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1.
Article in Chinese | WPRIM | ID: wpr-888069

ABSTRACT

Sanguinarine is the main active component of the Papaver plants, and protopine-6-hydroxylase(P6 H), involved in the sanguinarine biosynthetic pathway, can oxidize protopine to 6-hydroxyprotopine. The investigation on the diversity of P6 H genes in the medicinal Papaver plants contributes to the acquirement of P6 H with high activity to increase the biosynthesis of sanguinarine. Five P6 H genes in P. somniferum, P. orientale, and P. rhoeas were discovered based on the re-sequencing data of the Papaver species, followed by bioinformatics analysis. With the elongation factor 1α(EF-1α), which exhibits stable expression in the root and stem, as the internal reference gene, the transcription levels of P6H genes in roots and stems of the Papaver plants were detected by real-time fluorescent quantitative PCR. As indicated by the re-sequencing results, there were two genotypes of P6H in P. somniferum and P. orientale, respectively, and only one in P. rhoeas. The bioinformatics analysis showed that the P6 H proteins of the three Papaver plants contained the conserved domain cl12078, which is the characteristic of p450 supergene family, and transmembrane regions. The existence of signal peptide remained verification. Real-time fluorescent quantitative PCR results revealed that the transcription level of P6 H in roots of P. somniferum was about 1.44 times of that in stems(α=0.05). The present study confirmed genetic diversity of P6 H in the three medicinal Papaver plants, which lays a basis for the research on the biosynthesis pathway and mechanism of sanguinarine in Papaver species.


Subject(s)
Benzophenanthridines , Berberine Alkaloids , Cytochrome P-450 Enzyme System/genetics , Genetic Variation , Papaver/genetics
2.
Acta Pharmaceutica Sinica ; (12): 207-210, 2015.
Article in Chinese | WPRIM | ID: wpr-251793

ABSTRACT

Ongoing study on the chemical constituents of the roots of Macleaya microcarpa led to the isolation of eight compounds of derivatives of triterpenes and organic acids in addition to some previously identified benzophenanthridines. The eight compounds were identified by spectroscopic methods as well as comparison with literature values as 1-oxo-2, 22 (30)-hopandien-29-oic acid (1), 3-oxo-12-oleanen-30-oic acid (2), 3α-hydroxy-12-oleanen-30-oic acid (3), 3β-hydroxy-12-oleanen-30-oic acid (4), ferulic acid (5), ferulic acid 4-O-β-D-glucoside (6), 3-O-feruloylquinic acid (7), and methyl 3-O-feruloylquinate (8). Of which, 1 is a new triterpenoid of hopanes and 2-8 are isolated from M microcarpa for the first time. In order to discover natural active compounds as potential agents of anti-ulcerative colitis (UC), an in vitro drug high-throughput screening model targeted x-box-binding protein 1 (xbp1) was employed to evaluate the activity of the major chemical constituents of M microcarpa. The result confirmed that two dihydrobenzophenanthridines, dihydrosanguinarine (9) and dihydrochelerythrine (10), showed a certain activity on activating the transcription of xbpl, a transcription factor (TF) associated with the occurrence, development, and potential treatment of UC, with their relative activating ratios being 1.76 and 1.77 times, respectively, as compared with control group.


Subject(s)
Anti-Ulcer Agents , Chemistry , Benzophenanthridines , Chemistry , DNA-Binding Proteins , Genetics , Isoquinolines , Chemistry , Papaveraceae , Chemistry , Plant Roots , Chemistry , Regulatory Factor X Transcription Factors , Transcription Factors , Genetics , Transcription, Genetic , Triterpenes , Chemistry
3.
Article in Chinese | WPRIM | ID: wpr-341867

ABSTRACT

Endophytic fungi were isolated from Macleaya cordata growing in Dabie Mountain by agar-block method, and then the endophytic fungi were grouped into different types based on their morphological characteristics, and thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) were employed to determine whether the metabolic substances contained sanguinarine or not, and then preliminarily identified by morphological method. The results showed that the leaves hosted the largest number of endophytes (96 isolates) followed by the stems (57 isolates) and finally the roots (28 isolates), respectively. Based on morphological characteristics the endophytic fungi were grouped into 26 types in our study. TLC and HPLC results showed that there was sanguinarine in the metabolic substances of BLH 51 strain. According to the morphological characteristic, the BLH 51 strain was identified as Fusarium proliferatum. All these indicated that the medicinal plant M. cordata harbors abundant endophytes, which could be a new source for the search of active secondary metabolites.


Subject(s)
Benzophenanthridines , Metabolism , Endophytes , Fungi , Isoquinolines , Metabolism , Papaveraceae , Metabolism , Microbiology , Plant Leaves , Microbiology , Plant Roots , Microbiology , Plant Stems , Microbiology
4.
Article in Chinese | WPRIM | ID: wpr-238652

ABSTRACT

Chelerythrine is a kind of benzo[c] phenanthridine alkaloids, with such pharmacological activities as antitumor, antibiosis and anti-inflammation, which is widely found in plant of Fumariaceae, Papaveraceae, Ranunculaceae and Rutaceae families. This article summarizes the advances in domestic and foreign studies on pharmacological effect of chelerythrine in the recent decade, in the expectation of providing scientific basis for the in-depth studies, development and utilization of chelerythrine.


Subject(s)
Animals , Anti-Inflammatory Agents , Pharmacology , Antineoplastic Agents, Phytogenic , Pharmacology , Benzophenanthridines , Pharmacology , Drugs, Chinese Herbal , Pharmacology , Humans , Pesticides , Pharmacology , Plants, Medicinal , Chemistry
5.
Acta Pharmaceutica Sinica ; (12): 933-939, 2013.
Article in Chinese | WPRIM | ID: wpr-259527

ABSTRACT

The objective of the present study was to establish a method based on principal component analysis (PCA) for the study of transdermal delivery of multiple components in Chinese medicine, and to choose the best penetration enhancers for the active fraction of Xiangfusiwu decoction (BW) with this method. Improved Franz diffusion cells with isolated rat abdomen skins were carried out to experiment on the transdermal delivery of six active components, including ferulic acid, paeoniflorin, albiflorin, protopine, tetrahydropalmatine and tetrahydrocolumbamine. The concentrations of these components were determined by LC-MS/MS, then the total factor scores of the concentrations at different times were calculated using PCA and were employed instead of the concentrations to compute the cumulative amounts and steady fluxes, the latter of which were considered as the indexes for optimizing penetration enhancers. The results showed that compared to the control group, the steady fluxes of the other groups increased significantly and furthermore, 4% azone with 1% propylene glycol manifested the best effect. The six components could penetrate through skin well under the action of penetration enhancers. The method established in this study has been proved to be suitable for the study of transdermal delivery of multiple components, and it provided a scientific basis for preparation research of Xiangfusiwu decoction and moreover, it could be a reference for Chinese medicine research.


Subject(s)
Administration, Cutaneous , Alkenes , Pharmacology , Animals , Azepines , Pharmacology , Benzophenanthridines , Pharmacokinetics , Berberine Alkaloids , Pharmacokinetics , Bridged-Ring Compounds , Pharmacokinetics , Coumaric Acids , Pharmacokinetics , Drug Combinations , Drug Synergism , Drugs, Chinese Herbal , Chemistry , Pharmacokinetics , Glucosides , Pharmacokinetics , In Vitro Techniques , Male , Monoterpenes , Pharmacokinetics , Permeability , Plants, Medicinal , Chemistry , Principal Component Analysis , Rats , Rats, Sprague-Dawley , Skin Absorption
6.
Journal of Shahrekord University of Medical Sciences. 2012; 14 (3): 40-50
in Persian | IMEMR | ID: emr-132505

ABSTRACT

Studies have recently shown that intermittent normobaric hyperoxia has a significant therapeutic effect on the treatment of acute ischemia. In this study intermittent normobaric effect of hyperoxia and protein kinase C [PKC] activity in the blood - brain barrier [BBB] permeability and behavioral assessment were evaluated. In this experimental study 36 wistar rat were divided to 6 groups as follow: normoxi [shem], normoxi+CHEL, normoxi+halt, normoxi+halt+CHEL, hyproxi+halt, hyperoxi+halt+CHEL, n=6 in each group. Chelerythrin chlorid [CHEL] was used as a systemically inhibitor of PKC. 24 hours later, rats were subjected to 60 min of right middle cerebral artery occlusion [MCAO]. The hyperoxia and normoxia groups were exposed to 95% and 21% respectively, for 4 h/day, 6 continuous days. After 24 h reperfusion, neurological deficit scores and BBB permeability was assessed. Data were analyzed using two way ANOVA and Bonferroni test. Preconditioning with intermittent normobaric hyperoxia decreased neurologic deficit scores and BBB permeability. Inhibition of PKC resulted in the increase of neurologic deficit scores; which improved with hyperoxia [P<0.001]. PKC inhibition, independent of hyperoxia improved the BBB function [P<0.001]. With the deployment of hyperoxia and specific subunits of PKC during the stroke, stability of BBB integrity and improvement of neurological deficit scores occur


Subject(s)
Animals, Laboratory , Hyperoxia , Protein Kinase C , Permeability , Ischemia , Rats, Wistar , Benzophenanthridines , Infarction, Middle Cerebral Artery
7.
Article in Chinese | WPRIM | ID: wpr-814787

ABSTRACT

OBJECTIVE@#To investigate the effect of intrathecal sufentanil and protein kinase C inhibitor on pain threshold and the expression of N-methyl-D-aspartate receaptors (NMDAR)/calcitonin generelated peptide (CGRP) in spinal dorsal horn in rats with neuropathic pain.@*METHODS@#Fifty-four healthy male Sprague-Dawley rats were randomly divided into 6 groups (9 in each group). The rats in the sham group(Group S) + spared nerve injury (SNI), SP+SNI, and P+SNI were intrathecally injected sufentanil (1 μg), sufentanil (1 μg) and chelerythrine chloride (11 μg), chelerythrine chloride (11 μg) followed by 10 μL normal saline once every day for 14 days postoperatively, respectively. Similarly, rats in the control group (Group C), the sham group (Group S), and SNI model group (Group SNI) were intrathecally injected 20 μL normal saline in the uniform interval. Pain behaviours were measured on Day 1 pre-surgery and on Day 1, 2, 7, and 14 after the intrathecal injection. The expressions of NMDAR and CGRP in the spinal dorsal horn of L5 segment were determined by immunohistochemistry on Day 2, 7, and 14 after the intrathecal injection.@*RESULTS@#Compared with Group C and Group S, mechanical allodynia threshold in group SNI was decreased after the surgery (P<0.01), and expressions of NMDAR and CGRP immunoreactive soma in the spinal dorsal horn was significantly increased (P<0.01). Mechanical stimulation pain threshold was elevated in Group S+SNI, Group P+SNI, and Group SP+SNI compared with Group SNI (P<0.01), while expressions of NMDAR and CGRP immunoreactive soma in Group S+SNI, Group P +SNI, and Group SP+SNI were significantly decreased (P<0.05 or 0.01).@*CONCLUSION@#Intrathecal administration of sulfentanil and protein kinase C inhibitor can provide significant antinociception in rats with neuropathic pain and obviously inhibit the upregulation of NMDAR and CGRP expressions in the spinal dorsal horn of SNI rat models.


Subject(s)
Animals , Benzophenanthridines , Calcitonin Gene-Related Peptide , Metabolism , Injections, Spinal , Male , Neuralgia , Drug Therapy , Metabolism , Pain Measurement , Posterior Horn Cells , Metabolism , Protein Kinase C , Rats , Rats, Sprague-Dawley , Receptors, N-Methyl-D-Aspartate , Metabolism , Sufentanil
8.
Article in English | WPRIM | ID: wpr-123276

ABSTRACT

The purpose of this study is to determine 1) whether morphine postconditiong (MPostC) can attenuate the intercellular adhesion molecules-1 (ICAM-1) expression after reoxygenation injury and 2) the subtype(s) of the opioid receptors (ORs) that are involved with MPostC. Human umbilical vein endothelial cells (HUVECs) were subjected to 6 hr anoxia followed by 12 hr reoxygenation. Three morphine concentrations (0.3, 3, 30 microM) were used to evaluate the protective effect of MPostC. We also investigated blockading the OR subtypes' effects on MPostC by using three antagonists (a micro-OR antagonist naloxone, a kappa-OR antagonist nor-binaltorphimine, and a delta-OR antagonist naltrindole) and the inhibitor of protein kinase C (PKC) chelerythrine. As results, the ICAM-1 expression was significantly reduced in the MPostC (3, 30 microM) groups compared to the control group at 1, 6, 9, and 12 hours reoxygenation time. As a consequence, neutrophil adhesion was also decreased after MPostC. These effects were abolished by coadministering chelerythrine, nor-binaltorphimine or naltrindole, but not with naloxone. In conclusion, it is assumed that MPostC could attenuate the expression of ICAM-1 on endothelial cells during reoxygenation via the kappa and delta-OR (opioid receptor)-specific pathway, and this also involves a PKC-dependent pathway.


Subject(s)
Animals , Benzophenanthridines/pharmacology , Endothelial Cells/cytology , Endothelium, Vascular/cytology , Humans , Intercellular Adhesion Molecule-1/genetics , Morphine/pharmacology , Naloxone/pharmacology , Naltrexone/analogs & derivatives , Narcotic Antagonists/pharmacology , Narcotics/pharmacology , Protein Isoforms/metabolism , Protein Kinase C/antagonists & inhibitors , Receptors, Opioid/metabolism , Reperfusion Injury/metabolism , Signal Transduction/physiology , Umbilical Veins/cytology
9.
Article in English | WPRIM | ID: wpr-341461

ABSTRACT

<p><b>OBJECTIVE</b>To explore the mechanism governing the reversal of multidrug resistance in human breast carcinoma cells by chelerythrine.</p><p><b>METHODS</b>Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to determine the expressions of protein kinase Cα (PKCα) and multidrug resistance-related genes ABCG2, ABCC1, MDR1, and P-glycoprotein (P-gp) in MCF-7Taxol cells after treatment with chelerythrine and phorbol-12-myristate-13-acetate (PMA). Also, the antitumor effect of PMA or chelerythrine and effects of PKCα activator or inhibitor in combination with paclitaxel or adriamycin on multidrug resistance in MCF-7Taxol cells were evaluated by MTT.</p><p><b>RESULTS</b>RT-PCR or Western blot showed that the expressions of MDR1 and P-gp were significantly higher in MCF-7Taxol cells exposed to PMA stimuli (both P0.05).</p><p><b>CONCLUSION</b>PKCα inhibitor chelerythrine can reverse multidrug resistance in breast carcinoma cells by inhibiting the expressions of MDR1 and P-gp expression in vitro.</p>


Subject(s)
ATP Binding Cassette Transporter, Subfamily B , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Metabolism , Benzophenanthridines , Pharmacology , Breast Neoplasms , Metabolism , Pathology , Cell Line, Tumor , Drug Resistance, Multiple , Drug Resistance, Neoplasm , Female , Humans
10.
Article in Chinese | WPRIM | ID: wpr-307931

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the apoptosis-inducing effect of nitidine chloride in human osteosarcoma MG-63 cells and explore its mechanism.</p><p><b>METHODS</b>The effect of nitidine chloride on the proliferation of MG-63 cells was detected by colorimetric MTT assay, and the morphological changes of cells treated with nitidine chloride were observed using fluorescence and electron microscope. Flow cytometry was performed to analyze the apoptotic rate of the cells, and the protein expression levels of caspase-3, caspase-9, Bcl-2 and Bax were detected by Western blotting.</p><p><b>RESULTS</b>Nitidine chloride inhibited the proliferation of MG-63 cells in a dose- and time-dependent manner. Fluorescence and electron microscopy revealed distinct apoptotic changes of the cells after nitidine chloride exposure. Flow cytometry indicated that nitidine chloride induced the apoptosis of MG-63 cells in a dose-dependent manner. Exposure to nitidine chloride, as shown by Western blotting, resulted in increased expressions of cleaved caspase-3, cleaved caspase-9 and Bax and decreased expressions of pro-caspase-3, pro-caspase-9 and Bcl-2.</p><p><b>CONCLUSION</b>Nitidine chloride can inhibit the proliferation of osteosarcoma cell line MG-63 by inducing cell apoptosis, the mechanism of which might be related with the activation of the caspase-dependent pathway.</p>


Subject(s)
Antineoplastic Agents, Phytogenic , Pharmacology , Apoptosis , Benzophenanthridines , Pharmacology , Bone Neoplasms , Pathology , Cell Line, Tumor , Dose-Response Relationship, Drug , Humans , Osteosarcoma , Pathology
11.
Article in English | WPRIM | ID: wpr-727344

ABSTRACT

We have shown that myosin light chain kinase (MLCK) was required for the off-contraction in response to the electrical field stimulation (EFS) of feline esophageal smooth muscle. In this study, we investigated whether protein kinase C (PKC) may require the on-contraction in response to EFS using feline esophageal smooth muscle. The contractions were recorded using an isometric force transducer. On-contraction occurred in the presence of NG-nitro-L-arginine methyl ester (L-NAME), suggesting that nitric oxide acts as an inhibitory mediator in smooth muscle. The excitatory composition of both contractions was cholinergic dependent which was blocked by tetrodotoxin or atropine. The on-contraction was abolished in Ca2+-free buffer but reappeared in normal Ca2+-containing buffer indicating that the contraction was Ca2+ dependent. 4-aminopyridine (4-AP), voltage-dependent K+ channel blocker, significantly enhanced on-contraction. Aluminum fluoride (a G-protein activator) increased on-contraction. Pertussis toxin (a Gi inactivator) and C3 exoenzyme (a rhoA inactivator) significantly decreased on-contraction suggesting that Gi or rhoA protein may be related with Ca2+ and K+ channel. ML-9, a MLCK inhibitor, significantly inhibited on-contraction, and chelerythrine (PKC inhibitor) affected on the contraction. These results suggest that endogenous cholinergic contractions activated directly by low-frequency EFS may be mediated by Ca2+, and G proteins, such as Gi and rhoA, which resulted in the activation of MLCK, and PKC to produce the contraction in feline distal esophageal smooth muscle.


Subject(s)
4-Aminopyridine , Aluminum , Aluminum Compounds , Atropine , Azepines , Benzophenanthridines , Contracts , Esophagus , Fluorides , GTP-Binding Proteins , Muscle, Smooth , Myosin-Light-Chain Kinase , NG-Nitroarginine Methyl Ester , Nitric Oxide , Pertussis Toxin , Protein Kinase C , rhoA GTP-Binding Protein , Tetrodotoxin , Transducers
12.
Article in Chinese | WPRIM | ID: wpr-340176

ABSTRACT

<p><b>OBJECTIVE</b>To determine the effect of activation of lambda-opioid receptor with U50, 488H, a selective kappa-opioid receptor agonist, on the changes in electrical coupling during prolonged ischemia and to explore the possible mechanism.</p><p><b>METHODS</b>The isolated rat heart was perfused in a Langendorff apparatus. The effect of U50, 488H on electrical coupling parameters including onset of uncoupling, plateau time, slope and fold increase in r(t) was observed in isolated perfused rat heart subjected to global no-flow ischemia. The effect of U50, 488H on connexin 43 (Cx43) expression of ventricular muscle during ischemia was determined by immunohistochemistry.</p><p><b>RESULTS</b>In the prolonged ischemia model, U50, 488H concentration dependently delayed the onset of uncoupling, increased time to plateau, and decreased the maximal rate of uncoupling during ischemia. The effect of U50, 488H on electrical uncoupling parameters during ischemia was abolished by a selective kappa-opioid receptor antagonist nor-BNI or a PKC inhibitor chelerythrine. The amount of Cx43 immunoreactive signal in ventricular muscle was greatly reduced after ischemia. U50, 488H markedly increased Cx43 expression during ischemia and its effect was also attenuated by nor-BNI or chelerythrine.</p><p><b>CONCLUSION</b>These results demonstrated that U50, 488H delayed the onset of uncoupling and plateau time, decreased the maximal rate of uncoupling and increased Cx43 expression of ventricular muscle during ischemia, and these effects of U50, 488H were mediated by kappa-opioid receptor, in which activation of PKC was involved. The effect of U50, 488H on electrical coupling during ischemia was probably correlated with preservation of Cx43 in cardiac muscle.</p>


Subject(s)
3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer , Pharmacology , Animals , Benzophenanthridines , Pharmacology , Connexin 43 , Metabolism , Female , Heart , In Vitro Techniques , Myocardial Ischemia , Metabolism , Myocardium , Metabolism , Naltrexone , Pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Opioid, kappa , Metabolism , Signal Transduction
13.
Article in Chinese | WPRIM | ID: wpr-291490

ABSTRACT

<p><b>OBJECTIVE</b>To identify the differentially expressed proteins in the liver of Oncomelania snails induced by Eomecon chinanthe sanguinarine.</p><p><b>METHODS</b>Sanguinarine was extracted and purified from the dry powder of Eomecon chinanthe. Oncomelania snails were immersed in 5 mg/L sanguinarine (50 Oncomelania snails per 500 ml) or pure water for 36 h (25°C) and the livers were isolated from live snails. Total liver proteins were extracted and separated by two-dimensional gel electrophoresis. Electrophoretogram was analyzed by Image Master 2D 5.0 software. The differentially expressed proteins between sanguinarine group and pure water group were selected and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and tandem mass spectrometry sequencing of tryptic peptides.</p><p><b>RESULTS</b>In terms of protein spots, 433 ± 14 and 385 ± 12 were observed in sanguinarine group and in water group respectively. The eleven identified differentially expressed proteins included tropomyosin, hypothetical protein XP_533132, actin 87E, keratin 6A, beta-tubulin, mitochondrial inner membrane protein isoform 4, keratin 2, allatostatin precursor, ENSANGP00000020184, actin-3 and ENSANGP00000013943. Among them, hypothetical protein XP_533132 and ENSANGP00000013943 were down-regulated and the other nine proteins were up-regulated in sanguinarine group.</p><p><b>CONCLUSION</b>Sanguinarine could alter the expression of proteins in livers of Oncomelania snails.</p>


Subject(s)
Animals , Benzophenanthridines , Pharmacology , Electrophoresis, Gel, Two-Dimensional , Isoquinolines , Pharmacology , Liver , Metabolism , Proteomics , Snails , Metabolism , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
14.
Article in Chinese | WPRIM | ID: wpr-262168

ABSTRACT

<p><b>OBJECTIVE</b>To develop a HPLC method for determining the content of protopine in Corydalis racemose.</p><p><b>METHOD</b>Analysis was performed on a Gemini C18 column (4.6 mm x 250 mm, 5 microm) eluted with acetonitrile-water containing 0.8% triethylamine and 3% acetic acid acetum (20:80) as the mobile phase. The flow rate was 1.0 mL x min(-1). The detection wavelength was 289 nm.</p><p><b>RESULT</b>The average content of protopine in Herb of Racemose Corydalis was 0.905%. The calibration curve of protopine was linear between 0.124-1.36 microg (r = 0.9999). The average recovery was 98.49% with RSD 1.9%.</p><p><b>CONCLUSION</b>This method is simple, reproducible and can be used to determine the content of protopine in C. racemose.</p>


Subject(s)
Benzophenanthridines , Berberine Alkaloids , Chromatography, High Pressure Liquid , Methods , Corydalis , Chemistry , Drugs, Chinese Herbal
15.
Acta Pharmaceutica Sinica ; (12): 115-120, 2009.
Article in Chinese | WPRIM | ID: wpr-232587

ABSTRACT

The purpose of this study is to investigate the effect of chelerythrine on the hypertrophy of cardiomyocytes of neonatal rats induced by different glucose levels and its mechanism. Using cultured neonatal ventricular myocytes as a model, groups were divided as: control (5 mmol x L(-1)); high glucose level (10, 15, 20, and 25.5 mmol x L(-1)); high glucose level (25.5 mmol x L(-1)) add different concentrations of chelerythrine (1 and 8 micromol x L(-1)); and control glucose level (5 mmol x L(-1)) add different concentrations of chelerythrine (1 and 8 micromol x L(-1)). Different groups of cardiomyocytes after adding corresponding treat factors were cultured for 48 hours. Cardiomyocytes' diameters and protein level were measured and the expression of PKC-alpha, PKC-beta2, p-PKC-alpha, and p-PKC-beta2 were measured by Western blotting. Compared with control group, neonatal myocytes cultured in high glucose levels showed increased cellular volumes, protein level and expression of PKC-alpha, PKC-beta2, p-PKC-alpha, p-PKC-beta2. When chelerythrine was added, cellular volumes, protein level and expression of PKC-alpha, PKC-beta2, p-PKC-alpha, p-PKC-beta2 were significantly reduced. But in 1 micromol x L(-1) chelerythrine group, the expression of PKC-beta2 was not significantly reduced. The result suggested that chelerythrine can reverse the hypertrophy induced by different glucose levels on the cardiac myocytes, it may have protective effect against diabetic cardiomyopathy via PKC passageway.


Subject(s)
Animals , Animals, Newborn , Benzophenanthridines , Pharmacology , Cells, Cultured , Diabetes Mellitus, Experimental , Drug Therapy , Metabolism , Dose-Response Relationship, Drug , Glucose , Hypertrophy , Pathology , Hypoglycemic Agents , Pharmacology , Myocytes, Cardiac , Pathology , Phosphorylation , Protein Kinase C , Metabolism , Protein Kinase C beta , Protein Kinase C-alpha , Metabolism , Rats , Rats, Sprague-Dawley
16.
Acta Pharmaceutica Sinica ; (12): 134-139, 2009.
Article in Chinese | WPRIM | ID: wpr-232584

ABSTRACT

This study is to observe the effect of gross saponins of Tribulus terrestris (GSTT) on protein kinase Cepsilon (PKCepsilon) and apoptosis-associated protein in the apoptosis of cultured cardiocyte apoptosis induced by hydrogen peroxide (H2O2), and to explore the mechanisms of GSTT against myocardial apoptosis. Primary cardiocytes were isolated and cultured. Myocardial apoptosis was induced by H2O2 and analyzed with flow cytometry. Protein content of phospho-PKCepsilon, Bcl-2, and Bax were detected with Western blotting analysis. Cleaved caspase-3 protein content was determined with immunocytochemical technique. After the pretreatment of 100 mg x L(-1) GSTT, compared with H2O2 group, GSTT could not only decrease the apoptotic percentage in cardiocytes damaged by H2O2 (P < 0.01), but also reduce protein contents of Bax and cleaved caspase-3 (P < 0.01), and increase protein content of phospho-PKCepsilon and Bcl-2 significantly (P < 0.01). PKC inhibitor chelerythrine (Che) could prevent partly the effect of GSTT against myocardial apoptosis (P < 0.05 and P < 0.01). Mechanisms of GSTT against myocardial apoptosis might be associated with inhibition of mitochondrial apoptosis pathway after PKCepsilon activation.


Subject(s)
Animals , Apoptosis , Benzophenanthridines , Pharmacology , Caspase 3 , Metabolism , Cells, Cultured , Dose-Response Relationship, Drug , Enzyme Activation , Female , Hydrogen Peroxide , Toxicity , Male , Myocytes, Cardiac , Cell Biology , Metabolism , Oxidative Stress , Phosphorylation , Plants, Medicinal , Chemistry , Protein Kinase C , Protein Kinase C-epsilon , Metabolism , Proto-Oncogene Proteins c-bcl-2 , Metabolism , Rats , Rats, Wistar , Saponins , Pharmacology , Tribulus , Chemistry , bcl-2-Associated X Protein , Metabolism
17.
Article in Chinese | WPRIM | ID: wpr-346998

ABSTRACT

<p><b>OBJECTIVE</b>To develop an HPLC method for determination of the content of chelerythrine in Chelidonium majus.</p><p><b>METHOD</b>Chelerythrine was extracted from the fine powder of the crade with drug methanol and determined by HPLC. The mobile phase was acetonitrile-1% triethylamine (25:75) (adjusted pH to 3 using phosphoric acid) and the detection wavelength was set at 268 nm.</p><p><b>RESULT</b>The linear range of calibration curve was 0.051 6-0.516 0 microg (r = 1.000). The average recovery (n = 6) was 103.0% with RSD of 1.2%. Chelerythrine in the sample solution was stable in 8 h and the ruggedness was perfect among 3 different chromatographic columns.</p><p><b>CONCLUSION</b>The method is accurate, sensitive and reliable.</p>


Subject(s)
Benzophenanthridines , Chelidonium , Chemistry , Chromatography, High Pressure Liquid , Methods , Drugs, Chinese Herbal
18.
Article in Chinese | WPRIM | ID: wpr-263001

ABSTRACT

<p><b>OBJECTIVE</b>A high performance liquid chromatography (HPLC) method was developed to determine the concentration of nitidine chloride in plasma and successfully applied to study pharmacokinetics after i.v. administration in rabbits.</p><p><b>METHOD</b>Twelve rabbits, randomized into 2 groups , were given i.v. at the dose of 4, 6 mg x kg(-1) respectively. Chloramphenicol was used as an internal standard. Nitidine chloride was extracted from plasma with ion pair reagent, and was determined by HPLC.</p><p><b>RESULT</b>The calibration curves of nitidine chloride was linear in the range of 0.03-2.04 mg x L(-1). Its recoveries were more than 95%, intra-day and inter-day precisions were lower than 6%. The concentration-time curve of nitidine chloride in rabbits after i.v. of 4 and 6 mg x kg(-1) were shown to fit a two-compartment model, the main pharmacokinetic parameters showed no significant difference between the low and high dosage, and the AUC values are directly relative to doses. T1/2alpha were (5.46 +/- 0.89), (4.76 +/- 0.33) min respectively, T1/2beta were (263.33 +/- 16.4), (274.71 +/- 16.52) min respectively, AUC were (46.56 +/- 1.80), (69.19 +/- 2.30) microg x min(-1) x mL(-1) respectively.</p><p><b>CONCLUSION</b>It is first time to establish the HPLC method to determine the concentration of nitidine chloride in rabbits plasma. The method is sensitive, accurate and reproducible. It is first time to study the pharmacokinetic characters of nitidine chloride in rabbits after i.v. administration, the elimination of nitidine chloride is linear pharmacokinetics.</p>


Subject(s)
Animals , Benzophenanthridines , Blood , Pharmacokinetics , Chromatography, High Pressure Liquid , Methods , Drugs, Chinese Herbal , Pharmacokinetics , Female , Male , Rabbits , Random Allocation
19.
Article in English | WPRIM | ID: wpr-728193

ABSTRACT

Although many studies show that thromboxane A2 (TXA2) has the action of gastrointestinal (GI) motility using GI muscle cells and tissue, there are no reports on the effects of TXA2 on interstitial cells of Cajal (ICC) that function as pacemaker cells in GI tract. So, we studied the modulation of pacemaker activities by TXA2 in ICC with whole cell patch-clamp technique. Externally applied TXA2 (5 micrometer) produced membrane depolarization in current-clamp mode and increased tonic inward pacemaker currents in voltage-clamp mode. The tonic inward currents by TXA2 were inhibited by intracellular application of GDP-beta-S. The pretreatment of ICC with Ca2+ free solution and thapsigargin, a Ca2+-ATPase inhibitor in endoplasmic reticulum, abolished the generation of pacemaker currents and suppressed the TXA2-induced tonic inward currents. However, chelerythrine or calphostin C, protein kinase C inhibitors, did not block the TXA2-induced effects on pacemaker currents. These results suggest that TXA2 can regulate intestinal motility through the modulation of ICC pacemaker activities. This modulation of pacemaker activities by TXA2 may occur by the activation of G protein and PKC independent pathway via extra and intracellular Ca2+ modulation.


Subject(s)
Animals , Benzophenanthridines , Endoplasmic Reticulum , Gastrointestinal Motility , Gastrointestinal Tract , GTP-Binding Proteins , Guanosine Diphosphate , Interstitial Cells of Cajal , Intestines , Membranes , Mice , Muscle Cells , Naphthalenes , Patch-Clamp Techniques , Protein Kinase C , Thapsigargin , Thionucleotides , Thromboxane A2
20.
Article in English | WPRIM | ID: wpr-728192

ABSTRACT

Lysophosphatidylcholine (LPC), a metabolite of membrane phospholipids by phospholipase A(2), has been considered responsible for the development of abnormal vascular reactivity during atherosclerosis. Ca2+ influx was shown to be augmented in atherosclerotic artery which might be responsible for abnormal vascular reactivity. However, the mechanism underlying Ca2+ influx change in atherosclerotic artery remains undetermined. The purpose of the present study was to examine the effects of LPC on L-type Ca2+ current (ICa(L)) activity and to elucidate the mechanism of LPC-induced change of ICa(L) in rabbit portal vein smooth muscle cells using whole cell patch clamp. Extracellular application of LPC increased ICa(L) through whole test potentials, and this effect was readily reversed by washout. Steady state voltage dependency of activation or inactivation properties of ICa(L) was not significantly changed by LPC. Staurosporine (100 nanometer) or chelerythrine (3 micrometer, which is a potent inhibitor of PKC, significantly decreased basal ICa(L), and LPC-induced increase of ICa(L) was significantly suppressed in the presence of PKC inhibitors. On the other hand, application of PMA, an activator of PKC, increased basal ICa(L) significantly, and LPC-induced enhancement of ICa(L) was abolished by pretreatment of the cells with PMA. These findings suggest that LPC increased ICa(L) in vascular smooth muscle cells by a pathway that involves PKC, and that LPC-induced increase of ICa(L) might be, at least in part, responsible for increased Ca2+ influx in atherosclerotic artery.


Subject(s)
Arteries , Atherosclerosis , Benzophenanthridines , Dependency, Psychological , Hand , Lysophosphatidylcholines , Membranes , Muscle, Smooth , Muscle, Smooth, Vascular , Myocytes, Smooth Muscle , Phospholipases , Phospholipids , Portal Vein , Protein Kinase C , Protein Kinases , Staurosporine
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