Carnosic acid prevents dextran sulfate sodium-induced acute colitis associated with the regulation of the Keap1/Nrf2 pathway.

Crohn's disease and ulcerative colitis are inflammatory bowel diseases (IBDs) with high prevalence in humans. Carnosic acid (CA) has been reported to possess antioxidative properties; however, its role in IBDs has not been determined. In the present study, we found that CA significantly prevented the loss of body weight and shortening of colon length in acute colitis induced by dextran sodium sulfate (DSS). Pronounced infiltration of immune cells and a loss of crypt architecture and goblet cells were ameliorated by CA. CA significantly decreased the activity of MPO and infiltration of F4/80+ macrophages in the colon. DSS-induced pro-inflammatory cytokine mRNA and protein levels in the colon were also attenuated by CA. CA decreased the activation of p65 and c-Jun signalling. CA inhibited DSS-induced NLRP3 inflammasome activation by reducing caspase 1 activity. In addition, CA increased the level of Nrf2 and prevented the degradation of Nrf2 via ubiquitination by blocking the interaction between Cullin3 and Keap1, which resulted in the decrease of Nrf2 target genes. Finally, GSH levels and SOD activity were increased after CA treatment, while MDA and iNOS levels were significantly reduced. Taken together, our data showed that CA may be useful as a potential therapeutic candidate for IBDs.

In vitro interaction of clopidogrel and its hydrolysate with OCT1, OCT2 and OAT1.

Clopidogrel (CP) is metabolized by CYPs to the active metabolite, or hydrolyzed by esterase to clopidogrel carboxylate (CPC) in liver, and CPC is partly excreted from urine. Therefore, the objective of the present study was to evaluate the interactions of CP and CPC with organic cation transporter 1 (OCT1) (in liver), and CPC with organic cation transporter 2 (OCT2) and organic anion transporter 1 (OAT1) (in kidney). Both CP and CPC inhibited the uptake of 1-methyl-4-phenylpyridinium (MPP(+)) and metformin, typical substrates of OCT1, in MDCK-hOCT1 cells with low IC50 (0.307-14.0 µM). CPC (100 µM) reduced the uptake of MPP(+) and metformin mediated by OCT2 in MDCK-hOCT2 cells to 60.8% and 33.6% of the control, CPC (500 µM) decreased the uptake of 6-carboxyfluorescein (6-CFL) and para-aminohippuric acid (PAH), substrates of OAT1, in MDCK-hOAT1 cells to 64.6% and 79.4% of the control. CP and CPC were also found to inhibit other drugs of OCT1 substrates, such as lamivudine and amantadine, in MDCK-hOCT1 cells with the IC50 of 1.97-4.15µM, except CPC on amantadine (IC50>100 µM). The inhibition of CP and CPC on lamivudine uptake in primary rat hepatocytes was also confirmed with the IC50 of 2.91 and 1.25µM, respectively. Additionally, CP and CPC were not substrates of OCT1 and OCT2, whereas CPC was a substrate of OAT1 with the Km of 5.61 µM. In conclusion, CP and CPC are strong inhibitors of OCT1, but weak inhibitors of OCT2 and OAT1, and CPC is a high affinity substrate of OAT1.

Population pharmacokinetics of digoxin in elderly patients.

This study was aimed at determining the population pharmacokinetics of digoxin and identifying factors that explain pharmacokinetic variability in elderly patients. The data of 142 elderly patients and 448 samples were collected after repetitive oral digoxin. Blood samples were drawn at various times after administration. Population pharmacokinetic analysis was performed using nonlinear mixed effects modelling program (NONMEM). A one-compartment model with first-order absorption and elimination was selected as the base model. The influence of demographic characteristics, biochemical and haematological indices as well as other commonly used co-medications were explored. The typical values with interindividual variability for apparent clearance (CL/F) and apparent volume of distribution (V/F) were 8.9 L h(-1) (43.2 %) and 420 L (65.8 %), respectively. The residual variability was 31.6 %. CL/F decreased significantly with renal function, total body weight, calcium channel blockers or spironolactone co-therapy and symptom with congestive heart failure. The median parameter estimates from a nonparametric bootstrap procedure were comparable and within 5 % of the estimates from NONMEM. These results provide important information for clinicians to optimize digoxin regimens in elderly patients.

Different effects of dihydropyridine calcium channel antagonists on CYP3A4 enzyme of human liver microsomes.

The present study investigated inhibitory effects of 1,4-dihydropyridines (1,4-DHPs) calcium channel antagonists (1,4-DHP-CCAs) on cytochromeP450 3A4 (CYP3A4) of human liver microsomes and further explored importance of 1,4-DHPs molecular structural descriptors. Partial Least Squares method was applied to probe the quantitative relationships between the 1,4-DHPs molecular structural descriptors and its inhibitory actions, which demonstrated that different 1,4-DHP-CCAs could inhibit CYP3A4 enzyme's activity differently. The K (i) values of nicardipine, lercandipine, cilnidipine, nitrendipine, lacidipine, nifedipine, felodipine were 10.13, 10.17, 11.44, 23.90, 29.34, 29.06 and 32.64 µmol L⁻¹, respectively. It is suggested that the 1,4-DHPs molecular structural descriptors are the most important for its inhibitory effects based on the quantitative structure-activity relationship (QSAR) formula. The LogP was positively correlated to the K (i), whereas molecular weight and molecule volume were negatively correlated. It is concluded that analysis of K (i) of 1,4-DHPs derivatives on the CYP3A4 activity may apply for the QSAR formula at the initial stage of clinical application of new drugs.

[Pharmacokinetics of luteolin from Elsholtzia blanda extracts in rats].

An RP-HPLC method for determination of luteolin from Elsholtzia blanda Benth. extracts in rats' plasma was established and the pharmacokinetics of luteolin in rats was studied. Drug blood samples from caudal vein were gotten after oral administration of luteolin. Plasma samples were determined by RP-HPLC after being deproteinized with trichloroacetic acid and extracted with ethyl acetate. The calibration curve was linear in the range of 0.37-47.27 microg x mL(-1). The limit of quantification was 0.37 microg x mL(-1). The method recovery of luteolin was 93%-99%. The extract recovery was 75%-85%. RSDs of intra-and inter-day precisions were less than 5%. The concentration-time curve of luteolin after oral administration of Elsholtzia blanda Benth. extracts was fitted to two compartment open model. Two factors analysis of variance were adopted in the evaluation of gender and time spots for collection of blood. The result suggested that the gender-based difference in blood-drug concentrations had statistical significance. The metabolite in blood was identified as galcuronide. The method is sensitive, specific, accurate, and is appropriate for determination of luteolin in vivo.

In vitro metabolism of BYZX in human liver microsomes and the structural elucidation of metabolite by liquid chromatography-mass spectrometry method.

In vitro phase I metabolism of BYZX, a novel central-acting cholinesterase inhibitor for the treatment of the symptoms of Alzheimer's disease, was studied in human liver microsomes (HLM) and the metabolite formation pathways were investigated by chemical inhibition experiments and correlation analysis. The residual concentration of substrate and the metabolite formed in incubate were determined by HPLC method. The calibration curves of BYZX were linear over the concentration range from 5.07 microM to 200.74 microM. The relative standard deviations of within day and between day were less than 5% (n=5). The limit of detection (LOD) was 0.18 microg/mL (S/N=3) and the limit of quantification (LOQ) was 0.55 microg/mL (R.S.D.=5.2%, n=5). The determination recoveries of BYZX were in the range of 98.2-104.8%. The apparent K(m) of BYZX in HLM was 53.25+/-17.2 microM, the V(max) was 0.94+/-0.77 microM/min/mg protein, and the intrinsic clearance value (Cl(int)) was 0.018+/-0.02 mL/min/mg protein. Ketoconazole and cyclosporin A were the most potent inhibitors on BYZX metabolism in HLM with IC(50) being 0.89 microM and 18.17 microM, respectively. And the inhibition constant (K(i)) of ketoconazole was 0.42 microM. The metabolite of BYZX was N-des-ethyl-BYZX elucidated by LC-MS-MS. The results demonstrated that the developed HPLC method was reliability, simple technique, and was applicable to be used for the researches of in vitro metabolism of BYZX. CYP3A4 was the major isozyme responsible for BYZX metabolism; N-dealkylation was the major metabolic pathway of BYZX. The predominant metabolite of BYZX was N-des-ethyl-BYZX detected in vitro phase I metabolism in HLM.

[Interaction between (E)-2-(4-(diethylamino methyl) benzylidene)-5,6-dimethoxy-2,3-dihydroinden-one and P-glycoprotein].

Cell lines of Bcap37 and Bcap37/MDR1 (the high P-glycoprotein (P-gp) expressing cell line) were used as model to investigate the different accumulations of (E)-2-(4-(diethylamino methyl) benzylidene)-5,6-dimethoxy-2,3-dihydroinden-one (BYZX) in the two kinds of cells. It was authenticated that whether BYZX was the substrate of P-gp. Meanwhile, the inhibitive effects of BYZX on the P-gp were investigated by determining the fluorescence intensity of rhodamine 123 in the model cells, with and without BYZX. A reversed-phase high-performance liquid chromatography (RP-HPLC) method was used to determine the accumulations of BYZX in the two cells. The results showed that the amount of BYZX accumulation in Bcap37/MDR1 cells were as many as those in Bcap37 cells (P > 0.05), and the concentrations of BYZX accumulated in the Bcap37/MDR1 cells did not increase when co-incubated with P-gp inhibitor verapamil. Furthermore, different concentrations of BYZX also had no effects on the efflux of rhodamine 123 (P > 0.05). These results indicated that there were no interactions between BYZX and P-gp. BYZX will not be pumped out of the cells, and it also not inhibited the P-gp. It was the useful advantage for its absorption.