The Volatile Oil of Nardostachyos Radix et Rhizoma Induces Endothelial Nitric Oxide Synthase Activity in HUVEC Cells

<div><p>Nardostahyos Radix et Rhizoma (NRR; the root and rhizome of <i>Nardostachys jatamansi</i> DC.) is a widely used medicinal herb. Historically, NRR is being used for the treatment of cardiovascular and neurological diseases. To search for active ingredients of NRR, we investigated the vascular benefit of NRR volatile oil in (i) the vasodilation in rat aorta ring, and (ii) the release of nitric oxide (NO) and the phosphorylation of endothelial NO synthase (eNOS) in cultured human umbilical vein endothelial cells (HUVECs). By measuring the fluorescence signal in cultures, application of NRR volatile oil resulted in a rapid activation of NO release as well as the phosphorylation of eNOS: both inductions were markedly reduced by L-NAME. In parallel, the phosphorylation level of Akt kinase was markedly increased by the oil treatment, which was partially attenuated by PI3K/Akt inhibitor LY294002. This inhibitor also blocked the NRR-induced NO production and eNOS phosphorylation. In HUVECs, application of NRR volatile oil elevated the intracellular Ca²⁺ level, and BAPTA-AM, a Ca²⁺ chelator, reduced the Ca²⁺ surge: the blockage were also applied to NRR-induced eNOS phosphorylation and NO production. These findings suggested the volatile oil of NRR was the major ingredient in triggering the vascular dilatation, and which was mediated via the NO production.</p></div

Major chemical composition of volatile oil from NRR.

<p>Major chemical composition of volatile oil from NRR.</p

NRR volatile oil induces Ca²⁺ mobilization in HUVECs.

<p>Cultured HUVECs were labeled with fluorescent Ca²⁺ indicator Fluo-4 AM for 30 min. Fluorimetric measurement was performed after the treatment of NRR volatile oil (25 µg/mL), A23187 (1 µM, positive control) or control (untreated culture) (upper panel). Bar = 100 µm. Quantification of Ca²⁺ mobilization was displayed as a ratio of fluorescence intensity at 5 min (F5) to the control at time 0 (F0) in the cultures (lower panel). Mean ± SEM, <i>n</i> = 3, each with triplicate samples. **<i>p</i><0.01.</p

NRR volatile oil induces vasodilation of rat aortic ring.

<p><b>(A):</b> Rat aortic ring was isolated with or without intact endothelium, the vasoconstriction was induced by the applied phenylephrine (Phe, 0.5 µM); acetylcholine (ACh, 1 µM) was then added (left panel). <b>(B):</b> The contraction of aortic ring was tested similar to (A). Different concentrations of NRR volatile oil (1, 3, 10, 30 and 100 µg/mL) were added to induce the relaxation. Also, L-NAME (100 µM) was applied for 30 min, and then different concentrations of NRR volatile oil were added. Values are expressed as percentage of Phe tone as comparing to the control resting tension (right panel). Mean ± SEM, <i>n</i> = 3.</p