磷脂酶D在脂多糖及抗原引爆大鼠外周血白细胞中的作用

Role of phospholipase D in priming of rat peripheral leukocytes by lipopolysaccharide and antigen

目的 :探讨脂多糖 (L PS)与卵白蛋白 (OA)是否引爆大鼠外周血白细胞 ,致敏过程对大鼠外周血白细胞引爆的影响以及磷脂酶 D在大鼠外周血白细胞引爆中的作用。方法 :采用正常及主动致敏的大鼠外周血白细胞为实验材料 ,以 L PS或抗原 (OA)为引爆剂 ,f MLP为激动剂处理细胞 ,测定白细胞弹性蛋白酶释放和髓过氧化物酶(MPO)活性作为细胞激活指标 ,同时 ,用酶偶联比色法测定白细胞 PL D活性。结果 :LPS和 f ML P共同处理正常大鼠外周血白细胞 ,引起弹性蛋白酶和 MPO释放高于 f ML P单独处理 (P<0 .0 5 ) ;OA和 f ML P共同处理正常大鼠外周血白细胞 ,引起上述两种酶的释放与 f MLP单独处理无明显差别。 LPS和 f MLP共同作用于致敏大鼠外周血白细胞 ,引起弹性蛋白酶和 MPO释放与 f MLP单独作用无明显差别 ;OA和 f ML P共同作用于致敏大鼠外周血白细胞 ,引起上述两种酶的释放高于 f MLP单独作用 (P<0 .0 5 )。正常大鼠中 ,PLD活性与弹性蛋白酶释放和 MPO活性的相关系数 ,分别为 0 .5 1和 0 .73(P<0 .0 1)。致敏大鼠中 ,PLD活性与弹性蛋白酶释放和 MPO活性的相关系数 ,分别为 0 .4 8(P<0 .0 1)和 0 .37(P<0 .0 5 )。结论 :1LPS能引爆正常大鼠外周血白细胞。 2特异性抗原 OA能引爆致敏大鼠外周血白细胞。

Objective: To investigate whether or not lipopolysaccharide(LPS) and ovalbumin(OA) prime rat peripheral leukocytes, the effect of sensitization on priming and the role of phospholipase D in priming. Methods: The peripheral leukocytes were separated and purified from sensitized or unsensitized rats. LPS or OA was used as a priming agent and formylmethionylphenylalanine (fMLP) as an activating agent. Degradation of leukocyte was determined by measurement of elastase release and myeloperoxidase (MPO) activity. Phospholipase D (PLD) activity was assayed by the generation of choline,which was measured by choline-oxidase-catalyzed formation of H 2O 2 and Trinder reaction. Results: Compared with cells treated by fMLP alone,leukocytes from unsensitized rat challenged with fMLP after incubated with LPS released more elastase and MPO(P<0.05). But there was no significant difference between leukocytes challenged with fMLP after incubated with OA and fMLP treated alone. In sensitized rat,there was no difference between leukocytes challenged with fMLP after incubated with LPS and fMLP treated alone. But leukocytes challenged with fMLP after incubated with OA released significantly more elastase and MPO than fMLP treated alone(P<0.05). A significant correlation was obtained between the release of elastase and PLD activity(r s=0.51,P<0.01),and also between the release of MPO and PLD activity(r s=0.73,P<0.01) in unsensitized rat. In sensitized rat,it was 0.48(P<0.01) and 0.37(P<0.05), respectively. Conclusion: ①LPS primes peripheral leukocytes from unsensitized rats; ②OA primes peripheral leukocytes from actively sensitized rats; ③PLD plays a role in priming of rat peripheral leukocytes.