Pathologic characteristics of immunologic injury in primary cultured rat hepatocytes and protective effect of glycyrrhizin in vitro
Abstract
AIM: To explore the pathologic characteristics of immunologic injury in primary cultured rat hepatocytes, and to evaluate the protective effect of glycyrrhizin (Grz) in vitro.
METHODS: Sprague-Dawley rats were initiated with BCG vaccine in vivo, and then the primary cultured rat hepatocytes were stimulated with lipopolysaccharides (LPS) 10 mg/L in vitro (BCG+LPS treatment), the hepatoprotection by Grz was evaluated in the hepatocytes treated as above. Supernatant AST and LDH activities were measured, and nitric oxide (NO) was evaluated by Giress reaction. Aminoguanidine was used to confirm the target of Grz action. Expression of intercellular adhesion molecular-1 (ICAM-1) was determined by immunocytochemistry. Percentage of apoptosis was detected with fluorescence microscope and flow cytometer.
RESULTS: In LPS-treated group, supernatant AST was not increased compared with that in control, while supernatant AST and LDH in BCG+LPS treatment were increased significantly (P<0.05). The enhancements of AST and LDH were inhibited by co-culture with Grz (P<0.05). Both supernatant NO and ICAM-1 expression, and percentage of apoptosis in hepatocytes were elevated by BCG+LPS treatment (P<0.01), and these elevations could be decreased by Grz co-incubation either (P<0.05). The NO generation could be decreased by aminoguarnidine treatment (P<0.05).
CONCLUSION: The elevations of supernatant NO, ICAM-1 expression, and apoptosis in hepatocytes could be taken as three important pathologic changes in the immunologic hepatotoxicity induced by BCG+LPS treatment in primary cultured rat hepatocytes. NO synthesis mechanism was involved in the immunologic hepatotoxic process. Grz could downregulate both supernatant NO and hepatocyte apoptosis in the culture system.
Keywords:
METHODS: Sprague-Dawley rats were initiated with BCG vaccine in vivo, and then the primary cultured rat hepatocytes were stimulated with lipopolysaccharides (LPS) 10 mg/L in vitro (BCG+LPS treatment), the hepatoprotection by Grz was evaluated in the hepatocytes treated as above. Supernatant AST and LDH activities were measured, and nitric oxide (NO) was evaluated by Giress reaction. Aminoguanidine was used to confirm the target of Grz action. Expression of intercellular adhesion molecular-1 (ICAM-1) was determined by immunocytochemistry. Percentage of apoptosis was detected with fluorescence microscope and flow cytometer.
RESULTS: In LPS-treated group, supernatant AST was not increased compared with that in control, while supernatant AST and LDH in BCG+LPS treatment were increased significantly (P<0.05). The enhancements of AST and LDH were inhibited by co-culture with Grz (P<0.05). Both supernatant NO and ICAM-1 expression, and percentage of apoptosis in hepatocytes were elevated by BCG+LPS treatment (P<0.01), and these elevations could be decreased by Grz co-incubation either (P<0.05). The NO generation could be decreased by aminoguarnidine treatment (P<0.05).
CONCLUSION: The elevations of supernatant NO, ICAM-1 expression, and apoptosis in hepatocytes could be taken as three important pathologic changes in the immunologic hepatotoxicity induced by BCG+LPS treatment in primary cultured rat hepatocytes. NO synthesis mechanism was involved in the immunologic hepatotoxic process. Grz could downregulate both supernatant NO and hepatocyte apoptosis in the culture system.