Platycodin D reverses tumor necrosis factor-α-induced endothelial dysfunction by increasing nitric oxide through G protein-coupled estrogen receptor-mediated eNOS activity

Platycodin D (PCD) is a saponin extracted from the roots of Platycodon grandiflorum (Jacq.) A. DC., a medicinal plant used in Eastern traditional medicine for many years. PCD is known to exert anti-cancer, antioxidant, and anti-obesity effects, and its preventive effects on Cardiovascular Disease, including anti-arteriosclerosis properties are being explored. This study examined the protective effects of PCD against endothelial dysfunction in EA.hy926 endothelial cells, particularly in the context of nitric oxide (NO) production mediated by endothelial nitric oxide synthase (eNOS) activity, and explored the underlying molecular mechanisms. The effects of PCD on tumor necrosis factor-α (TNF-α)-triggered endothelial dysfunction were examined using MTT and Lactate Dehydrogenase, quantitative PCR, western blotting, monocyte adhesion, immunofluorescence, intracellular CA²⁺, and NO generation assays. PCD alleviated TNF-α-induced monocyte-endothelial cell adhesion via downregulation of VCAM-1 and ICAM-1. PCD also increased NO production and eNOS activity by promoting the phosphorylation of CA²⁺/calmodulin-dependent protein kinase β (CaMKKβ), CA²⁺/calmodulin-dependent protein kinase Ⅱα (CaMKⅡα), and AMP-activated protein kinase (AMPK). Importantly, blocking G protein-coupled Estrogen receptor (GPER) suppressed NO production and PCD-triggered eNOS activity by reducing the phosphorylation of three kinases, CaMKKβ, AMPK, and CaMKⅡα. Overall, PCD alleviates TNF-α-induced endothelial dysfunction by enhancing NO production, facilitated by eNOS activity. This upregulation is mediated by GPER-dependent CA²⁺/CaMKKβ/AMPK and CA²⁺/CaMKⅡα signaling pathways. These findings indicate the potential of PCD as a therapeutic agent for preventing endothelial dysfunction and improving cardiovascular health.

Endothelial dysfunction; Endothelial nitric oxide synthase; G protein-coupled estrogen receptor; Nitric oxide; Platycodin D.