Microglia‐Derived Elastase Produces a Low‐Molecular‐Weight Plasminogen that Enhances Neurite Outgrowth in Rat Neocortical Explant Cultures

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Abstract
In the course of analysis of plasminogen in microglial conditioned medium (Mic-CM), novel low-molecular-weight (LMW) zymogen with a molecular mass of approximately 36 kDa was detected by casein-urokinase zymography. Because this form was produced when rat native plasminogen was incubated with Mic-CM, a specific protease in the Mic-CM was thought to be responsible for the production of LMW plasminogen. The production of LMW plasminogen was strongly inhibited by elastase inhibitors. Furthermore, elastase (pancreatic or leukocyte) was also found to produce LMW zymogen from native plasminogen. These results indicate that LMW plasminogen is produced through limited proteolysis by an elastase-like protease in Mic-CM. To determine the biochemical characteristics of LMW plasminogen, rat native plasminogen was cleaved by pancreatic elastase, and the fragments (LMW plasminogen and nonzymogen fragments) were purified by several kinds of column chromatography. Amino acid sequence analysis revealed that LMW plasminogen is a carboxy-terminal region that contains the fifth kringle domain and a protease active site, and the amino acid sequence is identical to that of LMW plasminogen produced by Mic-CM. On the other hand, the nonzymogen fragment was the amino-terminal region containing four kringle domains. The effects of native plasminogen and the fragments on neurite outgrowth of rat brain explant were examined. LMW plasminogen promoted neurite outgrowth as well as did native plasminogen, whereas nonzymogen fragments did not. These results suggest that LMW plasminogen, which is produced from native plasminogen by elastase, may be a physiologically active molecule that mediates the intercellular interaction between microglia and neurons.