Ethanol Extract of Lycopodium serratum Thunb. Attenuates Lipopolysaccharide-Induced C6 Glioma Cells Migration via Matrix Metalloproteinase-9 Expression

Objective

To elucidate how ethanol extract of L. serratum (ELS) could exert anti-migratory effects on glioma with the suppression of nuclear factor kappa B (NF-κB) downstream pathway.

Methods

Cell viability of ELS on C6 glioma was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Nitric oxide (NO) assay and 2′,7′-dichlorofluorescin diacetate (DCFH-DA) assay were applied to measure NO production and reactive oxygen species (ROS) generation on lipopolysaccharide (LPS)-induced C6 glioma cells. NF-κB, mitogen-activated protein kinase (MAPK), inducible nictric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein were determined by Western blot. Wound healing assay was used to investigate the inhibitory effect of ELS on fetal bovine serum (FBS)-induced migration and matrix metalloproteinase (MMP)-9 and -2 activity was examined by zymography.

Results

ELS suppressed LPS-induced phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 through inhibiting the expression of chemokine CCL2 (or monocyte chemoattractant protein-1, MCP-1). In addition, ELS inhibited the expression of iNOS, COX-2, and the production of NO by LPS in C6 glioma cells. ELS also significantly decreased serum-induced migration of C6 glioma cells in scratch wound healing in a dose-dependent manner (P<0.01). The activity of MMP-9 and -2 were also significantly attenuated by ELS with LPS treatment (P<0.01).

Conclusions

Our results suggest that downregulation of MMP-9 gene expression might be involved in the anti-migration effect of ELS against LPS-induced C6 glioma cells.