Erratum: Lee et al. Ginseng Extracts, GS-KG9 and GS-E3D, Prevent Blood-Brain Barrier Disruption and Thereby Inhibit Apoptotic Cell Death of Hippocampal Neurons in Streptozotocin-Induced Diabetic Rats. Nutrients 2020, 12, 2383.

The authors have requested that the following changes be made to their paper [...].

Pectin-Lyase-Modified Ginseng Extract and Ginsenoside Rd Inhibits High Glucose-Induced ROS Production in Mesangial Cells and Prevents Renal Dysfunction in db/db Mice.

Diabetes increases the incidence rate of chronic renal disease. Pectin-lyase-modified ginseng (GS-E3D), with enhanced ginsenoside Rd content, has been newly developed. In this study, renal protective roles of GS-E3D in type-2 diabetic db/db mice were investigated. The generation of reactive oxygen species (ROS) induced by high glucose (25 mM) was reduced by ES-E3D (75%) and ginsenoside Rd (60%). Diabetic db/db mice received 100 or 250 mg/kg/day of GS-E3D daily via oral gavage for 6 weeks. Albuminuria and urinary 8-hydroxy-2'-deoxyguanosine (8-OhdG, an oxidative stress marker) levels were increased in db/db mice and the levels recovered after GS-E3D treatment. In renal tissues, TUNEL-positive cells were decreased after GS-E3D treatment, and the increased apoptosis-related protein expressions were restored after GS-E3D treatment. Therefore, GS-E3D has a potent protective role in diabetes-induced renal dysfunction through antioxidative and antiapoptotic activities. These results may help patients to select a dietary supplement for diabetes when experiencing renal dysfunction.

A randomized, double-blind, placebo-controlled pilot study to assess the effects of protopanaxadiol saponin-enriched ginseng extract and pectinase-processed ginseng extract on the prevention of acute respiratory illness in healthy people.

BACKGROUND: GS-3K8 and GINST, both of which are modified ginseng extracts, have never been examined in terms of their effectiveness for the prevention of acute respiratory illness (ARI) in humans. We conducted a pilot study to assess the feasibility of performing a large-scale, randomized, controlled trial. METHODS: This study was a randomized, double-blind, placebo-controlled, pilot study at a single center from October 2014 to March 2015. The 45 healthy applicants were randomly divided into the GS-3K8 (n = 15), GINST (n = 15), and placebo groups (n = 15). The study drug was administered as a capsule (500 mg/cap and 3000 mg/day). GS-3K8 contained 6.31 mg/g of Rg1, 15.05 mg/g of Re, 30.84 mg/g of Rb1, 15.02 mg/g of Rc, 12.44 mg/g of Rb2, 6.97 mg/g of Rd, 1.59 mg/g of Rg3, 3.25 mg/g of Rk1, and 4.84 mg/g of Rg5. GINST contained 7.54 mg/g of Rg1, 1.87 mg/g of Re, 5.42 mg/g of Rb1, 0.29 mg/g of Rc, 0.36 mg/g of Rb2, 0.70 mg/g of Rd, and 6.3 mg/g of compound K. The feasibility criteria were the rates of recruitment, drug compliance, and successful follow-up. The primary clinical outcome measure was the incidence of ARI. The secondary clinical outcome measures were the duration of symptoms. RESULTS: The rate of recruitment was 11.3 participants per week. The overall rate of completed follow-up was 97.8%. The mean compliance rate was 91.64 ± 9.80%, 95.28 ± 5.75%, and 89.70 ± 8.99% in the GS-3K8, GINST, and placebo groups, respectively. The incidence of ARI was 64.3% (9/14; 95% confidence interval [CI], 31.4-91.1%), 26.7% (4/15; 95% CI, 4.3-49.0%), and 80.0% (12/15; 95% CI, 54.8-93.0%) in the GS-3K8, GINST, and placebo groups, respectively. The average days of symptoms were 3.89 ± 4.65, 9.25 ± 7.63, and 12.25 ± 12.69 in the GS-3K8, GINST, and placebo groups, respectively. CONCLUSION: The results support the feasibility of a full-scale trial. GS-3K8 and GINST appear to have a positive tendency toward preventing the development of ARI and reducing the symptom duration. A randomized controlled trial is needed to confirm these findings.

Antioxidant and hepatoprotective effects of Korean ginseng extract GS-KG9 in a D-galactosamine-induced liver damage animal model.

BACKGROUND/OBJECTIVES: This study was designed to investigate the improvement effect of white ginseng extract (GS-KG9) on D-galactosamine (Ga1N)-induced oxidative stress and liver injury. SUBJECTS/METHODS: Sixty Sprague-Dawley rats were divided into 6 groups. Rats were orally administrated with GS-KG9 (300, 500, or 700 mg/kg) or silymarin (25 mg/kg) for 2 weeks. The rats of the GS-KG9- and silymarin-treated groups and a control group were then intraperitoneally injected Ga1N at a concentration of 650 mg/kg for 4 days. To investigate the protective effect of GS-KG9 against GalN-induced liver injury, blood liver function indicators, anti-oxidative stress indicators, and histopathological features were analyzed. RESULTS: Serum biochemical analysis indicated that GS-KG9 ameliorated the elevation of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) in GalN-treated rats. The hepatoprotective effects of GS-KG9 involved enhancing components of the hepatic antioxidant defense system, including glutathione, glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT). In addition, GS-KG9 treatment inhibited reactive oxygen species (ROS) production induced by GalN treatment in hepatocytes and significantly increased the expression levels of nuclear factor erythroid-2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) proteins, which are antioxidant proteins. In particular, by histological analyses bases on hematoxylin and eosin, Masson's trichrome, α-smooth muscle actin, and transforming growth factor-ß1 staining, we determined that the administration of 500 mg/kg GS-KG9 inhibited hepatic inflammation and fibrosis due to the excessive accumulation of collagen. CONCLUSIONS: These findings demonstrate that GS-KG9 improves GalN-induced liver inflammation, necrosis, and fibrosis by attenuating oxidative stress. Therefore, GS-KG9 may be considered a useful candidate in the development of a natural preventive agent against liver injury.

Effectiveness and Safety of Panax ginseng Extract on Hepatic Dysfunction: A Randomized, Double-Blind, Placebo-Controlled Clinical Trial.

BACKGROUND: The purpose of this study was to evaluate the efficacy and safety of Panax ginseng extract (GS-KG9) in the treatment of hepatic dysfunction. METHODS: A randomized, double-blind, placebo-controlled clinical trial was conducted from December 2017 to January 2019. The trial included 60 subjects between the ages of 19 and 70 who had higher alanine transaminase (ALT) levels than the normal upper limit. The subjects were randomly divided into two groups: GS-KG9 (n = 30) and placebo (n = 30). The former was administered three GS-KG9 capsules (3 g/day) and the latter three placebo capsules (3 g/day) twice each day orally after meals in the morning and evening for 12 weeks. The primary goal was to observe the changes in ALT and gamma-glutamyl transferase (GGT) levels. The safety of the treatment was assessed and adverse events (AEs) were recorded. RESULTS: Out of 60 subjects, nine were excluded from the efficacy analysis because they met the exclusion criteria. Therefore, a total of 51 subjects were evaluated for the effectiveness of the treatment (26 in the GS-KG9 group and 25 in the placebo group). After 12 weeks of treatment, the ALT levels were significantly reduced in the GS-KG9 group compared to the placebo group (p=0.009). The GGT level of the GS-KG9 group was significantly lower than that of the placebo group (p=0.036). Mild AEs, such as diarrhea, occurred during the study. There were no significant differences between the two groups. CONCLUSION: The results of this trial suggest that GS-KG9 might be an effective and safe option for mild hepatic dysfunction. This trial is registered with KCT0004080.

Pectin lyase-modified red ginseng extract improves glucose homeostasis in high fat diet-fed mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Red ginseng has long been used as a traditional folk medicine for various diseases including diabetes. Recently, a preparation of red ginseng extract by pectin lyase modification has been developed and named as GS-E3D. AIM OF THE STUDY: The aim of this study is to evaluate the preventive effect of GS-E3D on hyperglycemia induced by feeding a high fat diet (HFD) in mice. MATERIALS AND METHODS: GS-E3D was orally administered to C57BL/6J mice at different doses (250, 500, or 1000 mg/kg/day) for 6 weeks while on a HFD. Body weight and blood glucose were monitored weekly, and oral glucose tolerance test (OGTT) was performed at 5th week of the experiment. Glycemic indications and metabolic parameters were further measured in serum. RESULTS: Six weeks of GS-E3D treatment to mice significantly inhibited HFD-induced body weight gain, hyperglycemia, hyperinsulinemia and hypertriglyceridemia. Notably, GS-E3D treated mice at doses of 250, 500 and 1000 mg/kg showed 41.8%, 45.0% and 55.1% reduction in insulin resistance index, respectively, compared to HFD control mice. OGTT revealed that GS-E3D markedly prevented steep rise of blood glucose and insulin levels after glucose challenge and ameliorated HFD-induced glucose and insulin intolerance. The histological analysis showed enlarged adipocytes in HFD-fed mice whereas the adipocyte hypertrophy was prevented in GS-E3D treated mice in a dose-dependent manner. Furthermore, when peripheral glucose uptake level was assessed by total and membranous glucose transporter type 4 (GLUT4) protein contents, GS-E3D restored GLUT4 protein expression to the levels of regular diet fed mice, and dose-dependently translocated them to the plasma membrane. CONCLUSION: The results collectively show that GS-E3D ameliorates obesity-related impaired glucose tolerance by improving insulin sensitivity in the epidydimal adipose tissue.

Acute and genetic toxicity of GS-E3D, a new pectin lyase-modified red ginseng extract.

Korean red ginseng and its extract have been used as traditional medicines and functional foods in countries worldwide. Pectin lyase-modified red ginseng extract (GS-E3D) was newly developed as a dietary supplement for obesity, diabetes-related renal dysfunction, etc. In this study, the safety of GS-E3D on acute toxicity and genotoxicity was evaluated. For acute study, Sprague-Dawley rats were administrated by oral gavage at a dose of 5000 mg/kg GS-E3D. To evaluate genotoxicity of GS-E3D, we conducted three-battery tests, which are Ames test using Escherichia coli (WP2uvrA pKM101) and Salmonella typhimurium strains (TA98, TA100, TA1535 and TA1537), chromosomal aberration test -using Chinese hamster lung cells, and micronucleus test using ICR mice. In acute toxicity studies, there were no dead animals or abnormal necropsy findings in the control group and GS-E3D (5000 mg/kg) treated group. GS-E3D did not induce mutagenicity in the bacterial test, chromosomal aberrations in Chinese hamster lung cells and micronuclei in bone marrow cells of mice. Conclusively, the approximate lethal dose of GS-E3D was greater than 5000 mg/kg bw and GS-E3D has no genotoxic potential in the three-battery tests on genotoxicity.

Ginseng Extract Modified by Pectin Lyase Inhibits Retinal Vascular Injury and Blood-Retinal Barrier Breakage in a Rat Model of Diabetes.

GS-E3D is an enzymatically modified ginseng extract by pectin lyase. In this study, we evaluated the preventive effects of GS-E3D on blood-retinal barrier (BRB) leakage in a rat model of diabetes. To produce diabetes, rats were injected with streptozotocin. GS-E3D was orally gavaged at 25, 50, and 100 mg/kg body weight for 6 weeks. We then compared the effect of GS-E3D with that of an unmodified ginseng extract (UGE) on retinal vascular leakage. The administration of GS-E3D significantly blocked diabetes-induced BRB breakdown. Immunofluorescence staining showed that GS-E3D reduced the loss of occludin in diabetic rats. In TUNEL staining, the number of apoptotic retinal microvascular cells was dose dependently decreased by GS-E3D treatment. GS-E3D decreased the accumulations of advanced glycation end products in the retinal vessels. In addition, the inhibition potential of GS-E3D on BRB breakage was stronger compared with UGE. These results indicate that GS-E3D could be a beneficial treatment option for preventing diabetes-induced retinal vascular injury.

GS-KG9 ameliorates diabetic neuropathic pain induced by streptozotocin in rats.

BACKGROUND: Diabetic neuropathy is one of the most devastating ailments of the peripheral nervous system. Neuropathic pain develops in ∼30% of diabetics. Here, we examined the suppressive effect of GS-KG9 on neuropathic pain induced by streptozotocin (STZ). METHODS: Hyperglycemia was induced by intraperitoneal injection of STZ. Rats showing blood glucose level > 250 mg/dL were divided into five groups, and treatment groups received oral saline containing GS-KG9 (50 mg/kg, 150 mg/kg, or 300 mg/kg) twice daily for 4 wk. The effects of GS-KG9 on pain behavior, microglia activation in the lumbar spinal cord and ventral posterolateral (VPL) nucleus of the thalamus, and c-Fos expression in the dorsal horn of the lumbar spinal cord were examined. RESULTS: The development of neuropathic pain began at Day 5 and peaked at Week 4 after STZ injection. Mechanical and thermal pains were both significantly attenuated in GS-KG9-treated groups from 10 d after STZ injection as compared to those in the STZ control. GS-KG9 also repressed microglia activation in L4 dorsal horn and VPL region of the thalamus. In addition, increase in c-Fos-positive cells within L4 dorsal horn lamina I and II of the STZ control group was markedly alleviated by GS-KG9. CONCLUSION: These results suggest that GS-KG9 effectively relieves STZ-induced neuropathic pain by inhibiting microglial activation in the spinal cord dorsal horn and VPL region of the thalamus.

GS-E3D, a new pectin lyase-modified red ginseng extract, inhibited diabetes-related renal dysfunction in streptozotocin-induced diabetic rats.

BACKGROUND: GS-E3D is a newly developed pectin lyase-modified red ginseng extract. The purpose of this study was to investigate the therapeutic effects of GS-E3D on diabetes-related renal dysfunction in streptozotocin-induced diabetic rats. METHOD: GS-E3D (25, 50, and 100 mg/kg body weight per day) was administered for 6 weeks. The levels of blood glucose and hemoglobin A1c, and of urinary albumin, 8-hydroxy-2'-deoxyguanosine (8-OHdG), and advanced glycation end-products (AGEs) were determined. Kidney histopathology, renal accumulation of AGEs, and expression of α-smooth muscle actin (α-SMA) were also examined. RESULTS: Administration of GS-E3D for 6 weeks reduced urinary levels of albumin, 8-OHdG, and AGEs in diabetic rats. Mesangial expansion, renal accumulation of AGEs, and enhanced α-SMA expression were significantly inhibited by GS-E3D treatment. Oral administration of GS-E3D dose-dependently improved all symptoms of diabetic nephropathy by inhibiting renal accumulation of AGEs and oxidative stress. CONCLUSION: The results of this study indicate that the use of GS-E3D as a food supplement may provide effective treatment of diabetes-induced renal dysfunction.

Pectin lyase-modified red ginseng extract exhibits potent anti-glycation effects in vitro and in vivo.

PURPOSE: GS-E3D is a newly developed pectin lyase-modified red ginseng extract. The purpose of this study was to evaluate the inhibitory effects of GS-E3D against advanced glycation end products. METHODS: In this study, we evaluated the inhibitory effects of GS-E3D on the formation of advanced glycation end products (AGEs) and their cross-linking with collagen in vitro and in streptozotocin-induced diabetic rats. RESULTS: An in vitro assay for the glycation of bovine serum albumin by methylglyoxal showed that GS-E3D inhibited AGE formation at an IC50 value of 19.65 ± 4.35 µg/mL. In addition, GS-E3D showed a potent inhibitory effect (IC50 = 0.42 ± 0.08 mg/mL) on the cross-linking of AGEs with collagen. However, GS-E3D showed no effect on preformed AGEs cross-linked with collagen in the breakdown assay. To determine whether GS-E3D inhibits AGE formation and their cross-linking with proteins in vivo, streptozotocin induced diabetic rats were treated with GS-E3D (25, 50, and 100 mg/kg/day) for 6 weeks. The administration of GS-E3D decreased serum levels of AGEs and their cross linking with proteins in diabetic rats. CONCLUSION: The inhibitory effects of this agent on advanced glycation in vitro and in vivo suggested that it may have a potential therapeutic role in controlling diabetes-induced AGE burden in various tissues.

White Ginseng Protects Mouse Hippocampal Cells Against Amyloid-Beta Oligomer Toxicity.

Amyloid-beta oligomer (AßO) is a soluble oligomer form of the Aß peptide and the most potent amyloid-beta form that induces neuronal damage in Alzheimer's disease. We investigated the effect of dried white ginseng extract (WGE) on neuronal cell damage and memory impairment in intrahippocampal AßO (10 µM)-injected mice. Mice were treated with WGE (100 and 500 mg/kg/day, p.o.) for 12 days after surgery. WGE improved memory impairment by inhibiting hippocampal cell death caused by AßO. In addition, AßO-injected mice treated with WGE showed restoration of reduced synaptophysin and choline acetyltransferase intensity and lower levels of ionized calcium-binding adaptor molecule 1 in the hippocampus compared with those of vehicle-treated controls. These results suggest that WGE reverses memory impairment in Alzheimer's disease by attenuating neuronal damage and neuroinflammation in the AßO-injected mouse hippocampus. Copyright © 2017 John Wiley & Sons, Ltd.

Differential Effects of Quercetin and Quercetin Glycosides on Human α7 Nicotinic Acetylcholine Receptor-Mediated Ion Currents.

Quercetin is a flavonoid usually found in fruits and vegetables. Aside from its antioxidative effects, quercetin, like other flavonoids, has a various neuropharmacological actions. Quercetin-3-O-rhamnoside (Rham1), quercetin-3-O-rutinoside (Rutin), and quercetin- 3-(2(G)-rhamnosylrutinoside (Rham2) are mono-, di-, and tri-glycosylated forms of quercetin, respectively. In a previous study, we showed that quercetin can enhance α7 nicotinic acetylcholine receptor (α7 nAChR)-mediated ion currents. However, the role of the carbohydrates attached to quercetin in the regulation of α7 nAChR channel activity has not been determined. In the present study, we investigated the effects of quercetin glycosides on the acetylcholine induced peak inward current (IACh) in Xenopus oocytes expressing the α7 nAChR. IACh was measured with a two-electrode voltage clamp technique. In oocytes injected with α7 nAChR copy RNA, quercetin enhanced IACh, whereas quercetin glycosides inhibited IACh. Quercetin glycosides mediated an inhibition of IACh, which increased when they were pre-applied and the inhibitory effects were concentration dependent. The order of IACh inhibition by quercetin glycosides was Rutin≥Rham1>Rham2. Quercetin glycosides-mediated IACh enhancement was not affected by ACh concentration and appeared voltage-independent. Furthermore, quercetin-mediated IACh inhibition can be attenuated when quercetin is co-applied with Rham1 and Rutin, indicating that quercetin glycosides could interfere with quercetin-mediated α7 nAChR regulation and that the number of carbohydrates in the quercetin glycoside plays a key role in the interruption of quercetin action. These results show that quercetin and quercetin glycosides regulate the α7 nAChR in a differential manner.

Chloroform fraction of Euphorbia maculata has antiplatelet activity via suppressing thromboxane B2 formation.

Euphorbia maculata (EM) is a traditionally used antidiarrheal, antibacterial, antifungal and antioxidant agent. However, the effects of EM on platelet activity remain to be elucidated. Therefore, the present study investigated the antiplatelet effect of various EM extract fractions on platelet aggregation in rats. The antiplatelet activity of the EM fractions on collagen or adenosine diphosphate (ADP)­induced platelet aggregation was evaluated in vitro and ex vivo. Thromboxane B2 (TXB2) formation, rat­tail bleeding time and coagulation time were also measured. Among the fractions, the chloroform fraction of EM (CFEM) significantly inhibited ADP­induced platelet aggregation in vitro. Furthermore, oral administration of 50 mg/kg CFEM to rats significantly reduced ADP­induced platelet aggregation without increasing the tail bleeding time or coagulation time. In addition, EM significantly inhibited the level of TXB2 formation in a dose­dependent manner. These results suggest that CFEM exhibits antiplatelet activity, without causing bleeding, via the suppression of TXB2 formation. CFEM may be a type of food which has the potential for preventing cardiovascular disease.

Differential effects of quercetin glycosides on GABAC receptor channel activity.

Quercetin, a representative flavonoid, is a compound of low molecular weight found in various colored plants and vegetables. Quercetin shows a wide range of neuropharmacological activities. In fact, quercetin naturally exists as monomer-(quercetin-3-O-rhamnoside) (Rham1), dimer-(Rutin), or trimer-glycosides [quercetin-3-(2(G)-rhamnosylrutinoside)] (Rham2) at carbon-3 in fruits and vegetables. The carbohydrate components are removed after ingestion into gastrointestinal systems. The role of the glycosides attached to quercetin in the regulation of γ-aminobutyric acid class C (GABAC) receptor channel activity has not been determined. In the present study, we examined the effects of quercetin glycosides on GABAC receptor channel activity by expressing human GABAC alone in Xenopus oocytes using a two-electrode voltage clamp technique and also compared the effects of quercetin glycosides with quercetin. We found that GABA-induced inward current (I GABA ) was inhibited by quercetin or quercetin glycosides. The inhibitory effects of quercetin and its glycosides on I GABA were concentration-dependent and reversible in the order of Rutin ≈ quercetin ≈ Rham 1 > Rham 2. The inhibitory effects of quercetin and its glycosides on I GABA were noncompetitive and membrane voltage-insensitive. These results indicate that quercetin and its glycosides regulate GABAC receptor channel activity through interaction with a different site from that of GABA, and that the number of carbohydrate attached to quercetin might play an important role in the regulation of GABAC receptor channel activity.

Effects of pectin lyase-modified red ginseng extracts in high-fat diet-fed obese mice.

Red ginseng and its extracts have been used as traditional medicines and functional foods in countries worldwide. The aim of this study was to examine the bioavailability of pectin lyase-modified red ginseng extracts (GS-E3D), and the effects of GS-E3D on adipogenesis of 3T3-L1 adipocytes, as well as on metabolic disorders such as hyperglycemia, dyslipidemia, and fatty liver in high-fat diet fed obese C57BL/6 mice. Mice were divided into 5 groups: normal diet group, high fat diet-vehicle group, high fat diet + 0.1 g/kg GS-E3D (0.1-GS-E3D), high fat diet + 0.3 g/kg (0.3-GS-E3D), high fat diet + 1.0 g/kg (1.0-GS-E3D). Treatment of GS-E3D reduced differentiation of 3T3-L1 adipocytes with low cytotoxicity. In the animal model, compared to the high fat diet control, serum glucose, total cholesterol, LDL-cholesterol, HDL-cholesterol, TG, and leptin level were reduced in treatment animals in a dose-dependent manner. In addition, we found that GS-E3D could decrease total hepatic lipid droplets. These results suggest that GS-E3D, as a dietary supplement, has beneficial effects on obesity and may have useful effects in health-care products.

Gintonin, newly identified compounds from ginseng, is novel lysophosphatidic acids-protein complexes and activates G protein-coupled lysophosphatidic acid receptors with high affinity.

Recently, we isolated a subset of glycolipoproteins from Panax ginseng, that we designated gintonin, and demonstrated that it induced [Ca2+]i transients in cells via G protein-coupled receptor (GPCR) signaling pathway(s). However, active components responsible for Ca2+ mobilization and the corresponding receptor(s) were unknown. Active component(s) for [Ca2+]i transients of gintonin were analyzed by liquid chromatography-electrospray ionization-tandem mass spectrometry and ion-mobility mass spectrometry, respectively. The corresponding receptor(s)were investigated through gene expression assays. We found that gintonin contains LPA C18:2 and other LPAs. Proteomic analysis showed that ginseng major latex-like protein and ribonuclease-like storage proteins are protein components of gintonin. Gintonin induced [Ca2+]i transients in B103 rat neuroblastoma cells transfected with human LPA receptors with high affinity in order of LPA2 >LPA5 > LPA1 > LPA3 > LPA4. The LPA1/LPA3 receptor antagonist Ki16425 blocked gintonin action in cells expressing LPA1 or LPA3. Mutations of binding sites in the LPA3 receptor attenuated gintonin action. Gintonin acted via pertussis toxin (PTX)-sensitive and -insensitive G protein-phospholipase C (PLC)-inositol 1,4,5-trisphosphate (IP3)-Ca2+ pathways. However, gintonin had no effects on other receptors examined. In human umbilical vein endothelial cells (HUVECs) gintonin stimulated cell proliferation and migration. Gintonin stimulated ERK1/2 phosphorylation. PTX blocked gintonin-mediated migration and ERK1/2 phosphorylation. In PC12 cells gintonin induced morphological changes, which were blocked by Rho kinase inhibitorY-27632. Gintonin contains GPCR ligand LPAs in complexes with ginseng proteins and could be useful in the development of drugs targeting LPA receptors.

Effects of quercetin on α9α10 nicotinic acetylcholine receptor-mediated ion currents.

Quercetin, one of the flavonoids, is a low molecular weight substance found in fruits and vegetables. Quercetin, like other flavonoids, has a wide range of neuropharmacological actions and antioxidant effects. The α9α10 nicotinic acetylcholine receptor is one of the numerous nicotinic acetylcholine receptors that exist as a heteropentameric form between efferent olivocochlear fibers and hair cells of the cochlea. In this study, we report the effects of quercetin on rat α9α10 nicotinic acetylcholine receptor-mediated ion currents using the two-electrode voltage-clamp technique. Treatment with acetylcholine evoked inward currents (I(ACh)) in oocytes heterologously expressing the α9α10 nicotinic acetylcholine receptor. Quercetin blocked I(ACh) in concentration-dependent and reversible manners, and the blocking effect on I(ACh) was stronger with pre-application than co-application of quercetin. The half maximal inhibitory concentration (IC(50)) of quercetin was 45.4±10.1µM. Quercetin-mediated I(ACh) inhibition was not affected by acetylcholine concentration and was independent of membrane-holding potential. Although the inhibitory effect of quercetin was significantly attenuated in the absence of extracellular Ca(2+), the action of quercetin was independent of extracellular Ca(2+) concentration, indicating that the presence of extracellular Ca(2+) might be needed for quercetin-related effects and might play an important role in quercetin-mediated regulation of the α9α10 nicotinic acetylcholine receptor. These results indicate that quercetin-mediated regulation of the α9α10 nicotinic acetylcholine receptor could provide a molecular basis for quercetin actions at the cellular level.

A simple method for the preparation of crude gintonin from ginseng root, stem, and leaf.

Ginseng has been used as a general tonic agent to invigorate the human body as an adaptogenic agent. In a previous report, we have shown that ginseng contains a novel glycolipoprotein called gintonin. The main function of gintonin is to transiently enhance intracellular free Ca(2+) [Ca(2+)]i levels in animal cells. The previous method for gintonin isolation included multiple steps using organic solvents. In the present report, we developed a simple method for the preparation of crude gintonin from ginseng root as well as stem and leaf, which produced a higher yield of gintonin than the previous one. The yield of gintonin was 0.20%, 0.29%, and 0.81% from ginseng root, stem, and leaf, respectively. The apparent molecular weight of gintonin isolated from stem and leaf through sodium dodecyl sulfate polyacrylamide gel electrophoresis was almost same as that from root but the compositions of amino acids, carbohydrates or lipids differed slightly between them. We also examined the effects of crude gintonin from ginseng root, stem, and leaf on endogenous Ca(2+)-activated Cl- channel (CaCC) activity of Xenopus oocytes through mobilization of [Ca(2+)]i. We found that the order of potency for the activation of CaCC was ginseng root > stem > leaf. The ED50 was 1.4±1.4, 4.5±5.9, and 3.9±1.1 µg/mL for root, stem and leaf, respectively. In the present study, we demonstrated for the first time that in addition to ginseng root, ginseng stem and leaf also contain gintonin. Gintonin can be prepared from a simple method with higher yield of gintonin from ginseng root, stem, and leaf. Finally, these results demonstrate the possibility that ginseng stem and leaf could also be utilized for ginstonin preparation after a simple procedure, rather than being discarded.

Effects of quercetin on human α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor-mediated ion currents.

Quercetin is a low molecular weight flavonoid found in dietary fruits and vegetables. Quercetin, like other flavonoids, has demonstrated neuroprotective effects in vitro and in vivo. However, relatively little is known about how quercetin achieves its neuroprotective abilities. The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor is one of several excitatory receptors, which play an important role in postsynaptic neurotransmission. Over-stimulation of ionotropic glutamate receptor including AMPA receptors is closely associated with excitatory neurotoxicities. In the present study, we investigated the effects of quercetin on the glutamate-induced inward current (IGlu) in Xenopus oocytes that heterologously express human AMPA receptor and stargazin, an auxiliary subunit of AMPA receptor. IGlu was measured using the two-electrode voltage clamp technique. In oocytes injected with cRNAs coding AMPA receptor (GluR1) and stargazin, quercetin inhibited IGlu in a reversible and concentration-dependent manner. The IC50 was 84.9+/-15.0 microM. Quercetin action on IGlu was attenuated by increasing glutamate concentration, and was membrane holding potential-dependent. These results show a possibility that quercetin interacts with AMPA receptor, which was heterologously expressed in Xenopus oocytes and that quercetin action on IGlu of AMPA receptor could be one of contributions of quercetin-mediated neuroprotections.