The protective effect of Smilax glabra extract on advanced glycation end products-induced endothelial dysfunction in HUVECs via RAGE-ERK1/2-NF-κB pathway

Ethnopharmacological relevance: Smilax glabra Roxb. (SGR) is a traditional Chinese herb that has been used in folk for the treatment of diabetic vascular complications. Advanced glycation end products (AGEs)-induced endothelial dysfunction has been thought to be a major cause of diabetic vascular complications. The present study was conducted to investigate the protective effect of SGR extract on AGEs-induced endothelial dysfunction and its underlying mechanisms.

Materials and methods: Human umbilical vein endothelial cells (HUVECs) were exposed to 200 μg/ml AGEs to induce endothelial dysfunction. Acridine orange/ethidium bromide (AO/EB) fluorescence assay and Annexin-V/PI double-staining were performed to determine endothelium apoptosis. Dihydroethidium (DHE) fluorescence probe, SOD and MDA kits were used to evaluate oxidative stress. The effect of SGR extract on AGEs-induced TGF-beta1 expression was determined by immunocytochemistry and western blotting. Attenuations of SGR extract on receptor for AGEs (RAGE) expression, extracellular regulated protein kinases (ERK1/2) activation and NF-κB phosphorylation were determined by immunofluorescence assay and western blotting. The blockade assays for RAGE and ERK1/2 were carried out using a specific RAGE-antibody (RAGE-Ab) or a selective ERK1/2 inhibitor PD98059 in immunofluorescence assay.

Results: The pretreatment of SGR extract (0.125, 0.25 and 0.5 mg crude drug/ml) significantly attenuated AGEs-induced endothelium apoptosis, and down-regulated TGF-beta1 protein expression in HUVECs. It was also well shown that SGR extract could down-regulate dose-dependently ROS over-generation, MDA content, TGF-beta1 expression, ERK1/2 and NF-κB activation whereas increase significantly SOD activity. Furthermore, the AGEs-induced ERK1/2 activation could be attenuated by the blockade of RAGE-Ab (5 μg/ml) while the NF-κB activation was ameliorated by ERK1/2 inhibitor PD98059 (10 μM).

Conclusion: These results indicated that SGR extract could attenuate AGEs-induced endothelial dysfunction via RAGE-ERK1/2-NF-κB pathways. Our findings suggest that SGR extract may be beneficial for attenuating endothelial dysfunction in diabetic vascular complications.