We characterized and compared the characteristics of Ca2+ movements through the sarcoplasmic reticulum of inferior oblique muscles in the various conditions including primary inferior oblique overaction (IOOA), secondary IOOA, and controls, so as to further understand the pathogenesis of primary IOOA. Of 15 specimens obtained through inferior oblique myectomy, six were from primary IOOA, 6 from secondary IOOA, and the remaining 3 were controls from enucleated eyes. Ryanodine binding assays were performed, and Ca2+ uptake rates, calsequestrins and SERCA levels were determined. Ryanodine bindings and sarcoplasmic reticulum Ca2+ uptake rates were significantly decreased in primary IOOA (p < 0.05). Western blot analysis conducted to quantify calsequestrins and SERCA, found no significant difference between primary IOOA, secondary IOOA, and the controls. Increased intracellular Ca2+ concentration due to reduced sarcoplasmic reticulum Ca2+ uptake may play a role in primary IOOA.
Sarcoplasmic Reticulum Calcium-Transporting ATPases ; Sarcoplasmic Reticulum/*metabolism ; Ryanodine Receptor Calcium Release Channel/metabolism ; Ryanodine/metabolism ; Oxalates/metabolism ; Oculomotor Muscles ; Ocular Motility Disorders/*metabolism/*pathology ; Muscles/*pathology ; Models, Statistical ; Middle Aged ; Male ; Humans ; Female ; Child, Preschool ; Child ; Calsequestrin/metabolism ; Calcium-Transporting ATPases/metabolism ; Calcium/metabolism/*pharmacokinetics ; Blotting, Western ; Aged ; Adult ; Adolescent

OBJECTIVETo explore the potential biochemical mechanisms of neuromuscular junction transmission (NMJT) dysfunction induced by organophosphorus insecticides.

METHODSTen rats were dosed with phoxim (1,144 mg/kg) and 5 of them developed myasthenia. The NMJT function was evaluated by the mean consecutive differences (MCD) measured by stimulation single fiber electromyography (SSFEMG) with the frequency of stimuli at 20 Hz. The activity of Ca(2+)-ATPase, Na(+), K(+)-ATPase, cAMP dependent protein kinase (PKA), Ca(2+)/phospholipid dependent protein kinase (PKC), and tyrosine protein kinase (TPK) were determined.

RESULTSIn comparison with the control and non-myasthenic rats, the results in myasthenic rats showed that: (1) the MCDs increased; (2) the activities of Ca(2+)-ATPase, Na(+), K(+)-ATPase and PKA decreased and were negatively correlated with MCD; (3) the activities of PKC and TPK increased, and were positively correlated with MCD; (4) the phosphorylation of serine residuals in sarcolemma was weaker and the phosphorylation of tyrosine residuals was stronger.

CONCLUSIONSThe NMJT dysfunction is likely associated with the decrease in Ca(2+)-ATPase and Na(+), K(+)-ATPase activity. The acceleration of desensitization and prolongation of resensitization of nicotinic acetylcholine receptors occur following the increase in tyrosine phosphorylation and the decrease in serine phosphorylation induced by OPs poisoning.

Animals ; Calcium-Transporting ATPases ; metabolism ; Cyclic AMP-Dependent Protein Kinases ; Electromyography ; drug effects ; Insecticides ; pharmacokinetics ; toxicity ; Myasthenia Gravis ; chemically induced ; enzymology ; Organothiophosphorus Compounds ; pharmacokinetics ; toxicity ; Protein Kinase C ; metabolism ; Protein-Serine-Threonine Kinases ; metabolism ; Rats

OBJECTIVETo explore the mechanism of cytotoxic effects of the organophosphates (OPs) with delayed neurotoxicity on human neuroblastoma cells.

METHODSThe proliferation of neuroblastoma SH-SY5Y cells was determined by MTT spectrometry. (45)Ca uptake was determined by adding (45)CaCl(2) and tri-o-cresyl phosphate (TOCP) or methamidophos into the cultured medium for the SH-SY5Y cells. The cells were incubated and then lysed and finally counted in a Beckman LS 6000 liquid scintillation spectrometer.

RESULTSMethamidophos stimulated the cell proliferation of SH-SY5Y at its lower concentrations (7 x 10(-7) mol/L to 7 x 10(-6) mol/L), with an increase by 28% at 7 x 10(-7) mol/L; however, it inhibited the proliferation at higher ones (7 x 10(-4) mol/L to 7 x 10(-3) mol/L) with 62% inhibition at 7 x 10(-3) mol/L. TOCP only inhibited the cell proliferation at high concentration (with 34% inhibition at 7 x 10(-3) mol/L) and markedly inhibited calcium uptake of the cells up to 55% at higher concentrations (1 x 10(-6) mol/L to 1 x 10(-4) mol/L); while the uptake was stimulated by OPs up to 241% of increase at lower concentrations (1 x 10(-9) mol/L to 1 x 10(-7) mol/L).

CONCLUSIONThe interference of growth in nerve cells and disturbance of calcium homeostasis may be involved in the mechanisms of neurotoxicity of OPs.

Calcium Radioisotopes ; metabolism ; pharmacokinetics ; Cell Division ; drug effects ; Cell Line, Tumor ; drug effects ; Humans ; Insecticides ; administration & dosage ; Neuroblastoma ; metabolism ; pathology ; Organophosphorus Compounds

OBJECTIVETo explore the effects of 2A-1-1 (purified component from Panax notoginsengs saponins) on the aggregation of and Ca2+ influx into human platelets.

METHODSThe aggregation of platelets was tested by nephelometry, Fura-2 fluorescent technique was used for detecting cell [Ca2+]i. The effects of 2A-1-1, nifedipine and SK&F96365 on Ca(2+) influx into human platelets induced by ADP or CPA were observed separately.

RESULTSNifedipine (< 20 micromol/L) could not inhibit platelet aggregation induced by ADP or the Ca(2+) influx induced by ADP or CPA. SK&F96365 at 20 micromol/L could inhibit the maximal aggregation of platelets induced by ADP with a inhibitory rate of 59.83%, at 15 micromol/L could inhibit the Ca2+ influx induced by CPA or ADP. 2A-1-1 (5, 10 and 20 micromol/L) could inhibit the maximal aggregation of platelets induced by ADP with the inhibitory rates of 47.06%, 53.47% and 71.52%, respectively. 2A-1-1 at 10 and 20 micromol/L could inhibit the Ca2+ influx induced by CPA or ADP.

CONCLUSIONS2A-1-1 can inhibit platelets aggregation, block the ROC (Receptor-dependent Ca2+ channels) and inhibit Ca2+ influx of human platelets.

Adenosine Diphosphate ; pharmacology ; Adult ; Blood Platelets ; cytology ; drug effects ; metabolism ; Calcium ; metabolism ; pharmacokinetics ; Calcium Channel Blockers ; pharmacology ; Dose-Response Relationship, Drug ; Female ; Ginsenosides ; pharmacology ; Humans ; Imidazoles ; pharmacology ; Indoles ; pharmacology ; Male ; Nifedipine ; pharmacology ; Platelet Aggregation ; drug effects ; Platelet Aggregation Inhibitors ; pharmacology

AIMTo study the metabolic kinetics of MN9202 in Beagle dog liver microsome.

METHODSBeagle dog liver microsomes were prepared by using ultracentrifuge method. After incubating 0.4 micromol x L(-1) MN9202 with 1 g x L(-1) microsomes for 30 min at 37 degrees C, the reaction was terminated by adding 0.5 mL alkalization. The RP-HPLC was used to determine the drug in the incubation mixture. The Michaelis-Menten parameters Km, and Vmax in Beagle dog liver microsomes were initially estimated by analyzing Lineweave-Brurk plot. Various selective CYP inhibitors were used to investigate their inhibitory effect on the metabolism of MN9202.

RESULTSThe Km, Vmax and CLint of MN9202 were (22.6 +/- 8.0) micromol x L(-1), (0.54 +/- 0.17) micromol x g(-1) x min(-1) and (0.0242 +/- 0.0009) L x g(-1) x min(-1), respectively. The metabolism of MN9202 was significantly inhibited by ketoconazole (Ket) and troleandomycin (Tro) in Beagle dog liver microsomes. Tranylcypromine (Tra) could inhibit the metabolism of drug as well. While other inhibitors showed little inhibitory effect on the metabolism of MN9202.

CONCLUSIONIt was shown that CYP3A and CYP2C19 were involved in MN9202 metabolism. The inhibitors of human CYP3A and CYP2C19 may have potential interaction with MN9202, and this can reduce the metabolism rate and increase the toxicity of MN9202.

Animals ; Aryl Hydrocarbon Hydroxylases ; antagonists & inhibitors ; Calcium Channel Blockers ; metabolism ; pharmacokinetics ; Cytochrome P-450 CYP2C19 ; Cytochrome P-450 CYP3A Inhibitors ; Dihydropyridines ; metabolism ; pharmacokinetics ; Dogs ; Ketoconazole ; pharmacology ; Microsomes, Liver ; metabolism ; Mixed Function Oxygenases ; antagonists & inhibitors ; Nitrobenzenes ; metabolism ; pharmacokinetics ; Tranylcypromine ; pharmacology ; Troleandomycin ; pharmacology

OBJECTIVEIn order to find a method for improving the salt resistance of seeds and seedlings for Perilla frutescens under NaCl stress, seed germination and physiological characteristics of P. frutescens seedlings were studied.

METHODSeveral physiological indexes of P. frutescens seeds treated by Ca2+ and sodium nitroprusside (SNP) under NaCl stress like the germination vigor, germination rate, germination index and vigor index were measured. And other indexes like the biomass of the seedlings, the content of malondialdehyde (MDA) in leaves, the activities of superoxide (SOD), peroxidase (POD) and catalase (CAT) were also measured.

RESULTThe germination of P. frutescens seeds under NaCl stress was inhibited obviously. But after the treatment with Ca2+ and SNP, all of the germination indexes increased. And the seeds that treated with SNP + Ca2+ had the most significantly increase in all indexes. The germination vigor was 65.1%, the germination rate was 89.3%, the germination index and vigor index were 13.9 and 0.1109, respectively. The content of MDA decreased after the treatment. The activities of three enzymes include SOD, POD and CAT were increased by the treatment and get the maximin 0.84, 5.71 and 4.92 U x mg(-1) respectively. And the EGTA showed an obvious inhibition to the effect of SNP on P. frutescens.

CONCLUSIONSNP (0.1 mmol x L(-1)) and Ca2+ (10 mmol x L(-1)) could significantly alleviate the damages to the seeds and seedlings of P. frutescens under NaCl stress, and promote the salt resistance of the seeds and seedlings. These results might suggest that exogenous NO might enhance P. frutescens salt resistance and alleviate salt injury possible by enhancing Ca2+ influx by activating Ca2+ channels and improving concentration of Ca2+ of P. frutescens seedlings.

Calcium ; pharmacology ; Catalase ; metabolism ; Germination ; drug effects ; Nitroprusside ; pharmacokinetics ; Perilla ; drug effects ; enzymology ; physiology ; Peroxidases ; metabolism ; Plant Proteins ; metabolism ; Seedlings ; drug effects ; enzymology ; metabolism ; physiology ; Sodium Chloride ; metabolism ; Stress, Physiological

Reasonable sampling scheme is the important basis for establishing reliable population pharmacokinetic model. It is an effective method for estimation of population pharmacokinetic parameters with sparse data to perform population pharmacokinetic analysis using the nonlinear mixed-effects models. We designed the sampling scheme for amlodipine based on D-optimal sampling strategy and Bayesian estimation method. First, optimized sample scenarios were designed using WinPOPT software according to the aim, dosage regimen and visit schedule of the clinical study protocol, and the amlodipine population model reported by Rohatagi et al. Second, we created a NONMEM-formatted dataset (n = 400) for each sample scenario via Monte Carlo simulation. Third, the estimation of amlodipine pharmacokinetic parameters (clearance (CL/F), volume (V/F) and Ka) was based on the simulation results. All modeling and simulation exercises were conducted with NONMEM version 7.2. Finally, the accuracy and precision of the estimated parameters were evaluated using the mean prediction error (MPE) and the mean absolute error (MAPE), respectively. Among the 6 schemes, schemes 6 and 3 have good accuracy and precision. MPE is 0.1% for scheme 6 and -0.6% for scheme 3, respectively. MAPE is 0.7% for both schemes. There is no significant difference in MPE and MAPE of volume among them. Therefore, we select scheme 3 as the final sample scenario because it has good accuracy and precision and less sample points. This research aims to provide scientific and effective sampling scheme for population pharmacokinetic (PK) study of amlodipine in patients with renal impairment and hypertension, provide a scientific method for an optimum design in clinical population PK/PD (pharmacodynamics) research.
Adult ; Age Factors ; Alanine Transaminase ; blood ; Amlodipine ; pharmacokinetics ; pharmacology ; Antihypertensive Agents ; pharmacokinetics ; pharmacology ; Bayes Theorem ; Body Weight ; Calcium Channel Blockers ; pharmacokinetics ; pharmacology ; Humans ; Hypertension ; metabolism ; Metabolic Clearance Rate ; Middle Aged ; Models, Biological ; Monte Carlo Method ; Nonlinear Dynamics ; Renal Insufficiency ; metabolism ; Software

Gambogic acid (GA) is an anticancer agent in phase ‖b clinical trial in China but its mechanism of action has not been fully clarified. The present study was designed to search the possible target-related proteins of GA in cancer cells using proteomic method and establish possible network using bioinformatic analysis. Cytotoxicity and anti-migration effects of GA in MDA-MB-231 cells were checked using MTT assay, flow cytometry, wound migration assay, and chamber migration assay. Possible target-related proteins of GA at early (3 h) and late stage (24 h) of treatment were searched using a proteomic technology, two-dimensional electrophoresis (2-DE). The possible network of GA was established using bioinformatic analysis. The intracellular expression levels of vimentin, keratin 18, and calumenin were determined using Western blotting. GA inhibited cell proliferation and induced cell cycle arrest at G2/M phase and apoptosis in MDA-MB-231 cells. Additionally, GA exhibited anti-migration effects at non-toxic doses. In 2-DE analysis, totally 23 possible GA targeted proteins were found, including those with functions in cytoskeleton and transport, regulation of redox state, metabolism, ubiquitin-proteasome system, transcription and translation, protein transport and modification, and cytokine. Network analysis of these proteins suggested that cytoskeleton-related proteins might play important roles in the effects of GA. Results of Western blotting confirmed the cleavage of vimentin, increase in keratin 18, and decrease in calumenin levels in GA-treated cells. In summary, GA is a multi-target compound and its anti-cancer effects may be based on several target-related proteins such as cytoskeleton-related proteins.
Antineoplastic Agents ; pharmacokinetics ; Apoptosis ; drug effects ; Breast Neoplasms ; drug therapy ; metabolism ; Calcium-Binding Proteins ; genetics ; Cell Line, Tumor ; Cell Migration Assays ; Cell Migration Inhibition ; drug effects ; Cell Proliferation ; drug effects ; Computational Biology ; methods ; Cytoskeleton ; metabolism ; Electrophoresis, Gel, Two-Dimensional ; Flow Cytometry ; Gene Expression ; Humans ; Keratin-18 ; genetics ; Oxidation-Reduction ; Protein Biosynthesis ; drug effects ; Protein Transport ; Proteomics ; methods ; Transcription, Genetic ; drug effects ; Ubiquitin-Specific Proteases ; pharmacokinetics ; Vimentin ; genetics ; Xanthones ; pharmacokinetics