Role of platelet-activating factor in functional alterations induced by xenoreactive antibodies in porcine endothelial cells

Background, Platelet-activating factor (PAF) is a phospholipid mediator of inflammation which has been implicated in rejection. The interaction of ant i-alpha -galactosyl natural antibodies (anti-alpha gal Abs) with endothelia l cells is the initial step for the development of xenograft rejection, In our study, we stimulated porcine aorthic endothelial cells (PAEC) with anti -alpha gal IgG to investigate the synthesis of PAF from PAEC and its biolog ical consequences, Methods and Results. PAF was extracted and chromatografically purified from cultured PAEC stimulated with baboon anti-alpha gal Abs, The Abs induced a dose-dependent synthesis of PAF peaking after 30 min of incubation, and de creasing thereafter. Concomitant cell shape change, motility, and cytoskele ton redistribution were observed. These events were prevented by addition o f a panel of PAF-receptor antagonists. An SV40 T-large antigen-immortalized PAEC line was engineered to express PAF acetyl-hydrolase (PAF-AH) cDNA, th e major PAF-inactivating enzyme, These transfected cells exposed to anti-al pha gal Abs showed reduced cell contraction and motility compared with empt y vector-transfected cells, Moreover, in PAEC stimulated with anti-alpha ga l Abs, the synthesis of]PAF promoted the adhesion of a monocytic cell line as shown by the inhibitory effect of PAF-receptor antagonists and of PAF-AN expression. Finally, studies on cell monolayer demonstrated an enhanced pe rmeability 48 hr after exposure to anti-alpha gal Abs, and this increase wa s prevented by PAF-inactivation and by PAF-receptor blockade. Conclusions. These results demonstrate that on stimulation with anti-alpha gal Abs, PAEC synthetize PAF which can contribute to several vascular event s involved in xenograft rejection.