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Find video protocols related to scientific articles indexed in Pubmed.
The Extract of Ziziphus jujuba Fruit (Jujube) Induces Expression of Erythropoietin Via Hypoxia-Inducible Factor-1? in Cultured Hep3B Cells.
Planta Med.
PUBLISHED: 09-03-2014
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The fruit of Ziziphus jujuba Mill., known as jujube or Chinese date, is commonly consumed as health supplement or herbal medicine worldwide. To study the beneficial role of jujube in enhancing hematopoietic function, we investigated its roles on the expression of erythropoietin in cultured Hep3B human hepatocellular carcinoma cells. Application of chemically standardized jujube water extract stimulated erythropoietin expression in a dose-dependent manner, with the highest response by ~?100?% of increase. A plasmid containing hypoxia response element, a critical regulator for erythropoietin transcription, was transfected into Hep3B cells. Application of jujube water extract onto the transfected cells induced the transcriptional activity of the hypoxia response element. To account for its transcriptional activation, the expression of hypoxia-inducible factor-1? was increased after treatment with jujube water extract: the increase was in both mRNA and protein levels. These results confirmed the hematopoietic function of jujube in the regulation of erythropoietin expression in liver cells.
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Inhibiting ?-Amyloid-Associated Alzheimer's Pathogenesis In Vitro and In Vivo by a Multifunctional Dimeric Bis(12)-hupyridone Derived from Its Natural Analogue.
J. Mol. Neurosci.
PUBLISHED: 09-01-2014
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Fibrillar aggregates of ?-amyloid protein (A?) is the main constituent of senile plaques and considered to be one of the causative events in the pathogenesis of Alzheimer's disease (AD). Compounds that could inhibit the formation of A? fibrils and block A? fibrils-associated toxicity may have therapeutic potential to combat AD. Bis(12)-hupyridone (B12H) is a multifunctional homodimer derived from huperzine A, which is an anti-AD drug in China. In the current study, the inhibitory effect of B12H on the formation of A? fibrils and their associated toxicity was investigated both in vitro and in vivo. By using Thioflavin T fluorescence assay, we found that B12H (0.3-3 ?M) directly inhibited A? fibrils formation following co-incubation of B12H and A?1-40 at 37 °C for 6 days in vitro. However, huperzine A, at the same concentrations, did not show significant inhibitory effect on A?1-40 fibrils formation. Moreover, B12H markedly reduced A?1-40-induced cytotoxicity in cultured SH-SY5Y cells, as evidenced by the increase in cell viability, the decrease in lactate dehydrogenase release, and the reduction of apoptotic nuclei. Most importantly, B12H (0.2 and 0.4 mg/kg) reduced intracerebroventricular A?1-40 infusion-induced cognitive and memory impairments in rats, as evidenced by the decrease in escape latency and the increase in the spatial bias in Morris water maze test along with increasing choline acetyltransferase activity and decreasing acetylcholinesterase activity. Collectively, our study provided novel sights into the potential application of B12H in AD treatment.
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Evaluation of Hemostatic and Anti-inflammatory Activities of Extracts from Different Lagochilus Species in Experimental Animals: Comparison of Different Extractives and Sources.
Phytother Res
PUBLISHED: 08-30-2014
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Different members of Lagochilus genus have been used in folkloric medicine to treat hemorrhages and inflammation. However, only a few species of them have received scientific attention supporting their efficacy. Here, the hemostatic and antiinflammatory activities of five Lagochilus species were determined and compared by using in vivo assays. The results showed that the extracts of Lagochilus lanatonodus and Lagochilus diacanthophyllus showed better hemostatic activities among five species. The high doses of L.?lanatonodus extracts were able to shorten the values of thrombin time, activated partial thromboplastin time and prothrombin time in a rat model. Moreover, the extracts of L.?lanatonodus and L.?diacanthophyllus showed strong inhibitory effects on the acute phase of inflammation in both xylene-induced ear edema mouse model and carrageenan-induced paw edema rat model. In parallel, the treatment of these extracts modulated the expressions of those inflammatory parameters, that is, nitric oxide, prostaglandin E2 , inducible nitric oxide synthase, malondialdehyde and superoxide dismutase. L.?lanatonodus and L.?diacanthophyllus showed better hemostatic and antiinflammatory activities in several test models: these results therefore supported the folkloric utilization. L.?lanatonodus was found to be the most active Lagochilus species. Copyright © 2014 John Wiley & Sons, Ltd.
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The sulfur-fumigation reduces chemical composition and biological properties of Angelicae Sinensis Radix.
Phytomedicine
PUBLISHED: 07-31-2014
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Angelica Sinensis Radix (roots of Angelica sinensis; ASR) is a popular herbal supplement in China for promoting blood circulation. Today, sulfur-fumigation is commonly used to treat ASR as a means of pest control; however, the studies of sulfur-fumigation on the safety and efficacy of ASR are very limited. Here, we elucidated the destructive roles of sulfur-fumigation on ASR by chemical and biological assessments. After sulfur-fumigation, the chemicals in ASR were significantly lost. The biological activities of anti-platelet aggregation, induction of NO production and estrogenic properties were compared between the water extracts of non-fumigated and sulfur-fumigated ASR. In all cases, the sulfur-fumigation significantly reduced the biological properties of ASR. In addition, application of water extract deriving from sulfur-fumigated ASR showed toxicity to cultured MCF-7 cells. In order to ensure the safety and to achieve the best therapeutic effect, it is recommended that sulfur-fumigation is an unacceptable approach for processing herbal materials.
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An application of target profiling analyses in the hepatotoxicity assessment of herbal medicines: comparative characteristic fingerprint and bile acid profiling of Senecio vulgaris L. and Senecio scandens Buch.-Ham.
Anal Bioanal Chem
PUBLISHED: 06-11-2014
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The toxicity assessment of herbal medicines is important for human health and appropriate utilization of these medicines. However, challenges have to be overcome because of the complexity of coexisting multiple components in herbal medicines and the highly interconnected organismal system. In this study, a target profiling approach was established by combining the characteristic fingerprint analysis of herbal chemicals with potential toxicity through a precursor ion scan-based mass spectroscopy and the target profiling analysis of biomarkers responsible for the toxicity. Through this newly developed approach, the comparative hepatotoxicity assessment of two herbal medicines from the same genus, Senecio vulgaris L. and Senecio scandens Buch.-Ham, was performed. Significant differences were found between the two species in their chemical markers (i.e., pyrrolizidine alkaloids) and biomarkers (i.e., bile acids) responsible for their toxicities. This result was consistent with the conventional toxicity assessment conducted by histopathological examination and clinical serum index assay on experimental animal models. In conclusion, this study provided a new approach for the hepatotoxicity assessment of herbal medicines containing pyrrolizidine alkaloids, which are widely distributed in various herbal medicines. The target profiling approach may shed light on the toxicity assessment of other herbal medicines with potential toxicity.
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Identification of Angelica oil as a suppressor for the biological properties of Danggui Buxue tang: a Chinese herbal decoction composes of Astragali Radix and Angelica Sinensis Radix.
J Ethnopharmacol
PUBLISHED: 05-05-2014
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Danggui Buxue Tang (DBT), a Chinese herbal decoction commonly used in treating women?s ailments, contains two herbs: Angelica Sinensis Radix (ASR) and Astragali Radix (AR). Traditionally, ASR had to be pre-treated with yellow wine before the herbal preparation, which reduced the amount of volatile oil in water extract of ASR and DBT, and meanwhile the volatile oil-reduced DBT processed better bioactivities in cell cultures. The present study aimed to investigate the effect of volatile oil from ASR (Angelica oil) on the solubility of AR-derived ingredients and the biological properties of DBT.
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A Chemically Standardized Extract of Ziziphus jujuba Fruit (Jujube) Stimulates Expressions of Neurotrophic Factors and Anti-oxidant Enzymes in Cultured Astrocytes.
Phytother Res
PUBLISHED: 04-15-2014
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The fruit of Ziziphus jujuba Mill., known as jujube or Chinese date, is commonly consumed as a health supplement worldwide. To study the role of jujube in brain benefits, the expression of neurotrophic factors and anti-oxidant enzymes in the jujube-treated cultured astrocytes was determined. Application of a chemical standardized water extract of jujube in cultured astrocytes for 24?h stimulated the expressions of nerve growth factor, brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor in a concentration-dependent manner. The pre-treatment with H89, a protein kinase A inhibitor, attenuated the jujube-induced expression of neurotrophic factors. In parallel, the treatment of jujube water extract induced the transcriptional expressions of the enzymes responsible for anti-oxidation, i.e.
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Functions of Danggui Buxue Tang, a Chinese Herbal Decoction Containing Astragali Radix and Angelicae Sinensis Radix, in Uterus and Liver are Both Estrogen Receptor-Dependent and -Independent.
Evid Based Complement Alternat Med
PUBLISHED: 04-09-2014
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Danggui Buxue Tang (DBT), a herbal decoction containing Astragali Radix (AR) and Angelicae Sinensis Radix (ASR), has been used in treating menopausal irregularity in women for more than 800 years in China. Pharmacological results showed that DBT exhibited significant estrogenic properties in vitro, which therefore suggested that DBT could activate the nuclear estrogen receptors. Here, we assessed the estrogenic properties of DBT in an ovariectomized in vivo rat model: DBT was applied to the ovariectomized rats for 3 days. The application of DBT did not alter the weight of uterus and liver, as well as the transcript expression of the proliferation markers including the estrogen receptors ? and ?. However, DBT stimulated the transcript expression of the estrogen responsive genes. In addition, the inductive role of DBT on the expression of members of the aryl hydrocarbon receptor family in uterus and liver of ovariectomized rats was confirmed. These responses of DBT however were clearly distinct from the response pattern detectable here for 17?-estradiol. Therefore, DBT exhibited weak, but significant, estrogenic properties in vivo; however, some of its activities were independent of the estrogen receptor. Thus, DBT could be an exciting Chinese herbal decoction for an alternative treatment of hormone replacement therapy for women in menopause without subsequent estrogenic side effects.
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Metabolomic and genomic evidence for compromised bile acid homeostasis by senecionine, a hepatotoxic pyrrolizidine alkaloid.
Chem. Res. Toxicol.
PUBLISHED: 04-01-2014
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Pyrrolizidine alkaloids (PAs) are among the most hepatotoxic natural products that produce irreversible injury to humans via the consumption of herbal medicine and honey, and through tea preparation. Toxicity and death caused by PA exposure have been reported worldwide. Metabolomics and genomics provide scientific and systematic views of a living organism and have become powerful techniques for toxicology research. In this study, senecionine hepatotoxicity on rats was determined via a combination of metabolomic and genomic analyses. From the global analysis generated from two omics data, the compromised bile acid homeostasis in vivo was innovatively demonstrated and confirmed. Serum profiling of bile acids was altered with significantly elevated conjugated bile acids after senecionine exposure, which was in accordance with toxicity. Similarly, the hepatic mRNA levels of several key genes associated with bile acid metabolism were significantly changed. This process included cholesterol 7-? hydroxylase, bile acid CoA-amino acid N-acetyltransferase, sodium taurocholate cotransporting polypeptide, organic anion-transporting polypeptides, and multidrug-resistance-associated protein 3. In conclusion, a cross-omics study provides a comprehensive analysis method for studying the toxicity caused by senecionine, which is a hepatotoxic PA. Moreover, the change in bile acid metabolism and the respective transporters may provide a new PA toxicity mechanism.
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The volatile oil of Nardostachyos Radix et Rhizoma inhibits the oxidative stress-induced cell injury via reactive oxygen species scavenging and Akt activation in H9c2 cardiomyocyte.
J Ethnopharmacol
PUBLISHED: 03-03-2014
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Nardostachyos Radix et Rhizoma (NRR; the root and rhizome of Nardostachys jatamansi DC.) is a well-known medicinal herb widely used in Chinese, Uyghur and Ayurvedic medicines for the treatment of cardiovascular disorders. The oxidative stress-induced cardiomyocyte loss is the major pathogenesis of heart disorders. Here, the total volatile oil of NRR was isolated, and its function in preventing the cell death of cardiomyocyte was demonstrated.
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Metabonomic analysis of water extracts from different angelica roots by ¹H-nuclear magnetic resonance spectroscopy.
Molecules
PUBLISHED: 02-25-2014
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Angelica Radix, the roots of the genus Angelica, has been used for more than 2,000 years as a traditional medicine in Eastern Asia. The Chinese Pharmacopoeia records more than 100 herbal formulae containing Angelica roots. There are two common sources of Angelica roots, Angelica sinensis from China and A. gigas from Korea. The two species of Angelica roots differ in their chemical compositions, pharmacological properties and clinical efficacy. ¹H-NMR metabolic profiling has recently emerged as a promising quality control method for food and herbal chemistry. We explored the use of ¹H-NMR metabolic profiling for the quality control of Angelica Radix. Unlike previous work, we performed the metabolic profiling on hot water extracts, so as to mimic the clinically relevant preparation method. Unsupervised principle component analyses of both the full spectral profile and a selection of targeted molecules revealed a clear differentiation of three types of Angelica roots. In addition, the levels of 13 common metabolites were measured. Statistically significant differences in the levels of glucose, fructose and threonine were found between different sources of Angelica. Ferulic acid, a marker commonly used to evaluate Angelica root, was detected in our samples, but the difference in ferulic acid levels between the samples was not statistically significant. Overall, we successfully applied ¹H-NMR metabolic profiling with water extraction to discriminate all three sources of Angelica roots, and obtained quantitative information of many common metabolites.
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The standardized extract of Ziziphus jujuba fruit (jujube) regulates pro-inflammatory cytokine expression in cultured murine macrophages: suppression of lipopolysaccharide-stimulated NF-?B activity.
Phytother Res
PUBLISHED: 02-15-2014
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The fruit of Ziziphus jujuba Mill., known as jujube or Chinese date, is commonly consumed as a health supplement or herbal medicine worldwide. To study the beneficial role of jujube in regulating immune response, we investigated its roles on the expressions of pro-inflammatory cytokines in cultured macrophages. Application of chemically standardized jujube water extract for 24?h stimulated the transcriptional expression of interleukin (IL)-1?, IL-6, and tumor necrosis factor (TNF)-? in cultured RAW 264.7 macrophages. In contrast, the pretreatment with jujube water extract suppressed the expression of IL-1? and IL-6, but not for TNF-? in lipopolysaccharide (LPS)-stimulated macrophages. The IL-1? and IL-6 cytokines in LPS-induced macrophages were suppressed by jujube water extract in both mRNA and protein levels. In parallel, the inhibition of jujube water extract on the transcriptional activity of nuclear factor-kappa B was revealed in LPS-induced macrophages. These results verified the bidirectional immune-modulatory roles of jujube by regulating the expressions of pro-inflammatory cytokines in macrophages. Copyright © 2014 John Wiley & Sons, Ltd.
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Synergistic inhibition on acetylcholinesterase by the combination of berberine and palmatine originally isolated from Chinese medicinal herbs.
J. Mol. Neurosci.
PUBLISHED: 02-12-2014
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Alzheimer's disease is a multi-factorial neurodegenerative disorder devastatingly affecting the aged population worldwide. Previous studies have shown that medicinal herbs used in traditional Chinese medicine might be benefit to Alzheimer's disease patients. Berberine and palmatine, two isoquinoline alkaloids found in several medicinal herbs, were used for memory enhancement in China. In this study, the inhibitory effects of combined berberine and palmatine on acetylcholinesteras were evaluated using recombinant human acetylcholinesterase. Our results showed that the combination of berberine and palmatine inhibited acetylcholinesterase in a mixed competitive pattern. By the median-effect principle, the calculated combination index of the combination was less than 1, suggesting that berberine and plamatine inhibited acetylcholinesterase synergistically. Furthermore, the drug-reducing index of berberine and palmatine were 2.98 and 2.66, respectively. Taken together, the results showed that the combination of the two alkaloids might potentially be developed as a more effective therapeutic strategy for Alzheimer's disease patients.
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A standardized extract of the fruit of Ziziphus jujuba (Jujube) induces neuronal differentiation of cultured PC12 cells: a signaling mediated by protein kinase A.
J. Agric. Food Chem.
PUBLISHED: 02-12-2014
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The fruit of Ziziphus jujuba Mill., known as Chinese date or jujube, is consumed as a health supplement worldwide. To study the role of jujube in brain benefits, its effects on neuronal differentiation of PC12 cells were studied. Application of jujube water extract induced neurite outgrowth of PC12 cells, >25% of which were differentiated; this effect was similar to that of nerve growth factor. In parallel, the expressions of neurofilaments (NFs) in jujube-treated cultures showed a dose-dependent increase, with the highest inductions by ?150% for NF68 and NF160 and by ?100% for NF200. Application of H89, a protein kinase A inhibitor, attenuated jujube-induced neurite outgrowth of the cultures. Besides, using jujube extract induced the phosphorylation of cAMP responsive element binding protein on PC12 cells, which was blocked by H89. These results support the use of jujube as a food supplement for the prevention of neurodegenerative diseases in which neurotrophin deficiency is involved.
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Hydrolysis of Glycosidic Flavonoids during the Preparation of Danggui Buxue Tang: An Outcome of Moderate Boiling of Chinese Herbal Mixture.
Evid Based Complement Alternat Med
PUBLISHED: 02-07-2014
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Chemical change during boiling of herbal mixture is a puzzle. By using Danggui Buxue Tang (DBT), a herbal decoction that contains Astragali Radix (AR) and Angelicae Sinensis Radix (ASR), we developed a model in analyzing the hydrolysis of flavonoid glycosides during the boiling of herbal mixture in water. A proper preparation of DBT is of great benefit to the complete extraction of bioactive ingredients. Boiling of DBT in water increased the solubility of AR-derived astragaloside IV, calycosin, formononetin, calycosin-7-O- ? -D-glucoside, and ononin in a time- and temperature-dependent manner: the amounts of these chemicals reached a peak at 2?h. The glycosidic resides of AR, calycosin-7-O- ? -D-glucoside, and ononin could be hydrolyzed during the moderate boiling process to form calycosin and formononetin, respectively. The hydrolysis efficiency was strongly affected by pH, temperature, and amount of herbs. Interestingly, the preheated herbs were not able to show this hydrolytic activity. The current results supported the rationality of ancient preparation of DBT in boiling water by moderate heat.
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Ferulic acid enhances the chemical and biological properties of astragali radix: a stimulator for danggui buxue tang, an ancient Chinese herbal decoction.
Planta Med.
PUBLISHED: 01-31-2014
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Danggui buxue tang, an ancient formula composed of astragali radix and Angelicae sinensis radix, has been used for treating menopausal irregularity in women for more than 800 years in China. In danggui buxue tang, the complete functions of astragali radix require the assistance of Angelicae sinensis radix, and both herbs have to work harmoniously in order to achieve the maximal therapeutic purposes. In order to analyze the relationship of the two herbs, the role of ferulic acid, a major chemical within Angelicae sinensis radix, in chemical and biological properties of astragali radix was determined. Using ferulic acid in the extraction of astragali radix, the amounts of astragaloside IV, calycosin, and formononetin were increased in the final extract; however, the astragali radix polysaccharide showed a minor increase. The chemical-enriched astragali radix extract showed robust induction in osteogenic and estrogenic activities in cultured osteosarcoma MG-63 and breast MCF-7 cells. However, ferulic acid itself did not show such biological responses. The current results strongly suggest that Angelicae sinensis radix-derived ferulic acid is a positive regulator for danggui buxue tang, which enhanced the solubilities of active ingredients derived from astragali radix, and which therefore increased the biological efficacies of danggui buxue tang.
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Presenilin-1 influences processing of the acetylcholinesterase membrane anchor PRiMA.
Neurobiol. Aging
PUBLISHED: 01-26-2014
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Presenilin-1 (PS1) is the catalytic component of the ?-secretase complex. In this study, we explore if PS1 participates in the processing of the cholinergic acetylcholinesterase (AChE). The major AChE variant expressed in the brain is a tetramer (G(4)) bound to a proline-rich membrane anchor (PRiMA). Overexpression of the transmembrane PRiMA protein in Chinese hamster ovary cells expressing AChE and treated with the ?-secretase inhibitor N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester have enabled us to study whether, through its ?-secretase activity, PS1 participates in the processing of PRiMA-linked AChE. ?-Secretase inhibition led to a notable increase in the level of PRiMA-linked AChE, suggesting that ?-secretase is involved in the cleavage of PRiMA. We demonstrate that cleavage of PRiMA by ?-secretase results in a C-terminal PRiMA fragment. Immunofluorescence labeling allowed us to identify this PRiMA fragment in the nucleus. Moreover, we have determined changes in the proportion of the raft-residing AChE-PRiMA in a PS1 conditional knockout mouse. Our results are of interest as both enzymes have therapeutic relevance for Alzheimer's disease.
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Rapid identification of plant materials by wooden-tip electrospray ionization mass spectrometry and a strategy to differentiate the bulbs of Fritillaria.
Anal. Chim. Acta
PUBLISHED: 01-19-2014
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The counterfeit plant products, especially by using incorrect plant materials in pharmaceutical industry, have become a global problem. The plant materials belonging to closely related species but differing in medicinal properties are difficult to be identified. Here, a novel and generally applicable approach to identify the sources of plant materials was developed, which was based on the use of wooden-tip electrospray ionization mass spectrometry (wooden-tip ESI-MS) and multivariate statistical analysis of unidentified MS features (non-targeted). Using this approach, six officinal species of Fritillariae Cirrhosae Bulbus had been successfully differentiated. In addition, Fritillariae Pallidiflorae Bulbus, a common adulterant of Fritillariae Cirrhosae Bulbus, was also identified by using the strategy reported here. Compared with DNA phylogenetic trees, our approach provided finer resolution in distinguishing the closely related Fritillaria species. By combining wooden-tip ESI-MS and multivariate statistical analysis, a useful method was developed here for rapid identification of the sources of herbs, which showed promising perspectives in tracking the supply chain of pharmaceutical suppliers.
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Authentication of Bulbus Fritillariae Cirrhosae by RAPD-derived DNA markers.
Molecules
PUBLISHED: 01-17-2014
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Bulbus Fritillariae is the most commonly used antitussive herb in China. Eleven species of Fritillaria are recorded as Bulbus Fritillariae in the Chinese Pharmacopoeia. Bulbus Fritillariae Cirrhosae is a group of six Fritillaria species with higher efficiency and lower toxicity derived mainly from wild sources. Because of their higher market price, five other Fritillaria species are often sold deceptively as Bulbus Fritillariae Cirrhosae in the herbal market. To ensure the efficacy and safety of medicinal herbs, the authentication of botanical resources is the first step in quality control. Here, a DNA based identification method was developed to authenticate the commercial sources of Bulbus Fritillariae Cirrhosae. A putative DNA marker (0.65 kb) specific for Bulbus Fritillariae Cirrhosae was identified using the Random Amplified Polymorphic DNA (RAPD) technique. A DNA marker representing a Sequence Characterized Amplified Region (SCAR) was developed from a RAPD amplicon. The SCAR marker was successfully applied to differentiate Bulbus Fritillariae Cirrhosae from different species of Fritillaria. Additionally, the SCAR marker was also useful in identifying the commercial samples of Bulbus Fritillariae Cirrhosae. Our results indicated that the RAPD-SCAR method was rapid, accurate and applicable in identifying Bulbus Fritillariae Cirrhosae at the DNA level.
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Quality Control of Danggui Buxue Tang, a Traditional Chinese Medicine Decoction, by (1)H-NMR Metabolic Profiling.
Evid Based Complement Alternat Med
PUBLISHED: 01-15-2014
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Danggui Buxue Tang (DBT) is one of the simplest traditional Chinese medicine (TCM) decoctions, first described in China in 1247 AD. DBT is composed of 2 herbs, Astragali Radix (AR) and Angelica Sinensis Radix (ASR), boiled together in a 5?:?1 ratio. Clinically, DBT is prescribed to women as a remedy for menopausal symptoms. Here, H-NMR metabolic profiling was conducted for DBT and the water extracts of AR or ASR, to evaluate the potential of this chemical profiling method for quality control of the herbal decoction. Principal component analysis (PCA) showed that DBT could be readily distinguished from the water extracts of its constituent herbs by the metabolic profiles. More interestingly, the metabolic profile of DBT was not a simple sum of that of AR and ASR. Asparagine was found at significantly higher concentration in DBT than that in either AR or ASR extract, contributing mainly to the discrimination of DBT sample. In addition, we employed the same method to profile a commercial DBT powder, verifying its authenticity as compared to our prepared DBT. This study is the first to employ H-NMR metabolic profiling for the quality control of traditional Chinese medicine decoctions.
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Quantification of the transcripts encoding different forms of AChE in various cell types: real-time PCR coupled with standards in revealing the copy number.
J. Mol. Neurosci.
PUBLISHED: 01-03-2014
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Acetylcholinesterase (AChE) is encoded by a single gene, and the alternative splicing at the 3' end produces different isoforms, including tailed (AChET), read-through (AChER), and hydrophobic (AChEH). Different forms of this enzyme exist in different cell types. Each AChE form has been proposed to have unique function, and all of them could be found in same cell type. Thus, the splicing process of different AChE forms remains unclear. Here, we aimed to establish a quantification method in measuring the absolute amount of each AChE splicing variants within a cell type. By using real-time PCR coupled with standard curves of defined copy of AChE variants, the copies of AChET transcript per 100 ng of total RNA were 5.7?×?10(4) in PC12 (rat neuronal cell), 1.3?×?10(4) in Caco-2 (human intestinal cell), 0.67?×?10(4) in TF-1 (human erythropoietic precursor), 133.3 in SH-SY5Y (human neuronal cell), and 56.7 in human umbilical vein endothelial cells (human endothelial cells). The copies of AChEH in these cell types were 0.3?×?10(4), 3.3?×?10(4), 2.7?×?10(4), 133.3, and 46.7, respectively, and AChER were 0.07?×?10(4), 0.13?×?10(4), 890, 3.3, and 2.7, respectively. Furthermore, PC12 and TF-1 cells were chosen for the analysis of AChE splicing pattern during differentiation. The results demonstrated a selective increase in AChET mRNA but not AChER or AChEH mRNAs in PC12 upon nerve growth factor-induced neuronal differentiation. PC12 cells could therefore act as a good cell model for the study on alternative splicing mechanism and regulation of AChET.
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Yu Ping Feng San, an ancient Chinese herbal decoction, regulates the expression of inducible nitric oxide synthase and cyclooxygenase-2 and the activity of intestinal alkaline phosphatase in cultures.
PLoS ONE
PUBLISHED: 01-01-2014
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Yu Ping Feng San (YPFS), a Chinese herbal decoction comprising Astragali Radix (AR; Huangqi), Atractylodis Macrocephalae Rhizoma (AMR; Baizhu), and Saposhnikoviae Radix (SR; Fangfeng), has been used clinically to treat inflammatory bowel diseases (IBD). Previously, we demonstrated a dual role of YPFS in regulating cytokine release in cultured macrophages. In this study, we elucidated the anti-inflammatory effect of YPFS that is mediated through modulating the expression of three key enzymes involved in IBD: inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and intestinal alkaline phosphatase (IALP). In a lipopolysaccharide (LPS)-induced chronic-inflammation model of cultured murine macrophages, YPFS treatment suppressed the activation of iNOS and COX-2 expression in a dose-dependent manner. Conversely, application of YPFS in cultured small intestinal enterocytes markedly induced the expression of IALP in a time-dependent manner, which might strengthen the intestinal detoxification system. A duality of YPFS in modulating the expression of iNOS and COX-2 was determined here. The expression of iNOS and COX-2 in macrophages was induced by YPFS, and this activation was partially blocked by the NF-?B-specific inhibitor BAY 11-7082, indicating a role of NF-?B signaling. These YPFS-induced changes in gene regulation strongly suggest that the anti-inflammatory effects of YPFS are mediated through the regulation of inflammatory enzymes.
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Flavonoids induce the expression of synaptic proteins, synaptotagmin, and postsynaptic density protein-95 in cultured rat cortical neuron.
Planta Med.
PUBLISHED: 11-15-2013
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Flavonoids, a family of phenolic compounds, are widely present in our daily diet and exist in traditional Chinese medicines, in which they act as the major active functional ingredients. Different lines of evidence indicate that flavonoids have positive impacts on human health. Here, different subclasses of flavonoids were analyzed for their inductive roles in promoting the expression of synaptic proteins, synaptotagmin, and post-synaptic density protein-95 in cultured rat cortical neurons. Among the screened 65 flavonoids, (-)-catechin, luteolin, and isorhamnetin, in micromolar concentration, were found to induce the expression of synaptic proteins in a dose-dependent manner: the induction values were from 2- to 8-fold that of the control. Similar results were revealed in the flavonoid-treated hippocampal neurons. The identification of these synapse-promoting flavonoids could be very useful in finding potential drugs, or food supplements, for treating various neurodegenerative diseases, including Alzheimers disease and depression.
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Flavonoids, derived from traditional chinese medicines, show roles in the differentiation of neurons: Possible targets in developing health food products.
Birth Defects Res. C Embryo Today
PUBLISHED: 10-24-2013
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Flavonoids, a family of phenolic compounds, are distributed in a variety of fruits, vegetables, tea, and wine. More importantly, many flavonoids are served as the active ingredients in traditional Chinese herbal medicines, which in general do not have side effects. Several lines of evidence support that flavonoids have impacts on many aspects of human health, including anti-tumor, anti-oxidation, and anti-inflammation. Recently, there is significant attention focused on the neuronal beneficial effects of flavonoids, including the promotion of nervous system development, neuroprotection against neurotoxin stress, as well as the promotion of memory, learning, and cognitive functions. Here, the activities of flavonoids on the development of nervous system are being summarized and discussed. The flavonoids from diverse herbal medicines have significant effects in different developmental stages of nervous systems, including neuronal stem cell differentiation, neurite outgrowth, and neuronal plasticity. These findings imply that flavonoids are potential candidates for the development of health supplements in preventing birth defects and neuronal diseases. Birth Defects Research (Part C) 99:292-299, 2013. © 2013 Wiley Periodicals, Inc.
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Acetylcholinesterase Protein Level Is Preserved in the Alzheimers Brain.
J. Mol. Neurosci.
PUBLISHED: 10-18-2013
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Acetylcholinesterase (AChE) is a key enzyme in the cholinergic nervous system and is one of the most studied proteins in the field of Alzheimers disease (AD). Moreover, alternative functions of AChE unrelated with the hydrolysis of acetylcholine are suspected. Until now, the majority of investigations on AChE in AD pathology have been focused on the determination of its enzymatic activity level, which is depleted in the AD brain. Despite this overall decrease, AChE activity increases at the vicinity of the two hallmarks of AD, the amyloid plaques and the neurofibrillary tangles (NFT). In fact, AChE may directly interact with A? in a manner that increases the deposition of A? to form plaques. In the context of protein-protein interactions, we have recently reported that AChE can interact with presenilin-1, the catalytic component of ?-secretase, influencing its expression level and also its activity. However, the alteration of AChE protein in the AD brain has not been determined. Here, we demonstrated by Western blotting and immunohistochemistry that a prominent pool of enzymatically inactive AChE protein existed in the AD brain. The potential significance of these unexpected levels of inactive AChE protein in the AD brain was discussed, especially in the context of protein-protein interactions with ?-amyloid and presenilin-1.
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Characterizations of Cholinesterases in Golden Apple Snail (Pomacea canaliculata).
J. Mol. Neurosci.
PUBLISHED: 08-30-2013
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Cholinesterases (ChEs) have been identified in vertebrates and invertebrates. Inhibition of ChE activity in invertebrates, such as bivalve molluscs, has been used to evaluate the exposure of organophosphates, carbamate pesticides, and heavy metals in the marine system. The golden apple snail (Pomacea canaliculata) is considered as one of the worst invasive alien species harmful to rice and other crops. The ChE(s) in this animal, which has been found recently, but poorly characterized thus far, could serve as biomarker(s) for environmental surveillance as well as a potential target for the pest control. In this study, the tissue distribution, substrate preference, sensitivity to ChE inhibitors, and molecular species of ChEs in P. canaliculata were investigated. It was found that the activities of both AChE and BChE were present in all test tissues. The intestine had the most abundant ChE activities. Both enzymes had fair activities in the head, kidney, and gills. The BChE activity was more sensitive to tetra-isopropylpyrophosphoramide (iso-OMPA) than the AChE. Only one BChE molecular species, 5.8S, was found in the intestine and head, whereas two AChE species, 5.8S and 11.6S, were found there. We propose that intestine ChEs of this snail may be potential biomarkers for manipulating pollutions.
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Chemical and biological assessment of Ziziphus jujuba fruits from China: different geographical sources and developmental stages.
J. Agric. Food Chem.
PUBLISHED: 07-22-2013
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Chinese date, the fruit of Ziziphus jujuba Mill., has thousands of years cultivation history, and about 700 cultivars of dates in China. Two types of dates are commonly found in the market: (i) fresh immature dates consumed as fruits, and (ii) dried mature dates used as Chinese medicines. Here, chemical and biological properties of these dates were revealed. Different sources of dates showed similar chemical profiles; however, the amounts of identified chemicals showed a great variation. The amount of nucleotides, flavonoids and polysaccharides in dates could be affected by its maturity and drying process. In parallel, the antioxidative functions of their extracts were compared. The date extracts protected PC12 cells against tBHP-induced cytotoxicity, and which also stimulated the transcriptional activity of antioxidant response element. The antioxidative effects were varied among different dates. The current results suggested the optimization of sources and specific usage of different maturity dates.
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Dual-index evaluation of character changes in Panax ginseng C. A. Mey stored in different conditions.
J. Agric. Food Chem.
PUBLISHED: 06-25-2013
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Panax ginseng C. A. Mey has been used as a traditional medicine and functional food in Asia for thousands of years for its improvement of human immunity and metabolism and its antitumor and antifatigue activities. This study reports the impact of storage conditions and storage period on the quality of P. ginseng. The contents of four major ginsenosides in P. ginseng and phosphorylation activities of Akt of ginseng extracts were affected by both storage conditions and storage period. In contrast, the ATP generation capacity of ginseng extracts was affected by storage conditions, but not by storage period. The results showed that the quality of P. ginseng could be well maintained at a relative humidity between 70% and 90%, and dry conditions might decrease the quality of P. ginseng. Through dual-index evaluation, the present study extended our knowledge on the changes of ginsenosides and bioactivities in P. ginseng with respect to different storage conditions and storage periods.
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Kai-xin-san, a chinese herbal decoction containing ginseng radix et rhizoma, polygalae radix, acori tatarinowii rhizoma, and poria, stimulates the expression and secretion of neurotrophic factors in cultured astrocytes.
Evid Based Complement Alternat Med
PUBLISHED: 06-07-2013
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Kai-xin-san (KXS), a Chinese herbal decoction prescribed by Sun Simiao in Beiji Qianjin Yaofang about 1400 years ago, contains Ginseng Radix et Rhizoma, Polygalae Radix, Acori Tatarinowii Rhizoma, and Poria. In China, KXS has been used to treat stress-related psychiatric diseases with the symptoms of depression and forgetfulness. Although animal study has supported the antidepression function of KXS, the mechanism in cellular level is still unknown. Here, a chemically standardized water extract of KXS was applied onto cultured astrocytes in exploring the action mechanisms of KXS treatment, which significantly stimulated the expression and secretion of neurotrophic factors, including NGF, BDNF, and GDNF, in a dose-dependent manner: the stimulation was both in mRNA and protein levels. In addition, the water extracts of four individual herbs did not significantly stimulate the expression of neurotrophic factors, which could explain the optimized effect of KXS in a herbal decoction. The KXS-induced expression of neurotrophic factors did not depend on signaling mediated by estrogen receptor or protein kinase. The results suggested that the antidepressant-like action of KXS might be mediated by an increase of expression of neurotrophic factors in astrocytes, which fully supported the clinical usage of this decoction.
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Chemical changes of Angelicae Sinensis Radix and Chuanxiong Rhizoma by wine treatment: chemical profiling and marker selection by gas chromatography coupled with triple quadrupole mass spectrometry.
Chin Med
PUBLISHED: 05-26-2013
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Angelicae Sinensis Radix (ASR) and Chuanxiong Rhizoma (CR) can be treated with wine to promote their biological functions in Chinese medicine. Both ASR and CR contain similar volatile chemicals that could be altered after wine treatment. This study aims to identify the differential chemical profiles and to select marker chemicals of ASR and CR before and after wine treatment.
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Song bu li decoction, a traditional uyghur medicine, protects cell death by regulation of oxidative stress and differentiation in cultured PC12 cells.
Evid Based Complement Alternat Med
PUBLISHED: 05-21-2013
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Song Bu Li decoction (SBL) is a traditional Uyghur medicinal herbal preparation, containing Nardostachyos Radix et Rhizoma. Recently, SBL is being used to treat neurological disorders (insomnia and neurasthenia) and heart disorders (arrhythmia and palpitation). Although this herbal extract has been used for many years, there is no scientific basis about its effectiveness. Here, we aimed to evaluate the protective and differentiating activities of SBL in cultured PC12 cells. The pretreatment of SBL protected the cell against tBHP-induced cell death in a dose-dependent manner. In parallel, SBL suppressed intracellular reactive oxygen species (ROS) formation. The transcriptional activity of antioxidant response element (ARE), as well as the key antioxidative stress proteins, was induced in dose-dependent manner by SBL in the cultures. In cultured PC12 cells, the expression of neurofilament, a protein marker for neuronal differentiation, was markedly induced by applied herbal extract. Moreover, the nerve growth factor- (NGF-) induced neurite outgrowth in cultured PC12 cells was significantly potentiated by the cotreatment of SBL. In accord, the expression of neurofilament was increased in the treatment of SBL. These results therefore suggested a possible role of SBL by its effect on neuron differentiation and protection against oxidative stress.
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Synergistic Action of Flavonoids, Baicalein, and Daidzein in Estrogenic and Neuroprotective Effects: A Development of Potential Health Products and Therapeutic Drugs against Alzheimers Disease.
Evid Based Complement Alternat Med
PUBLISHED: 05-15-2013
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Despite the classical hormonal effect, estrogen has been reported to mediate neuroprotection in the brain, which leads to the searching of estrogen-like substances for treating neurodegenerative diseases. Flavonoids, a group of natural compounds, are well known to possess estrogenic effects and used to substitute estrogen, that is, phytoestrogen. Flavonoid serves as one of the potential targets for the development of natural supplements and therapeutic drugs against different diseases. The neuroprotection activity of flavonoids was chosen for a possible development of anti-Alzheimers drugs or food supplements. The estrogenic activity of two flavonoids, baicalein and daidzein, were demonstrated by their strong abilities in stimulating estrogen receptor phosphorylation and transcriptional activation of estrogen responsive element in MCF-7 breast cells. The neuroprotection effects of flavonoids against ? -amyloid (A ? ) were revealed by their inhibition effects on in vitro A ? aggregation and A ? -induced cytotoxicity in PC12 neuronal cells. More importantly, the estrogenic and neuroprotective activities of individual flavonoid could be further enhanced by the cotreatment in the cultures. Taken together, this synergistic effect of baicalein and daidzein might serve as a method to improve the therapeutic efficacy of different flavonoids against A ? , which might be crucial in developing those flavonoidsin treating Alzheimers disease in the future.
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Flavonoids induce the synthesis and secretion of neurotrophic factors in cultured rat astrocytes: a signaling response mediated by estrogen receptor.
Evid Based Complement Alternat Med
PUBLISHED: 04-23-2013
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Neurotrophic factors are playing vital roles in survival, growth, and function of neurons. Regulation of neurotrophic factors in the brain has been considered as one of the targets in developing drug or therapy against neuronal disorders. Flavonoids, a family of multifunctional natural compounds, are well known for their neuronal beneficial effects. Here, the effects of flavonoids on regulating neurotrophic factors were analyzed in cultured rat astrocytes. Astrocyte is a major secreting source of neurotrophic factors in the brain. Thirty-three flavonoids were screened in the cultures, and calycosin, isorhamnetin, luteolin, and genistein were identified to be highly active in inducing the synthesis and secretion of neurotrophic factors, including nerve growth factor (NGF), glial-derived neurotrophic factor (GDNF), and brain-derived neurotrophic factor (BDNF). The inductions were in time- and dose-dependent manners. In cultured astrocytes, the phosphorylation of estrogen receptor was triggered by application of flavonoids. The phosphorylation was blocked by an inhibitor of estrogen receptor, which in parallel reduced the flavonoid-induced expression of neurotrophic factors. The results proposed the role of flavonoids in protecting brain diseases, and therefore these flavonoids could be developed for health food supplement for patients suffering from neurodegenerative diseases.
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Activation of UTP-sensitive P2Y2 receptor induces the expression of cholinergic genes in cultured cortical neurons: a signaling cascade triggered by Ca2+ mobilization and extracellular regulated kinase phosphorylation.
Mol. Pharmacol.
PUBLISHED: 04-16-2013
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ATP functions as an extracellular signaling molecule that is costored and coreleased with neurotransmitters at central and peripheral neuronal synapses. Stimulation by ATP upregulates the expression of synaptic genes in muscle-including the genes for nicotine acetylcholine receptor (?-, ?-, and ?-subunits) and acetylcholinesterase (AChE)-via the P2Y receptor (P2YR), but the trophic response of neurons to the activation of P2YRs is less well understood. We reported that cultured cortical neurons and the developing rat brain expressed different types of P2YRs, and among these the UTP-sensitive P2Y2R was the most abundant. P2Y2R was found to exist in membrane rafts and it colocalized with the postsynaptic protein PSD-95 in cortical neurons. Notably, agonist-dependent stimulation of P2Y2R elevated the neuronal expression of cholinergic genes encoding AChE, PRiMA (an anchor for the globular form AChE), and choline acetyltransferase, and this induction was mediated by a signaling cascade that involved Ca(2+) mobilization and extracellular regulated kinases 1/2 activation. The importance of P2Y2R action was further shown by the receptors synergistic effect with P2Y1R in enhancing cholinergic gene expression via the robust stimulation of Ca(2+) influx. Taken together our results revealed a developmental function of P2Y2R in promoting synaptic gene expression and demonstrated the influence of costimulation of P2Y1R and P2Y2R in neurons.
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Distribution and expression of Kirre, an IgSF molecule, during postnatal development of rat cerebellum.
Neurosci. Lett.
PUBLISHED: 03-11-2013
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Immunoglobulin superfamily (IgSF) molecules are actively involved in cell-cell adhesion, neuronal migration, axonal guidance and synapse formation in the nervous system. Kirre, as a member of this family, has been implicated in mammalian neuronal differentiation and development. Although the distribution of rKirre (a rat homologue of Drosophila Kirre) mRNA was previously analyzed in adult rat cerebellum by in situ hybridization, the expression levels of transcript and protein were not well studied. Here, we showed that the expressions of rKirre mRNA and protein significantly increased during postnatal development of rat cerebellum. rKirre mRNA was mainly expressed in the granular layers and Purkinje cell layer in the developing cerebellum, revealing a possible involvement of rKirre in granule cell migration and Purkinje cell development. An essential relationship between rKirre and Purkinje cells was implied by the co-localization of rKirre and NF-200 on the cell bodies of Purkinje cells. These results suggest that rKirre may play a potential role in postnatal developing rat cerebellum.
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Importance of wine-treated Angelica Sinensis Radix in Si Wu Tang, a traditional herbal formula for treating womens ailments.
Planta Med.
PUBLISHED: 03-01-2013
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Si Wu Tang (Four Agents Decoction), a traditional Chinese decoction composed of Angelica Sinensis Radix, Chuanxiong Rhizoma, Paeoniae Radix Alba, and Rehmanniae Radix Praeparata in a ratio of 1 : 1 : 1 : 1, has been used to treat womens diseases for more than a thousand years. According to the original description of Si Wu Tang, Angelica Sinensis Radix should be treated with wine. However, the importance of this wine-treated Angelica Sinensis Radix in Si Wu Tangs function has not been identified. In this article, the chemical and biological properties of two decoctions processed in different ways (Si Wu Tang with crude Angelica Sinensis Radix and Si Wu Tang with wine-treated Angelica Sinensis Radix) were compared for examination. The herbal decoction Si Wu Tang prepared from wine-treated Angelica Sinensis Radix contained much different amounts of its active compounds. Compared with Si Wu Tang using crude Angelica Sinensis Radix, Si Wu Tang prepared from wine-treated Angelica Sinensis Radix had better biological responses. Therefore, these findings accentuate the functional importance of herbs treated with wine in the Chinese decoction.
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Chemical and biological assessment of angelica roots from different cultivated regions in a chinese herbal decoction danggui buxue tang.
Evid Based Complement Alternat Med
PUBLISHED: 02-05-2013
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Roots of Angelica sinensis (Danggui) have been used in promoting blood circulation as herbal medicine for over 2000 years in China. Another species of Angelica roots called A. gigas is being used in Korea. To reveal the efficiency of different Angelica roots, the chemical and biological properties of Angelica roots from different cultivated regions were compared. Roots of A. sinensis contained higher levels of ferulic acid, Z-ligustilide, and senkyunolide A, while high amounts of butylphthalide and Z-butylenephthalide were found in A. gigas roots. The extracts deriving from A. gigas roots showed better effects in osteogenic and estrogenic properties than that of A. sinensis from China. However, this difference was markedly reduced when the Angelica roots were being prepared in a Chinese herbal decoction together with Astragali Radix as Danggui Buxue Tang. In contrast, the herbal decoction prepared from A. sinensis roots showed better responses in cell cultures. In addition, the extracts of A. gigas roots showed strong cell toxicity both as single herb and as Danggui Buxue Tang. This result revealed the distinct properties of Angelica roots from China and Korea suggesting the specific usage of herb in preparing a unique herbal decoction.
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Optimizing combinations of flavonoids deriving from astragali radix in activating the regulatory element of erythropoietin by a feedback system control scheme.
Evid Based Complement Alternat Med
PUBLISHED: 02-04-2013
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Identifying potent drug combination from a herbal mixture is usually quite challenging, due to a large number of possible trials. Using an engineering approach of the feedback system control (FSC) scheme, we identified the potential best combinations of four flavonoids, including formononetin, ononin, calycosin, and calycosin-7-O- ? -D-glucoside deriving from Astragali Radix (AR; Huangqi), which provided the best biological action at minimal doses. Out of more than one thousand possible combinations, only tens of trials were required to optimize the flavonoid combinations that stimulated a maximal transcriptional activity of hypoxia response element (HRE), a critical regulator for erythropoietin (EPO) transcription, in cultured human embryonic kidney fibroblast (HEK293T). By using FSC scheme, 90% of the work and time can be saved, and the optimized flavonoid combinations increased the HRE mediated transcriptional activity by ~3-fold as compared with individual flavonoid, while the amount of flavonoids was reduced by ~10-fold. Our study suggests that the optimized combination of flavonoids may have strong effect in activating the regulatory element of erythropoietin at very low dosage, which may be used as new source of natural hematopoietic agent. The present work also indicates that the FSC scheme is able to serve as an efficient and model-free approach to optimize the drug combination of different ingredients within a herbal decoction.
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Can Hedysari Radix replace Astragali Radix in Danggui Buxue Tang, a Chinese herbal decoction for woman aliment?
Phytomedicine
PUBLISHED: 01-25-2013
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Astragali Radix (AR) has been used for over 2000 years in China for the enrichment of "Qi". Hedysari Radix (HR), a herb having similar chemical composition with AR, has been commonly used as a substitute of AR in herbal decoction. In order to evaluate the possible replacement of HR for AR in Chinese herbal decoction, systematic comparison of AR and HR was done by chemical and biological assessments. The water extract of AR contained higher levels of calycosin, calycosin-glucoside, ononin, astragaloside III and astragaloside IV, while higher amount of formononetin was found in the HR extract. The estrogenic, erythropoetic and osteogenic effects were compared between the water extracts of AR and HR, and in all cases AR extract showed higher biological activities. Danggui Buxue Tang (DBT) is a very common herbal decoction for woman aliment, and which contains AR and Angelica Sinensis Radix. Here, we generated two forms of DBT having either AR or HR as the major herbs. Chemically, AR-contained DBT showed higher amounts of various active chemicals, except formononetin that was higher in HR-contained DBT. In parallel, the estrogenic, osteogenic and erythropoetic effects of DBT containing AR showed better activities than that of DBT having HR. Thus, AR and HR showed distinct differences in terms of chemical and biological properties. In order to achieve the best therapeutical effect, as well as to guarantee the safety, AR is recommended here to be used for making DBT.
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Yu ping feng san, an ancient chinese herbal decoction containing astragali radix, atractylodis macrocephalae rhizoma and saposhnikoviae radix, regulates the release of cytokines in murine macrophages.
PLoS ONE
PUBLISHED: 01-01-2013
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Yu Ping Feng San (YPFS), a Chinese herbal decoction, is composed of Astragali Radix (AR; Huangqi), Atractylodis Macrocephalae Rhizoma (AMR; Baizhu) and Saposhnikoviae Radix (SR; Fangfeng) in a weight ratio of 1?2?1. Clinically, YPFS has been widely used to regulate immune functions; however, the action mechanism of it is not known. Here, we addressed this issue by providing detail analyses of chemical and biological properties of YPFS. By using rapid resolution liquid chromatography coupled with mass spectrometry, fifteen chemicals deriving from different herbs of YPFS were determined, and which served as a control for the standardization of the herbal extract of YPFS. In general, the amounts of chosen chemical markers were higher in a preparation of YPFS as compared to that of single herb or two-herb compositions. In order to reveal the immune functions of YPFS, the standardized extract was applied onto cultured murine macrophages. The treatment of YPFS stimulated the mRNA and protein expressions of pro-inflammatory cytokines via activation of NF-?B by enhancing I?B? degradation. In contrast, the application of YPFS suppressed the expressions of pro-inflammatory cytokines significantly in the lipopolysaccharide (LPS)-induced chronic inflammation model. In addition, YPFS could up regulate the phagocytic activity in cultured macrophages. These results therefore supported the bi-directional immune-modulatory roles of YPFS in regulating the releases of cytokines from macrophages.
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Fo Shou San, an ancient herbal decoction prepared from Angelicae Sinensis Radix and Chuanxiong Rhizoma, induces erythropoietin expression: a signaling mediated by the reduced degradation of hypoxia-inducible factor in cultured liver cells.
Planta Med.
PUBLISHED: 11-17-2011
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Fo Shou San (FSS) is an ancient herbal decoction composed of Angelicae Sinensis Radix (ASR; Danggui) and Chuanxiong Rhizoma (CR; Chuanxiong) in a ratio of 3:2. FSS is mainly prescribed for patients having a deficiency of blood supply, and it indeed has been shown to stimulate the production of erythropoietin (EPO) in cultured cells. In order to reveal the mechanism of this FSS-induced EPO gene expression, the upstream regulatory cascade, via hypoxia-induced signaling, was revealed here in cultured hepatocellular carcinoma cell line Hep3B. The induction of EPO gene expression, triggered by FSS, was revealed in cultured hepatocytes by: (i) the increase of EPO mRNA; and (ii) the activation of the hypoxia response element (HRE), an upstream regulator of the EPO gene. The FSS-induced EPO gene expression was triggered by an increased expression of hypoxia-inducible factor-1 ? (HIF-1 ?) protein; however, the mRNA expression of HIF-1 ? was not altered by the treatment of FSS. The increased HIF-1 ? was a result of reduced protein degradation after the FSS treatment. The current results therefore provide one of the molecular mechanisms of this ancient herbal decoction for its hematopoietic function.
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Phenolic compounds from the rhizomes of Dioscorea bulbifera.
Chem. Biodivers.
PUBLISHED: 11-16-2011
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One new bibenzyl, 7, and one new diarylheptanone, diobulbinone A (18), together with sixteen known compounds, 1-6 and 8-17, have been isolated form the rhizomes of Dioscorea bulbifera. Their structures were elucidated by NMR and MS analyses. Compound 7 showed high antioxidant capacity in FRAP assay and DPPH radical-scavenging activity.
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Molecular Assembly and Biosynthesis of Acetylcholinesterase in Brain and Muscle: the Roles of t-peptide, FHB Domain, and N-linked Glycosylation.
Front Mol Neurosci
PUBLISHED: 07-30-2011
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Acetylcholinesterase (AChE) is responsible for the hydrolysis of the neurotransmitter, acetylcholine, in the nervous system. The functional localization and oligomerization of AChE T variant are depending primarily on the association of their anchoring partners, either collagen tail (ColQ) or proline-rich membrane anchor (PRiMA). Complexes with ColQ represent the asymmetric forms (A(12)) in muscle, while complexes with PRiMA represent tetrameric globular forms (G(4)) mainly found in brain and muscle. Apart from these traditional molecular forms, a ColQ-linked asymmetric form and a PRiMA-linked globular form of hybrid cholinesterases (ChEs), having both AChE and BChE catalytic subunits, were revealed in chicken brain and muscle. The similarity of various molecular forms of AChE and BChE raises interesting question regarding to their possible relationship in enzyme assembly and localization. The focus of this review is to provide current findings about the biosynthesis of different forms of ChEs together with their anchoring proteins.
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The assembly of proline-rich membrane anchor (PRiMA)-linked acetylcholinesterase enzyme: glycosylation is required for enzymatic activity but not for oligomerization.
J. Biol. Chem.
PUBLISHED: 07-27-2011
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Acetylcholinesterase (AChE) anchors onto cell membranes by a transmembrane protein PRiMA (proline-rich membrane anchor) as a tetrameric form in vertebrate brain. The assembly of AChE tetramer with PRiMA requires the C-terminal "t-peptide" in AChE catalytic subunit (AChE(T)). Although mature AChE is well known N-glycosylated, the role of glycosylation in forming the physiologically active PRiMA-linked AChE tetramer has not been studied. Here, several lines of evidence indicate that the N-linked glycosylation of AChE(T) plays a major role for acquisition of AChE full enzymatic activity but does not affect its oligomerization. The expression of the AChE(T) mutant, in which all N-glycosylation sites were deleted, together with PRiMA in HEK293T cells produced a glycan-depleted PRiMA-linked AChE tetramer but with a much higher K(m) value as compared with the wild type. This glycan-depleted enzyme was assembled in endoplasmic reticulum but was not transported to Golgi apparatus or plasma membrane.
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Trillin, a steroidal saponin isolated from the rhizomes of Dioscorea nipponica, exerts protective effects against hyperlipidemia and oxidative stress.
J Ethnopharmacol
PUBLISHED: 06-10-2011
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Numerous efforts have been conducted in searching for effective agents against cardiovascular diseases, in particular from herbal medicines. The rhizome of Dioscorea nipponica (Dioscoreae Nipponicae Rhizoma) is a traditional Chinese herb being prescribed to improve the blood circulation. Here, we identified a steroidal saponin trillin from Dioscorea nipponica, which showed robust anti-hyperlipidemic effects.
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Baicalin, a flavone, induces the differentiation of cultured osteoblasts: an action via the Wnt/beta-catenin signaling pathway.
J. Biol. Chem.
PUBLISHED: 06-07-2011
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Flavonoids, a group of natural compounds found in a variety of vegetables and herbal medicines, have been intensively reported on regarding their estrogen-like activities and particularly their ability to affect bone metabolism. Here, different subclasses of flavonoids were screened for their osteogenic properties by measuring alkaline phosphatase activity in cultured rat osteoblasts. The flavone baicalin derived mainly from the roots of Scutellaria baicalensis showed the strongest induction of alkaline phosphatase activity. In cultured osteoblasts, application of baicalin increased significantly the osteoblastic mineralization and the levels of mRNAs encoding the bone differentiation markers, including osteonectin, osteocalcin, and collagen type 1?1. Interestingly, the osteogenic effect of baicalin was not mediated by its estrogenic activity. In contrast, baicalin promoted osteoblastic differentiation via the activation of the Wnt/?-catenin signaling pathway; the activation resulted in the phosphorylation of glycogen synthase kinase 3? and, subsequently, induced the nuclear accumulation of the ?-catenin, leading to the transcription activation of Wnt-targeted genes for osteogenesis. The baicalin-induced osteogenic effects were fully abolished by DKK-1, a blocker of Wnt/?-catenin receptor. Moreover, baicalin also enhanced the mRNA expression of osteoprotegerin, which could regulate indirectly the activation of osteoclasts. Taken together, our results suggested that baicalin could act via Wnt/?-catenin signaling to promote osteoblastic differentiation. The osteogenic flavonoids could be very useful in finding potential drugs, or food supplements, for treating post-menopausal osteoporosis.
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Chemical and biological assessment of Angelicae Sinensis Radix after processing with wine: an orthogonal array design to reveal the optimized conditions.
J. Agric. Food Chem.
PUBLISHED: 05-05-2011
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The roots of Angelica sinensis [Angelica Sinensis Radix (ASR)] have been used as a common health food supplement for womens care for thousands of years in China. According to Asian tradition, ASR could be processed with the treatment of wine, which subsequently promoted the biological functions of ASR. By chemical and biological assessments, an orthogonal array design was employed here to determine the roles of three variable parameters in the processing of ASR, including oven temperature, baking time, and flipping frequency. The results suggested that oven temperature and baking time were two significant factors, while flipping frequency was a subordinate factor. The optimized condition of processing with wine therefore was considered to be heating in an oven at 80 °C for 90 min with flipping twice per hour. Under the optimized processing conditions, the solubilities of ferulic acid and Z-ligustilide from ASR were markedly increased and decreased, respectively. In parallel, the biological functions of processed ASR were enhanced in both anti-platelet aggregation and estrogenic activation; these increased functions could be a result of the altered levels of ferulic acid and Z-ligustilide in wine-processed ASR. Thus, the chemical and biological assessment of the processed ASR was in full accordance with the Chinese old tradition.
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Flavonoids from Radix Astragali induce the expression of erythropoietin in cultured cells: a signaling mediated via the accumulation of hypoxia-inducible factor-1?.
J. Agric. Food Chem.
PUBLISHED: 02-10-2011
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Radix Astragali (RA) is commonly used as a health food supplement to reinforce the body vital energy. Flavonoids, including formononetin, ononin, calycosin, and calycosin-7-O-?-d-glucoside, are considered to be the major active ingredients within RA. Here, we provided different lines of evidence that the RA flavonoids stimulated the expression of erythropoietin (EPO), the central regulator of red blood cell mass, in cultured human embryonic kidney fibroblasts (HEK293T). A plasmid containing hypoxia response element (HRE), a critical regulator for EPO transcription, was tagged upstream of a firefly luciferase gene, namely, pHRE-Luc, which was being transfected into fibroblasts. The application of RA flavonoids onto the transfected cells induced the transcriptional activity of HRE. To account for the transcriptional activation after the treatment of flavonoids, the expression of hypoxia-inducible factor-1? (HIF-1?) was markedly increased: The increase was in both mRNA and protein levels. In addition, the degradation of HIF-1? was reduced under the effect of flavonoids. The regulation of HIF-1? therefore could account for the activation of EPO expression mediated by the RA flavonoids. The current results therefore reveal the function of this herb in enhancing hematopoietic functions.
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The establishment of a sensitive method in determining different neurotransmitters simultaneously in rat brains by using liquid chromatography-electrospray tandem mass spectrometry.
J. Chromatogr. B Analyt. Technol. Biomed. Life Sci.
PUBLISHED: 02-07-2011
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An effective way to determine the amount of different neurotransmitters is vital to the study of brain function. Here, a highly sensitive HPLC-MS/MS method was developed to simultaneously measure ?-aminobutyric acid, dopamine, epinephrine, norepinepherine, glutamate and serotonin in one sample. The quantification of the neurotransmitters was achieved by a tandem mass spectrometer using the selected reaction monitoring scan mode. The method validation included selectivity, linearity, accuracy, precision, stability, recovery and matrix effect. For the six neurotransmitters, the linear regression analysis was calibrated by deuterated internal standards with a R(2) of over 0.991, and the limit of detection (LOD) and the limit of quantification (LOQ) were from 2.5 to 500 pg/mg and 7.5 to 1000 pg/mg, respectively. This method was employed here to reveal different types and amounts of neurotransmitters simultaneously in adult and embryonic rat brains. Here, the change of dopamine concentration in embryonic and adult brain was from 0.071 to 0.760 ng/mg of brain tissue, GABA was from 207.643 to 445.148 ng/mg, glutamate was from 679.535 to 1408.920 ng/mg, serotonin was from 0.058 to 0.485 ng/mg and norepinepherine was from 0.054 to 0.290 ng/mg. For epinephrine, it was only detected in embryonic stage but not in adult, with the concentration at 0.241 ng/mg.
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Expression of the IgSF protein Kirre in the rat central nervous system.
Life Sci.
PUBLISHED: 01-14-2011
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Immunoglobulin superfamily (IgSF) proteins play a critical role in development of the nervous system. Here, a new member of IgSF gene family was cloned from rat brain, which was subsequently identified as rat homolog of Drosophila Kirre. This new molecule was named as rat Kirre (rKirre). We aimed to reveal the developmental expression of rKirre, both at mRNA and protein levels, in the central nervous system. The deduced amino acid sequence of rKirre showed a putative PDZ binding motif at the C-terminus, which provided a rationale for analyzing the co-localization of rKirre and post-synaptic density protein 95 (PSD-95) in cultured rat cortical neurons.
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The extract of Rhodiolae Crenulatae Radix et Rhizoma induces the accumulation of HIF-1? via blocking the degradation pathway in cultured kidney fibroblasts.
Planta Med.
PUBLISHED: 12-14-2010
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Rhodiolae Crenulatae Radix et Rhizoma (Rhodiola), the root and rhizome of Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba, has been used as a traditional Chinese medicine (TCM) to increase the body resistance against hypoxia in mountain sickness. The mechanism of this adaptogenic property deriving from Rhodiola, however, has not been revealed. Erythropoietin (EPO) is an erythrocyte-specific hematopoietic hormone that increases the production of red blood cells: this hormone is a crucial factor in regulating the body balance in responding to hypoxia. In cultured kidney fibroblasts (HEK293T), application of water extract deriving from Rhodiola induced the expression of EPO both in mRNA and protein levels. The activation of the Hypoxia Response Element (HRE) located on the promoter region of the EPO gene is one of the mechanisms accounting for transcriptional activation. In addition, the Rhodiola-induced EPO expression was triggered by an increase of hypoxia-inducible factor-1 ? (HIF-1 ?) protein, via the reduction of HIF-1 ? degradation but not the induction of HIF-1 ? mRNA. Moreover, the same EPO induction effect by Rhodiola was also observed in cultured liver cells since liver is another vital organ to provide EPO regulation apart from the kidney. These results therefore elucidate one of the molecular mechanisms of this herb in mediating the anti-hypoxia function.
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Quality assessment of a formulated Chinese herbal decoction, Kaixinsan, by using rapid resolution liquid chromatography coupled with mass spectrometry: A chemical evaluation of different historical formulae.
J Sep Sci
PUBLISHED: 11-16-2010
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Kaixinsan is an ancient Chinese herbal decoction mainly prescribed for patients suffering from mental depression. This decoction was created by Sun Si-miao of Tang Dynasty (A.D. 600) in ancient China, and was composed of four herbs: Radix and Rhizome Ginseng, Radix Polygalae, Rhizoma Acori Tatarinowii and Poria. Historically, this decoction has three different formulations, each recorded at a different point in time. In this study, the chemical compositions of all three Kaixinsan formulae were analyzed. By using rapid resolution LC coupled with a diode-array detector and an ESI triple quadrupole tandem MS (QQQ-MS/MS), the Radix and Rhizome Ginseng-derived ginsenosides including Rb(1), Rd, Re, Rg(1), the Radix Polygalae-derived 3,6-disinapoyl sucrose, the Rhizoma Acori Tatarinowii-derived ?- and ?-asarone and the Poria-derived pachymic acid were compared among the three different formulations. The results showed variations in the solubility of different chemicals between one formula and the others. This systematic method developed could be used for the quality assessment of this herbal decoction.
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ATP induces synaptic gene expressions in cortical neurons: transduction and transcription control via P2Y1 receptors.
Mol. Pharmacol.
PUBLISHED: 09-16-2010
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Studies in vertebrate neuromuscular synapses have revealed previously that ATP, via P2Y receptors, plays a critical role in regulating postsynaptic gene expressions. An equivalent regulatory role of ATP and its P2Y receptors would not necessarily be expected for the very different situation of the brain synapses, but we provide evidence here for a brain version of that role. In cultured cortical neurons, the expression of P2Y(1) receptors increased sharply during neuronal differentiation. Those receptors were found mainly colocalized with the postsynaptic scaffold postsynaptic density protein 95 (PSD-95). This arises through a direct interaction of a PDZ domain of PSD-95 with the C-terminal PDZ-binding motif, D-T-S-L of the P2Y(1) receptor, confirmed by the full suppression of the colocalization upon mutation of two amino acids therein. This interaction is effective in recruiting PSD-95 to the membrane. Specific activation of P2Y(1) (G-protein-coupled) receptors induced the elevation of intracellular Ca(2+) and activation of a mitogen-activated protein kinase/Raf-1 signaling cascade. This led to distinct up-regulation of the genes encoding acetylcholinesterase (AChE(T) variant), choline acetyltransferase, and the N-methyl-d-aspartate receptor subunit NR2A. This was confirmed, in the example of AChE, to arise from P2Y(1)-dependent stimulation of a human ACHE gene promoter. That involved activation of the transcription factor Elk-1; mutagenesis of the ACHE promoter revealed that Elk-1 binding at its specific responsive elements in that promoter was induced by P2Y(1) receptor activation. The combined findings reveal that ATP, via its P2Y(1) receptor, can act trophically in brain neurons to regulate the gene expression of direct effectors of synaptic transmission.
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The establishment of a highly sensitive method in detecting ketamine and norketamine simultaneously in human hairs by HPLC-Chip-MS/MS.
Forensic Sci. Int.
PUBLISHED: 08-12-2010
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An effective way to reveal the history of drug abuse is to determine the parental drug and its metabolites in hair. Here, a quantitative HPLC-Chip-MS/MS method was developed for simultaneous measurement of ketamine and its metabolite norketamine in human hair. Ketamine and norketamine were extracted from hair by acid hydrolysis, and then enriched by organic solvent extraction. The chromatographic separation was achieved in 15 min, with the drug identification and quantification by a tandem mass spectrometer. The linear regression analysis was calibrated by deuterated internal standards with a R(2) of over 0.996. The limit of detection (LOD) and the limit of quantification (LOQ) for ketamine and norketamine were 0.5 and 1 pg/mg of hair, respectively. The standard curves were linear from the value of LOQ up to 100 pg/mg of hair. The validation parameters including selectivity, accuracy, precision, stability and matrix effect were also determined. In conclusion, this method was able to reveal the present of ketamine and norketamine with less hair from the drug abusers, and which had the sensitivity of ?1000-fold higher than the conventional method. In addition, the amount of ketamine and norketamine being detected in different hair segments would be useful in revealing the historical record of ketamine uptake in the drug abusers.
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The PRiMA-linked cholinesterase tetramers are assembled from homodimers: hybrid molecules composed of acetylcholinesterase and butyrylcholinesterase dimers are up-regulated during development of chicken brain.
J. Biol. Chem.
PUBLISHED: 06-21-2010
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Acetylcholinesterase (AChE) is anchored onto cell membranes by the transmembrane protein PRiMA (proline-rich membrane anchor) as a tetrameric globular form that is prominently expressed in vertebrate brain. In parallel, the PRiMA-linked tetrameric butyrylcholinesterase (BChE) is also found in the brain. A single type of AChE-BChE hybrid tetramer was formed in cell cultures by co-transfection of cDNAs encoding AChE(T) and BChE(T) with proline-rich attachment domain-containing proteins, PRiMA I, PRiMA II, or a fragment of ColQ having a C-terminal GPI addition signal (Q(N-GPI)). Using AChE and BChE mutants, we showed that AChE-BChE hybrids linked with PRiMA or Q(N-GPI) always consist of AChE(T) and BChE(T) homodimers. The dimer formation of AChE(T) and BChE(T) depends on the catalytic domains, and the assembly of tetramers with a proline-rich attachment domain-containing protein requires the presence of C-terminal "t-peptides" in cholinesterase subunits. Our results indicate that PRiMA- or ColQ-linked cholinesterase tetramers are assembled from AChE(T) or BChE(T) homodimers. Moreover, the PRiMA-linked AChE-BChE hybrids occur naturally in chicken brain, and their expression increases during development, suggesting that they might play a role in cholinergic neurotransmission.
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Antihyperlipidemic effect of protodioscin, an active ingredient isolated from the rhizomes of Dioscorea nipponica.
Planta Med.
PUBLISHED: 05-27-2010
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Developing drugs against metabolic-related disorders, including obesity and hyperlipidemia, is of importance for human health. Here, a bioactive phytochemical, protodioscin, isolated from the rhizomes of Dioscorea nipponica (Rhizoma Dioscoreae Nipponicae), was identified for its anti-hyperlipidemic effect. In hyperlipidemic rats, the post-administration of protodioscin significantly reduced the time of blood coagulation. In addition, the blood levels of triglyceride, cholesterol, low-density and high-density lipoproteins were also changed accordingly. Taken together, this was the first report of an antihyperlipidemic effect of protodioscin. Its great availability in various vegetables and medicinal plants will be useful in developing health food supplement(s) and/or drug(s) against hyperlipidemia.
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The expression of erythropoietin triggered by danggui buxue tang, a Chinese herbal decoction prepared from radix Astragali and radix Angelicae Sinensis, is mediated by the hypoxia-inducible factor in cultured HEK293T cells.
J Ethnopharmacol
PUBLISHED: 05-17-2010
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Danggui buxue tang (DBT), a Chinese medicinal decoction that is being commonly used as hematopoietic medicine to treating woman menopausal irregularity, contains two herbs: radix Astragali and radix Angelicae Sinensis. Pharmacological results indicate that DBT can stimulate the production of erythropoietin (EPO), a specific hematopoietic growth factor, in cultured cells.
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Galangin, a flavonol derived from Rhizoma Alpiniae Officinarum, inhibits acetylcholinesterase activity in vitro.
Chem. Biol. Interact.
PUBLISHED: 04-28-2010
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Acetylcholinesterase (AChE) inhibitors are widely used for the treatment of Alzheimers disease (AD). Several AChE inhibitors, e.g. rivastigmine, galantamine and huperzine are originating from plants, suggesting that herbs could potentially serve as sources for novel AChE inhibitors. Here, we searched potential AChE inhibitors from flavonoids, a group of naturally occurring compounds in plants or traditional Chinese medicines (TCM). Twenty-one flavonoids, covered different subclasses, were tested for their potential function in inhibiting AChE activity from the brain in vitro. Among all the tested flavonoids, galangin, a flavonol isolated from Rhizoma Alpiniae Officinarum, the rhizomes of Alpiniae officinarum (Hance.) showed an inhibitory effect on AChE activity with the highest inhibition by over 55% and an IC(50) of 120 microM and an enzyme-flavonoid inhibition constant (K(i)) of 74 microM. The results suggest that flavonoids could be potential candidates for further development of new drugs against AD.
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Anti-oxidative effects of the biennial flower of Panax notoginseng against H2O2-induced cytotoxicity in cultured PC12 cells.
Chin Med
PUBLISHED: 04-05-2010
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Radix notoginseng is used in Chinese medicine to improve blood circulation and clotting; however, the pharmacological activities of other parts of Panax notoginseng have yet to be explored. The present study reports the anti-oxidative effects of various parts of Panax notoginseng.
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Differential gene expression profiling on the muscle of acetylcholinesterase knockout mice: a preliminary analysis.
Chem. Biol. Interact.
PUBLISHED: 03-25-2010
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Acetylcholinesterase (AChE) (EC. 3.1.1.7) is the acetylcholine-hydrolyzing enzyme that plays an essential role on cholinergic neurotransmission at the synapses of the brain and at the neuromuscular junctions. In order to gain insight into the molecular mechanisms of neuromuscular dysfunction associated with AChE deficiency, we have compared the RNA expression profiles of the muscles of AChE knockout mice with those of the wild-type siblings. Total RNA from the leg muscle of the mice of the wild-type and the AChE nullizygous mice were subjected to microarray analyses with Affymetrix GeneChip((R)) Mouse Gene 1.0 ST Array. The pair-wise comparison of gene expression levels of the 28,853 mRNA transcripts showed that 303 genes were either up- or down-regulated by more than 2.0 folds in the AChE knockout mice. The interaction study of these differentially regulated genes indicated that some of these genes are clustered in biological functions that are related to lipid metabolism and the skeletal-muscular functions.
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Fo Shou San, an ancient herbal decoction prepared from Rhizoma Chuanxiong and Radix Angelicae Sinensis, stimulates the production of hemoglobin and erythropoietin in cultured cells.
Planta Med.
PUBLISHED: 03-22-2010
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Fo Shou San (FSS) is an ancient herbal decoction comprised of Rhizoma Chuanxiong (RC; Chuanxiong) and Radix Angelicae Sinensis (RAS; Danggui) in a ratio of 2?:?3. It is mainly prescribed for patients having a blood deficiency. This combination is considered the most popular herb pair among Chinese medicines; however, the rationale of having these two chemically similar herbs within the decoction has historically not been made clear. Here, we attempted to reveal the chemical and biological properties of this decoction as a means to deduce its mechanism of action. The effects of FSS were determined in different cell culture models. With respect to stimulation of blood circulation, FSS inhibited ADP-mediated platelet aggregation in a dose-dependent manner. In order to reveal the hematopoietic effect of this decoction, FSS was applied onto cultured K562 human leukemia cells and Hep3B human hepatocellular carcinoma cells. Application of FSS in cultured K562 cells inhibited cell proliferation and subsequently induced the production of hemoglobin. Additionally, the mRNA expression of erythropoietin (EPO) was induced in a dose-dependent manner when FSS was applied to Hep3B cells. The current results reveal the effects of FSS in different cell models, paving a direction for mechanistic studies.
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Targeting acetylcholinesterase to membrane rafts: a function mediated by the proline-rich membrane anchor (PRiMA) in neurons.
J. Biol. Chem.
PUBLISHED: 02-10-2010
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In the mammalian brain, acetylcholinesterase (AChE) is anchored in cell membranes by a transmembrane protein PRiMA (proline-rich membrane anchor). We present evidence that at least part of the PRiMA-linked AChE is integrated in membrane microdomains called rafts. A significant proportion of PRiMA-linked AChE tetramers from rat brain was recovered in raft fractions; this proportion was markedly higher at low rather than at high concentrations of cold Triton X-100. The detergent-resistant fraction increased during brain development. In NG108-15 neuroblastoma cells transfected with cDNAs encoding AChE(T) and PRiMA, PRiMA-linked G(4) AChE was found in membrane rafts and showed the same sensitivity to cold Triton X-100 extraction as in the brain. The association of PRiMA-linked AChE with rafts was weaker than that of glycosylphosphatidylinositol-anchored G(2) AChE or G(4) Q(N)-H(C)-linked AChE. It was found to depend on the presence of a cholesterol-binding motif, called CRAC (cholesterol recognition/interaction amino acid consensus), located at the junction of transmembrane and cytoplasmic domains of both PRiMA I and II isoforms. The cytoplasmic domain of PRiMA, which differs between PRiMA I and PRiMA II, appeared to play some role in stabilizing the raft localization of G(4) AChE, because the Triton X-100-resistant fraction was smaller with the shorter PRiMA II isoform than that with the longer PRiMA I isoform.
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PRiMA directs a restricted localization of tetrameric AChE at synapses.
Chem. Biol. Interact.
PUBLISHED: 02-08-2010
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Acetylcholinesterase (AChE), a highly polymorphic enzyme with various splicing variants and molecular isoforms, plays an essential role in the cholinergic neurotransmission by hydrolyzing acetylcholine into choline and acetate. The AChE(T) variant is expressed in the brain and muscle: this subunit forms non-amphiphilic tetramers with a collagen tail (ColQ) as asymmetric AChE (A(12) AChE) in muscle, and amphiphilic tetramers with a proline-rich membrane anchor (PRiMA) as globular AChE (G(4) AChE) in the brain and muscle. During the brain development, the expression of amphiphilic G(4) AChE is up regulated and becomes the predominant form of AChE there. This up-regulation of G(4) AChE can be attributed to the increased expressions of both AChE(T) and PRiMA. A significant portion of this membrane-bound G(4) AChE is localized at the membrane rafts of the cell membranes derived from the brain. This raft association could be directed by PRiMA via its CRAC (cholesterol recognition/interaction amino acid consensus) motif and C-terminus. In cultured cortical neurons and muscles, the PRiMA-linked AChE was clustered and partially co-localized with synaptic proteins. The restricted localizations suggest that the raft association of PRiMA-linked AChE could account for its synaptic localization and function.
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An induction effect of heat shock on the transcript of globular acetylcholinesterase in NG108-15 cells.
Chem. Biol. Interact.
PUBLISHED: 02-05-2010
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Heat shock response, an induced transcription of a set of genes in response to high temperature, occurs in all organisms. In neurons, the catalytic subunit of acetylcholinesterase (AChE(T)) interacts with proline-rich membrane anchor (PRiMA) to form a globular tetrameric form (G(4) form). In this study, we examined the effects of heat shock on the transcription and protein assembly of AChE(T) in cultured NG108-15 cells. The transcription of AChE(T) was rapidly induced by heat shock at 40 degrees C, reaching a 15-fold increase in 3h and decreasing thereafter. On the other hand, the level of PRiMA mRNA was not affected after the heat shock. In parallel with AChE(T) mRNA, the enzymatic activity of cellular AChE, in terms of G(1) and G(2) forms, was increased after heat shock; however, the PRiMA-linked G(4) remained unchanged. These results suggest that heat shock can induce the expression level of AChE(T) by the regulation of AChE(T) transcripts in NG108-15 cells.
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Ligustilide suppresses the biological properties of Danggui Buxue Tang: a Chinese herbal decoction composed of radix astragali and radix angelica sinensis.
Planta Med.
PUBLISHED: 10-21-2009
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Danggui Buxue Tang (DBT), a herbal decoction composed of Radix Astragali (RA) and Radix Angelica sinensis (RAS), has been used for treating menopausal irregularity in women for more than 800 years in China. According to the old tradition, RAS had to be processed with yellow wine before DBT preparation, which markedly reduced the amount of ligustilide in RAS and DBT, as well as enhanced the bioactivities of DBT. Here, we hypothesized that ligustilide would be an ingredient that possessed suppressive effects on DBTs functions. In the presence of ligustilide, the amount of astragaloside IV, calycosin, formononetin, and total polysaccharides extracted from RA were decreased. An increase of ligustilide caused a decrease of DBTs osteogenic activity in stimulating proliferation and differentiation of cultured bone cells. In addition, in the presence of a high level of ligustilide, DBT caused a side effect inducing the proliferation of breast MCF-7 cells. The current results strongly suggest that ligustilide is a negative regulator that hinders DBT to achieve its biological efficacy, which supports the traditional practice of preparing DBT using the ethanol-treated RAS.
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Two new compounds from the roots of Ligusticum chuanxiong.
J Asian Nat Prod Res
PUBLISHED: 08-28-2009
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Two new compounds, named ligusticoside A (1), a novel phthalide derivative with a lactone ring, and 4-pentylcyclohex-3-ene-1alpha,2beta-diol (2), were isolated from the rhizomes of Ligusticum chuanxiong. Their structures were determined by spectral methods (MS, IR, UV, 1D and 2D NMR). An X-ray diffraction experiment was performed to confirm the structure of compound 1.
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Quality evaluation of Rhizoma Belamcandae (Belamcanda chinensis (L.) DC.) by using high-performance liquid chromatography coupled with diode array detector and mass spectrometry.
J Chromatogr A
PUBLISHED: 07-28-2009
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A high-performance liquid chromatography coupled with diode array detector and mass spectrometry (HPLC-DAD-MS) method was developed to evaluate the quality of Rhizoma Belamcandae (Belamcanda chinensis (L.) DC.) through establishing chromatographic fingerprint and simultaneous determination of seven phenolic compounds. The analysis was achieved on an Alltima C(18) analytical column (250 mm x 4.6 mm i.d. 5 microm) using linear gradient elution of acetonitrile-0.1% trifluoroacetic acid. The correlation coefficients of similarity were determined from the HPLC fingerprints, and they shared a close similarity. By using an online APCI-MS/MS, twenty phenols were identified. In addition, seven of these phenols including mangiferin, 7-O-methylmangiferin, tectoridin, resveratrol, tectorigenin, irigenin and irisflorentin were quantified by the validated HPLC-DAD method. These phenols are considered to be major constituents in Rhizoma Belamcandae, and are generally regarded as the index for quality assessment of this herb. This developed method by having a combination of chromatographic fingerprint and quantification analysis could be applied to the quality control of Rhizoma Belamcandae.
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Expression and Localization of PRiMA-linked globular form acetylcholinesterase in vertebrate neuromuscular junctions.
J. Mol. Neurosci.
PUBLISHED: 07-06-2009
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Acetylcholinesterase (AChE) is well known to process different molecular forms via the distinct interacting partners. Proline-rich membrane anchor (PRiMA)-linked tetrameric globular AChE (G4 AChE) is mainly found in the vertebrate brain; however, recent studies from our laboratory have suggested its existence at neuromuscular junctions (nmjs). Both muscle and motor neuron express AChE at the nmjs. In muscle, the expression of PRiMA-linked AChE is down-regulated during myogenic differentiation and by motor neuron innervation. As compared with muscle, spinal cord possessed higher total AChE activity and contained PRiMA-linked AChE forms. The spinal cord expression of this form increased during development. More importantly, PRiMA-linked G4 AChE identified as aggregates localized at nmjs. These findings suggest that the restricted localization of PRiMA-linked G4 AChE at the nmjs could be contributed by the pre-synaptic motor neuron and/or the post-synaptic muscle fiber.
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Estrogenic and neuroprotective properties of scutellarin from Erigeron breviscapus: a drug against postmenopausal symptoms and Alzheimers disease.
Planta Med.
PUBLISHED: 06-16-2009
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Besides the classical hormonal effect, estrogen possesses neuroprotective effects in the brain, which leads to the searching of novel treatments for neurodegenerative diseases such as Alzheimers disease. Scutellarin is a major flavone derived from Herba Erigerontis, a Chinese medicine derived from Erigeron breviscapus, which has been shown here to possess both estrogenic and neuroprotective properties. Scutellarin showed the estrogenic effects by activating the estrogen responsive elements and phosphorylation of estrogen receptor alpha in cultured MCF-7 cells: the activation was in a dose-dependent manner. On the other hand, scutellarin inhibited the aggregation of beta-amyloid in vitro, and prevented the cell death mediated by beta-amyloid when applied to cultured neuronal PC12 cells. These results therefore suggested that Herba Erigerontis and its component scutellarin might have therapeutic effects against postmenopausal symptoms and Alzheimers disease.
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