INTERACTION OF PLATELET-ACTIVATING-FACTOR WITH RAT HEPATOCYTES - UPTAKE, TRANSLOCATION, METABOLISM, AND EFFECTS ON PAF-ACETYLHYDROLASE SECRETION AND PROTEIN-TYROSINE PHOSPHORYLATION
Si. Svetlov et al., INTERACTION OF PLATELET-ACTIVATING-FACTOR WITH RAT HEPATOCYTES - UPTAKE, TRANSLOCATION, METABOLISM, AND EFFECTS ON PAF-ACETYLHYDROLASE SECRETION AND PROTEIN-TYROSINE PHOSPHORYLATION, Archives of biochemistry and biophysics, 327(1), 1996, pp. 113-122
In the present study the interaction of the phospholipid mediator plat
elet-activating factor (PAF) with rat hepatocytes in primary culture w
as examined, Following exposure to hepatocytes, exogenous [H-3]alkyl-P
AF was metabolized rapidly to [SH]lyso-PAF, the content of which was r
aised in the outer leaflet of the plasma membrane within the initial 5
min of incubation, Thereafter [H-3]lyso-PAF was translocated into cel
ls with concomitant reacylation to [H-3]alkyl-acyl-glycerophosphocholi
ne, A portion of untransformed [H-3]PAF accumulated in the outer leafl
et, and only a small amount of the [H-3]PAF was translocated into the
inner leaflet of the plasma membrane, Detectable levels of [H-3]lyso-P
AF were found in the medium of hepatocyte cultures at all times of inc
ubation, These findings suggest that at least a portion of the cellula
r PAF-acetylhydrolase (PAF-AH) activity is located in the outer leafle
t of the plasma membrane and can be secreted into the medium, Indeed,
rat hepatocytes in culture released PAF-AH into the medium in a time d
ependent fashion, Incubation of hepatocytes with exogenous PAF increas
ed secretion of PAF-AH, whereas lyso-PAF and the nonhydrolyzable analo
g methylcarbamyl-PAF significantly reduced secretion, The structurally
related PAF receptor antagonist CV 3988 markedly inhibited the activi
ty of PAF-AH and also diminished its release by hepatocytes, In contra
st, BN 50739 and WEB 2170, thienotriazolodiazepine PAF receptor antago
nists, did not affect the PAF-AH activity, but increased its secretion
by the cells, A full-length 3,8-kb mRNA encoding the cell surface PAF
receptor was absent in hepatocytes as indicated by Northern blot anal
ysis using the rat PAF receptor cDNA, whereas PAF receptor mRNA was re
adily detected in Kupffer cells, Upon incubation with hepatocytes, PAF
induced tyrosine phosphorylation of proteins with molecular masses of
120-130 and 160-180 kDa and dephosphorylation of 80- to 90-kDa protei
ns; these responses were not inhibited by WEB 2170 and BN 50739, The p
rotein tyrosine kinase inhibitor genistein abolished the release of fr
ee arachidonic acid, suggesting a crucial role for tyrosine phosphoryl
ation in PAF-induced phospholipase Az activation in rat hepatocytes. T
aken together, our data indicate that the interaction of PAF with rat
hepatocytes is dependent upon its metabolism, involves protein tyrosin
e phosphorylation/dephosphorylation and arachidonic acid release, and
does not involve the heteromeric G-protein-coupled PAF receptor which
has been characterized in Kupffer cells, This metabolically regulated
mechanism for PAF action on hepatocytes may be of potential biological
importance in the liver under normal and pathological conditions. (C)
1996 Academic Press, Inc.