星形孢菌素诱导大鼠腹腔中性粒细胞产生中性粒细胞趋化因子

Induction of neutrophil chemotactic factor production by staurosporine in rat peritoneal neutrophils.

作者信息

Edamatsu T, Xiao Y Q, Tanabe J, Mue S, Ohuchi K

机构信息

Department of Pathophysiological Biochemistry, Faculty of Pharmaceutical Sciences, Tohoku University, Miyagi, Japan.

出版信息

Br J Pharmacol. 1997 Aug;121(8):1651-8. doi: 10.1038/sj.bjp.0701322.

DOI:10.1038/sj.bjp.0701322

PMID:9283699

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC1564877/

Abstract

Incubation of rat peritoneal neutrophils in medium containing various concentrations of staurosporine (6.4-64 nM) increased the neutrophil chemotactic activity in the conditioned medium in a time- and concentration-dependent manner. 2. Separation of the neutrophil chemotactic activity in the conditioned medium by isoelectric focusing revealed that staurosporine (64 nM) stimulated the production of basic (pH > 8) neutrophil chemotactic factors, while TPA (12-O-tetradecanoylphorbol 13-acetate, 49 nM) stimulated the production of both basic (pH > 8) and acidic (pH 5) neutrophil chemotactic factors. 3. Determination by immunoassay of cytokine-induced neutrophil chemoattractant (CINC)-1, -2 alpha, -2 beta and -3 in the conditioned medium at 4 h revealed that staurosporine (64 nM) and TPA (49 nM) strongly stimulated the production of CINC-3 (staurosporine, 133.0 +/- 3.8; TPA, 26.7 +/- 1.0; control, 0.32 +/- 0.01 ng ml-1, means +/- s.e.mean from four samples) compared to CINC-1 (staurosporine, 55.0 +/- 1.2; TPA, 12.2 +/- 0.3; control, 0.56 +/- 0.01 ng ml-1), and CINC-2 alpha (staurosporine, 1.09 +/- 0.03; TPA, 0.90 +/- 0.02; control, < 0.10 ng ml-1). CINC-2 beta was below the detectable amount (< 0.078 ng ml-1). 4. The level of CINC-3 mRNA in the peritoneal neutrophils was determined by reverse transcription-polymerase chain reaction. Staurosporine (64 nM) and TPA (49 nM) enhanced the level of CINC-3 mRNA time-dependently, but had no effect on GAPDH mRNA levels. 5. Production of staurosporine-induced neutrophil chemotactic factor was inhibited by the protein kinase C inhibitors, H-7 (IC50, 12.3 microM), calphostin C (IC50, 0.77 microM) and Ro 31-8425 (24.3% inhibition at 10 microM), and by the tyrosine kinase inhibitor, genistein (IC50, 68.5 microM). Production of TPA-induced neutrophil chemotactic factor was also inhibited by both inhibitors. 6. Both the staurosporine- and the TPA-induced increase in CINC-3 mRNA levels were suppressed by H-7 and genistein.

摘要

将大鼠腹腔中性粒细胞置于含有不同浓度（6.4 - 64 nM）星形孢菌素的培养基中孵育，条件培养基中的中性粒细胞趋化活性会随时间和浓度呈依赖性增加。2. 通过等电聚焦分离条件培养基中的中性粒细胞趋化活性发现，星形孢菌素（64 nM）刺激碱性（pH > 8）中性粒细胞趋化因子的产生，而佛波酯（12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯，49 nM）刺激碱性（pH > 8）和酸性（pH 5）中性粒细胞趋化因子的产生。3. 4小时时通过免疫测定法测定条件培养基中细胞因子诱导的中性粒细胞趋化因子（CINC）-1、-2α、-2β和 -3，结果显示与CINC -1（星形孢菌素，55.0±1.2；佛波酯，12.2±0.3；对照，0.56±0.01 ng/ml）和CINC -2α（星形孢菌素，1.09±0.03；佛波酯，0.90±0.02；对照，<0.10 ng/ml）相比，星形孢菌素（64 nM）和佛波酯（49 nM）强烈刺激CINC -3的产生（星形孢菌素，133.0±3.8；佛波酯，26.7±1.0；对照，0.32±0.01 ng/ml，四个样本的平均值±标准误均值）。CINC -2β低于可检测量（<0.078 ng/ml）。4. 通过逆转录 - 聚合酶链反应测定腹腔中性粒细胞中CINC -3 mRNA的水平。星形孢菌素（64 nM）和佛波酯（49 nM）随时间依赖性增强CINC -3 mRNA的水平，但对甘油醛 - 3 - 磷酸脱氢酶（GAPDH）mRNA水平无影响。5. 蛋白激酶C抑制剂H - 7（IC50，12.3 μM）、钙泊三醇C（IC50，0.77 μM）和Ro 31 - 8425（10 μM时抑制24.3%）以及酪氨酸激酶抑制剂染料木黄酮（IC50，68.5 μM）可抑制星形孢菌素诱导的中性粒细胞趋化因子的产生。佛波酯诱导的中性粒细胞趋化因子的产生也受到这两种抑制剂的抑制。6. H - 7和染料木黄酮可抑制星形孢菌素和佛波酯诱导的CINC -3 mRNA水平的升高。