Rosmarinic Acid Attenuates the Activation of Murine Microglial N9 Cells through the Downregulation of Inflammatory Cytokines and Cleaved Caspase-3.

OBJECTIVE: The present study evaluated the ability of rosmarinic acid (RA) to inhibit microglia activation induced by lipopolysaccharide (LPS) in the N9 murine microglial cell line, and investigated the putative mechanisms involved in this process. METHODS: In all tests, N9 murine microglial cells were pretreated with RA (0.1, 1.0, and 10 µM) for 20 h and exposed to LPS (1 µM/mL) for 4 h. Cell viability was measured by Trypan blue exclusion assay. Flow cytometry was used to detect reactive oxygen species (ROS), quantify cleaved caspase-3, and analyze the mitochondrial electrochemical potential. iNOS, Arg-1, TNF-α, IL-1ß, and IL-6 proteins were analyzed by Western blotting, and their antigens were detected using the chemiluminescence technique. The effect of RA on DNA was evaluated by the Comet assay. RESULTS: RA attenuated the expression of the M1 marker iNOS and the levels of proinflammatory factors, including TNF-α, IL-1ß, and IL-6; it increased the expression of the M2 marker Arg-1, and inhibited, at least in part, ROS generation and loss of mitochondrial outer membrane permeabilization through the inhibition of cleaved caspase-3 activation. RA also inhibited DNA damage, reassuring cell protection. CONCLUSIONS: The results suggested a protective effect of RA through downregulation of inflammatory cytokines and cleaved caspase-3.

Gamma-decanolactone inhibits iNOS and TNF-alpha production by lipopolysaccharide-activated microglia in N9 cells.

Activated microglia that produce reactive nitrogen species (RNS), inflammatory factors, reactive oxygen species (ROS), and other neurovirulent factors may lead to the development of neurodegenerative diseases. Certain compounds can inhibit the activation of microglia. However, these mechanisms remain unclear. In the present study, we investigated the inhibitory effect of Gamma-decanolactone (GD) on the production of reactive oxygen species and inducible nitric oxide synthase (iNOS) in lipopolysaccharide (LPS) - stimulated N9 murine microglial cells through the p38 MAPK signaling pathway. The results showed that GD attenuated the activation of N9 cells and inhibited intracellular reactive oxygen species and the expression of iNOS and TNF-α induced by LPS in the cells. In addition, GD blocked the phosphorylation of p38 and inhibited cleaved caspase-9 and DNA damage. These data indicate that GD has therapeutic potential for the treatment of neurodegenerative diseases, and that it exerts its effects by inhibiting inflammation.