title: Induction of neutrophil chemotactic factor production by staurosporine in rat peritoneal neutrophils.
abstract: 1. 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.
authors.affiliation: Department of Pathophysiological Biochemistry, Faculty of Pharmaceutical Sciences, Tohoku University, Miyagi, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Edamatsu
authors.forename: Y Q
authors.initials: YQ
authors.lastname: Xiao
authors.forename: J
authors.initials: J
authors.lastname: Tanabe
authors.forename: S
authors.initials: S
authors.lastname: Mue
authors.forename: K
authors.initials: K
authors.lastname: Ohuchi
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000818:Animals; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors / Q000096:biosynthesis* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D006133:Growth Substances / Q000096:biosynthesis* / Q000235:genetics; D036341:Intercellular Signaling Peptides and Proteins; D008297:Male; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D011493:Protein Kinase C / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D019311:Staurosporine / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors; C513297:Cxcl1 protein, rat; C513294:Cxcl3 protein, rat; D006133:Growth Substances; D036341:Intercellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate