Clarifying the pharmacological mechanisms of action of Shenfu Decoction on cardiovascular diseases using a network pharmacology approach.

Since the molecular mechanisms underlying in the pathogenesis of cardiovascular diseases (CVD) are extremely complex and have not yet been elucidated in detail, CVD remain the leading cause of death worldwide. Traditional Chinese medicine involves the treatment of disease from an overall perspective, and its therapeutic effects on CVD have been demonstrated. However, the mechanisms contributing to the multiscale treatment of cardiovascular diseases at the systematic level remain unclear. Network pharmacology methods and a gene chip data analysis were integrated and applied in the present study, which was conducted to investigate the potential target genes and related pathways of Shenfu Decoction (SFD) for the treatment of myocardial injury. The gene chip analysis was initially performed, followed by network pharmacology to identify differentially expressed genes (DEG) and a functional enrichment analysis. Protein-protein networks were constructed and a module analysis was conducted. A network analysis was used to identify the target genes of SFD. Regarding the results obtained, 1134 DEG were identified using the STRING website. The module analysis revealed that nine hub genes exhibited ubiquitin-protein ligase activity. Therefore, SFD significantly alters the expression of ubiquitination-related genes and, thus, plays an important therapeutic role in the treatment of heart failure. In conclusion, hub genes may provide a more detailed understanding of the molecular mechanisms of action of as well as candidate targets for SFD therapy.

Use of an electrophysiological technique for stepwise detection of trace agonist constituents of Hochuekkito in Xenopus oocytes injected with serotonin 2C receptor mRNA.

An electrophysiological bioassay was used to isolate the active compound from Hochuekkito (HET), which the current authors previously described as having potent agonist action against serotonin 2C receptors (5-HT2CR). Synthetic 5-HT2CR mRNA was injected into Xenopus oocytes to specifically express these receptors. Crude extracts and purified products were subjected to an electrophysiological bioassay using the voltage clamp method. HET stimulated a 5-HT2CR-induced current response, whereas Juzentaohoto (JTT), which has anti-depressive action similar to that of HET, did not. Current responses were not observed with an extract mixed with five types of herbal medicines common to HET and JTT but were detected with an extract with the five types of herbal medicines found in HET alone (Hoc5). When the responses to each of the five types of Hoc5 were examined, current responses were noted with Cimicifugae rhizoma (CR) and Citrus unshiu Markovich extracts. Since efficacy and the EC50 value were higher for CR, its constituents were separated using three-dimensional high-performance liquid chromatography and the current response at each of the isolated peaks was examined. One constituent displayed a strong response and was identified as a single substance with a molecular weight of 283.1393 based on liquid chromatography/mass spectrometry. These results will contribute to the isolation of 5-HT2CR-stimulating constituents in HET and the identification of trace constituents with agonist action.

The Wakan-yaku Universe: A Useful Authorized Traditional Concept for Developing Novel Therapeutic Categories and Medicinal Drugs.

This review article mentions about the following points, and proposes its importance and positive thinking. 1) Wakan-yaku (Japanese oriental medicines) is covered by the national health insurance system in Japan as therapeutic drugs to be actively used in medical practice to treat illness. 2) Applications of Wakan-yaku is accomplished based on the reliable own theories which are established with long histories. 3) Promotion of studies based on these theories will be highly expected to find novel view points which breaks conventional concepts and to novel standards for developing new medicinal drugs. Although studies based on the reliable Wakan-yaku theories are not advancing satisfactorily till now, the possibilities to obtain the advanced resources for drugs and novel viewpoints for experiments by studies about Wakan-yaku theories are discussed in this review.

Comparing the effects of kamikihito in Japan and kami-guibi-tang in Korea on memory enhancement: working towards the development of a global study.

Traditional medicine is widely used in East Asia, and studies that demonstrate its usefulness have recently become more common. However, formulation-based studies are not globally understood because these studies are country-specific. There are many types of formulations that have been introduced to Japan and Korea from China. Establishing whether a same-origin formulation has equivalent effects in other countries is important for the development of studies that span multiple countries. The present study compared the effects of same-origin traditional medicine used in Japan and Korea in an in vivo experiment. We prepared drugs that had the same origin and the same components. The drugs are called kamikihito (KKT) in Japan and kami-guibi-tang (KGT) in Korea. KKT (500 mg extract/kg/day) and KGT (500 mg extract/kg/day) were administered to ddY mice, and object recognition and location memory tests were performed. KKT and KGT administration yielded equivalent normal memory enhancement effects. 3D-HPLC showed similar, but not identical, patterns of the detected compounds between KKT and KGT. This comparative research approach enables future global clinical studies of traditional medicine to be conducted through the use of the formulations prescribed in each country.

Evidence of Antidepressive Effects of a Wakan-yaku, Hochuekkito, in Depression Model Mice with Learned-Helplessness Behavior.

Wakan-yaku is a type of Japanese and Sino traditional, systematized medical care that has been practiced for hundreds of years. This medicinal system includes many antidepressive prescriptions. One of the candidates is Hochuekkito, although experimental evidence has not yet been established clearly. To obtain evidence, a depression model of learned-helplessness (LH) mice was used. Based on the score of escape failure, an index of the depression degree, mice with a depressive condition were selected to assess Hochuekkito's effects. This selection was significant and effective in the following two points: evaluation of the drug effect under disease conditions and minimization of the number of animals. Treatment with Hochuekkito (1 and 5 g/kg p.o.; estimated galenical amount) for 14 days significantly decreased the depression index, the number of escape failures, and desipramine (10 mg/kg p.o.) suggesting that Hochuekkito has an antidepressive effect.

γ-Mangostin increases serotonin 2A/2C, muscarinic, histamine and bradykinin receptor mRNA expression.

AIM OF THE STUDY: γ-Mangostin is a xanthone found in the fruit hulls of Garcinia mangostana L., which have long been used in Southeast Asia as a traditional medicine for the treatment of abdominal pain, dysentery, wound infections, fever and convulsions. Recent studies have revealed that γ-mangostin exhibits a variety of pharmacological activities, including serotonin 2 (5-HT(2)) receptor antagonism, anti-inflammatory effects and analgesic effects. To explore the mechanism of γ-mangostin responsible for these pharmacological activities, especially its effects on some related receptors, we investigated the effects of γ-mangostin on 5-HT(2), histamine (H(1)) and bradykinin (BK(2)) receptor gene expression in neuroblastoma (NG 108-15) cells in vitro. Additionally, to extend the study of the pharmacological properties, we examined the effect of γ-mangostin on the muscarinic (M(4)) receptor. MATERIALS AND METHODS: NG 108-15 cells were cultured in vitro and treated with γ-mangostin or a 5-HT(2) receptor antagonist (either imipramine or ketanserin). Then, the levels of mRNA for 5-HT(2A/2C) receptors were evaluated by semi-quantitative RT-PCR. The preventive effect of serotonin on the enhancement effects was also revealed. Additionally, the effects of γ-mangostin on the muscarinic, histamine and bradykinin receptors were determined. RESULTS: Chronic application of γ-mangostin at a concentration of 0.1 µM induced a significant increase in the level of 5-HT(2A/2C) receptor mRNA. These effects were prevented by serotonin. Moreover, γ-mangostin up-regulated the M(4), H(1) and BK(2) receptors. CONCLUSION: The ability of γ-mangostin to enhance the expression of 5-HT(2A/2C), muscarinic, histamine and bradykinin receptor mRNA suggests that this compound has antagonistic effects. These pharmacological properties may partly account for the benefits of using mangosteen in the treatment of inflammation, pain and neuropsychiatric symptoms.

BNIP-3: a novel candidate for an intrinsic depression-related factor found in NG108-15 cells treated with Hochu-ekki-to, a traditional oriental medicine, or typical antidepressants.

Wakan-yaku is a type of Japanese and Sino traditional, systematized medical care that has been practiced for hundreds of years. To search for novel intrinsic factors related to the action of antidepressants, we used Hochu-ekki-to (HET), a Wakan-yaku medicine with antidepressive effects. First, we verified the quality of the HET by three-dimensional high-performance liquid chromatography and a cytotoxicity check in NG108-15 cells. We performed a DNA microarray analysis of the gene expression in cells treated with 50 micro/ml HET for more than 20 days. HET enhanced the expression of 125 (2.9%) genes and decreased the expression of 255 (6.0%) genes among the 4277 genes that were tested. The concentration-dependent increase in the expression of BCL2/adenovirus E1B 19-kDa protein-interacting protein 3 (BNIP-3) mRNA was particularly remarkable. A concentration-dependent increase in the expression of BNIP-3 mRNA was also observed when cells were treated with imipramine, mianserin, or milnacipran. These results suggest that BNIP-3 is a candidate for an intrinsic factor related to antidepressive effects and that Wakan-yaku theory may be useful for the identification of other intrinsic functional molecules.

Chotosan enhances macrophage colony-stimulating factor mRNA expression in the ischemic rat brain and C6Bu-1 glioma cells.

Macrophage colony stimulating factor (M-CSF) is a cytokine which has been recently reported to have a neuroprotective effect on ischemic rat brain. In this study, we investigated the effect of chotosan, an oriental medicine, which has been clinically demonstrated to be effective for the treatment of vascular dementia, on M-CSF gene expression in rats with permanent occlusion of bilateral common carotid arteries (P2VO) in vivo and in a C6Bu-1 glioma cell line in vitro. The expression level of M-CSF mRNA in the cerebral cortices of P2VO rats was significantly higher than that in the cerebral cortices of sham-operated animals. Repeated treatment of P2VO rats with chotosan (75 mg/kg per day) for 4 d after P2VO significantly increased the expression level of M-CSF mRNA in the cortex but it had no effect on the expression of beta-actin, granulocyte colony stimulating factor (G-CSF), granulocyte/macrophage colony stimulating factor (GM-CSF) mRNAs. Moreover, the present in vitro studies revealed that chotosan treatment (10-100 mug/ml) of C6Bu-1 glioma cells dose-dependently enhanced M-CSF mRNA expression without affecting the expression of G-CSF, GM-CSF, and inducible nitric oxide synthase mRNAs. The effect of chotosan was reversed by Ro 31-8220 (1 muM), a selective protein kinase C (PKC) inhibitor, but not by H-89 (10 muM), a selective protein kinase A (PKA) inhibitor. These findings suggest that the upregulatory effect of chotosan on M-CSF mRNA expression involves PKC and may play an important role in the anti-vascular dementia action of this formula.

Chotosan, a kampo formula, ameliorates chronic cerebral hypoperfusion-induced deficits in object recognition behaviors and central cholinergic systems in mice.

We previously demonstrated that the Kampo formula chotosan (CTS) ameliorated spatial cognitive impairment via central cholinergic systems in a chronic cerebral hypoperfusion (P2VO) mouse model. In this study, the object discrimination tasks were used to determine if the ameliorative effects of CTS on P2VO-induced cognitive deficits are a characteristic pharmacological profile of this formula, with the aim of clarifying the mechanisms by which CTS enhances central cholinergic function in P2VO mice. The cholinesterase inhibitor tacrine (THA) and Kampo formula saikokeishito (SKT) were used as controls. P2VO impaired object discrimination performance in the object recognition, location, and context tests. Daily administration of CTS (750 mg/kg, p.o.) and THA (2.5 mg/kg, i.p.) improved the object discrimination deficits, whereas SKT (750 mg/kg, p.o.) did not. In ex vivo assays, tacrine but not CTS or SKT inhibited cortical cholinesterase activity. P2VO reduced the mRNA expression of m(3) and m(5) muscarinic receptors and choline acetyltransferase but not that of other muscarinic receptor subtypes in the cerebral cortex. Daily administration of CTS and THA but not SKT reversed these expression changes. These results suggest that CTS and THA improve P2VO-induced cognitive impairment by normalizing the deficit of central cholinergic systems and that the beneficial effect on P2VO-induced cognitive deficits is a distinctive pharmacological characteristic of CTS.

Prevention of kainic acid-induced changes in nitric oxide level and neuronal cell damage in the rat hippocampus by manganese complexes of curcumin and diacetylcurcumin.

Curcumin is a natural antioxidant isolated from the medicinal plant Curcuma longa Linn. We previously reported that manganese complexes of curcumin (Cp-Mn) and diacetylcurcumin (DiAc-Cp-Mn) exhibited potent superoxide dismutase (SOD)-like activity in an in vitro assay. Nitric oxide (NO) is a free radial playing a multifaceted role in the brain and its excessive production is known to induce neurotoxicity. Here, we examined the in vivo effect of Cp-Mn and DiAc-Cp-Mn on NO levels enhanced by kainic acid (KA) and L-arginine (L-Arg) in the hippocampi of awake rats using a microdialysis technique. Injection of KA (10 mg/kg, i.p.) and L-Arg (1000 mg/kg, i.p.) significantly increased the concentration of NO and Cp-Mn and DiAc-Cp-Mn (50 mg/kg, i.p.) significantly reversed the effects of KA and L-Arg without affecting the basal NO concentration. Following KA-induced seizures, severe neuronal cell damage was observed in the CA1 and CA3 subfields of hippocampal 3 days after KA administration. Pretreatment with Cp-Mn and DiAc-Cp-Mn (50 mg/kg, i.p.) significantly attenuated KA-induced neuronal cell death in both CA1 and CA3 regions of rat hippocampus compared with vehicle control, and Cp-Mn and DiAc-Cp-Mn showed more potent neuroprotective effect than their parent compounds, curcumin and diacetylcurcumin. These results suggest that Cp-Mn and DiAc-Cp-Mn protect against KA-induced neuronal cell death by suppression of KA-induced increase in NO levels probably by their NO scavenging activity and antioxidative activity. Cp-Mn and DiAc-Cp-Mn have an advantage to be neuroprotective agents in the treatment of acute brain pathologies associated with NO-induced neurotoxicity and oxidative stress-induced neuronal damage such as epilepsy, stroke and traumatic brain injury.

Preventive effect of chotosan, a Kampo medicine, on transient ischemia-induced learning deficit is mediated by stimulation of muscarinic M1 but not nicotinic receptor.

We have previously shown using a water maze task that transient 2 vessel occlusion (T2VO) induced learning deficit in mice and that the deficit was prevented by pre-treatment of mice with chotosan, a Kampo prescription. In this study, we investigated the mechanism underlying the preventive effect of chotosan on T2VO-induced learning deficit. Chotosan administration 1 h before T2VO operation prevented learning impairment. The extract of Uncaria, a major constituent of chotosan, also had a protective effect on learning impairment in T2VO mice, whereas Uncaria-free chotosan had no beneficial effect on maze performance of T2VO mice. The ameliorative effect of chotosan was blocked by pirenzepine, a muscarinic M1 antagonist, but not by mecamylamine, a nicotinic receptor antagonist. Acetylcholine (ACh) content in the hippocampus of T2VO mice was significantly lower than that in the hippocampus of sham-operated control mice. Chotosan and Uncaria administration attenuated T2VO-induced reduction of ACh levels in the brain. These results suggest that the preventive effect of chotosan on transient ischemia-induced learning impairment is mainly attributable to the effect of Uncaria and that the ameliorative effect is mediated by stimulation of muscarinic M1 receptor.

Social isolation stress-induced oxidative damage in mouse brain and its modulation by majonoside-R2, a Vietnamese ginseng saponin.

Stressors with a physical factor such as immobilization, electric foot shock, cold swim, etc., have been shown to produce oxidative damage to membrane lipids in the brain. In this study, we investigated the effect of protracted social isolation stress on lipid peroxidation activity in the mouse brain and elucidated the protective effect of majonoside-R2, a major saponin component of Vietnamese ginseng, in mice exposed to social isolation stress. Thiobarbituric acid reactive substance levels, one of the end products of lipid peroxidation reaction, were increased in the brains of mice subjected to 6-8 weeks of social isolation stress. Measurements of nitric oxide (NO) metabolites (NO(x)(-)) also revealed a significant increase of NO production in the brains of socially isolated mice. Moreover, the depletion of brain glutathione content, an endogenous antioxidant, in socially isolated animals occurred in association with the rise in lipid peroxidation. The intraperitoneal administration of majonoside-R2 (10-50 mg/kg) had no effect on thiobarbituric acid reactive substances (TBARS), NO, or glutathione levels in the brains of group-housed control mice but it significantly suppressed the increase in TBARS and NO levels and the decrease in glutathione levels caused by social isolation stress. These results suggest that mice subjected to 6-8 weeks of social isolation stress produces oxidative damage in the brain partly via enhancement of NO production, and that majonoside-R2 exerts a protective effect by modulating NO and glutathione systems in the brain.

Suppressive effects of isorhynchophylline on 5-HT2A receptor function in the brain: behavioural and electrophysiological studies.

Isorhynchophylline is a major oxindole alkaloid found in Uncaria species which have long been used in traditional Chinese medicine. Here, we investigated the effects of isorhynchophylline and isorhynchophylline-related alkaloids on 5-hydroxytryptamine (5-HT) receptor-mediated behavioural responses in mice and 5-HT-evoked current responses in Xenopus oocytes expressing 5-HT2A or 5-HT2C receptors. Isorhynchophylline dose-dependently inhibited 5-HT2A receptor-mediated head-twitch but not 5-HT1A receptor-mediated head-weaving responses evoked by 5-methoxy-N,N-dimethyltryptamine. Pretreatment with reserpine, a monoamine-depleting agent, enhanced the head-twitching, but did not influence the effect of isorhynchophylline on the behavioural response. Isocorynoxeine, an isorhynchophylline-related alkaloid in which the configuration of the oxindole moiety is the same as in isorhynchophylline, also reduced the head-twitch response in reserpinized mice over the same dose range as isorhynchophylline, while both rhynchophylline and corynoxeine, stereoisomers of isorhynchophylline and isocorynoxeine, did not. None of the alkaloids tested had an effect on meta-chlorophenylpiperazine-induced hypolocomotion, a 5-HT2C receptor-mediated behavioural response. In experiments in vitro, isorhynchophylline and isocorynoxeine dose-dependently and competitively inhibited 5-HT-evoked currents in Xenopus oocytes expressing 5-HT2A receptors, but had less of a suppressive effect on those in oocytes expressing 5-HT2C receptors. These results indicate that isorhynchophylline and isocorynoxeine preferentially suppress 5-HT2A receptor function in the brain probably via a competitive antagonism at 5-HT2A receptor sites and that the configuration of the oxindole moiety of isorhynchophylline is essential for their antagonistic activity at the 5-HT2A receptor.

Choto-san, a Kampo formula, improves chronic cerebral hypoperfusion-induced spatial learning deficit via stimulation of muscarinic M1 receptor.

A recent double-blind and placebo-controlled study demonstrated a beneficial effect of Choto-san, a Kampo (traditional medicine of Japan) formula, on cognitive impairment in patients with vascular dementia. However, the neuronal mechanism underlying the therapeutic effects of this formula remains to be clarified. Using a chronic cerebral hypoperfusion model, we investigated the effect of Choto-san on cognitive dysfunction in mice to clarify its mechanism of actions. Chronic cerebral hypoperfusion was induced by permanent occlusion of both the common carotid arteries (2VO). Choto-san and Uncaria, a major constituent of Choto-san, caused an improvement in 2VO-induced learning deficits, whereas Uncaria-free Choto-san did not. The effects of Choto-san and Uncaria were blocked by pirenzepine, a selective muscarinic M1 antagonist. In a tube-dominance test, 2VO induced increased rates of assertive behavior in mice. 2VO mice administered Choto-san showed significantly reduced rates of assertive behavior compared to vehicle-treated controls, whereas Uncaria-free Choto-san and Uncaria had little effect on 2VO-induced assertive behavior. 2VO caused a significant decrease in the level of acetylcholine (ACh) contents in the brain, and the daily administration of Choto-san or Uncaria raised the ACh level to that in the sham-operated controls. These results suggest that Choto-san has an ameliorating effect on the spatial memory deficit caused by chronic hypoperfusion, and that the effect is mainly attributable to Uncaria. Moreover, it was suggested that the effects of Choto-san and Uncaria are at least partly mediated by stimulation of the muscarinic M1 receptor.

Corymine potentiates NMDA-induced currents in Xenopus oocytes expressing NR1a/NR2B glutamate receptors.

Previous studies demonstrated that corymine, an indole alkaloid isolated from the leaves of Hunter zeylanica, dose-dependently inhibited strychnine-sensitive glycine-induced currents. However, it is unclear whether this alkaloid can modulate the function of the N-methyl-D-aspartate (NMDA) receptor on which glycine acts as a co-agonist via strychnine-insensitive glycine binding sites. This study aimed to evaluate the effects of corymine on NR1a/NR2B NMDA receptors expressed in Xenopus oocytes using the two-electrode voltage clamp technique. Corymine significantly potentitated the NMDA-induced currents recorded from oocytes on days 3 and 4 after cRNA injection but it showed no effect when the current was recorded on days 5 and 6. The potentiating effect of corymine on NMDA-induced currents was induced in the presence of a low concentration of glycine (< or =0.1 microM). Spermine significantly enhanced the potentiating effect of corymine observed in the oocytes on days 3 and 4, while the NMDA-receptor antagonist 2-amino-5-phosphonopentanone (AP5) and the NMDA-channel blocker 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801) reversed the effect of corymine. On the other hand, the nonspecific chloride channel blocker 4,4-di-isothiocyano stilbene-2,2-disulfonoc acid (DIDS) had no effect on the corymine potentiation of NMDA currents. There was no good correlation between corymine- and spermine-induced potentiation of the NMDA-current response in Xenopus oocytes. These results suggest that corymine potentiates the NMDA-induced currents by interacting with a site different from the spermine binding site.

The effects of Choto-san on the mRNA expression of Alzheimer's disease related factors in the permanent ischemic rat brain.

Choto-san is a Kampo medicines that has been used clinically for the treatment of dementia. We measured the mRNA expressions of some factors related to Alzheimer's disease in a dementia model rat brain. The expressions of beta-amyloid precursor protein, gamma-secretase, alpha7 nicotinic acetylcholine receptor, neprilysin, and insulin degrading enzyme (IDE) were significantly increased on day 4 after permanent occlusion of the bilateral common carotid arteries (2VO). Choto-san inhibited the enhancement of IDE expression caused by 2VO, although it failed to show any effects on the expressions of the other molecules. These results suggest that Choto-san may produce a state in which it is not necessary to induce IDE expression to demonstrate the anti-dementia effects.

Effects of Choto-san and its related constituents on endogenous antioxidant systems.

We previously reported that Choto-san acts as an antioxidant and cytoprotective agents against H2O2-induced oxidative damage in NG108-15 cells, and the effect is due at least partly to the phenolic compounds. To further investigate the detail mechanisms of this cytoprotection effects of Choto-san and related compounds on enzyme activities of antioxidant systems were examined. Choto-san (5-100 microg/ml) and Chotoko (5-100 mg/ml) stimulated the activity of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPX). These also increased the level of glutathione. Although Choto-san without Chotoko (w/o CKO) did not show the effects on SOD and catalase, GPX activity and glutathion content also, but weakly, stimulated by w/o CKO. The effects of phenolic compounds, epicatechin, caffeic acid and quercetin were also investigated. Epicatechin stimulated catalase, GPX and glutathion content, but not SOD. On the other hand, caffeic acid stimulated SOD activity but had no effects on others. Quercetin stimulated all, although intensities were different among. These results suggest that simultaneous induction of cellular antioxidant defense systems by Choto-sam and its related constituents may be an important mechanisms underlying the protective effects of Choto-san on ischemia-induced neuronal cells injury, and the characteristics of the stimulative effects of phenolic compounds were depend on enzymes.

Antioxidant and free radical-scavenging activity of Choto-san and its related constituents.

The antioxidant properties of Choto-san and its related constituents such as Chotoko and Choto-san without Chotoko, and phenolic compounds contained in Chotoko such as epicatechin, caffeic, acid and quercetin were evaluated. In the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging assay, the scavenging activity of Chotoko (IC(50) 14.3 microg/ml) was found to be higher than that of Choto-san (IC(50) 206.2 microg/ml) and Choto-san without Chotoko (IC(50) 244.3 microg/ml). Epicatechin (IC(50) 10.4 microM), caffeic acid (IC(50) 13.8 microM), and quercetin (IC(50) 7.1 microM) also revealed scavenging activity against DPPH radicals. Choto-san (IC(50) 67.7 microg/ml) exhibited stronger inhibitory activity against superoxide anion formation than Choto-san without Chotoko (IC(50) 92.4 microg/ml) but weaker activity than Chotoko (IC(50) 18.3 microg/ml). The generation of superoxide anion was also inhibited by epicatechin (IC(50) 175.2 microM), caffeic acid (IC(50) 141.7 microM), and quercetin (IC(50) 18.7 microM). In a hydroxyl radical-scavenging experiment, Choto-san (IC(50) 2.4 mg/ml), Chotoko (IC(50) 2.2 mg/ml), Choto-san without Chotoko (IC(50) 2.8 mg/ml), epicatechin (IC(50) 3.9 mM), caffeic acid (IC(50) 3.6 mM), and quercetin (IC(50) 1.9 mM) exhibited activity. In NG108-15 cells, when added simultaneously with H(2)O(2) (500 microM), Choto-san (250 microg/ml), Chotoko (250 microg/ml), Choto-san without Chotoko (500 microg/ml), epicatechin (200 microM), caffeic acid (200 microM), and quercetin (200 microM) effectively protected cells from oxidative damage. In conclusion, the present results provide evidence that Choto-san acts as an antioxidant and cytoprotective agent against oxidative damage, which is due at least partly to the phenolic compounds contained in Chotoko.

Manganese complexes of curcumin and its derivatives: evaluation for the radical scavenging ability and neuroprotective activity.

In this study, three manganese complexes of curcumin (Cp) and related compounds, diacetylcurcumin (AcylCp) and ethylenediamine derivative (CpED), were synthesized and evaluated in vitro for antilipid peroxidation and superoxide dismutase activity. The manganese complexes exhibited a great capacity to protect brain lipids against peroxidation with IC50 of 6.3-26.3 microM. All manganese complexes showed much greater SOD activity than their corresponding antioxidant ligands as well as trolox with IC50 values of 8.9-29.9 microM. AcylCp and curcumin manganese complexes (AcylCpCpx and CpCpx) also gave the highest inhibitory activity to H2O2-induced cell damage (oxidative stress) at 0.1 microg/ml (< 0.2 microM) in NG108-15 cells, which were more potent than curcumin and related compounds. The neuropharmacological tests in mice supported the idea that the SOD mimicking complexes were able to penetrate to the brain as well as their role in the modulation of brain neurotransmitters under the aberrant conditions. The complexes significantly improved the learning and memory impairment induced by transient ischemic/reperfusion. AcylCpCpx, CpCpx, and CpEDCpx showed significant protection at 6.25, 25, and 50 mg/kg (i.p.), respectively, whereas manganese acetate and curcumin had no effect at doses of 50 mg/kg. In addition, treatment of AcylCpCpx and curcumin significantly attenuated MPTP-induced striatal dopamine depletion in mice, which was in accordance with the increase in the density of dopaminergic neurons when compared with MPTP-treated mice. These results support the important role of manganese in importing SOD activity and consequently, the enhancement of radical scavenging activity. AcylCpCpx and CpCpx seem to be the most promising neuroprotective agents for vascular dementia.

Pharmacological evidence for antidementia effect of Choto-san (Gouteng-san), a traditional Kampo medicine.

To clarify the clinical efficacy of one of the traditional medicines in the treatment of patients with vascular dementia, we investigated the pharmacological activities of Choto-san in animal models. Pretreatment with Choto-san (0.75-6.0 g/kg po), a component herb, Chotoko (75-600 mg/kg po), and indole alkaloids and phenolic fractions of Chotoko prevented ischemia-induced impairment of spatial learning behaviour in water maze performance of mice. A single administration of Choto-san (0.5 to 6.0 g/kg po) or Chotoko (Uncaria genus) produced a dose-dependent antihypertensive effect in spontaneously hypertensive rats (SHR) and partly inhibited the induction of the apoplexy in stroke-prone SHR (SHR-SP). Choto-san, Chotoko, and its phenolic constituents, (-)epicatechin and caffeic acid, significantly protected NG108-15 cells from injury induced by H(2)O(2) exposure in vitro and also inhibited lipid peroxidation in the brain homogenate. Indole alkaloids, rhynchophylline and isorhynchophylline (1-100 microM), reversibly reduced N-methyl-D-aspartate (NMDA)-induced current concentration dependently in NMDA receptor-expressed Xenopus oocytes. These results suggest that antidementia effects of Choto-san are due to antihypertensive, free radical scavenging and antiexcitotoxic effects, which are attributed at least partly to phenolic compounds and indole alkaloids contained in Chotoko.