Effects of lipoxygenase on primary periodontitis

<目的b>研究表明，花生四烯酸的12-脂氧合酶（12-HETE）代谢物12-羟基二十碳四烯酸（12-HETE）在牙龈组织中产生。12-HETE，具有许多生物活性，如脱颗粒、趋化活性和骨吸收。在本研究中，我们研究了12-HETE对人白细胞的影响，特别是12-HETE诱导细胞内Ca^<2+>浓度升高的机制（[Ca2,]i）。方法与结果采用薄层自谱法测定大鼠各组织>12-脂氧合酶活性。采用改良Boyden室法检测12-HETE对人外周血多形核白细胞（PMNS）的趋化动力学活性。用荧光探针fura-2.12脂氧合酶检测大鼠牙龈细胞内游离钙的活性。12-HETE和n -甲酰基-蛋氨酸-亮氨酸-苯丙氨酸（FMLP）均以剂量依赖性的方式刺激hurffan PMNs的趋化动力学运动。然而，12-HETE以剂量依赖的方式抑制fMLP刺激的趋化动力学运动。hete诱导人PMNs中[Ca^<2+>]i呈剂量依赖性升高，抑制fMLP诱导的[Ca^<2+>]i升高。FMLP受体拮抗剂N-BOC-MLP（10^<-5>M）抑制FMLP对[Ca^<2+>]i的增加，但对12-HETE没有抑制作用。12- hete诱导[Ca^<2+>]升高，经百日咳毒素预处理消除。特异性磷脂酶C（PLC）抑制剂U-73122可抑制12-HETE诱导的[Ca^<2+>，]i的升高。12-HETE刺激肌醇1,4,5三磷酸的生成。这些结果表明，12-HETE对人pmn具有趋化动力学活性，并与另一种趋化因子相互作用。12-HETE增加[Ca^<2+>]i的机制可能涉及通过鸟嘌呤核苷酸结合蛋白激活PLC，导致磷脂酰肌醇周转增加。

<Purpose>It has been demonstrated that 12-hydroxyeicosatetraenoic acid(12-HETE), which is 12-lipoxygenase(12-HETE)metabolites of arachidonic acid, is produced in gingival tissues. 12-HETE, which has many biological activities, i. e. degranulation, chemotactic activity and bone resorption. In this study, we investisted the effects of 12-HETE on human leukocytes, especially, the mechanisms of 12-HETE induced increase of intracellular Ca^<2+> concentration([Ca2, ]i).<Methods & Results>12-Lipoxygenase activity of various tissues in rat was measured by thin layer autoradlogram. The chemokinetic activity of 12-HETE for human peripheral blood polymorphonuclear leukocytes(PMNS)was examined in modified Boyden chamber assay. Intracellular free calcium was monitored using the fluorescent probe fura-2.12-Lipoxygenase activity in rat gingiva was much higher than any other tissues. Both 12-HETE and N-formyl-methionyl-leucyl-phenylalanine(FMLP)stimulated chemokinetic movement of hurffan PMNs in a dose dependent manner. However, 12-HETE suppressed chemokinetic movement stimulated by fMLP in a dose dependent manner. 12HETE induced the increase of[Ca^<2+>i in human PMNs in a dose dependent manner and suppressed the[Ca^<2+>]i increase induced by fMLP. N-BOC-MLP(10^<-5>M), FMLP receptor antagonist, inhibited the increase of[Ca^<2+>]i by fMLP, but not by 12-HETE. 12-HETEinduced the increase of[Ca^<2+>], Was abolished by pretreatment with pertussis toxin. The increase in[Ca^<2+>, ]i induced by 12-HETE was inhibited by U-73122, a specific phospholipase C(PLC)inhibitor. 12-HETE stimulated the production of inositol 1, 4, 5trisphosphate.<Conclusions>These results suggest that 12-HETE has chemokinetic activity for human PMNs and have interaction with another chemotactic factor. The mechanisms for 12-HETE to increase[Ca^<2+>]i may involve PLC activation through the guanin-nucleotide-binding protein, resulting in an increase in the phosphatidylinositol turnover.