Effects of naofen on enzyme activities of serine proteases and matrix metallo-p roteinases

The aim of this study was to investigate whether naofen affected on the activities of metallo and serine proteases. Naofen, found in both intra- and extra-cellular spaces, increased in the livers especially under pathological conditions such as CCl₄-induced cirrhosis of rats. Moreover, naofen seemed to be digested into fragments which might be closely correlated to the pathological alterations of proliferations and fibrosis. Recent studies showed that metallo and serine proteases degrade the fibrous tissues. Therefore, we investigated possible influences of naofen fragment (s) on the activities of metallo-protease, gelatinase/collagenase and serine protease, trypsin, in vitro by using quenching fluorescence method. It was found that 1.2×10^-1 and 4×l0^-8 M naofen C-fragment had inhibitory effect on trypsin but not gelatinase/collagenase activity. Naofen N-fragment of 1.2×l0^-7 and 4×^-7M did not change gelatinase/collagenase activity but did enhance trypsin activity in a dose-dependent manner. Kunitz type serine protease inhibitor, bikunin inhibited both gelatinase/collagenase and trypsin activities, at bikunin concentrations of 1.2×l0^-7 and 1.2×l0^-6 g mL^-1. Interestingly, naofen N-fragment depleted the reduction ability of bikunin on trypsin from 80 to 50%. These findings indicated that naofen C-fragments may be an endogenous serine protease inhibitor like bikunin, whereas naofen N-fragment may be an enhancer of serine protease and further counteracts the action of the inhibitor, bikunin. Therefore, naofen may be a precursor for active fragments which interacts with serine proteases.