Glossogyne tenuifolia Extract Increases Nitric Oxide Production in Human Umbilical Vein Endothelial Cells

The vascular nitric oxide (NO) system has a protective effect in atherosclerosis. NO is generated from the conversion of L-arginine to L-citrulline by the enzymatic action of endothelial NO synthase (eNOS). Compounds with the effect of enhancing eNOS expression are considered to be candidates for the prevention of atherosclerosis. In this study, extracts from the aerial, root, and whole plant of Glossogyne tenuifolia (GT) were obtained with ethanol, n-hexane, ethyl acetate (EA), and methanol extraction, respectively. The effects of these GT extracts on the synthesis of NO and the expression of eNOS in human umbilical vein endothelial cells (HUVECs) were investigated. NO production was determined as nitrite by colorimetry, following the Griess reaction. The treatment of HUVECs with EA extract from the root of GT and n-hexane, methanol, and ethanol extract from the aerial, root, and whole plant of GT increased NO production in a dose-dependent manner. When at a dose of 160 μg/mL, NO production increased from 0.9 to 18.4-fold. Among these extracts, the methanol extract from the root of GT (R/M GTE) exhibited the most potent effect on NO production (increased by 18.4-fold). Furthermore, using Western blot and RT–PCR analysis, treatment of HUVECs with the R/M GTE increased both eNOS protein and mRNA expression. In addition, Western blot analysis revealed that the R/M GTE increased eNOS phosphorylation at serine1177 as early as 15 min after treatment. The chemical composition for the main ingredients was also performed by HPLC analysis. In conclusion, the present study demonstrated that GT extracts increased NO production in HUVECs and that the R/M GTE increased NO production via increasing eNOS expression and activation by phosphorylation of eNOS at serine1177.

Glossogyne tenuifolia Attenuates Proliferation and Migration of Vascular Smooth Muscle Cells

The proliferation and migration of vascular smooth muscle cells (VSMCs) are essential in the pathogenesis of various vascular diseases, such as atherosclerosis and restenosis. Among the mediators of VSMC during atherosclerosis development, platelet-derived growth factor (PDGF)-BB is a potent mitogen for VSMCs and greatly contributes to the intimal accumulation of VSMCs. Glossogyne tenuifolia (GT, Xiang-Ru) is a traditional antipyretic and hepatoprotective herb from Penghu Island, Taiwan. This study evaluated the inhibitory effect of GT ethanol extract (GTE) and GT water extract (GTW) on proliferative and migratory activities in PDGF-BB-induced VSMCs. The experimental results demonstrated that GTE significantly inhibited the PDGF-BB-stimulated VSMC proliferation and migration, as shown by MTT, wound healing, and Boyden chamber assays. GTE was found to have a much more potent inhibitory activity than GTW. Based on the Western blot analysis, GTE significantly blocked the PDGF-BB-induced phosphorylation of NF-κB and mitogen-activated protein kinase (MAPK) pathways, including extracellular signal-regulated kinase (ERK), p38, and JNK, in VSMCs. In addition, GTE retarded the PDGF-BB-mediated migration through the suppression of matrix metalloproteinase (MMP)-2 and MMP-9 expression in VSMCs. Three main ingredients of GT—chlorogenic acid, luteolin-7-glucoside, and luteolin—all showed significant anti-proliferative effects on PDGF-BB-induced VSMCs. As a whole, our findings indicated that GTE has the potential to be a therapeutic agent to prevent or treat restenosis or atherosclerosis.