Platelet-activating factor biosynthesis induced by various stimuli in human HaCaT keratinocytes.
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.
Travers
J B
JB
Department of Dermatology, University of Colorado Health Sciences Center, Denver, USA.
Harrison
K A
KA
Johnson
C A
CA
Clay
K L
KL
Morelli
J G
JG
D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D019332:Endothelin-1 / Q000494:pharmacology; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D015603:Keratinocytes / Q000378:metabolism*; D010972:Platelet Activating Factor / Q000031:analogs & derivatives / Q000096:biosynthesis* / Q000494:pharmacology; D013268:Stimulation, Chemical
D019332:Endothelin-1; D010972:Platelet Activating Factor; D000001:Calcimycin; C083283:1-O-hexadecyl-2-N-methylcarbamol -sn-glycerol-3-phosphocholine; D002118:Calcium