Binding of carbamyl-platelet-activating factor to the raji lymphoblast platelet-activating factor receptor

Abstract

Carbamyl-platelet-activating factor (1-hexadecyl-2- N-methylcarbamyl-glycero-3-phosphocholine; CPAF) is an analog of platelet-activating factor (PAF) containing an N-methylcarbamyl moiety at the sn-2 position. CPAF was tested for effects on the Raji lymphoblast PAF receptor. Binding studies conducted at 4°C demonstrated specific binding that reached saturation within 60–80 min. Scatchard analysis of CPAF binding data revealed a single class of CPAF binding sites (14,800/cell) with a K = 2.9 ± 0.9 nM. Competition binding studies with PAF indicated that CPAF has about one-third the potency of native PAF. Unlike PAF, however, CPAF was not significantly metabolized by Raji lymphoblasts at 37°C. CPAF was shown to have PAF-agonistic qualities, since 100 pM to 1 μM CPAF increased free intracellular calcium in a dose-dependent manner. The structurally dissimilar PAF receptor antagonists CV-6209 and alprazolam inhibited the CPAF-induced calcium changes at doses that compared with CPAF binding. Treatment of Raji lymphoblasts with PAF or CPAF (10 pM - 1 μM) did not affect spontaneous proliferation, suggesting that the PAF receptor is not involved in the proliferation process in this cell line. These studies demonstrate that CPAF is a metabolically stable lymphoblast PAF receptor agonist that may provide a useful tool in the further elucidation of the role of PAF in lymphocyte function.