Gandhi C R, Hanahan D J, Olson M S
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284.
J Biol Chem. 1990 Oct 25;265(30):18234-41.
Stimulation of rat Kupffer cells in primary culture with platelet-activating factor (PAF) caused a rapid hydrolysis of phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 4-phosphate with a concomitant increase in the levels of myo-inositol 1,4,5-trisphosphate and myo-inositol 1,4-bisphosphate. This phospholipase C-mediated hydrolysis of polyphosphoinositides was independent of extracellular Ca2+ but was inhibited by the intracellular Ca2+ antagonist TMB-8. A second slower response to PAF was characterized by deacylation of PI leading to the accumulation of glycerophosphoinositol (GPI). PAF-induced GPI synthesis was not inhibited by TMB-8. These effects of PAF were accompanied by initial transient mobilization of Ca2+ from intracellular stores followed by a rather slow influx of Ca2+ from the extracellular medium. PAF-stimulated deacylation and phosphodiesteric hydrolysis of inositol lipids were differentially affected by cholera toxin and pertussis toxin. Pretreatment of the Kupffer cells with either of these toxins caused inhibition of phospholipase C activity. Pertussis toxin also inhibited PAF-stimulated deacylation. However, cholera toxin itself stimulated GPI release and addition of PAF to the cholera toxin-treated cells caused a further increase in GPI release. Phorbol ester inhibited PAF-induced phosphodiesteric hydrolysis of phosphoinositides, but not deacylation. PAF-induced metabolism of phosphoinositides was inhibited by the PAF antagonist, U66985. These results suggest that PAF-induced phosphodiesteric hydrolysis and deacylation of inositol phospholipids are regulated via distinct mechanisms involving activation of separate G-proteins in rat Kupffer cells. Also the regulation of phosphoinositide metabolism by Ca2+ mobilization from two separate Ca2+ pools is indicated by this study.
用血小板活化因子（PAF）刺激原代培养的大鼠枯否细胞，导致磷脂酰肌醇4,5-二磷酸和磷脂酰肌醇4-磷酸迅速水解，同时1,4,5-三磷酸肌醇和1,4-二磷酸肌醇水平升高。这种由磷脂酶C介导的多磷酸肌醇水解不依赖细胞外Ca2+，但被细胞内Ca2+拮抗剂TMB-8抑制。对PAF的第二种较慢反应的特征是PI去酰化，导致甘油磷酸肌醇（GPI）积累。PAF诱导的GPI合成不受TMB-8抑制。PAF的这些作用伴随着最初细胞内钙库中Ca2+的短暂动员，随后是细胞外介质中Ca2+相当缓慢的内流。PAF刺激的肌醇脂质去酰化和磷酸二酯水解受到霍乱毒素和百日咳毒素的不同影响。用这两种毒素中的任何一种预处理枯否细胞都会导致磷脂酶C活性受到抑制。百日咳毒素也抑制PAF刺激的去酰化。然而，霍乱毒素本身刺激GPI释放，将PAF添加到经霍乱毒素处理的细胞中会导致GPI释放进一步增加。佛波酯抑制PAF诱导的磷酸肌醇磷酸二酯水解，但不抑制去酰化。PAF诱导的磷酸肌醇代谢受到PAF拮抗剂U66985的抑制。这些结果表明，PAF诱导的肌醇磷脂磷酸二酯水解和去酰化是通过不同机制调节的，这些机制涉及大鼠枯否细胞中不同G蛋白的激活。此外，本研究还表明了从两个独立的Ca2+库动员Ca2+对磷酸肌醇代谢的调节作用。
