Cu, Zn-superoxide dismutase release from cultured rat hepatocytes]

Oxygen free radicals have been reported to play an important role in some pathologic conditions. Some kinds of cells were reported to release superoxide anion (O2-) when they were exposed to anoxia followed by reoxygenation or some stimuli. As it is thought that hepatocytes are also attacked by exogenous O2-, cultured hepatocytes were subjected to this study. Firstly the reaction of rat cultured hepatocytes to exogenous O2- was investigated with the use of electron spin resonance (ESR) spectroscopy. The hepatocytes rapidly released a O2(-)-scavenging factor and its activity was higher than that of fibroblasts. As the activity of the factor was inhibited by diethyldithiocarbamate (DDC), the factor was thought to be Cu, Zn-SOD. The release was not specifically to O2- and supposed to be caused by mechanical stress. Secondly the factor was analyzed by electrophoresis and Western blot analysis, and proved to be Cu, Zn-SOD. It was also confirmed by the inhibiting effects of DDC and potassium cyanide (KCN). Furthermore Cu, Zn-SOD release was observed for a period of 3 days, comparing with that of lactate dehydrogenase (LDH). The day-to-day release pattern of SOD was similar to that of LDH. But after replacement of medium on Day 1, Cu, Zn-SOD was released more rapidly than LDH. In conclusion the cultured rat hepatocytes released a substantial amount of Cu, Zn-SOD, capable of altering the levels of free radicals. Investigators should pay attention to this effect in an experimental study using cultured rat hepatocytes.