Abstract: This study was designed to investigate the role of glycyrrhizin acid in the infection of the mouse macrophage cell line RAW264.7 against Mycoplasma ovipneumoniae (MO).The optimal concentration of glycyrrhizic acid was determined by CCK-8 cell activity assay and the effects of glycyrrhizic acid on the activity of MO-infected macrophages were examined.The effect of glycyrrhizin acid on the growth cycle of RAW 264.7 macrophages was detected by flow cytometry.On the other hand,the effect of glycyrrhizin acid on the secretion of TNF-α by MO-infected macrophages was detected by ELISA.The expression of apoptosis factors Bax and Bad were detected by Western blot.RT-PCR was used to detect the expression of apoptosis and autophagy related genes.The results showed that glycyrrhizic acid at a concentration of 12 μmol/L significantly increased the activity of RAW264.7 (P=0.012 9).In cell cycle experiment,it is found that the number of cells treated with glycyrrhizic acid in the G1 phase decreased while the number of cells in the G2 phase increased.Glycyrrhizic acid (12 μmol/L) increased the proliferation rate of MO-infected RAW264.7 (P=0.034 0).The ELISA test resulted in an increase of TNF-α content in culture supernatant (P=0.015 2).Western Blot analysis to examine the expression level of proapoptotic protein was done and Bax protein was found increase in macrophages.For RT-PCR,it was found that the expression levels of Caspase 3 and Caspase 9 were significantly down-regulated (P<0.000 1),however the expression levels of autophagy-related genes Atg 7 and Beclin 1 were significantly increased (P<0.000 1).The results suggest that glycyrrhizic acid can play an immunoregulatory role by promoting proliferation,inhibiting apoptosis,promoting the expression of TNF-α and increasing autophagy in the case of MO-infected macrophages.

Key words: Mycoplasma ovipneumoniae, macrophages, glycyrrhizic acid, autophagy, immunoregulation