A. and Horrocks, L.A. Plasmalogens, platelet-activating factor, and other ether glycerolipids. In: Bioactive Lipids. pp. 107-134 (edited by A. Nicolaou and G. Kokotos, The Oily Press, Bridgwater) (2004).
  • McIntyre, T.M., Prescott, S.M. and Stafforini, D.M. The emerging roles of PAF acetylhydrolase. J. Lipid Res., 50, S255-S259 (2009) (DOI: 10.1194/jlr.R800024-JLR200).
  • McIntyre, T.M., Snyder, F. and Marathe, G.K. Ether-linked lipids and their bioactive species. In: Biochemistry of Lipids, Lipoproteins and Membranes (5th Edition), pp. 245-276 (edited by D.E. Vance and J.E. Vance, Elsevier Science) (2008).
  • Prescott, S.M. Zimmerman, G.A., Stafforini, D.M. and McIntyre, T.M. Platelet activating factor and related lipid mediators. Annu. Rev. Biochem., 69, 419-445 (2000) (DOI: 10.1146/annurev.biochem.69.1.419).
  • Snyder, F. Platelet-activating factor and its analogs: metabolic pathways and related intracellular processes. Biochim. Biophys. Acta, 1254, 231-249 (1995) (DOI: 10.1016/0005-2760(94)00192-2).
  • Yost, C.C., Weyrich, A.S. and Zimmerman, G.A. The platelet activating factor (PAF) signaling cascade in systemic inflammatory responses. Biochimie, 92, 692-697 (2010) (DOI: 10.1016/j.biochi.2010.02.011).

 

Updated July 1, 2014

pdf