Search

Spirocitromycetin, an antiosteoporotic polyketide bearing a unique spirocycle, was characterized from a human mucus sputum-derived

Epithelial-mesenchymal transformation(EMT) exists in embryonic development and is closely related to cell migration and invasion. The increased EMT level in tumors showed that E-cadherin was replaced by N-cadherin, and the expression of interstitial markers such as α-SMA and vimentin was up-regulated. It has been reported that lupeol can reduce the expression of matrix metalloproteinase-2(MMP-2), matrix metalloproteinase-9(MMP-9) and N-cadherin to inhibit the metastasis of osteoma cells. However lupeol has been less studied in liver cancer. Therefore, this paper investigated the effect of lupanol on invasion and metastasis of human hepatoma cell line HepG2 and SK-HEP-1 and its possible mechanism. MTT assay and Annexin V/PI double staining were used to investigate the effect of lupeol on activity and apoptosis of HepG2 cells and SK-HEP-1 cells. Moreover, the effect of lupeol on the invasion of HepG2 cells and SK-HEP-1 cells were evaluated by Transwell assay. The expressions of E-cadherin, N-cadherin, α-SMA, vimentin and MMP-9 were measured by Western blot. The model of subcutaneous transplantation of nude mice and the lung metastasis model of H22 hepatocellular carcinoma cells were established to evaluate the efficacy of lupeol in vivo on tumor growth and lung metastasis by HE staining combined with immunohistochemical assay. The results showed that lupeol inhibited the activity and invasion of HepG2 cells and SK-HEP-1 cells in a dose-dependent manner and induced apoptosis. Western blot showed that the expression of E-cadherin, a landmark protein for EMT, was induced by lupeol, and the expressions of N-cadherin, α-SMA, vimentin and MMP-9 were decreased. In vivo experiments showed that lupeol inhibited tumor growth in mice bearing xenograft. In addition, immunohistochemical experiments confirmed that lupeol could up-regulate the expression of E-cadherin in tumor tissues of nude mice, reduce the expression of N-cadherin, and inhibit the metastasis of liver cancer H22 cells in the lungs of mice. The above results indicated that the mechanism of lupeol inhibiting the invasion and metastasis of HCC cells may be related to the regulation of EMT process.

In present study, a comprehensive strategy integrating multiple chromatographic and chemometric methods to simultaneously characterize the volatile and non-volatile components was developed for the holistic quality evaluation of commercial Agastache rugosa (AR), a common edible and medicinal herb, collected in China. The volatile components and the non-volatile components were characterized by GC-MS and UPLC-QTOF-MS/MS, respectively. And the data were analyzed either independently or integratively by multivariate statistical analysis (MVS) for the quality assessment of commercial samples. The results revealed that the commercial AR samples were different in both the composition and the content of volatile components. However, the compositions of non-volatile components in commercial AR were generally similar, whereas the contents of some components were different. All the results indicated that the holistic quality of commercial AR was inconsistent, and the commercial samples collected could be classified into two main groups, the volatile components were majorly responsible for the classification. Whether or not the holistic quality variations affect the efficacy of AR deserves further investigation.

Acaulin A (1) and its macrolactone ring-opened congener acaulin B (2) were characterized from the culture of Acaulium sp. H-JQSF (an isopod-associated fungus) as architecturally undescribed trimeric macrodiolides, with the former being antiosteoporotic at 0.4 μM in the prednisolone-induced osteoporotic zebrafish. Identification of acaudiolic acid (3) as the monomeric macrodiolide precursor facilitated the proposal of the acaulin biosynthetic pathway.

Acaulide (1), a macrodiolide with an unprecedented framework, was characterized along with its shunt products-acaulones A (2) and B (3)-from the culture of Acaulium sp. H-JQSF associated with the isopod Armadillidium vulgare. The spiro-linked 14-, 14-, and 6-membered cycles of 1 arise likely from iterative intermolecular Michael addition reactions. Biological evaluation in the prednisolone-induced osteoporotic zebrafish demonstrated that 1 is antiosteoporotic at 0.4 and 2.0 μM.

Diterpenoid lactones (DLs) have been reported to be the main hepatotoxic constituents in Dioscorea bulbifera tubers (DBT), a traditional Chinese medicinal herb. The acquisition of early information regarding its metabolism is critical for evaluating the potential hepatotoxicity of DLs. We investigated, for the first time, the main metabolites of diosbulbin A (DIOA), diosbulbin C (DIOC), diosbulbin (DIOG), diosbulbin (DIOM) and diosbulbin (DIOF) in adult zebrafish. By using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF MS), 6, 2, 7, 5 and 4 metabolites of DIOA, DIOC, DIOF, DIOM and DIOG were identified in the zebrafish body and the aqueous solution, respectively. Both phase-I and phase-II metabolites were observed in the metabolic profiles and the metabolic pathways involved in hydroxyl reduction, glucuronidation, glutathione conjugation and sulfation. The above results indicated that hepatocytic metabolism might be the major route of clearance for DLs. This study provided important information for the understanding of the metabolism of DLs in DBT.

Based on metabolomics,the metabolites of larvae zebrafish with overdose of Panax notoginseng saponins( PNS) were compared with those in normal group of larvae zebrafish to investigate the possible toxicity mechanism of overdose PNS in larvae zebrafish. An experimental animal model of long-term toxicity induced by PNS overdose was established by administering 1-6 dpf at low,medium and high doses of PNS,respectively. The ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry( UPLC-Q-TOF-MS) technique was combined with principal component analysis( PCA) and orthogonal partial least squares discriminant analysis( OPLS-DA) to screen and identify biomarkers associated with toxicity,and then the MetaboAnalyst database was used to analyze metabolism-related pathways. The results showed that the metabolites of each group could be distinguished distinctly,and they deviated more from the normal group in a time and dose dependent manner. Twenty-nine potential biomarkers related to toxicity( VIP1,P0. 05) were identified preliminarily,mainly involving six metabolic pathways. From the metabonomics point of view,the toxicity mechanism of overdose PNS may be related to the disorders of lipid metabolism,amino acid metabolism and energy metabolism.

Diabetes is characterized by a loss and dysfunction of the β-cell. Glucagon-like peptide 1 receptor (GLP-1R) signaling plays an important role in β-cell survival and function. It is meaningful to identify promising agents from natural products which might activate GLP-1R signaling. In this study, puerarin, a diet isoflavone, was evaluated its beneficial effects on β-cell survival and GLP-1R pathway. We showed that puerarin reduced the body weight gain, normalized blood glucose, and improved glucose tolerance in high-fat diet-induced and db/db diabetic mice. Most importantly, increased β-cell mass and β-cell proliferation but decreased β-cell apoptosis were observed in puerarin-treated diabetic mice as examined by immunostaining of mice pancreatic sections. The protective effect of puerarin on β-cell survival was confirmed in isolated mouse islets treated with high glucose. Further mechanism studies showed that the circulating level of GLP-1 in mice was unaffected by puerarin. However, puerarin enhanced GLP-1R signaling by up-regulating expressions of GLP-1R and pancreatic and duodenal homeobox 1, which subsequently led to protein kinase B (Akt) activation but forkhead box O1 inactivation, and promoted β-cell survival. The protective effect of puerarin was remarkably suppressed by Exendin(9–39), an antagonist of GLP-1R. Our study demonstrated puerarin improved glucose homeostasis in obese diabetic mice and identified a novel role of puerarin in protecting β-cell survival by mechanisms involving activation of GLP-1R signaling and downstream targets.

Vacuum well point is a new but faint soft ground treatment method. This work focuses on the consolidation behavior of a reconstituted soft clayey specimen under vacuum well point combined with surcharge loading. The laboratory test was conducted through a vacuum-surcharge consolidation apparatus, and the vacuum loading scheme was adopted for vacuum pressure application to investigate the vacuum effect on soil consolidation. In the testing process, some key parameters such as vacuum pressure, pore water pressure and settlement deformation were timely recorded. Furthermore, the water content, void ratio and permeability coefficient of samples collected after loading were measured to reflect the consolidation characteristics. By comparing with the membrane system and membraneless system, something different was found for the vacuum well point method. The results indicate that the consolidation behavior of an axisymmetric single vacuum well point is almost identical to the behavior of vacuum preloading combined with prefabricated vertical drain (PVD), except for the distribution of the vacuum pressure along the well drain due to the structure of the vacuum well point. And the vacuum well point method may be useful for the improvement of soft clayey deposit in a certain depth.

Citrofulvicin (1), along with its early shunt product fulvionol (2), was characterized as a skeletally unprecedented antiosteoporotic agent from a human sputum-derived fungus Penicillium velutinum. The unique citrofulvicin framework is likely formed by a nonenzymatic intermolecular Diels–Alder cycloaddition between heptaketide-based intermediates. Citrofulvicin and fulvionol were demonstrated to be osteogenic at 0.1 μM in the prednisolone-induced osteoporotic zebrafish.

Diterpenoid lactones (DLs) have been reported to be the main hepatotoxic constituents in

The increasingly apparent liver injury problems of bone strengthening Chinese medicines have brought challenges for clinical application, and it is necessary to consider both effectiveness and safety in screening anti-osteoporosis Chinese medicines. Metabolic transformation is closely related to drug efficacy and toxicity, so it is significant to comprehensively consider metabolism-action/toxicity(M-Act/Tox) for screening anti-osteoporosis Chinese medicines. The current evaluation models and the number of compounds(including metabolites) severely restrict efficient screening in vivo. By referring to previous relevant research and domestic and abroad literature, zebrafish M-Act/Tox integrative method was put forward for efficiently screening anti-osteoporosis herb medicines, which has organically integrated zebrafish metabolism model, osteoporosis model and toxicity evaluation method. This method can break through the bottleneck and blind spots that trace compositions can't achieve efficient and integrated in vivo evaluation, and realize both efficient and comprehensive screening on anti-osteoporosis traditional medicines based on in vivo process taking both safety and effectiveness into account, which is significant to accelerate discovery of effective and safe innovative traditional Chinese medicines for osteoporosis.

Metabolites derived from herbal compounds are becoming promising sources for discovering new drugs. However, the rapid identification of metabolites from biological matrixes is limited by massive endogenous interference and low abundance of metabolites. Thus, by using zebrafish larvae as the biotransformation system, we herein proposed and validated an integrated strategy for rapid identification of metabolites derived from herbal compounds. Two pivotal steps involved in this strategy are to differentiate metabolites from herbal compounds and match metabolites with their parent compounds. The differentiation step was achieved by cross orthogonal partial least-squares discriminant analysis. Automatic matching analysis was performed on R Project based on a self-developed program, of which the number of matched ionic clusters and its corresponding percentage between metabolite and parent compound were taken into account to assess their similarity. Using this strategy, 46 metabolites screened from incubation water samples of zebrafish treated with total Epimedium flavonoids (EFs) could be matched with their corresponding parent compounds, 37 of them were identified and validated by the known metabolic pathways and fragmentation patterns. Finally, 75% of the identified EFs metabolites were successfully detected in urine samples of rats treated with EFs. These experimental results indicate that the proposed strategy using zebrafish larvae as the biotransformation system will facilitate the rapid identification of metabolites derived from herbal compounds, which shows promising perspectives in providing additional resources for pharmaceutical developments from natural products.

Ginsenosides F

Soil conditioning plays an important role in reducing soil abrasivity and wear of cutting tools in earth pressure balance shield tunneling, leading to service life extension of cutting tools. In this article, an experimental study to assess the abrasivity of conditioned soil with different conditioners, namely foam and slurry, was carried out using a soil abrasion tester. The abrasive characteristics of the tested soil samples were quantitatively assessed through the measurement of weight loss of the steel cubes fixed on the propeller. Preliminary test results indicate that both foam and slurry can modify the sandy gravel to a suitable state according to slump test and decrease soil abrasivity significantly. The torque, which reflects the interaction between the propeller and tested materials, is used to provide insights into the soil-tool contact. Potential relationships between tool wear and tool position are also investigated.

The safety of traditional Chinese medicine (TCM) has received the widespread attention in recent years. Hepatotoxicity of TCM is one of the key problems of the safety of TCM. This article summarized research progress and application prospect in the mechanism of TCM hepatotoxicity, biomarkers, toxic omics database, prevention of hepatotoxicity of the liver cell lines, subcellular fraction, three-dimensional cultivation models, the model animals, aiming to provide theoretical basis for TCM toxicity evaluation and technical guidelines, thus promoting the development of TCM toxicity studies. Hope for Chinese medicine liver toxicity evaluation method provides the theoretical foundation and technical guidelines, promote the development and improvement of TCM liver toxicity research system.

Background Gualou Xiebai Decoction (GXD) is a well-known traditional Chinese recipe. It has been used to treat cardiovascular disorders for nearly two thousand years. But there is a lack of reports on cardiac fibrosis and underlying mechanism. Methods Myocardial infarction was performed by ligation of left anterior descending coronary artery (LAD) in male Wistar rats. Rats with myocardial infarction were treated with GXD (1.14 g/kg, 4.53 g/kg) daily for 4 weeks. Cardiac function was evaluated by echocardiography. Hemodynamic parameters and infarct size were measured in each group. Myocardial enzymes were examined by biochemical tests. Inflammatory cytokines were assessed by ELISA, and interrelated proteins were detected by western blot. Results Cardiac function was significantly improved in GXD-treatment rats after myocardial infarction (MI), which was accompanied with decreased infarct size. Administration of GXD to myocardial fibrosis rats significantly ameliorated the activities of AST, LDH and CK-MB in serum. The increase in inflammatory factors (TNF-α, IL-1β) were markedly reduced upon GXD treatment. Furthermore, the inflammatory mediators (NF-κB p65, TNF-α, MCP-1) were down-regulated by GXD in the myocardial fibrosis rats. Conclusions Treatment with GXD improved cardiac function induced by myocardial fibrosis by inhibiting expression of inflammatory mediators associated with NF-κB.

Cavitation typically occurs when the fluid pressure is lower than the vapor pressure at a local thermodynamic state. The aim of this paper is a numerical investigation of the cryogenic cavitation flow characteristics, considering variable thermodynamic properties of liquid nitrogen and numerical simulation liquid nitrogen around hydrofoil cryogenic cavitation flow characteristics. Based on homogeneous flow model and Zwart cavitation model, calculates hydrofoil isothermal and cryogenic cavitation in liquid nitrogen steadily, updates the evaporation and condensation coefficients of Zwart cavitation model, gives the hydrofoil surface pressure profile, temperature depression and distribution of cavitation intensity, contrasts the isothermal and cryogenic cavitation flow characteristics. Numerical results show that thermodynamics effect in cryogenic liquid cavitation significantly. Meanwhile, the hydrofoil surface pressure and temperature numerical results with experimental data and more Hord compared to verify the validity of the numerical simulation.

Cutting tool wear is a critical problem for tunneling in soft ground with the earth pressure balance shield. In this study, a cutting tool tester was developed in the China University of Geosciences in Beijing, and a series of tool wear tests were conducted to investigate the characteristics of tool wear in granular soils. The characteristics of cutting tool wear can be quantitatively assessed through the ratio (M0 − M)/M0 (i.e., tool wear rate, %; M0 and M are the mass of the tested blocks before and after the wear test, respectively). The effects of five important factors, including mineralogy, moisture, soil conditioner, test time, and sand content, on tool wear were considered in various test conditions. According to the primary test results, it is observed that (1) high soil hardness produced significantly higher tool wear than low soil hardness in dry condition; (2) with increasing water content, the tool wear gradually increased to a maximum value and then decreased; (3) the foam conditioner mixed in sandy gravel reduced the tool wear dramatically regardless of the cutting tool material, e.g., cemented carbide and Q235 steel; (4) the tool wear increased with the test time in the dry sand mixture, and their relationship showed a good agreement with a power function; and (5) the tool wear decreased as the sand content increased in dry sand-gravel soil mixtures. The reasons for these phenomena were also explored.

During water-exit of underwater vehicle, the high pressure， which the local cavity collapse of underwater vehicle causes, poses a great challenge to the vehicle structure. The cavity collapse during vertical water exit of an underwater vehicle is simulated by a viscous flow solver with homogeneous multiphase flow model. The shrinkage and collapse of cavity zone and violent changes of the pressure field have been captured during the simulation. The changes of cavity shape and pressure field at different water exit velocity have been analyzed. It has been concluded that higher velocity may cause the delay of cavity total collapse and the max pressure point to transfer backwards.

Based on the movement characteristics of the supercavitating underwater vehicle, the tail impact force has been defined by summarizing related papers, then the dynamic responses of structure has been simulated by using Finite Element Method. The displacement and acceleration responses and corresponding spectrums have been gotten. The calculation results show that: the displacement response along impact force's direction oscillates periodically; the displacement response along body axis lengthens and shortens periodically; the displacement response along the 3rd axis increases over time. The acceleration response along impact force's direction and body axis oscillates periodically. The corresponding spectrums show that: accessories of the underwater vehicle should avoid some scaled frequencies in order to let them run reliably.

Caudatin-2,6-dideoxy-3-O-methy-β-D-cymaropyranoside (CDMC), the C-21 steroidal glycoside recently extracted from the traditional Chinese medicinal plant, the root of Cynanchum auriculatum Royle ex Wight (Asclepiadaceae), has been shown to possess potent antitumor properties. However, the bioactivities of CDMC are still largely unknown, especially the antitumor effect and its mechanism. This study investigated the CDMC antitumor effects on human hepatoma cell line SMMC7721 cells by analysis of cell viability, cell cycle phases and apoptosis. The results showed that CDMC inhibited the growth of SMMC7721 cells in a time- and dose-dependent manner and resulted in cell cycle arrest in G(0)/G(1) phase. Furthermore, CDMC induced SMMC7721 cell apoptosis rather than necrosis through caspase 3 activation, and a caspase 3 inhibitor, Ac-DEVD-CHO, could attenuate the apoptosis induced by CDMC. The results suggested that the anticancer activity of CDMC could be attributed partially to its inhibition of cell proliferation and induction of apoptosis associated with caspase 3 activation.

We experimentally study the cavity formation when heated spheres impact onto water at low and high subcooling. The observations present that the formation and appearance of the cavity are affected by the boiling modes and the heat transfer intensity. In the nucleate-boiling regime, a rough cavity can be formed at a rather low impact velocity, while at the same velocity, the cavity formed in the film-boiling regime may have a very smooth interface with a stable vapor layer around the sphere. We discuss the effects of the impact speed, water and sphere temperatures on the stability of the vapor layer. For low subcooled water, the stable vapor layer will be disturbed when increasing the impact velocity, leading to a disturbed cavity. For high subcooled water, the film boiling has a particular boiling model in which the vapor layer around the sphere cannot keep its stability. In this particular film-boiling regime, no cavities can be formed at low impact velocities and only broken cavities can be formed at high impact velocities.

Background and Objectives: Bone marrow derived mesenchymal stem cells (BMSCs) have shown to be an appealing source for cell therapy and tissue engineering. Previous studies have confirmed that the application of low-level laser irradiation (LLLI) could affect the cellular process. However, little is known about the effects of LLLI on BMSCs. The aim of this study was designed to investigate the influence of LLLI at different energy densities on BMSCs proliferation, secretion and myogenic differentiation. Study Design/Materials and Methods: BMSCs were harvested from rat fresh bone marrow and exposed to a 635 nm diode laser (60 mW; 0, 0.5, 1.0, 2.0, or 5.0 J/cm 2 ). The lactate dehydrogenase (LDH) release was used to assess the cytotoxicity of LLLI at different energy densities. Cell proliferation was evaluated by using 3-(4, 5-dimethylithiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) and 5-bromo-2 0 -deoxyuridine (BrdU) assay. Production of vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) were measured by enzyme-linked immunosorbent assay (ELISA). Myogenic differentiation, induced by 5-azacytidine (5-aza), was assessed by using immunocytochemical staining for the expression of sarcomeric a-actin and desmin. Results: Cytotoxicity assay showed no significant difference between the non-irradiated group and irradiated groups. LLLI significantly stimulated BMSCs proliferation and 0.5 J/cm 2 was found to be an optimal energy density. VEGF and NGF were identified and LLLI at 5.0 J/cm 2 significantly stimulated the secretion. After 5-aza induction, myogenic differentiation was observed in all groups and LLLI at 5.0 J/cm 2 dramatically facilitated the differentiation. Conclusions: LLLI stimulates proliferation, increases growth factors secretion and facilitates myogenic differentiation of BMSCs. Therefore, LLLI may provide a novel approach for the preconditioning of BMSCs in vitro prior to transplantation. Lasers Surg. Med. 40:726–733, 2008. 2008 Wiley-Liss, Inc.

A high-performance liquid chromatography method with evaporative light scattering detection was established for simultaneous determination of three major triterpenoid glycosides, i.e. asiaticoside, madecassoside and asiaticoside-B, in Centella asiatica extracts. The optimal chromatographic conditions were achieved on a COSMOSIL 5C(18)-MS-II column by constant elution with water (0.01% trifluoroacetic acid, v/v) and acetonitrile (1.0% methyl tert-butyl ether, 0.01% trifluoroacetic acid, v/v) (78:22) as mobile phase at a flow rate of 1.0 mL/min; the column temperature was 30 degrees C. The evaporative light scattering detector was set at an evaporating temperature of 40 degrees C and nitrogen gas pressure of 3.5 bar. The validation of the method included tests of linearity, sensitivity, precision, repeatability, stability and accuracy. All calibration curves showed good linear regression (r(2) > 0.9993) within test ranges. The established method showed good precision and accuracy with overall intra-day and inter-day variations of 1.73-3.06 and 3.89%-4.92%, respectively, and overall recoveries of 97.63-99.39% for the three compounds analyzed. The method developed was successfully applied to quantify the main triterpenoid glycosides in Centella asiatica extracts from different companies.

A method for predicting bioactive candidates in herbal medicines (HMs) was developed using four types of target cell extraction followed by high performance liquid chromatography coupled with diode array detection-mass spectrometry (HPLC-DAD-MS) analysis. Through the proposed method, some potential bioactive components in Radix Salviae miltiorrhizae, a widely used HM for the treatment of cardiovascular diseases, were screened in view of interacting with endothelial cells, myocardial cells, blood platelets and red cells. Specifically, eleven compounds were detected in the desorption eluate of endothelial cells, nine in myocardial cells, fifteen in blood platelets and five in red cells. Totally, sixteen compounds were detected and fourteen of them were identified qualitatively in terms of their MS spectra and in comparison with some of the reference compounds. The results indicate that the proposed method may be applied to prediction of the bioactive multi-compounds in HMs.

A herb-combined prescription, mainly derived from roots of Salvia miltiorrhiza and Panax notoginseng, has been widely used for improving coronary or cerebral circulation in China as well as in Western countries. Multiple commercially available preparations, known as Fufang Danshen preparations (FDPs), produced by various manufacturers with the raw materials from different sources, pose a serious challenge to the quality control of this herb medicine. Previous pharmacological studies identify three types of bioactive components correlated with the clinical effect of those herb preparations. Those mainly include four phenolic acids, four saponins and four diterpenoid quinones. In this report, by using high performance liquid chromatography (HPLC) coupled with diode array and evaporative light scattering detectors (DAD-ELSD), we developed an improved quality control method for those herb medicines. A simultaneous separation and quantification of the 12 components was performed on a C(18) column, in which the mobile phase consisted of (A) 0.1% aqueous formic acid and (B) acetonitrile using a gradient elution. The optimum detection wavelength was set at 281 nm, the drift tube temperature of ELSD was set at 113 degrees C, the nitrogen flow rate at 3.1L/min, and the gain=4. All calibration curves showed good linear regression (r(2)>0.9927) within test ranges. The method developed showed good precision and accuracy with overall intra- and inter-day variations of 0.64-4.79% and 0.69-4.96%, respectively, and the overall recoveries of 93.50-107.69% for the 12 compounds analyzed. This method was successfully applied to quantify the twelve components in ten commercial samples from three formulas by seven independent manufacturers. This readily available, low-cost and reliable HPLC-DAD-ELSD method improved the quality control of traditional Chinese medicinal preparations consisting of complex compounds with different structures such as FDPs.

The objective of this research was to establish a simple, practical and efficient method for routine quantitative analysis of Erigeron breviscapus and its extract injection to control their qualities. The reversed phase high performance liquid chromatographic method was adopted to determine simultaneously the contents of two major classes of constituents namely phenolic acids and flavonoids, which were usually ignored in previous studies of E. breviscapus. Under the optimum conditions, three flavonoids including scutellarin, scutellarein and apigenin and four phenolic acids including caffeic acid, chlorogenic acid, 3,4-O-dicaffeoylquinic acid and 3,5-O-dicaffeoylquinic acid were successfully separated on a Zorbax SB-C18 column (250 × 4.6 mm I.D., 5.0 μm particle size) at 25 °C. Of the three flavonoids, scutellatin is a flavone glucuronide. The mobile phase was a mixture of acetonitrile and 1.0% (v/v) aqueous acetic acid employing gradient elution at a flow rate of 1.0 mL min−1 and the detection wavelength was set at 330 nm. Regression equations revealed good linear relationship between the peak areas of the analytes and their concentrations (r 2 > 0.9990). The relative standard deviations of retention time and peak area were less than 0.33 and 1.45%. The intra- and inter-day precisions as determined from sample solutions were below 1.66 and 2.35%. And the recoveries ranged from 96.5 to 101.8%. The proposed method has been successfully applied to the simultaneous quantification of two major classes of constituents in E. breviscapus and its extract injection for the first time.

Bone marrow-derived mesenchymal stem cells (MSCs) are of great therapeutic potential after myocardial ischemic injury. However, little is known about the biological characteristics of MSCs in patients with coronary artery disease and their effects on infracted myocardium. The present study evaluated the biological characteristics of MSCs from patients with coronary artery disease and their effects after being transplanted into infarcted myocardium using a rat model.Sternal bone marrow aspirates were taken at the time of coronary artery bypass graft surgery. Mononuclear cells isolated from bone marrow were cultured based on plastic adherence. The morphology and growth characteristics of MSCs were observed in primary and successive passages. A myocardial infarction model was created in 27 adult rats. Two weeks later, animals were randomized into two groups: culture medium (group I, n=13) or MSCs (2x10(6)) from early passages labeled with BrdU (group II, n=14) were injected into the infarcted myocardium. Echocardiography, histological examination, and reverse transcription-polymerase chain reaction (RT-PCR) were performed four weeks after cell transplantation.Flow cytometry analyses demonstrated that adherent spindle cells from bone marrow are mesenchymal stem cells (positive for CD29 and CD44, but negative for CD34 and CD45). Growth curves showed that MSCs have great proliferative capability especially at early passages. MSCs implantation in the infarcted border zone improved left ventricular function significantly in group II compared with group I. However, despite improved left ventricular function, we did not observe significant regeneration of cardiac myocytes. Immunohistochemistry revealed only the expression of desmin in the engrafted MSCs, a marker of premature myocyte. Moreover, the improved left ventricular function in this study seemed to be secondary to the beneficial reverse remodeling induced by the increase of collagen in infarcted zone, the decrease in the adjacent myocardium, and the increase of neovascularization (capillary density: 192+/-7.8/mm2 in group II vs. 165+/-5.9/mm2 in group I, P0.05). Reverse transcription-polymerase chain reaction (RT-PCR) results showed the expression levels of collagen I, collagen III, SDF-1 (stromal cell-derived factor-1), and VEGF (vascular endothelia growth factor) in the infarcted border zone were significantly higher in the MSCs treated group.The MSCs from patients with coronary artery disease have a typical phenotype with highly proliferative potential and the engrafted MSCs may regulate extracellular collagens and cytokines to prevent the ventricular scar from pathologic thinning and attenuate the contractile dysfunction of the infarcted heart.

High-performance liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) methods were developed for the analysis of chemical and metabolic components in traditional Chinese medicinal combined prescription containing Radix Salvia miltiorrhiza and Radix Panax notoginseng (commonly known as Fufang Danshen prescription, FDP). The HPLC experiments used a reversed-phase Zorbax C(18) column with the column temperature at 30 degrees C and a binary mobile phase system consisting of aqueous formic acid (0.1%, v/v) and acetonitrile using a gradient elution at the flow rate of 1.0 mL/min. The ESI-MS was operated with a single-quadrupole mass spectrometer in both negative and positive ion modes. 36 major chromatographic peaks of FDP, including 14 saponins, 13 phenolic acids and nine diterpenoid quinones were characterized by their MS spectra and in comparison with some of the reference standards. In addition, after oral administration of extraction of FDP, the rat's plasma, urine and feces were also analyzed; 53 metabolic components including 30 original components and 23 transformative components of FDP were detected, and possible metabolic pathways of some components in FDP were given. The analysis of chemical and metabolic components in FDP by HPLC-MS methods could be a useful means of identifying the multi-components of FDP and to hint at their possible metabolic mechanism of action in the body.

Mononuclear cells (MNCs) isolated from peripheral blood by density gradient centrifugation were plated on human fibronectin-coated culture plates and cultured in EGM-2 medium. Attached spindle-shaped cells, reported as endothelial progenitor cells (EPCs) by some investigators, had elongated from adherent round cells, but had not proliferated from a small number of cells as supposed previously. The growth curve of the primary EPCs showed that the cells had little proliferative capacity. Flow cytometry analysis showed that the cells could express some of the endothelial lineage markers, while they could also express CD14, which is considered a marker of monocyte/macrophage lineages throughout culture. In endothelial function assays, the cells demonstrated a lower level of expression of eNOS than mature endothelial cells in the reverse transcription-polymerase chain reaction and did not show an ability to develop tube-like structures in angiogenesis assay in vitro. In this study, we identified the monocytoid function of EPCs by the combined Dil-labeled acetylated low-density lipoprotein (Dil-Ac-LDL) and Indian ink uptake tests. All the cells were double positive for Dil-Ac-LDL and Indian ink uptake at days 4, 14 and 28 of culture, which means the EPCs maintained monocytoid function throughout the culture. Therefore, although adult EPCs from peripheral MNCs have some endothelial lineage properties, they maintain typical monocytic function and have little proliferative capacity.

A high-performance liquid chromatography method was established for simultaneously determining seven major components, i.e. protocatechuic aldehyde, notoginsenoside R1, ginsenoside Rg1, salvianolic acid B, ginsenoside Rb1, cryptotanshinone and tanshinone IIA in Fufang Danshen tablet, a commonly used traditional Chinese medicinal combined prescription mainly derived from the roots of Salvia miltiorrhiza and Panax notoginseng. These seven compounds, belonging to the chemical types of phenolic acids, diterpenoid quinones and saponins, were simultaneously separated on Zorbax C18 column (250 × 4.6 mm, 5.0 µm) with the column temperature at 30°C. The mobile phase was composed of (A) aqueous phosphoric acid (0.1%, v/v) and (B) acetonitrile using a gradient elution of 7–17% B at 0–10 min, 17–20% B at 10–12 min, 20–21% B at 12–16 min, 21% B at 16–32 min, 21–29% B at 32–40 min, 29–35% B at 40–55 min, 35–65% B at 55–65 min and 65–80% B at 65–80 min; the flow rate was 1.0 mL/min. Detection wavelengths were set at 203 nm for notoginsenoside R1, ginsenoside Rg1 and ginsenoside Rb1, 281 nm for protocatechuic aldehyde, salvianolic acid B, and 270 nm for cryptotanshinone and tanshinone IIA. All calibration curves showed good linear regression (r2 > 0.9992) within test ranges. The established method showed good precision and accuracy with overall intra-day and inter-day variations of 0.15–4.35 and 0.61–5.17% respectively, and overall recoveries of 94.8–102.1% for the seven compounds analyzed. The developed method has been successfully applied to simultaneous evaluation of the intrinsic quality of both Danshen and Sanqi in Fufang Danshen tablets from different pharmaceutical companies. Copyright © 2006 John Wiley & Sons, Ltd.

Prednisolone-induced osteoporosis model using zebrafish was used to screen the antiosteoporotic active parts of Dipsacus Radix, in order to investigate the applicability and rationality of the zebrafish model of osteoporosis.Zebrafish larvae at 5 days post fertilization (dpf) were exposed with 25 micromol/L prednisolone and 0.5% DMSO for 48 h, then except one group of 25 micromol/L prednisolone, other groups of 25 micromol/L prednisolone were treated with a range of concentration (0.025, 0.25, 2.5, 25 microg crude drug/mL) of extract of Dipsacus Radix and its different concentration ethanol elution parts of macroporous resin with 25 micromol/L prednisolone. All groups were incubated in 24-well plates (28.5 degrees C) until 10 dpf. Zebrafish skeleton at 10 dpf were anesthetized and fixed for staining with alizarin red. Quantitative analysis of the stained area was performed by microscopic inspection and digital imaging methods to reflect the amount of zebrafish head skeleton mineralization.The results indicated that head skeleton mineral area and integrated optical density (IOD) of 25 micromol/L prednisolone model group were significantly decreased when compared with vehicle control group, and the extract of Dipsacus Radix and its 30%, 50%, 70% and 90% ethanol elution parts of macroporous resin rescued the further bone loss of zebrafish induced by prednisolone when compared with the model group. HPLC analysis indicated that components of 30%, 50%, 70% and 90% ethanol elution parts of macroporous resin containing saponins and nonsaponins components.Both saponins and nonsaponins can prevent bone loss of zebrafish induced by prednisolone. This novel osteoporosis zebrafish model was successfully used to screen antiosteoporotic active parts of Dipsacus Radix, which had advantages of simple, high efficiency and easy to perform.

This article firstly established a new efficient method for screening anti-osteoporosis ingredients, which used two-dimensional zebrafish model combined with hyphenated chromatographic techniques to evaluate anti-osteoporosis activities of epimedin A and its metabolite baohuoside I. Adult zebrafish was used for metabolism of epimedin A in 0.5% DMSO, and LC-MS was used for analysis of the metabolite, which was captured by HPLC, and prednisolone-induced osteoporosis model of zebrafish was used to evaluate the anti-osteoporotic activities of trace amounts of epimedin A and baohuoside I. The results indicated that epimedin A and baohuoside I can prevent prednisolone-induced osteoporosis in zebrafish. The developed method in this paper enables the separation, enrichment and analysis of micro-amount metabolite of epimedin A, and anti-osteoporosis activities in vivo of epimedin A and baohuoside I was simple and efficient screening resorting to zebrafish osteoporosis mode. This paper would provide new ideas and methods for a rapid and early discovery of anti-osteoporosis activities of micro-ingredients and its metabolite of traditional Chinese medicine.

To break through the restrictions of the evaluation model and the quantity of compounds by using the two-dimensional zebrafish model combined with chromatographic techniques, and establish a new method for the high-throughput screening of active anti-osteoporosis components.According to the research group-related studies and relevant foreign literatures, on the basis of the fact that the zebrafish osteoporosis model could efficiently evaluate the activity, the zebrafish metabolism model could efficiently enrich metabolites and the chromatographic techniques could efficiently separate and analyze components of traditional Chinese medicines, we proposed that the inherent combination of the three methods is expected to efficiently decode in vivo and in vitro efficacious anti-osteoporosis materials of traditional Chinese medicines.The method makes it simple and efficient in the enrichment, separation and analysis on components of traditional Chinese medicines, particularly micro-components and metabolites and the screening anti-osteoporosis activity, fully reflects that efficacious materials of traditional Chinese medicines contain original components and metabolites, with characteristic of "multi-components, multi-targets and integral effect", which provides new ideas and methods for the early and rapid discovery of active anti-osteoporosis components of traditional Chinese medicines.

Based on practice of Epimedium processing mechanism for many years and integrated multidisciplinary theory and technology, this paper initially constructs the research system for processing mechanism of traditional Chinese medicine based on chemical composition transformation combined with intestinal absorption barrier, which to form an innovative research mode of the " chemical composition changes-biological transformation-metabolism in vitro and in vivo-intestinal absorption-pharmacokinetic combined pharmacodynamic-pharmacodynamic mechanism". Combined with specific examples of Epimedium and other Chinese herbal medicine processing mechanism, this paper also discusses the academic thoughts, research methods and key technologies of this research system, which will be conducive to systematically reveal the modem scientific connotation of traditional Chinese medicine processing, and enrich the theory of Chinese herbal medicine processing.

As beneficial traditional Chinese medicine, Epimedium fried with suet oil can enhance the effect of warming kidney yang. Previous literature studies about processing of Epimedium mainly focused on changes in chemical composition and pharmacological effect. From the angle of flavonoids absorption and metabolism, our group innovatively study the processing mechanism of Epimedium based on active component transformation combined with intestinal absorption barrier. The processing effect of fried Epimedium can be divided into two key aspects of " heat" during processing and processing accessories "suet oil". Through continuous three National Natural Science Foundation items, the research group has clarified the scientific connotation of "heat" during processing with ADME, and explains the synergistic mechanism of processing accessories "suet oil" based on self-assembled micelles formation in vivo for the first time. This paper summarizes the research ideas and results of Epimedium processing mechanism of the project team for many years, and discusses the future research direction and content, in order to provide new ideas and new methods for modern Chinese medicine processing mechanism.

In this report, reporter gene β-galactosidase (LacZ) was chosen to compare two different intramuscular gene transfer methods, direct injection and gene suture. Evidence showed that gene suture can produce a higher foreign gene express efficiency in skeletal muscle compared with the direct injection method. The highly efficient eukaryotic expressing vectors of human atrial natriuretic factor (ANF) were constructed (pcD2/pAdVAntage TM /hANF and pcDNA3/hANF), and in vivo ANF gene delivery was performed by intramuscular gene suture. The effects of ANF gene transfer on blood pressure and renal sodium and water excretion were studied in three models of hypertensive animals. Results showed that a marked decrease of mean arterial pressure (MAP) and a significant increase of urine volume and urinary sodium excretion was produced in rats receiving the hANF construct due to the local expression of ANF and its secretion into plasma. Taken together, these results indicate that gene suture may represent a novel gene delivery modality in gene therapy.

Model organism zebrafish was used to study metabolism of asperosaponin VI from Dipsacus asper Wall. ex Henry for the first time. Metabolic components of asperosaponin VI after exposing to zebrafish for 24 h were identified by high performance liquid chromatography--electrospray mass spectrometry (HPLC-ESI-MS), the separation was performed with a Zorbax C18 column using a binary gradient elution of 0.05% formic acetonitrile--0.05% formic acid water. The quasi-molecular ions of compounds in both negative and positive mode were observed for molecule mass information, and the potential structures were identified by attentive study on the deglycosylation metabolites and one hydroxylation metabolite of asperosaponin VI. The results were highly in consistent with metabolism of asperosaponin VI in rat. It can be concluded that zebrafish model can wonderfully imitate current metabolic model with advantages of small amount of lower cost, far less amount compound, higher efficiency and more simple, and can reflect integrated metabolism results of in vivo method. Zebrafish metabolic model may become a novel organism model for quick predication on metabolism of even mircoamount compound, which can enrich the available models greatly.

Zebrafish was selected as model animal, and glucocorticoid dexamethasone was used as a model compound to establish a rapid and high efficient osteopenia model. Zebrafish larvae at 4 days post fertilization (dpf) were exposed to a serial concentrations of dexamethasone solutions, and 0.5% DMSO was selected as the vehicle control group. All groups were incubated in 24-well plates (28.5 degrees C) until 9 dpf. In addition, effects of 10 micromol x L(-1) dexamethasone on preventing against osteopenia induced by etidronate disodium were also investigated. Zebrafish bones at 9 dpf were stained with alizarin red. Quantitative analysis of the stained area was performed by microscopic inspection and digital imaging methods to reflect the amount of bone mineralization. Results showed that dexamethasone group at 2.5, 10 and 25 micromol x L(-1) can decrease the staining area and the staining optical density values of zebrafish head bones when compared with the vehicle control group (0.5% DMSO), which suggested that dexamethasone can significantly reduce the zebrafish mineralized bone and the bone mineral density. Results also showed that 15 and 30 microg x mL(-1) etidronate disodium can increase the mineralized matrix of zebrafish head bone and prevent against osteopenia induced by dexamethasone. In conclusion, the study indicated that zebrafish can be an idea osteopenia model induced by dexamethasone.