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Identification of Elastase as a Secretory Protease from Cultured Rat Microglia
Author(s) -
Nakajima Kazuyuki,
Shimojo Masato,
Hamanoue Makoto,
Ishiura Shoichi,
Sugita Hideo,
Kohsaka Shinichi
Publication year - 1992
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1992.tb11356.x
Subject(s) - protease , elastase , biochemistry , proteases , proteolysis , pancreatic elastase , microbiology and biotechnology , pmsf , biology , secretion , casein , extracellular , neutrophil elastase , chemistry , enzyme , inflammation , immunology
In the course of studying the secretory products of microglia, we detected protease activity in the conditioned medium. Various proteins (casein, histone, myelin basic protein, and extracellular matrix) were digested. The protease activity was characterized by using purified myelin basic protein as a substrate. Maximal activity was observed at neutral pH levels (7‐8), which was different from the optimum pH level of proteolytic activity observed in the cell homogenate. The activity was inhibited approximately 60 and 50% by 1 m M phenylmethylsulfonyl fluoride and 40 μ M elastatinal, respectively. In gel filtration, the major activity, which was inhibited in the presence of N ‐methoxysuccinyl‐Ala‐Ala‐Pro‐Val‐methyl chloride, eluted at a position corresponding to a molecular mass of ∼ 25 kDa. These results suggest that the major protease present in microglial conditioned medium is elastase or an elastase‐like protease. This suggestion was confirmed by the finding that the 25‐kDa protein band was stained with anti‐elastase antiserum by western blotting. De novo synthesis of elastase in microglia was supported by [ 35 S]methionine incorporation. In the presence of lipopoly‐saccharide, the secretory elastase decreased. These results demonstrate that microglia secrete proteases, one of which was identified as elastase. The significance of this enzyme production in physiological and pathological conditions is discussed.