The study aims to explore the herb-drug interaction between Xuefu Zhuyu Decoction(XFZY) and atorvastatin(AT). Reverse transcription polymerase chain reaction(RT-PCR) was used to analyze the transcription levels of proteins related to drug metabolism and transport in LS174T cells, detect the intracellular drug uptake under various substrate concentrations and incubation time, and optimize the model reaction conditions of transporter multidrug resistance protein 1(MDR1)-specific probe Rhodamine 123 and AT to establish a cell model for investigating the human intestinal drug interaction. The cell counting kit-8(CCK-8) method was adopted to evaluate the cytotoxicity of XFZY on LS174T cells. After a single and continuous 48 h culture with XFZY, AT or Rhodamine 123 was added for co-incubation. The effect and mechanism of XFZY on human intestinal absorption of AT were analyzed by measuring the intracellular drug concentrations and transcription levels of related transporters and metabolic enzymes. The results of in vitro experiments show that a single co-culture with a high concentration of XFZY significantly increases the intracellular concentrations of Rhodamine 123 and AT. A high concentration of XFZY co-culture for 48 h increases the AT uptake level, significantly induces the CYP3A4 and UGT1A1 gene expression levels, and inhibits the OATP2B1 gene expression level. To compare with the evaluation results of the in vitro human cell model, the pharmacokinetic experiment of XFZY combined with AT was carried out in rats. Sprague-Dawley(SD) rats were randomly divided into a blank control group and an XFZY group. After 14 days of continuous intragastric administration, AT was given in combination. The liquid chromatography-mass spectrometry(LC-MS)/MS method was used to detect the concentrations of AT and metabolites 2-hydroxyatorvastatin acid(2-HAT), 4-hydroxyatorvastatin acid(4-HAT), atorvastatin lactone(ATL), 2-hydroxyatorvastatin lactone(2-HATL), and 4-hydroxyatorvastatin lactone(4-HATL) in plasma samples, and the pharmacokinetic parameters were calculated. Pharmacokinetic analysis in rats shows that continuous administration of XFZY does not significantly change the pharmacokinetic characteristics of AT in rats, but the AUC_(0-6 h) values of AT and metabolites 2-HAT, 4-HAT, and 2-HATL increase by 21.37%, 14.94%, 12.42%, and 6.68%, respectively. The metabolic rate of the main metabolites shows a downward trend. The study indicates that administration combined with XFZY can significantly increase the uptake level of AT in human intestinal cells and increase the exposure level of AT and main metabolites in rats to varying degrees. The mechanism may be mainly due to the inhibition of intestinal MDR1 transport activity.
Animals ; Drugs, Chinese Herbal/administration & dosage* ; Atorvastatin/administration & dosage* ; Humans ; Rats ; Rats, Sprague-Dawley ; Male ; Intestines/cytology* ; Intestinal Mucosa/metabolism* ; Herb-Drug Interactions ; Cytochrome P-450 CYP3A/metabolism* ; Intestinal Absorption/drug effects*

BACKGROUND: To date, results on relationship between CYP3A4 gene polymorphism were limited and inconclusive, and no study focused on the influence of CYP3A4 gene-obesity interaction on breast cancer risk, especially in Chinese women. The purpose of this study was to evaluate the impact of four single nucleotide polymorphisms (SNPs) of CYP3A4 gene, the SNP-SNP and gene-environment interactions on the susceptibility to breast cancer in Chinese women. METHODS: Logistic regression was used to explore the relationship between four SNPs of CYP3A4 gene and the risk of breast cancer. Generalized multifactor dimensionality reduction (GMDR) was used to screen the best SNP-SNP and gene-abdominal obesity interaction combinations among four SNPs and abdominal obesity. Haplotype examination among 4 SNPs was conducted using the SHEsis web-based platform. RESULTS: Logistic regression analysis showed that carriers of rs2242480- T allele have significantly higher breast cancer risk, than those with rs2242480- CC genotype, adjusted OR (95%CI) was 1.68 (1.23-2.16) and 2.03 (1.53-2.58) for participants with CT genotype and TT genotype under additive model. We did not find any notable interactions between the four SNPs within the CYP3A4 gene. GMDR model found a significant association in a two-locus model involving rs2242480 and obesity, with a p-value of 0.018. Stratified analysis found that breast cancer risk was the highest in obese participants with rs2242480- CT or TT genotype, compared to those non-obese participants with rs2242480- CC genotype, OR (95%CI) was 3.02 (1.83-4.25). We found that all haplotype combinations were not correlated with breast cancer risk. CONCLUSIONS We found that the T allele of rs2242480 within the CYP3A4 gene and interaction between rs2242480 and obesity were associated with an increased risk of breast cancer. However, the results of this study were only applicable to the Han ethnic group and cannot be generalized to other ethnic groups in China, and more SNPs of CYP3A4 gene should been enrolled in the analysis in the future, to verify the results obtained in this study.
Adult ; Aged ; Female ; Humans ; Middle Aged ; Breast Neoplasms/etiology* ; China/epidemiology* ; Cytochrome P-450 CYP3A/metabolism* ; Gene-Environment Interaction ; Genetic Predisposition to Disease ; Haplotypes ; Obesity/epidemiology* ; Polymorphism, Single Nucleotide ; Risk Factors ; East Asian People

Gukang Capsules are often used in combination with drugs to treat fractures, osteoarthritis, and osteoporosis. Cytochrome P450(CYP450) mainly exists in the liver and participates in the oxidative metabolism of a variety of endogenous and exogenous substances and serves as an important cause of drug-metabolic interactions and adverse reactions. Therefore, it is of great significance to study the effect of Gukang Capsules on the activity and expression of CYP450 for increasing its clinical rational medication and improving the safety of drug combination. In this study, the Cocktail probe method was used to detect the changes in the activities of CYP1A2, CYP3A2, CYP2C11, CYP2C19, CYP2D4, and CYP2E1 in rat liver after treatment with high-, medium-and low-dose Gukang Capsules. The rat liver microsomes were extracted by the calcium chloride method, and protein expression of the above six CYP isoform enzymes was detected by Western blot. The results showed that the low-dose Gukang Capsules could induce CYP3A2 and CYP2D4 in rats, medium-dose Gukang Capsules had no effect on them, and high-dose Gukang Capsules could inhibit them in rats. The high-dose Gukang Capsules did not affect CYP2C11 in rats, but low-and medium-dose Gukang Capsules could induce CYP2C11 in rats. Gukang Capsules could inhibit CYP2C19 in rats and induce CYP1A2 in a dose-independent manner, but did not affect CYP2E1. If Gukang Capsules were co-administered with CYP1A2, CYP2C19, CYP3A2, CYP2C11, and CYP2D4 substrates, the dose should be adjusted to avoid drug interactions.
Rats ; Animals ; Cytochrome P-450 CYP1A2/metabolism* ; Cytochrome P-450 CYP2C19 ; Cytochrome P-450 CYP2E1/pharmacology* ; Rats, Sprague-Dawley ; Cytochrome P-450 Enzyme System/metabolism* ; Microsomes, Liver ; Liver ; Cytochrome P-450 CYP3A/metabolism*

BACKGROUND: The renal function of individuals is one of the reasons for the variations in therapeutic response to various drugs. Patients with renal impairment are often exposed to drug toxicity, even with drugs that are usually eliminated by hepatic metabolism. Previous study has reported an increased plasma concentration of indoxyl sulfate and decreased plasma concentration of 4β-hydroxy (OH)-cholesterol in stable kidney transplant recipients, implicating indoxyl sulfate as a cytochrome P450 (CYP) inhibiting factor. In this study, we aimed to evaluate the impact of renal impairment severity-dependent accumulation of indoxyl sulfate on hepatic CYP3A activity using metabolic markers. METHODS: Sixty-six subjects were enrolled in this study; based on estimated glomerular filtration rate (eGFR), they were classified as having mild, moderate, or severe renal impairment. The plasma concentration of indoxyl sulfate was quantified using liquid chromatography-mass spectrometry (LC-MS). Urinary and plasma markers (6β-OH-cortisol/cortisol, 6β-OH-cortisone/cortisone, 4β-OH-cholesterol) for hepatic CYP3A activity were quantified using gas chromatography-mass spectrometry (GC-MS). The total plasma concentration of cholesterol was measured using the enzymatic colorimetric assay to calculate the 4β-OH-cholesterol/cholesterol ratio. The correlation between variables was assessed using Pearson's correlation test. RESULTS: There was a significant negative correlation between MDRD eGFR and indoxyl sulfate levels. The levels of urinary 6β-OH-cortisol/cortisol and 6β-OH-cortisone/cortisone as well as plasma 4β-OH-cholesterol and 4β-OH-cholesterol/cholesterol were not correlated with MDRD eGFR and the plasma concentration of indoxyl sulfate. CONCLUSION: Hepatic CYP3A activity may not be affected by renal impairment-induced accumulation of plasma indoxyl sulfate.
Cholesterol ; Cytochrome P-450 CYP3A* ; Cytochrome P-450 Enzyme System* ; Cytochromes* ; Drug-Related Side Effects and Adverse Reactions ; Gas Chromatography-Mass Spectrometry ; Glomerular Filtration Rate ; Humans ; Indican ; Kidney ; Metabolism ; Plasma ; Spectrum Analysis ; Transplant Recipients

This paper aimed to study the six chemical components of Polygoni Multiflori Radix (gallic acid, quercetin, luteolin, kaempferol, resveratrol, apigenin). By the established pregnane X receptor (human pregnant X receptor, PXR) CYP3A4 mediated drug induced rapid screening technique, the effect of chemical components on the cell activity was detected by MTS cell method, and the value of IC₅₀ was calculated. The dual luciferase reporter system was used to co-transfect PXR reporter gene expression vector containing transcriptional regulation and CYP3A4 with HepG2 cells, with 10 μmol·L⁻¹ rifampicin (RIF) as a positive control, and 10 μmol·L⁻¹ of ketoconazole (TKZ) as negative control. Gallic acid, quercetin, luteolin, kaempferol, apigenin, resveratrol(5, 10, 20 μmol·L⁻¹) were used to incubate for 24 h, and the luciferase activity was detected. The results showed that when plasmid pcDNA3.1 was co-transfected with pGL4.17-CYP3A4, gallic acid and resveratrol had an inhibitory effect on the regulation of CYP3A4, and quercetin, luteolin, kaempferol had an inductive effect on CYP3A4; when pcDNA3.14-PXR was co-transfected with pGL4.17-CYP3A4, quercetin, luteolin, kaempferol, apigenin, resveratrol had an inductive effect. To sum up, the 6 reported liver injury components had inhibitory or activating effects on CYP3A4. After PXR plasmid was involved, 5 components had an inductive effect on CYP3A4, and the inductive effects of 2 components were significantly different. In this experiment, we found that 2 kinds of potential liver injury components in Polygoni Multiflori Radix had been induced by CYP3A4, which was achieved through PXR regulation. It suggested that attention shall be paid to potential drug interactions when combined with Polygoni Multiflori Radix, so as to improve the safety and efficacy.
Cytochrome P-450 CYP3A ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Hep G2 Cells ; Humans ; Liver ; Phytochemicals ; pharmacology ; Plant Roots ; chemistry ; Polygonum ; chemistry ; Pregnane X Receptor ; metabolism

Pharmacological activities and adverse side effects of ginkgolic acids (GAs), major components in extracts from the leaves and seed coats of Ginkgo biloba L, have been intensively studied. However, there are few reports on their hepatotoxicity. In the present study, the metabolism and hepatotoxicity of GA (17 : 1), one of the most abundant components of GAs, were investigated. Kinetic analysis indicated that human and rat liver microsomes shared similar metabolic characteristics of GA (17 : 1) in phase I and II metabolisms. The drug-metabolizing enzymes involved in GA (17 : 1) metabolism were human CYP1A2, CYP3A4, UGT1A6, UGT1A9, and UGT2B15, which were confirmed with an inhibition study of human liver microsomes and recombinant enzymes. The MTT assays indicated that the cytotoxicity of GA (17 : 1) in HepG2 cells occurred in a time- and dose-dependent manner. Further investigation showed that GA (17 : 1) had less cytotoxicity in primary rat hepatocytes than in HepG2 cells and that the toxicity was enhanced through CYP1A- and CYP3A-mediated metabolism.
Animals ; Cells, Cultured ; Cytochrome P-450 CYP1A2 ; metabolism ; Cytochrome P-450 CYP3A ; metabolism ; Ginkgo biloba ; chemistry ; Glucuronosyltransferase ; metabolism ; Hepatocytes ; chemistry ; drug effects ; enzymology ; metabolism ; Humans ; Kinetics ; Liver ; chemistry ; drug effects ; enzymology ; metabolism ; Microsomes, Liver ; chemistry ; drug effects ; enzymology ; metabolism ; Plant Extracts ; chemistry ; metabolism ; toxicity ; Rats ; Rats, Sprague-Dawley ; Salicylates ; chemistry ; metabolism ; toxicity

Cardiovascular disease (CVD) is the most common cause of death in patients with non-alcoholic fatty liver disease (NAFLD). New therapeutic strategies which have the potential for slowing down the evolution of NAFLD and reducing CVD-related mortality are urgently needed. Statins are well recognized in the treatment of dyslipidemia, but their use in the treatment of NAFLD is limited due to the safety concerns. Ilexgenin A (IA) is one of the main bioactive compounds in 'Shan-lv-cha', an herbal tea commonly used in China. In the present study, we investigated the possible synergistic therapeutic effects of IA and simvastatin (SV) on NAFLD. IA or SV showed beneficial effects on the rats with NAFLD by lowering the liver weight, liver index and plasma levels of alanine aminotransferase and aspartate aminotransferase, regulating abnormal metabolism of lipids and ameliorating steatosis in liver. IA significantly enhanced the hypolipidemic and anti-inflammation effects of SV. Furthermore, a sensitive, accurate, convenient and reproducible LC-MS method was developed to investigate the effects of IA on the pharmacokinetics of SV. No significant changes were observed in pharmacokinetic parameters of SV and simvastatin hydroxy acid in the IA plus SV co-treated group in comparison with those in the group treated with SV alone. The mRNA levels and activity of CYP3A1 were not altered by IA. In conclusion, the results obtained from the present study should be helpful for further clinical application of SV and IA alone or in combination.
Alanine Transaminase ; metabolism ; Animals ; Aspartate Aminotransferases ; metabolism ; Cytochrome P-450 CYP3A ; genetics ; metabolism ; Diet, High-Fat ; Disease Models, Animal ; Drug Synergism ; Drug Therapy, Combination ; Lipids ; blood ; Liver ; metabolism ; pathology ; physiopathology ; Male ; Molecular Structure ; Non-alcoholic Fatty Liver Disease ; blood ; drug therapy ; Rats ; Rats, Sprague-Dawley ; Simvastatin ; analogs & derivatives ; pharmacokinetics ; therapeutic use ; Transcription, Genetic ; Triterpenes ; chemistry ; therapeutic use

Ginsenoside Rh2 (Rh2) is one of the major bioactive ginsenosides in Panax ginseng. However, the oral bioavailability of Rh2 is low, with P-glycoprotein (P-gp) and CYP3A4 being reported to be the main factors. The purpose of the present study was to determine the enhancing effect of piperine on the oral bioavailability as well as bioactivity of Rh2. The inhibitory effect of piperine on P-gp and CYP3A4 was determined using a Caco-2 monolayer model and a recombinant CYP3A4 metabolic system, respectively. The pharmacokinetics of oral Rh2 (10 mg·kg) administered alone or in combination with piperine (10 and 20 mg·kg) was performed in rats. The immune boosting effect of Rh2 was assessed in rats by measuring IL-12 level after treated by Rh2 alone or co-administered with piperine. The results indicated that piperine significantly increased the permeability of Rh2 and inhibited the metabolism of Rh2. The pharmacokinetic study results showed that the AUC of Rh2 was significantly increased in combination with piperine at high dose (20 mg·kg) when compared to the control group, with relative bioavailability of 196.8%. The increase of Rh2 exposure led to increased serum levels of IL-12. In conclusion, piperine may be used as a bioenhancer to improve pharmacological effect of Rh2 when given orally.
Administration, Oral ; Alkaloids ; administration & dosage ; Animals ; Benzodioxoles ; administration & dosage ; Biological Availability ; Caco-2 Cells ; Cytochrome P-450 CYP3A ; metabolism ; Drugs, Chinese Herbal ; administration & dosage ; Ginsenosides ; administration & dosage ; pharmacokinetics ; Humans ; Interleukin-2 ; metabolism ; Panax ; chemistry ; Piperidines ; administration & dosage ; Polyunsaturated Alkamides ; administration & dosage ; Rats ; Rats, Sprague-Dawley

<p>OBJECTIVEConsuming botanical dietary supplements or herbal drugs along with prescription drugs may lead to potential pharmacokinetic-pharmacodynamic (PK-PD) herb-drug interactions (HDI). The present study focuses on the importance of and novel approach for assessing HDI in integrative medicine with case examples of two frequently-used Ayurvedic Rasayana botanicals.p><p>METHODSThe aqueous extracts of Asparagus racemosus (ARE) and Gymnema sylvester (GSE) were prepared as per Ayurvedic Pharmacopoeia of India. Chemoprofiling of these extracts was done using high-performance liquid chromatography (HPLC). Additionally, ARE was characterized for the presence of shatavarins IV and I using HPLC & mass spectroscopy respectively. Effects of ARE and GSE were investigated on rat liver microsome using testosterone probe drug assay. The changes in formation of metabolite (6-β hydroxy testosterone) were monitored on incubation of testosterone alone, testosterone with ketoconazole, ARE and GSE using HPLC. Half inhibitory concentration (IC50) was used to predict plausible HDI.p><p>RESULTSARE and GSE showed no inhibition with IC50 values >1 000 μg/mL while the standard inhibitor ketoconazole completely abolished CYP3A4-dependent activity at 0.531 μg/mL and IC50 was found to be 0.036 μg/mL.p><p>CONCLUSIONARE and GSE prepared as per Ayurvedic Pharmacopoeia of India were found to be safe for CYP3A4-mediated inhibitory HDI in rats. Our in vitro study suggests the need of further in vivo investigation for HDI in order to provide clinical relevance.p>
Animals ; Asparagus Plant ; Chromatography, High Pressure Liquid ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; pharmacology ; Gymnema sylvestre ; Herb-Drug Interactions ; Isoenzymes ; antagonists & inhibitors ; Plant Extracts ; pharmacology ; Rats ; Rats, Wistar

In adult liver, CYP3A4 plays an important role in the metabolism of a wide range of endogenous and exogenous compounds. To investigate whether there is a single nucleotide polymorphism (SNP) of CYP3A4 intron 2 in the liver and its effects on the mRNA expression and enzymatic activity of CYP3A4, genomic DNA was extracted from 96 liver tissue samples obtained from patients who had undergone liver surgery. An SNP of CYP3A4 intron 2 was identified by polymerase chain reaction (PCR)-single-strand confirmation polymorphism and DNA sequencing. The mRNA expression of CYP3A4 was determined by the fluorescence quantitative PCR technique. The enzymatic activity of CYP3A4 was measured using erythromycin and testosterone as probe substrates. Twelve patients were found to have the SNP/T4127G CYP3A4 within intron 2. The mRNA levels of CYP3A4 in wild-type and SNP/T4127G samples were 2.62±1.09 and 2.79±1.63, respectively (P>0.05). Erythromycin N-demethylase activity in wild-type and SNP/T4127G samples were 121.2±32.8 and 124.7±61.6 nmol·mg(-1)·min(-1), respectively (P>0.05). The activity of testosterone 6β-hydroxylase was significantly different between wild-type (648±173 pmol·mg(-1)·min(-1)) and SNP/T4127G samples (540±196 pmol·mg(-1)·min(-1); P<0.05). In conclusion, the SNP/T4127G of CYP3A4 intron 2 exists in the liver. This SNP does not affect the mRNA expression of CYP3A4 but significantly decreases the hepatic microsomal testosterone 6β-hydroxylase activity of CYP3A4. Furthermore, this study indicates that the appropriate selection of probe substrates is very important in studying the relationship between the genotype and phenotype of CYP3A4.
Adult ; Asian Continental Ancestry Group ; genetics ; China ; Cytochrome P-450 CYP3A ; genetics ; metabolism ; Erythromycin ; metabolism ; Genotype ; Humans ; Introns ; Liver ; enzymology ; Phenotype ; Polymorphism, Single Nucleotide ; RNA, Messenger ; genetics ; Sequence Analysis, DNA ; methods ; Testosterone ; metabolism