Identification and characterization of endogenous biomarkers for hepatic vectorial transport (OATP1B3-P-gp) function using metabolomics with serum pharmacology.

The organic anion-transporting polypeptide 1B3 and P-glycoprotein (P-gp) provide efficient directional transport (OATP1B3-P-gp) from the blood to the bile that serves as a key determinant of hepatic disposition of the drug. Unfortunately, there is still a lack of effective means to evaluate the disposal ability mediated by transporters. The present study was designed to identify a suitable endogenous biomarker for the assessment of OATP1B3-P-gp function in the liver. We established stably transfected HEK293T-OATP1B3 and HEK293T-P-gp cell lines. Results showed that azelaic acid (AzA) was an endogenous substrate for OATP1B3 and P-gp using serum pharmacology combined with metabolomics. There is a good correlation between the serum concentration of AzA and probe drugs of rOATP1B3 and rP-gp when rats were treated with their inhibitors. Importantly, after 5-fluorouracil-induced rat liver injury, the relative mRNA level and expression of rOATP1B3 and rP-gp were markedly down-regulated in the liver, and the serum concentration of AzA was significantly increased. These observations suggest that AzA is an endogenous substrate of both OATP1B3 and P-gp, and may serve as a potential endogenous biomarker for the assessment of the function of OATP1B3-P-gp for the prediction of changes in the pharmacokinetics of drugs transported by OATP1B3-P-gp in liver disease states.

Effects of Early Versus Delayed Feeding in Patients With Acute Pancreatitis: A Systematic Review and Meta-analysis.

BACKGROUND: The aim of this study was to summarize the optimal strategy for early feeding in patients with acute pancreatitis. METHODS: The search was undertaken in electronic databases, which compared early with delayed feeding in acute pancreatitis. The primary outcome was the length of hospital stay (LOHS). The second outcomes were intolerance of refeeding, mortality, and total cost of each patient. This meta-analysis followed the "Preferred Reporting Items for Systematic Reviews and Meta-analyses" guideline. Research is registered by PROSPERO, CRD42020192133. RESULTS: A total of 20 trials involving 2168 patients were included, randomly assigned to the early feeding group (N = 1033) and delayed feeding group (N = 1135). The LOHS was significantly lower in the early feeding group than the delayed feeding group (mean difference: -2.35, 95% CI: -2.89 to -1.80; P < 0.0001), no matter the mild or severe subgroup ( Pint = 0.69). The secondary outcome of feeding intolerance and mortality were no significant difference (risk ratio: 0.96, 0.40 to 2.16, P = 0.87 and 0.91, 0.57 to 1.46, P = 0.69; respectively). Moreover, the hospitalization cost was significantly less in the early feeding group, resulting in an average savings of 50%. In patients with severe pancreatitis, early feeding after 24 hours may be beneficial ( Pint = 0.001). CONCLUSION: Early oral feeding can significantly reduce the LOHS and hospitalization costs in patients with acute pancreatitis without increasing feeding intolerance or mortality. In patients with severe pancreatitis, early feeding after 24 hours may be beneficial.

Accumulation of Renal Fibrosis in Hyperuricemia Rats Is Attributed to the Recruitment of Mast Cells, Activation of the TGF-ß1/Smad2/3 Pathway, and Aggravation of Oxidative Stress.

Renal fibrosis is relentlessly progressive and irreversible, and a life-threatening risk. With the continuous intake of a high-purine diet, hyperuricemia has become a health risk factor in addition to hyperglycemia, hypertension, and hyperlipidemia. Hyperuricemia is also an independent risk factor for renal interstitial fibrosis. Numerous studies have reported that increased mast cells (MCs) are closely associated with kidney injury induced by different triggering factors. This study investigated the effect of MCs on renal injury in rats caused by hyperuricemia and the relationship between MCs and renal fibrosis. Our results reveal that hyperuricemia contributes to renal injury, with a significant increase in renal MCs, leading to renal fibrosis, mitochondrial structural disorders, and oxidative stress damage. The administration of the MCs membrane stabilizer, sodium cromoglycate (SCG), decreased the expression of SCF/c-kit, reduced the expression of α-SMA, MMP2, and inhibited the TGF-ß1/Smad2/3 pathway, thereby alleviating renal fibrosis. Additionally, SCG reduced renal oxidative stress and mitigated mitochondrial structural damage by inhibiting Ang II production and increasing renal GSH, GSH-Px, and GR levels. Collectively, the recruitment of MCs, activation of the TGF-ß1/Smad2/3 pathway, and Ang II production drive renal oxidative stress, ultimately promoting the progression of renal fibrosis in hyperuricemic rats.

Clinical Aspects of Drug-Drug Interaction and Drug Nephrotoxicity at Renal Organic Cation Transporters 2 (OCT2) and Multidrug and Toxin Exclusion 1, and 2-K (MATE1/MATE2-K).

The organic cation transporter 2 (OCT2) belongs to the SLC22 family, while the multidrug and toxin extrusion 1 and 2-K (MATE1/MATE2-K) belong to the SLC47 family, are localized to the basolateral and apical membrane of human renal proximal tubular epithelial cells, respectively. They are polyspecific transporters that enable the transit of structurally diversified drugs with overlapping selectivity across plasma membranes. OCT2 and MATE1/2-K are critically involved in renal secretion, pharmacokinetics (PK), and toxicity of cationic drugs. Drug-drug interactions (DDIs) at OCT2 and/or MATE1/2-K have been shown to result in clinical impacts on PK, therapeutic efficacy and are probably involved in the renal accumulation of drugs. Sites of OCT2 and MATE1/2-K expression and function play an essential role in the pharmacokinetics and toxicity of drugs, such as cisplatin. Thus, knowing the sites (basolateral vs. apical) of the interaction of two drugs at transporters is essential to understanding whether this interaction helps prevent or enhance drug-induced nephrotoxicity. In this work, an overview of OCT2 and MATE1/2-K is presented. Primary structure, membrane location, functional properties, and clinical impact of OCT2 and MATE1/2-K are presented. In addition, clinical aspects of DDIs in OCT2 and MATE1/2-K and their involvement in drug nephrotoxicity are compiled.

Enalapril increases the urinary excretion of metformin in rats by inducing multidrug and toxin excretion protein 1 in the kidney.

Two-thirds of patients with type 2 diabetes mellitus have hypertension, and thus the combination of two or more drugs to treat these diseases is common. It has been shown that the combination of metformin and enalapril has beneficial effects, but few studies have evaluated the interactions between these two drugs. This study investigated the effects of enalapril on the pharmacokinetics and urinary excretion of metformin in rats, with a focus on transporter-mediated drug interactions. Rats were dosed orally with metformin alone (100 mg/kg) or in combination with enalapril (4 mg/kg). The concentration of metformin was measured by high performance liquid chromatography and the level of organic cation transporters (rOCTs) and multidrug and toxin excretion protein 1 (rMATE1), which mediate the uptake and efflux of metformin, respectively, were evaluated by immunoblotting. After single and 7-day dosing, the plasma concentration of metformin in the co-administration group was significantly lower than that in the metformin-only group, and the CL/F and urinary excretion were increased in the co-administration group. Enalapril did not affect the Kp of metformin but reduced renal slice-uptake of metformin. The expression of rMATE1 was increased, whereas rOCT2 expression was decreased in rat kidney. Importantly, long-term co-administration of metformin and enalapril markedly decreased the level of lactic acid and uric acid in the blood. Enalapril increases the urinary excretion of metformin through the up-regulation of rMATE1. This reveals a new mechanism of drug interactions and provides a basis for drug dosage adjustment when these drugs are co-administered.

Investigating Intestinal Transporter Involvement in Rivaroxaban Disposition through Examination of Changes in Absorption.

PURPOSE: The involvement of the intestinally expressed xenobiotic transporters P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP) have been implicated in rivaroxaban disposition based on in vitro studies, similar to what had previously been proposed for apixaban. We recently showed that these efflux transporters were not clinically relevant for apixaban disposition and examine here their relevance for this second Factor Xa inhibitor. METHODS: Using recently published methodologies to discern metabolic- from transporter- mediated drug interactions, a critical evaluation was undertaken of 9 rivaroxaban studies reporting 12 DDIs, one study of food effects and one study of hepatic function. RESULTS: Rationale examination of these clinical studies using basic pharmacokinetic theory finds little support for the clinical significance of intestinal efflux transporters in rivaroxaban disposition. Drug-drug interactions are most likely adequately predicted based on the level of CYP 3A metabolism. CONCLUSION: These analyses indicate that inhibition of efflux transporters appears to have negligible, clinically insignificant effects on the rivaroxaban absorption process, which is consistent with the concern that predictions based on in vitro measures may not translate to a clinically relevant interaction in vivo. We emphasize the need to evaluate gastric emptying, dissolution and other processes related to absorption when using MAT changes to indicate efflux transporter inhibition.

Evidence-based Guideline for Therapeutic Drug Monitoring of Vancomycin: 2020 Update by the Division of Therapeutic Drug Monitoring, Chinese Pharmacological Society.

BACKGROUND: Clinical practice guidelines or recommendations often require timely and regular updating as new evidence emerges, because this can alter the risk-benefit trade-off. The scientific process of developing and updating guidelines accompanied by adequate implementation can improve outcomes. To promote better management of patients receiving vancomycin therapy, we updated the guideline for the therapeutic drug monitoring (TDM) of vancomycin published in 2015. METHODS: Our updated recommendations complied with standards for developing trustworthy guidelines, including timeliness and rigor of the updating process, as well as the use of the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) approach. We also followed the methodology handbook published by the National Institute for Health and Clinical Excellence and the Spanish National Health System. RESULTS: We partially updated the 2015 guideline. Apart from adults, the updated guideline also focuses on pediatric patients and neonates requiring intravenous vancomycin therapy. The guideline recommendations involve a broadened range of patients requiring TDM, modified index of TDM (both 24-hour area under the curve and trough concentration), addition regarding the necessity and timing of repeated TDM, and initial dose for specific subpopulations. Overall, 1 recommendation was deleted and 3 recommendations were modified. Eleven new recommendations were added, and no recommendation was made for 2 clinical questions. CONCLUSIONS: We updated an evidence-based guideline regarding the TDM of vancomycin using a rigorous and multidisciplinary approach. The updated guideline provides more comprehensive recommendations to inform rational and optimized vancomycin use and is thus of greater applicability.

Pyridoxal isonicotinoyl hydrazone inhibition of FXR is involved in the pathogenesis of isoniazid-induced liver injury.

Isoniazid (INH)-induced liver injury may be associated with inhibition of the liver farnesoid X receptor (FXR). However, the relationship between FXR and INH-induced liver injury remained unclear. The present study was performed to clarify the role of inhibition of FXR in the pathogenesis of INH-induced liver injury and to further identify potential inhibitors of FXR from INH and its metabolites. HepaRG cells were treated with INH (10 mM) plus mixed bile acids (BA) and rats were treated with INH (60-600 mg/kg p.o.) or INH plus obeticholic acid (OCA, 10 mg/kg), a potent FXR agonist, for seven days. INH can cause BA-dependent toxicity and apoptosis with elevated intracellular bile acids in vitro; indeed, in these studies, liver bile acids and mRNA levels for Cyp7a1, an FXR target gene were increased, while mRNA levels for FXR and Shp were significantly decreased, and these changes could be prevented by co-treatment with the FXR agonist OCA. In silico molecular docking studies showed that INH, acetyl isoniazid, isonicotinic acid and PIH may be potential FXR inhibitors, and a TR-FRET FXR-coactivator assay confirmed that PIH is a strong antagonist of FXR (IC50 = 52 nM). To further determine if PIH also inhibits FXR activity in vivo, rats were treated with PIH directly (5 mg/kg). Liver total bile acids were significantly increased while FXR expression was not changed, but Shp mRNA levels were significantly decreased and Cyp7a1 mRNA was significantly increased, consistent with PIH acting as an FXR antagonist. In summary, PIH inhibition of liver FXR function leading to bile acid accumulation in hepatocytes may be an early pathogenesis event in INH-induced liver injury.

Determination of Total and Unbound Meropenem, Imipenem/Cilastatin, and Cefoperazone/Sulbactam in Human Plasma: Application for Therapeutic Drug Monitoring in Critically Ill Patients.

BACKGROUND: Critically ill patients show several pathophysiological alterations that can complicate antibiotic dosing. Hence, there is a strong rationale to individualize anti-infective dosing in these patients by using therapeutic drug monitoring (TDM). The current study aimed to develop and validate a liquid chromatography-tandem mass spectrometry method for the simultaneous determination of total and unbound plasma concentrations of 3 commonly used antibiotics (meropenem, imipenem/cilastatin, and cefoperazone/sulbactam) in the treatment of infections in critically ill patients in China, which could be suitable for routine TDM in hospital laboratories. METHODS: The unbound drug was separated from the bound drug by ultrafiltration. Simple protein precipitation was used for sample preparation. Meropenem, imipenem/cilastatin, cefoperazone/sulbactam, and their corresponding internal standards were then resolved using the Waters CORTECS C18 column. All the compounds were detected using electrospray ionization in the positive/negative ion-switching mode. RESULTS: The calibration curves were linear for all compounds, with correlation coefficients (R) above 0.99 for total concentrations in human plasma and unbound concentrations in the plasma ultrafiltrate. For both total and unbound drugs, the relative errors and intra-assay/interassay relative standard deviations were below 15%. The limit of quantification was 0.05 mcg/mL for both total plasma concentrations and plasma ultrafiltrate concentrations of all compounds. CONCLUSIONS: The method was simple, rapid, and reliable and is currently being used to provide a TDM service to enhance the efficacious use of the 3 antibiotics.

Effect of high uric acid on the disposition of metformin: in vivo and in vitro studies.

Metformin is always used as the baseline antidiabetic therapy for patients with type 2 diabetes mellitus (T2DM) and hyperuricemia. Metformin is excreted into urine through active secretion mediated by rOCTs and rMATE1.The aim of this study was to identify the effects of high uric acid on the disposition and its mechanism. For the in vivo study, a hyperuricemic animal model was induced by intraperitoneal injection of potassium oxonate (250 mg/kg) in rats. Metformin (100 mg/kg) was administered orally to investigate the pharmacokinetics in control and hyperuricemic rats, respectively. For the in vitro study, HEK293 and HepaRG cells were used to investigate the effect of uric acid (15 mg/dl) on the expression of OCT1, OCT2 and MATE1 and the disposition of metformin, respectively. The in vivo study showed that the AUC0 â†’ 600 of metformin was significantly decreased by 33.3%, whereas the cumulative urinary excretion of metformin was increased by 25.4% in hyperuricemic rats compared with that in control rats. The renal rOCT1, rOCT2 and rMATE1 and hepatic rMATE1 levels were increased in hyperuricemic rats compared with those in control rats, respectively. The in vitro study showed that uric acid could upregulate the expression of OCT2 and MATE1 in HEK293 cells and MATE1 in HepaRG cells and increase the intracellular metformin concentration in these two cell lines. These results demonstrated that a high uric acid level promoted urinary metformin excretion and decreased the plasma metformin concentration; the in vivo and in vitro studies provided a possible explanation being that high uric acid could upregulate the expression of renal metformin transporters OCTs and MATE1.

Alteration of renal excretion pathways in gentamicin-induced renal injury in rats.

The kidney plays a major part in the elimination of many drugs and their metabolites, and drug-induced kidney injury commonly alters either glomerular filtration or tubular transport, or both. However, the renal excretion pathway of drugs has not been fully elucidated at different stages of renal injury. This study aimed to evaluate the alteration of renal excretion pathways in gentamicin (GEN)-induced renal injury in rats. Results showed that serum cystatin C, creatinine and urea nitrogen levels were greatly increased by the exposure of GEN (100 mg kg-1 ), and creatinine concentration was increased by 39.7% by GEN (50 mg kg-1 ). GEN dose-dependently upregulated the protein expression of rOCT1, downregulated rOCT2 and rOAT1, but not affected rOAT2. Efflux transporters, rMRP2, rMRP4 and rBCRP expressions were significantly increased by GEN(100), and the rMATE1 level was markedly increased by GEN(50) but decreased by GEN(100). GEN(50) did not alter the urinary excretion of inulin, but increased metformin and furosemide excretion. However, GEN(100) resulted in a significant decrease of the urinary excretion of inulin, metformin and p-aminohippurate. In addition, urinary metformin excretions in vivo were significantly decreased by GEN(100), but slightly increased by GEN(50). These results suggested that GEN(50) resulted in the induction of rOCTs-rMATE1 and rOAT3-rMRPs pathway, but not changed the glomerular filtration rate, and GEN(100)-induced acute kidney injury caused the downregulated function of glomerular filtration -rOCTs-rMATE1 and -rOAT1-rMRPs pathway.

Multicomponent determination of traditional Chinese medicine preparation yin-zhi-huang injection by LC-MS/MS for screening of its potential bioactive candidates using HepaRG cells.

Yin-zhi-huang (YZH) injection is an injectable multiherbal prescription derived from the ancient Chinese medicine formula of Yin-chen-hao-tang, which is widely used in the clinic for the treatment of jaundice and chronic liver diseases. To date, the systematic study of the components in this multiherbal prescription still lacks suitable analytical methods that are able to simultaneously detect a broad array of components at low concentrations. In this study, a new liquid chromatography-tandem mass spectrometry method using dynamic multiple reaction monitoring mode was developed to determine multiple peaks in traditional Chinese medicine preparation YZH injection. This simple, selective and sensitive method enabled the quantification of 22 components with standard materials with a lower limit of quantification of 1.46-12.5 ng/mL in cell lysates. This method was successfully applied to celluar uptake and binding investigation of components in YZH injection. The results indicated that this strategy might be a useful approach for rapidly screening of the potential bioactive candidates from YZH injection, and the discovered candidates could be used to investigate the pharmacodynamics in further studies.

Inhibition of invasion by N-trans-feruloyloctopamine via AKT, p38MAPK and EMT related signals in hepatocellular carcinoma cells.

N-trans-feruloyloctopamine (FO) isolated from Garlic skin was identified as the primary antioxidant constituents, however, the effect of which on HCC invasion is still unclear. Herein, the FO was synthesized and its antitumor activities were evaluated in HCC cell lines. Cellular functional analyses have revealed that the reformed FO owns strong abilities of inhibiting cell proliferation and invasion in HCC cells. Molecular data have further showed that FO could significantly decrease the phosphorylation levels of Akt and p38 MAPK. In addition, the expression of Slug was inhibited and the level of E-cadherin increased. Molecular docking analysis indicates that the H-bond and hydrophobic interactions were critical for FO and E-cadherin binding, but FO did not seem to act directly on phosphorylated Akt and p38 MAPK. We have thus concluded that reformed FO inhibits cell invasion might be directly through EMT related signals (E-cadherin) and indirectly through PI3K/Akt, p38 MAPK signaling pathways. FO might be a promising drug in HCC treatment and prognosis.

[Effects of metoprolol or/and pravastatin on the pharmacokinetics of metformin in rats].

This study investigates the effects of metoprolol (METO) or/and pravastatin (PRAV) on the pharmacokinetics of metformin (METF) in rats. Twenty-eight male SD rats were divided into METF group, METF+METO group, METF+PRAV group and METF+METO+PRAV group. Blood samples were collected at 10, 20, 40, 60, 90, 120, 180, 240, 360, 480 and 600 min after oral administration of metformin, and concentration of metformin in plasma was determined by HPLC. Compared to the METF group, Cmax of metformin was significantly decreased (P < 0.01) and MRT0−t , t1/2 and V were significantly increased in the METF+METO group; t1/2 was significantly decreased in the METF+PRAV group; Cmax was significantly decreased and MRT0−t was significantly increased in the METF+METO+PRAV group. Compared to the METF+METO group, MRT0−t of metformin was significantly decreased in the METF+METO+PRAV group. Compared to the METF+PRAV group, Cmax of metformin was significantly decreased (P < 0.01), and MRT0−t , t1/2 and V were significantly increased in the METF+METO+PRAV group. There exist multiple drug interactions of metformin, metoprolol and pravastatin in rats.

Metoprolol decreases the plasma exposure of metformin via the induction of liver, kidney and muscle uptake in rats.

Drug interactions are one of the commonest causes of side effects, particularly in long-term therapy. The aim of the current study was to investigate the possible effects of metoprolol on the pharmacokinetics of metformin in rats and to clarify the mechanism of drug interaction. In this study, rats were treated with metformin alone or in combination with metoprolol. Plasma, urine and tissue concentrations of metformin were determined by HPLC. Western blotting and real-time qPCR were used to evaluate the expression of rOCTs and rMATE1. The results showed that, after single or 7-day repeated administration, the plasma concentrations of metformin in the co-administration group were significantly decreased compared with that in the metformin group. However, the parameter V/F of metformin in the co-administration group was markedly increased compared with that in the metformin group. The hepatic, renal and muscular Kp of metformin were markedly elevated after co-administration with metoprolol. Consistently, metformin uptake in rat kidney slices was significantly induced by metoprolol. In addition, multiple administrations of metoprolol significantly reduced the expression of rMATE1 in rat kidney as well as the urinary excretion of metformin. Importantly, after long-term administration, lactic acid and uric acid levels in the co-administration group were increased by 25% and 26%, respectively, compared with that in the metformin group. These results indicate that metoprolol can decrease the plasma concentration of metformin via the induction of hepatic, renal and muscular uptake, and long-term co-administration of metformin and metoprolol can cause elevated lactic acid and uric acid levels. Copyright © 2016 John Wiley & Sons, Ltd.

Monoammonium glycyrrhizinate protects rifampicin- and isoniazid-induced hepatotoxicity via regulating the expression of transporter Mrp2, Ntcp, and Oatp1a4 in liver.

CONTEXT: Drug-induced liver injury (DILI) is associated with altering expression of hepatobiliary membrane transporters. Monoammonium glycyrrhizin (MAG) is commonly used for hepatic protection and may have a correlation with the inhibition effect of multidrug resistance-associated protein 2 (Mrp2). OBJECTIVE: This study evaluates the dynamic protective effect of MAG on rifampicin (RIF)- and isoniazid (INH)-induced hepatotoxicity in rats. MATERIALS AND METHODS: Male Wistar rats were randomly divided into four groups of 15 rats. Liver injury was induced by co-treatment with RIF (60 mg/kg) and INH (60 mg/kg) by gavage administration; MAG was orally pretreated at the doses of 45 or 90 mg/kg 3 h before RIF and INH. Rats in each group were sacrificed at 7, 14, and 21 d time points after drug administration. RESULTS: Liver function, histopathological analysis, and oxidative stress factors were significantly altered in each group. The expression of Mrp2 was significantly increased 230, 760, and 990% at 7, 14, and 21 time points, respectively, in RIF- and INH-treated rats. Compared with the RIF and INH groups, Mrp2 was reduced and Ntcp was significantly elevated by 180, 140, and 160% in the MAG high-dose group at the three time points, respectively. The immunoreaction intensity of Oatp1a4 was increased 170, 190, and 370% in the MAG low-dose group and 160, 290, and 420% in the MAG high-dose group at the three time points, respectively, compared with the RIF and INH groups. DISCUSSION AND CONCLUSION: These results indicated that MAG has a protective effects against RIF- and INH-induced hepatotoxicity. The underlying mechanism may have correlation with its effect on regulating the expression of hepatobiliary membrane transporters.

Serum serotonin reduced the expression of hepatic transporter Mrp2 and P-gp via regulating nuclear receptor CAR in PI-IBS rats.

Hepatic transporters and drug metabolizing enzymes (DMEs) play important roles in the pharmacological effects and (or) side-effects of many drugs, and are regulated by several mediators, including neurotransmitters. This work aimed to investigate whether serum levels of 5-hydroxytryptamine (5-HT) affected the expression of hepatic transporters or DMEs. The expression of hepatic transporters was assessed using the Western-blot technique in a 2,4,6-trinitrobenzenesulfonic-acid-induced rat model of post-infectious irritable bowel syndrome (PI-IBS), in which serum levels of 5-HT were significantly elevated. To further clarify the underlying mechanism, the 5-HT precursor 5-hydroxytryptophan (5-HTP) and the 5-HT depleting agent parachlorophenylalanine (pCPA) were applied to adjust serum levels of 5-HT. Serum levels of 5-HT were measured using LC-MS/MS; the expression of hepatic transporters, DMEs, and nuclear receptors were examined by Western-blot technique. Our results showed that in PI-IBS rats the expression of multidrug resistance protein 2 (Mrp2) was significantly decreased, while colonic enterochromaffin cell density and serum levels of 5-HT were all significantly increased. Moreover, 5-HTP treatment significantly increased serum levels of 5-HT and decreased the expression of Mrp2 and glycoprotein P (P-gp), whereas treatment with pCPA markedly decreased serum levels of 5-HT and increased the expression of Mrp2 and P-gp. Our results indicated that serum 5-HT regulates the expression of Mrp2 and P-gp, and the underlying mechanism may be related to the altered expression of the nuclear receptor constitutive androstane receptor (CAR).

Development of a LC-MS/MS method for simultaneous determination of metoprolol and its metabolites, α-hydroxymetoprolol and O-desmethylmetoprolol, in rat plasma: application to the herb-drug interaction study of metoprolol and breviscapine.

A simple, specific and sensitive LC-MS/MS method was developed and validated for the simultaneous determination of metoprolol (MET), α-hydroxymetoprolol (HMT) and O-desmethylmetoprolol (DMT) in rat plasma. The plasma samples were prepared by protein precipitation, then the separation of the analytes was performed on an Agilent HC-C18 column (4.6 × 250 mm, 5 µm) at a flow rate of 1.0 mL/min, and post-column splitting (1:4) was used to give optimal interface flow rates (0.2 mL/min) for MS detection; the total run time was 8.5 min. Mass spectrometric detection was achieved using a triple-quadrupole mass spectrometer equipped with an electrospray source interface in positive ionization mode. The method was fully validated in terms of selectivity, linearity, accuracy, precision, stability, matrix effect and recovery over a concentration range of 3.42-7000 ng/mL for MET, 2.05-4200 ng/mL for HMT and 1.95-4000 ng/mL for DMT. The analytical method was successfully applied to herb-drug interaction study of MET and breviscapine after administration of breviscapine (12.5 mg/kg) and MET (40 mg/kg). The results suggested that breviscapine have negligible effect on pharmacokinetics of MET in rats; the information may be beneficial for the application of breviscapine in combination with MET in clinical therapy.

[Study of pharmacokinetics of digoxin in ovariectomized rats model].

This study aims to investigate the change of plasma concentration of digoxin (DIG) in rats with ovariectomy. Twelve female SD rats were randomly assigned into ovariectomized group and sham group (n = 6). All rats plasma was collected after a single dose of 2 mg x kg(-1) DIG administrated orally, serum DIG concentration was determined by LC-MS/MS. The level of P-gp in the intestinal was analyzed by Western blotting. Pharmacokinetic calculations were performed on each individual using DAS 2.0 practical pharmacokinetic software. Compared with the sham group, C(max) of ovariectomized group decreased significantly (P < 0.01). There was no significant difference of AUC(0-t), and the level of P-gp was elevated in ovariectomized group. It was found that C(max) of DIG was significantly reduced after ovariectomy, and the change was associated with the decreased level of estrogen, which contributes to the increased level of P-gp.

Effect of Yin-Zhi-Huang on up-regulation of Oatp2, Ntcp, and Mrp2 proteins in estrogen-induced rat cholestasis.

CONTEXT: Yin-Zhi-Huang (YZH), a prescription of traditional Chinese medicine, is widely used to treat neonatal jaundice or cholestasis. OBJECTIVE: This study investigates the regulatory effect of YZH on the localization and expression of organic anion transporting polypeptides 2 (Oatp2), Na(+)-taurocholate co-transporting polypeptide (Ntcp), multidrug-resistance-associated protein 2 (Mrp2), and bile salt export pump (Bsep) in estrogen-induced cholestasis rats. MATERIAL AND METHODS: Cholestasis model rats were induced via subcutaneous injection of estradiol benzoate (EB, 5 mg/kg/d) for 5 d. Other EB-induced rats were treated with saline (2 ml) or YZH (1.5 g/kg, two times a day) for 7, 14, and 21 d. The biochemical and pathologic examinations were performed. Moreover, the localization and expression of Oatp2, Ntcp, Mrp2, and Bsep were determined by immunohistochemisty and Western blotting, respectively. RESULTS: YZH treatment could significantly decrease the serum total bile acids (TBA) (4.9 ± 0.6-2.8 ± 0.8) and direct bilirubin (DBIL) (2.6 ± 0.7-1.0 ± 0.1) levels, improve the histological disorganization, and, respectively, increase the expression of Oatp2 and Ntcp by 46% and 28% compared with saline-treated (p < 0.05) rats at 14 d. The expression of Mrp2 increased by 45% was observed in YZH treated compared with saline-treated (p < 0.05) rats at 7 d. However, there was a little change in the expression of Bsep (p > 0.05) after YZH treatment for 7, 14, and 21 d. DISCUSSION AND CONCLUSION: In conclusion, the therapeutic effect of YZH to cholestasis could be attributed to the regulation of Oatp2, Ntcp, Mrp2, and Bsep.