Effect of R-(+)-lipoic acid on growth, proliferation and related mechanism in human HepG2 cells

To study the effect of antioxidants R-(+)-lipoic acid(R-LA)on cells growth, proliferation and related mechanisms in human HepG2 cells lines. MTT was used to measure cells growth and proliferation. Reactive oxygen species(ROS)kit was used to analyze ROS level. Cell apoptosis and cell morphological changes were observed by flow cytometry and Hoechst 33258 test. Protein expression levels of apoptosis, autophagy and related pathway were analyzed through Western blot, including Bax, Bcl-2, caspase 3, PARP, ATG5, ATG7, LC3, Beclin1, mTOR, P70S6K, P38, P53, ERK and Akt etc. Results showed that cell growth and proliferation were inhibited in a dose- and time-dependent manner after being treated by lipoic acid. R-LA could increase ROS production, pro-apoptosis proteins Bax levels, activated caspase family and PARP. Meanwhile, R-LA could up-regulate the levels of autophagy-related proteins including ATG5, ATG7, Beclin1 and LC3, and down-regulate phospho-mTOR and P70S6K levels. Signal pathway results showed that R-LA could up-regulate phospho-P38 and phospho-JNK levels, and decrease phospho-Akt and phospho-ERK. When adding 3-methyladenine, autophagy was inhibited. Thus, R-LA might activate autophagy and induce apoptosis by P38/AMPK-JNK, PI3K/AKT and Ras/Raf/MEK/ERK pathways.