Abstract:

Objective To investigate the regulation role and mechanism of ginsenoside Rg1 on autophagy and apoptosis in Raw 264.7 macrophages stimulated with serum free. Methods Raw 264.7 macrophages were cultured and treated differently in vitroand randomly divided into the control group, the serum free group (12, 24, 36, 48 and 60 hours) in which ginsenoside Rg1 (50μmol/L) was added with serum free treatment group (24, 36 and 48 hours), pretreatment with autophagy inhibitor 3-methyladennine (3-MA) (5mmol/L) for 1 hour then added ginsenoside Rg1 (50μmol/L) with serum free treatment group and corresponding control group. The protein expressions of microtubule-associated protein 1 light chain 3 (LC3), Atg5, Beclin 1, cleaved Caspase-3, Bcl-2 and Bax were detected by Western blotting respectively. The variation of protein expression level of LC3 was measured by double immunofluorescence labeling. The morphology of cell nucleus was measured by Hochest 33342/PI double fluorescent double staining. Results Different time (12, 24, 36, 48 and 60 hours) of the serum free groups induced autophagy. Compared with different time of the serum free groups, the ginsenoside Rg1 (50μmol/L) was added with serum free group up-regulated the protein expression of LC3, Atg 5 and Beclin 1. Compared with the ginsenoside Rg1 (50μmol/L) group, pretreatment with 3-MA (5mmol/L) inhibited the protein expression of Bcl-2 and up-regulated the protein expression of cleaved Caspase-3 and Bax and the quantity of apoptosis in Raw 264.7 macrophages. Conclusion Ginsenoside Rg1 effectively attenuates serum free-induced apoptosis by inducing the autophagy in Raw 264.7 macrophages.

Key words: Ginsenoside Rg1, Autophagy, Raw 264.7 macrophage, Western blotting, Double immunofluorescent labeling, Hochest 33342/PI fluorescent double staining, Mouse