Platelet-Activating Factor Biosynthesis Induced by Various Stimuli in Human HaCaT Keratinocytes

Abstract

Platelet-activating factor (PAF) is a potent inflammatory mediator that is thought to play a role in cutaneous inflammation. These studies used mass spectrometry to examine the molecular species of PAF precursor glycerophosphocholine lipids (GPC) as well as the biosynthesis of PAF and other sn-2 acetyl-GPC in a human keratinocyte-derived cell line (HaCaT keratinocytes). Approximately 28% of HaCaT keratinocyte GPC consisted of 1-alkyl species, and the relative amounts of the sn-1 alkyl constituents of the PAF precursor 1-alkyl-2-acyl-GPC were as follows: hexadecyl > octadecenyl > octadecyl. Ionophore (A23187)-stimulated HaCaT keratinocytes synthesized both PAF (1-hexadecyl, 1-octadecenyl, and 1-octadecyl species) and less potent 1-acyl analogs (1-palmitoyl, 1-oleoyl, and 1-stearoyl species). PAF production was rapid and maximal by 10 min. The major species of sn-2acetyl-GPC at 2.5 min were 1-hexadecyl-2-acetyl-GPC (2.2 ng/10(6) cells) and 1-palmitoyl-2-acetyl-GPC (2.4 ng/10(6) cells). HaCaT keratinocytes also synthesized PAF and 1-acyl PAF analogs when stimulated with the peptide growth factor endothelin-1 and the nonhydrolyzable PAF receptor agonist carbamyl-PAF. Both 1-hexadecyl-2- acetyl-GPC and 1-palmitoyl-2-acetyl-GPC stimulated intracellular calcium mobilization in HaCaT cells, indicating that these sn-2 acetyl-GPC act in autocrine fashion. These studies revealed that the human keratinocyte-derived cell line HaCaT can synthesize significant amounts of PAF and 1-acyl analogs in vitro from both nonspecific (A23187) and specific (endothelin-1, carbamyl-PAF) stimulation, suggesting a role for this inflammatory lipid mediator in keratinocyte pathophysiology.

Oxidized glycerophosphocholines as biologically active mediators for ultraviolet radiation-mediated effects

“…1-alkyl GPC with a polyunsaturated fatty (such as arachidonate) esterified to the sn-2 position is found in reasonable abundance in epithelial cells 9 . These molecules are precursors of bioactive lipid platelet-activating factor (1-alkyl-2-acetyl GPC), which is formed upon the actions of enzymes phospholipase A 2 and PAF acetyltransferase.…”

PAF, a Putative Mediator of Oral Inflammation

“…Moreover, both acyl-linked and alkyl-linked acetylated phospholipids have been identified in stimulated human keratinocytes in culture (Travers et al, 1996(Travers et al, , 1997 (1) PAF has direct and potent effects upon cardiac function and has been implicated in cardiovascular (patho)physiology (Evangelou, 1994;Prescott et al, 1996Prescott et al, ,1997Feuerstein et al, 1997;Yamada et al, 1998); (2)PAF-like phospholipids (oxidized-PC) and PAF-AH may be involved in the pathogenesis of atherosclerosis (see above); (3)salivary PAF levels are significantly increased in subjects with periodontal disease (Fig. 5); (4)salivary PAF levels are transiently increased in subjects presenting with myocardial infarction (Jones et al, 1994); and (5)periodontitis appears to be a risk factor for cardiovascular disease (including accelerated or severe atherosclerosis, myocardial infarction, and stroke) (Beck et al, 1996(Beck et al, , 1998.…”

Staphylococcal lipoteichoic acid inhibits delayed-type hypersensitivity reactions via the platelet-activating factor receptor

“…Through its ability to both attract and activate leukocytes, this mediator has potent proinflammatory effects. PAF is produced in a wide variety of cell types; in keratinocytes its synthesis is stimulated by physical agents including ionophores, ultraviolet radiation, and heat/cold trauma (14)(15)(16)(17). PAF exerts its effects via the PAF receptor, a G protein-coupled receptor found on leukocytes as well as keratinocytes.…”