Increased expression of matrix metalloproteinase-9 in neutrophils in glycogen-induced peritoneal inflammation of guinea pigs

Objective: Matrix metalloproteinase (MMP)-9 plays an important role in neut rophil extravasation and migration by its ability to degrade the major comp onents of basement membrane. To evaluate the expression of neutrophil MMP-9 under inflammatory conditions, we examined the levels of MMP-9 and its mRN A in neutrophils of glycogen-induced peritoneal inflammation. Materials and Methods: Male Hartley guinea pigs weighing 250-300 g were int raperitoneally injected with 0.17% glycogen solution, and 13-15 h after the injection, blood and peritoneal neutrophils were isolated. The levels of M MP-9 and its mRNA were analyzed by gelatin zymography and Northern blotting , respectively. Furthermore, MMP-9 activities in the peritoneal supernatant s were measured. Results: MMP-9 level in peritoneal neutrophils was essentially the same as that in blood neutrophils, although peritoneal neutrophils were assumed to have extracellularly released MMP-9 from the granules during infiltration i nto the peritoneal cavity. Interestingly, MMP-9 mRNA was expressed more abu ndantly in peritoneal neutrophils than in blood neutrophils (p < 0.01). Mor eover, MMP-9 levels in blood and peritoneal neutrophils were reduced to 30- 45 % of non-treated controls by actinomycin D (500 mu g/kg) or cycloheximid e (10 mg/kg)treatment (p<0.05). In contrast, MMP-9 activity increased in th e peritoneal supernatants of glycogen-injected animals was not significantl y affected by actinomycin D- or cycloheximide-treatment. In addition, when blood neutrophils of non-injected animals were stimulated with 10(-7) M N-f ormyl-Met-Leu-Phe, 10 mu g/ml lipopolysaccharide, 10 ng/ml phorbol 12-myris tate 13-acetate, 10(-8) M IL-8 or 100 U/ml tumor necrosis factor-alpha, exp ression of MMP-9 mRNA was markedly increased (p < 0.05). Conclusions: The present observations indicate that MMP-9 gene is transcrib ed, and MMP-9 protein is synthesized in neutrophils during glycogen-injecte d peritoneal inflammation. Moreover, it is likely that MMP-9 protein in the peritoneal supernatants is mostly derived from preformed MMP-9 which is st ored in the neutrophil granules and extracellularly released during infiltr ation of neutrophils into the peritoneal cavity. Finally, the transcription of MMP-9 gene can be upregulated in neutrophils by stimulation with inflam matory mediators, even after neutrophils have been matured.