Yokukansan, a kampo medicine, protects PC12 cells from glutamate-induced death by augmenting gene expression of cystine/glutamate antiporter system Xc-.

Hitomi Kanno,Yoshio Kase,Kazushige Mizoguchi,Zenji Kawakami,Yasushi Ikarashi,Hiroyoshi Ariga

doi:10.1371/journal.pone.0116275

Abstract

Effects of the kampo medicine yokukansan on gene expression of the cystine/glutamate antiporter system Xc−, which protects against glutamate-induced cytotoxicity, were examined in Pheochromocytoma cells (PC12 cells). Yokukansan inhibited glutamate-induced PC12 cell death. Similar cytoprotective effects were found in Uncaria hook. Experiments to clarify the active compounds revealed that geissoschizine methyl ether, hirsuteine, hirsutine, and procyanidin B1 in Uncaria hook, had cytoprotective effects. These components enhanced gene expressions of system Xc− subunits xCT and 4F2hc, and also ameliorated the glutamate-induced decrease in glutathione levels. These results suggest that the cytoprotective effect of yokukansan may be attributed to geissoschizine methyl ether, hirsuteine, hirsutine, and procyanidin B1 in Uncaria hook.

Highlights

Glutamate-mediated toxicity is an important mechanism of neuronal death in various pathologic conditions including ischemia [1], trauma [2], epileptic seizures [3], and neurodegenerative disorders such as Alzheimer’s, Parkinson’s, and Huntington’s diseases [4, 5]
YKS is composed of seven dried medicinal herbs: 19.5% Atractylodes Lancea rhizome (ALR; rhizome of Atractylodes lancea De Candolle, Compositae), 19.5% Poria sclerotium (PS; sclerotium of Poria cocos Wolf, Polyporaceae), 14.6% Cnidium rhizome (CR; rhizome of Cnidium officinale Makino, Umbelliferae), 14.6% Japanese Angelica root (JAR; root of Angelica acutiloba Kitagawa, Umbelliferae), 9.8% Bupleurum root (BR; root of Bupleurum falcatum Linne, Umbelliferae), 7.3% Glycyrrhiza (GR; root and stolon of Glycyrrhiza uralensis Fisher, Leguminosae), and 14.6% Uncaria hook (UH; hook of Uncaria rhynchophilla Miquel, Rubiaceae)
In other set of experiment, we examined the inhibitory effect of a system Xc2 inhibitor, CPG, on cytoprotection of YKS and four UH-derived components (GM, HTE, HIR, and procyanidin B1 (PCB1)) on glutamate-induced PC12 cell death to verify whether these cytoprotective effects are related to system Xc2 (Fig. 12)

Summary

Introduction

Glutamate-mediated toxicity is an important mechanism of neuronal death in various pathologic conditions including ischemia [1], trauma [2], epileptic seizures [3], and neurodegenerative disorders such as Alzheimer’s, Parkinson’s, and Huntington’s diseases [4, 5]. Pheochromocytoma cells (PC12 cells) are demonstrated to express system Xc2, but do not exhibit the normal NMDA receptor profile [10,11,12,13,14]. Cytoprotection by Yokukansan via System Xc2 demonstrated that PC12 cells lack NR2A and NR2B subunits in the NMDA receptor, system Xc2, consisting of xCT and 4F2hc subunits, is expressed in PC12 cells as well as in primary cultured neurons [15]. These findings suggest that the PC12 cell is a valuable tool for selective evaluation of test substances on system Xc2

Methods

Results

Conclusion