Chengtao Lu

Safflower yellow B suppresses HepG2 cell injury induced by oxidative stress through the AKT/Nrf2 pathway.

Oxidative stress plays an important role in the pathogenesis of various liver diseases. Safflower yellow B (SYB) has been reported to protect the brain against damage induced by oxidative stress; however, whether SYB can also protect hepatocytes from oxidative stress remains unknown. In the present study, to determine whether pre-treatment with SYB reduces hydrogen peroxide (H2O2)-induced oxidative stress in HepG2 cells, we investigated H2O2-induced oxidative damage to HepG2 cells treated with or without SYB. Cell viability was measured by MTT assay and cytotoxicity was evaluated by lactate dehydrogenase (LDH) assay. The activities of the antioxidant enzymes, glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) were determined using respective kits. Intracellular reactive oxygen species (ROS) accumulation in the HepG2 cells was monitored using the fluorescent marker, 2′,7′-dichlorodihydrofluorescein diacetate (H2DCF-DA). Cell apoptosis was evaluated by determining the activity of caspase-3 and by Annexin V/propidium iodide (PI) double staining. Protein expression levels were measured by western blot analysis, and the levels of related cellular kinases were also determined. H2O2 induced pronounced injury to the HepG2 cells, as evidenced by increased levels of malondialdehyde (MDA) and ROS, the decreased activity of SOD and GSH-Px, the increased activitation of caspase-3 and cell apoptosis, and the loss of mitochondrial membrane potential. SYB significantly inhibited the damaging effects of H2O2, indicating that it protected the cells against H2O2-induced oxidative damage. Moreover, pre-treatment with SYB increased the expression of nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase 1 (HO-1) and NAD(P)H dehydrogenase, quinone 1 (NQO1) which are peroxiredoxins. SYB also significantly increased the phosphorylation of AKT. However, this inductive effect was blunted in the presence of the AKT inhibitor, LY294002. The findings of our study suggest that the activation of the AKT/Nrf2 pathway is involved in the cytoprotective effects of SYB against oxidative stress. Our findings provide new insight into the cytoprotective effects of SYB and the possible mechanisms underlying these effects. Thus, SYB may prove to be of therapeutic value for the treatment of various liver diseases.

Synergistic efficacy of meropenem and rifampicin in a murine model of sepsis caused by multidrug-resistant Acinetobacter baumannii

Multidrug-resistant Acinetobacter baumannii becomes an increasing challenge due to the overuse of antibiotics. Combination therapies are considered as effective options to overcome this matter. The present study was to investigate the synergistic activity of meropenem combined with other antibiotics in vitro and in vivo. Checkerboard assay and time-kill assay were performed to study the combination effects in vitro. For the animal model, a murine sepsis model injected with inoculums intraperitoneally was used. Susceptibility test showed that all the twelve strains in this study were resistant to most of the antibiotics except rifampicin. In combination, meropenem plus rifampicin exhibited synergistic activity against six of twelve strains. In the sepsis model, meropenem monotherapy had no therapeutic effect in this model while it can enhance the activity of rifampicin in both survival rate and bacterial clearance from blood. Moreover, combination therapy significantly reduced plasma IL-6 levels compared with rifampicin monotherapy. Pharmacokinetic analysis of rifampicin was also performed in this study. These data above showed that there was synergistic activity between meropenem and rifampicin against multidrug-resistant Acinetobacter baumannii both in vitro and for experimental model of sepsis. It suggested that combining meropenem with rifampicin may be appropriate in treating multidrug-resistant Acinetobacter baumannii infections.

Hydroxysafflor Yellow A Ameliorates Renal Fibrosis by Suppressing TGF-β1-Induced Epithelial-to-Mesenchymal Transition

Objective Renal fibrosis is the common pathological foundation of many chronic kidney diseases (CKDs). The aim of this study was to investigate whether Hydroxysafflor yellow A (HSYA) can preserve renal function by inhibiting the progression of renal fibrosis and the potential mechanisms. Methods Renal fibrosis was induced by unilateral ureteral obstruction (UUO) performed on 7-week-old C57BL/6 mice. HSYA (10, 50 and 100 mg/kg) were intragastrically administered. Sham group and model group were administered with the same volume of vehicle. Serum and kidney samples were collected 14 days after the UUO surgery. Serum biochemical indicators were measured by automatic biochemical analyzer. Histological changes were evaluated by HE and Masson staining. In vitro, the anti-fibrotic effect of HSYA was tested on human recombinant transforming growth factor-β1 (TGF-β1) stimulated HK-2 cells. The protein levels of α-SMA, collagen-I and fibronectin in kidney tissue andHK-2 cells were measured by immunohistochemistry and immunofluorescence. The protein levels of apoptosis-relative and TGF-β1/Smad3 signaling were detected by western blot. Results HSYA slowed the development of renal fibrosis both in vivo and in vitro. In UUO rats, renal function index suggested that HSYA treatment decreased the level of serum creatinine (Scr) and blood urea nitrogen (BUN) rose by UUO (P<0.05). HE staining and Masson staining demonstrated that kidney interstitial fibrosis, tubular atrophy, and inflammatory cell infiltration were notably attenuated in the high-dose HSYA group compared with the model group. The expressions of α-SMA, collagen-I and fibronectin were decreased in the UUO kidney and HK-2 cells of the HSYA-treatment group. Moreover, HSYA reduced the apoptotic rate of HK-2 cells stimulated by TGF-β1. Further study revealed that HSYA regulated the TGF-β1/Smads signaling pathway both in kidney tissue and HK-2 cells. Conclusions These results suggested that HSYA had a protective effect against fibrosis in renal cells, at least partly, through inhibiting TGF-β1/smad3-mediated Epithelial–mesenchymal transition signaling pathway.

Application of a liquid chromatographic/tandem mass spectrometric method to a urinary excretion study of rabeprazole and two of its metabolites in healthy human urine

To study urinary excretion properties of rabeprazole and two of its metabolites, i.e. rabeprazole thioether and desmethyl rabeprazole thioether in human urine, a sensitive, selective, accurate and precise method for the quantification of rabeprazole and two of its metabolites using a liquid chromatographic/tandem mass spectrometric method has been developed and validated. Starting with a 200 μL urine aliquot, a general sample preparation was performed using protein precipitation with methanol. Analytes were separated on a Dikma Inspire™ C18 column (150 mm × 2.1mm, 5 μm) using a mixture of methanol and aqueous 10mM ammonium acetate buffer containing 0.05% formic acid (55:45, v/v) as mobile phase. Linearity was obtained over the concentration range of 0.1446-96.38 ng/mL, 0.3198-319.8 ng/mL and 0.05160-82.53 ng/mL for rabeprazole, rabeprazole thioether, desmethyl rabeprazole thioether in human urine, respectively. The fully validated method was applied to a urine excretion study of rabeprazole sodium administered as a 30 min intravenous infusion for the first time. The calculated cumulative urinary recovery just reached 0.04745‰, 1.272‰ and 0.1631‰ of dose within 24h post-dose for rabeprazole, rabeprazole thioether, and desmethyl rabeprazole thioether, respectively, after intravenous infusion administration, indicating that rabeprazole and its two main metabolites undergo substantial non-renal elimination in healthy Chinese volunteers.

Relative Bioavailability Study of a Novel Prodrug of Tenofovir, Tenofovir Dipivoxil Fumarate, in Healthy Male Fasted Volunteers

AbstractBackground and Objective: Tenofovir dipivoxil fumarate (9-[(R)-2-[[bis (pivaloyloxymethoxy) phosphinoyl] methoxy] propyl] adenine fumarate) is a novel ester prodrug of tenofovir. It has been developed as an anti-hepatitis B virus clinical candidate. The purpose of this study was to determine the pharmacokinetic parameters of tenofovir dipivoxil fumarate (test) and the commercially available tenofovir disoproxil fumarate (Viread®, reference). In addition, the bioavailability of tenofovir dipivoxil fumarate was evaluated in healthy male fasted subjects after a single comparatively equivalent dose. Methods: This single-dose, randomized, two-way, open-label, crossover study was conducted at Xijing Hospital, Xi’an, China. The study drug was administered under fasted conditions. Serial blood samples were obtained over 72 hours after oral administration of each treatment. Bioavailability of the drug was assessed in accordance with the State Food and Drug Administration bioequivalence criteria. Results: Eighteen subjects were enrolled and completed the study, with all study treatments being generally well tolerated. The geometric mean ratios (90% confidence interval [CI]) for maximum plasma concentration (Cmax), area under the plasma concentration-time curve from time zero to the last quantifiable concentration (AUClast), and area under the plasma concentration-time curve from time zero extrapolated to infinity (AUC∞) were 124.49% (90% CI 115.85, 133.79), 122.85% (90% CI 115.55, 130.62), and 122.43% (90% CI 115.71, 129.54), respectively. Conclusion: The relative bioavailability of tenofovir dipivoxil was 20% higher compared with tenofovir disoproxil fumarate; this result was consistent with the preclinical data. Background and Objective: Tenofovir dipivoxil fumarate (9-[(R)-2-[[bis (pivaloyloxymethoxy) phosphinoyl] methoxy] propyl] adenine fumarate) is a novel ester prodrug of tenofovir. It has been developed as an anti-hepatitis B virus clinical candidate. The purpose of this study was to determine the pharmacokinetic parameters of tenofovir dipivoxil fumarate (test) and the commercially available tenofovir disoproxil fumarate (Viread®, reference). In addition, the bioavailability of tenofovir dipivoxil fumarate was evaluated in healthy male fasted subjects after a single comparatively equivalent dose. Methods: This single-dose, randomized, two-way, open-label, crossover study was conducted at Xijing Hospital, Xi’an, China. The study drug was administered under fasted conditions. Serial blood samples were obtained over 72 hours after oral administration of each treatment. Bioavailability of the drug was assessed in accordance with the State Food and Drug Administration bioequivalence criteria. Results: Eighteen subjects were enrolled and completed the study, with all study treatments being generally well tolerated. The geometric mean ratios (90% confidence interval [CI]) for maximum plasma concentration (Cmax), area under the plasma concentration-time curve from time zero to the last quantifiable concentration (AUClast), and area under the plasma concentration-time curve from time zero extrapolated to infinity (AUC∞) were 124.49% (90% CI 115.85, 133.79), 122.85% (90% CI 115.55, 130.62), and 122.43% (90% CI 115.71, 129.54), respectively. Conclusion: The relative bioavailability of tenofovir dipivoxil was 20% higher compared with tenofovir disoproxil fumarate; this result was consistent with the preclinical data.

High performance liquid chromatographic method for the determination of cinepazide maleate and its application to a pharmacokinetic study in rats.

A simple and reliable high performance liquid chromatographic (HPLC) method has been developed and validated to quantify cinepazide maleate, a calcium blocker, in rat plasma. Cinepazide maleate and Tinidazole (internal standard) have been extracted by a simple liquid-liquid extraction before injection into chromatographic system. Chromatographic separation was achieved on a reversed phase C18 column with a mobile phase consisted of a water mixture of 10mM potassium dihydrogen phosphate (pH=4.5):methanol (40:60, v/v), pumped at flow rate of 1.0mL/min, and detected at 303nm. The method exhibited a linear range of 0.12-120μg/mL in blank rat plasma, with the lower detection limit of 0.06μg/mL. The method was statistically validated for linearity, accuracy, precision, selectivity and stability following FDA guidelines. The intra- and inter-assay coefficients of variation did not exceed ±15% from the nominal concentration. The accuracy of cinepazide maleate was within ±15% of the theoretical value. The assay has been applied successfully in a pharmacokinetic study of cinepazide maleate after a single intravenous at three doses in rat. And cinepazide maleate injection can improve the bioavailability of cinepazide maleate greatly, and has a dose-dependence profile in rats.

Impact on L-carnitine Homeostasis of Short-term Treatment with the Pivalate Prodrug Tenofovir Dipivoxil

Pivalate‐generating prodrugs have been suggested to cause clinically significant hypocarnitinaemia. Tenofovir dipivoxil, a novel ester prodrug of tenofovir, can be used for treatment for hepatitis B and HIV infection and it was necessary to evaluate the effect of its treatment on carnitine homeostasis. We sought to investigate the effect of Class 1 drug tenofovir dipivoxil on endogenous L‐carnitine level during a 72‐hr test in healthy Chinese volunteers and to establish a suitable dose of L‐carnitine nutritional supplement for patients who were administered short‐term tenofovir dipivoxil tablets for treatment for hepatitis B and herpes simplex virus infection. Tenofovir dipivoxil was administered in one of eight dosing regimens (single dose 150, 300 and 600 mg, multiple dose 300, 450, and 600 mg, multiple dose 450 (600) mg tenofovir dipivoxil and 0.5 g L‐carnitine) to gender‐balanced groups of 84 healthy Chinese volunteers. Plasma concentrations of L‐carnitine were quantified before, during and after treatment. Plasma L‐carnitine concentrations fell during tenofovir dipivoxil dosing. The nadir in L‐carnitine concentration was dependent on the dose of tenofovir dipivoxil and it decreased from 6.1 ± 0.6 to 4.4 ± 0.8 μg/ml, 6.1 ± 1.8 to 3.3 ± 1.2 μg/ml, 6.2 ± 0.6 to 2.5 ± 0.5 μg/ml for single doses of 150, 300, 600 mg tenofovir dipivoxil tablets and from 6.0 ± 1.4 to 2.1 ± 1.5 μg/ml, 6.2 ± 0.4 to 0.9 ± 0.5 μg/ml for multiple doses of 450, 600 mg tenofovir dipivoxil tablets, respectively. Short‐term administration of tenofovir dipivoxil results in hypocarnitinaemia and increased losses of carnitine in resulting of minor adverse events of decreased food appetite, nausea, abdominal distention and muscle weakness.

Effects of Amlodipine on the Oral Bioavailability of Cephalexin and Cefuroxime Axetil in Healthy Volunteers

In this study, the authors compared the effects of amlodipine (AML) on the bioavailability of cephalexin (LEX) and cefuroxime axetil (CXM). Twenty‐four healthy men were randomized to 4 treatments according to a crossover design with a 14‐day washout. After an overnight fast, they were administered orally LEX 500 mg alone, LEX 500 mg 2 hours after oral administration of AML 5 mg, CXM 500 mg alone, and CXM 500 mg 2 hours after oral administration of AML 5 mg. All participants completed the whole study without side effects being observed. Pharmacokinetic data were analyzed by noncompartmental modeling with WinNonlin software. The geometric mean (GM) ratios were 1.38 (90% confidence interval [CI], 1.32‐1.45) for the area under the concentration‐time curve (AUC) for LEX and 1.27 (1.18‐1.36) for the maximum concentration of drug in serum (Cmax) for LEX followed by AML versus alone. In contrast, no significant differences were found in the pharmacokinetic parameters of CXM between treatments (P < .05). They authors conclude that AML possesses an enhancement effect in β‐lactam antibiotic bioavailability (in this case, LEX), and this interaction may be specific to the peptidomimetic β‐lactam antibiotics.

Pharmacokinetics and bioavailability in healthy Chinese volunteers of a novel rabeprazole sterile powder formulation for injection.

Rabeprazole sterile powder for injection (RSPI) is a new formulation, compared with rabeprazole enteric coated tablets. The aim of this study was to evaluate the pharmacokinetic properties and bioavailability of RSPI in healthy Chinese volunteers. Pharmacokinetic studies included an ascending single dose of 10, 20, 40 mg, and a multiple doses of 20 mg. A bioavailability study was evaluated following a single dose of 20 mg between RSPI and Pariet® Pharmacokinetic parameters of rabeprazole given in each treatment period were calculated using non-compartmental analysis. In the PK study, after a single intravenous dose of 10, 20, and 40 mg, the main pharmacokinetic parameters for rabeprazole were as follows: Cmax 566.88, 897.23, 2,171.6 ng/mL; AUClast/794.31, 1,122.76, 2,446.85 ng×h/mL, respectively. After multiple doses of 20 mg, the main pharmacokinetic parameters for rabeprazole were Cmax 991.90 ng/mL, AUClast 1,261.08 ng×h/mL. In the BA study, after a single oral dose of Pariet® 20 mg, the main pharmacokinetic parameters for rabeprazole were Cmax 582.74 ng/mL, AUClast 1,135.5 ng×h/mL. RSPI produced a less-than-proportional increase in exposure with increasing dose in healthy subjects. The accumulation ratio was 1.0, suggesting RSPI displayed no accumulation after repeated administration. The bioavailability of RSPI was increased by ~ 11% as measured by AUClast compared with Pariet® after a single oral administration.

Effect of food on the pharmacokinetics of triflusal and its major active metabolite, 2-hydroxy-4-trifluoromethyl benzoic acid, in healthy subjects.

OBJECTIVE The objective of this study was to evaluate the pharmacokinetic parameters of triflusal and its major active metabolite, 2-hydroxy-4-trifluoromethyl benzoic acid (HTB), following a single oral dose of 900 mg in healthy subjects under fed and fasting conditions. METHODS The study participants (n=12) were randomized to receive one 900 mg triflusal capsule in a fasting condition (no food for 12 hours) or a fed condition (after a high-fat meal); after a 2-week washout period, participants received the same dose of triflusal capsule under the converse condition. Pharmacokinetic parameters were calculated using WinNonlin 6.2 software. Safety was evaluated through assessment of adverse events, standard laboratory evaluations, vital signs, and 12-lead electrocardiography. RESULTS The mean Cmax of triflusal and HTB were 13.96, 110.2 ug/mL for the fasting state and 9.546, 97.15 ug/mL for the fed state, respectively. The AUC0-144 of triflusal and HTB were 19.66, 5,572 hxμg/mL for the fasting state and 22.20, 5,038 hxμg/mL for the fed state, the AUC0-∞ of triflusal and HTB were 19.79, 6,333 hxμg/mL for the fasting state and 22.44, 5,632 hxμg/mL for the fed state, respectively. The results showed that Cmax and AUCs for triflusal were outside the bioequivalency (BE) interval after food intake, but there was no statistically significant change for HTB. CONCLUSION High-fat food intake may affect the pharmacokinetics of triflusal capsule in healthy subjects.