Tomoko Takano

Platelet-Activating Factor Receptor

Platelet-activating factor (PAF) is a pro-inflammatory lipid mediator possessing a unique 1-O-alkyl glycerophospholipid (GPC) backbone (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholin). Cloned PAF receptor, which belongs to the G protein-coupled receptor superfamily, transduces pleiotropic functions including cell motility, smooth muscle contraction, and synthesis and release of mediators and cytokines via multiple heterotrimeric G proteins. Pharmacological studies have suggested that PAF functions in a variety of settings including allergy, inflammation, neural functions, reproduction, and atherosclerosis. Establishment of PAFR(-/-) mice confirmed that the PAF receptor is responsible for pro-inflammatory responses, but that its roles in other settings remain to be clarified.

DEMONSTRATION OF PLATELET ACTIVATING FACTOR RECEPTOR IN GUINEA PIG KIDNEY

Distribution of platelet activating factor (PAF) receptor was examined in the guinea pig kidney. Northern blot analysis showed a single band electrophoresed just below the 28S rRNA, and the mRNA was richest in the cortex with lesser amounts in the outer and then inner medulla. Scatchard analysis of membrane fraction using [3H]WEB 2086, a specific PAF receptor antagonist, revealed a single binding site with Bmax of 522, 228, 58 fmol/mg protein for the cortex, outer medulla and inner medulla, respectively. Kd values were in the same order of magnitude (10(-8) M). These results indicate the presence of a single class of PAF receptor in the guinea pig kidney which is most abundant in the cortex.

Expression of platelet activating factor receptor in renal tubular cell line (LLC-PK1)

Northern blot analyses of poly(A)+ RNA extracted from cloned renal tubule cells, LLC-PK1, MDCK and JTC-12 cells, were performed using an isolated cDNA of guinea pig platelet activating factor (PAF) receptor as a probe. The results showed the expression of single entity of PAF receptor mRNA in LLC-PK1, but not in MDCK or JTC-12 cells. PAF also induced a dose-dependent elevation of intracellular calcium in LLC-PK1 cells as assessed by Fura-2 fluorescence signals, which was completely blocked by a PAF receptor antagonist, WEB 2170. These results indicate a PAF receptor-effector system in a renal tubular cell line LLC-PK1, providing a possible model system for studying physiological roles of PAF receptors in renal tubules.