The aim of this study is to evaluate the efficacy of total saponins of Dioscorea (TSD), an extract of the Chinese herbal Bi Xie, on hyperuricemia and to elucidate the underlying mechanisms. The rat hyperuricemia model was established by administration of adenine. Thirty-two rats were randomly allocated into 4 groups: model group, low/high-dose TSD-treated groups, and allopurinol-treated group. Meanwhile, 8 rats were used as normal controls. Serum uric acid (UA), blood urea nitrogen (BUN), serum creatinine (Scr), and organic anion transporting polypeptide 1A1 (OATP1A1) levels were measured. Comparison between the model group and treatment (allopurinol and TSD) groups showed the serum UA levels were significantly decreased in treatment groups. TSD had similar effects to allopurinol. It was found that the OATP1A1 protein expression levels in treatment groups were higher than in model group and normal controls. And different from the allopurinol-treated groups, TSD-treated group had elevated OATP1A1 expression levels in the stomach, liver, small intestine and large intestine tissues. It was suggested that TSD may facilitate the excretion of UA and lower UA levels by up-regulating OATP1A1 expression.
Animals ; Creatinine ; blood ; Dioscorea ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Hyperuricemia ; drug therapy ; Intestines ; drug effects ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Organic Anion Transporters, Sodium-Independent ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Saponins ; pharmacology ; therapeutic use ; Stomach ; drug effects ; metabolism ; Up-Regulation ; Uric Acid ; blood

Organic anion transporting polypeptide 1B1 (OATP1B1) is an important liver-specific uptake transporter, which mediates transport of numerous endogenous substances and drugs from blood into hepatocytes. To identify and investigate potential modulators of OATP1B1 from natural products, the effect of 21 frequently used natural compounds and extracts on OATP1B1-mediated fluorescein methotrexate transport was studied by using Chinese hamster ovary cells stably expressing OATP1B1 (CHO-OATP1B1) in 96-well plates. This method could be used for the screening of large compound libraries. Our studies showed that some flavonoids (e.g., quercetin, quercitrin, rutin, chrysanthemum flavonoids and mulberrin) and triterpenoids (e.g., glycyrrhetinic acid and glycyrrhizic acid) were inhibitors of OATP1B1 with IC50 values less than 16 µmol · L(-1). The IC50 value of glycyrrhetinic acid on OATP1B1 was comparable to its blood concentration in clinics, indicating an OATPlB1-mediated drug-drug interaction could occur. Structure-activity relationship analysis showed that flavonoids had much higher inhibitory activity than their glycosides. Furthermore, the type and length of saccharides had a significant effect on their activity. In addition, we used OATP1B1 substrates fluvastatin and rosuvastatin as probe drugs to investigate the substrate-dependent effect of several natural compounds on the function of OATP1B1 in vitro. Our results demonstrated that the effect of these natural products on the function of OATPlB1 was substrate-dependent. In summary, this study would be conducive to predicting and avoiding potential OATP1B1-mediated drug-drug and drug-food interactions and thus provide the experimental basis and guidance for rational drug use.
Animals ; Biological Products ; CHO Cells ; Cricetulus ; Drug Interactions ; Fatty Acids, Monounsaturated ; pharmacology ; Flavonoids ; pharmacology ; Indoles ; pharmacology ; Inhibitory Concentration 50 ; Organic Anion Transporters ; genetics ; metabolism ; Rosuvastatin Calcium ; pharmacology ; Structure-Activity Relationship

Hepatitis B virus (HBV) is the prototype of hepatotropic DNA viruses (hepadnaviruses) infecting a wide range of human and non-human hosts. Previous studies with duck hepatitis B virus (DHBV) identified duck carboxypeptidase D (dCPD) as a host specific binding partner for full-length large envelope protein, and p120 as a binding partner for several truncated versions of the large envelope protein. p120 is the P protein of duck glycine decarboxylase (dGLDC) with restricted expression in DHBV infectible tissues. Several lines of evidence suggest the importance of dCPD, and especially p120, in productive DHBV infection, although neither dCPD nor p120 cDNA could confer susceptibility to DHBV infection in any cell line. Recently, sodium taurocholate cotransporting polypeptide (NTCP) has been identified as a binding partner for the N-terminus of HBV large envelope protein. Importantly, knock down and reconstitution experiments unequivocally demonstrated that NTCP is both necessary and sufficient for in vitro infection by HBV and hepatitis delta virus (HDV), an RNA virus using HBV envelope proteins for its transmission. What remains unclear is whether NTCP is the major HBV receptor in vivo. The fact that some HBV patients are homozygous with an NTCP mutation known to abolish its receptor function suggests the existence of NTCP-independent pathways of HBV entry. Also, NTCP very likely mediates just one step of the HBV entry process, with additional co-factors for productive HBV infection still to be discovered. NTCP offers a novel therapeutic target for the control of chronic HBV infection.
Animals ; Carboxypeptidases/genetics/*metabolism ; Gene Products, pol/genetics/metabolism ; Heparan Sulfate Proteoglycans/metabolism ; Hepatitis B virus/*physiology ; Hepatocytes/metabolism/virology ; Organic Anion Transporters, Sodium-Dependent/antagonists & inhibitors/genetics/metabolism ; RNA Interference ; Symporters/antagonists & inhibitors/genetics/metabolism ; Viral Envelope Proteins/metabolism ; Virus Internalization

OBJECTIVETo study the effect of Zhusha Anshen pill, cinnabar, HgS, HgCl2 and MeHg on the gene expression of renal transporters in mice.

METHODHealthy male mice were given equivalent physiological saline, Zhusha Anshen pill (1.8 g · kg(-1), containing 0.17 g · kg(-1) of mercury), cinnabar (0.2 g · kg(-1), containing 1.7 g · kg(-1) of mercury), high dose cinnabar (2 g · kg(-1), containing 1.7 g · kg(-1) of mercury), HgS (0.2 g · kg(-1), containing 0.17 g · kg(-1) of mercury), HgCl2 (0.032 g · kg(-1), containing 0. 024 g · kg(-1) of mercury), MeHg (0.026 g · kg(-1), containing 0.024 g · kg(-1) of mercury), once daily, for 30 d, measuring body mass gain. 30 days later, the mice were sacrificed. The mercury accumulation in kidneys was detected with atomic fluorescence spectrometer. Expressions of Oat1, Oat2, Oat3, Mrp2, Mrp4, Urat1 were detected with RT-PCR.

RESULTCompared with the normal control group, a significant accumulation of Hg in kidney in HgCl2 and MeHg groups was observed (P <0.05), but these changes were not found in other groups. Compared with normal control group, mRNA expressions of Oat1 and Oat2 were evidently lower in HgCl2 and MeHg groups, but mRNA expressions of Mrp2 were apparently higher in HgCl2 group (P <0.05), mRNA expression of Mrp4 was significant higher in HgCl2 and MeHg groups, and mRNA expression of Urat1 was apparently lower in MeHg group.

CONCLUSIONHgCl2 and MeHg groups show significant difference from the normal group in mercury accumulation in kidneys and gene expression of kidney transporters, but with no difference between other groups and the normal group. Compared with HgCl2 and MeHg, cinnabar and its compounds could cause lower renal toxicity to mice.

Animals ; Carrier Proteins ; genetics ; Drugs, Chinese Herbal ; toxicity ; Gene Expression ; drug effects ; Kidney ; drug effects ; metabolism ; Male ; Mercuric Chloride ; toxicity ; Mercury Compounds ; toxicity ; Methylmercury Compounds ; toxicity ; Mice ; Multidrug Resistance-Associated Proteins ; genetics ; Organic Anion Transport Protein 1 ; genetics ; Organic Anion Transporters, Sodium-Independent ; genetics

OBJECTIVETo explore roles of mRNA and protein expressions of organic anion transporting polypeptide (oatp2b1) of rats with high fat diet and overstrain induced Pi deficiency syndrome in the transporting of damp turbidity.

METHODSTotally 24 SD rats were randomly divided into three groups, i.e., the normal group, the overstrain group, and the high fat diet group, 8 in each group. After successful modeling, one piece of tissues such as spleen, kidney, liver, lung, stomach, small intestine, and large intestine was taken from each rat. Rats of the overstrain group were bonded by specially made bondage cylinder, 3 h each time on odd days, and forced to swim in cold water (10 +/- 1) degrees C for 7 min on even days alternatively for twelve weeks. Rats in the model group and the normal group were fed with standard routine granular forage for 12 weeks. Rats in the high fat diet group were fed with high fat forage for twelve weeks. All rats drank and ate freely. The mRNA and protein expressions of oatp2b1 were detected in the seven tissues using RT-PCR and Western blot.

RESULTSThe mRNA expression of oatp2b1 in liver and kidney tissues of rats in the high fat diet group was higher when compared with that of the normal group and the overstrain group (P < 0.01, P < 0.05). The oatp2b1 mRNA expression in the normal group was sequenced from high to low as liver > lung > spleen > larger intestine > small intestine > kidney > stomach. The oatp2b1 mRNA expression in the overstrain group was sequenced from high to low as liver > lung > larger intestine > spleen > kidney > stomach > small intestine. The oatp2b1 mRNA expression in the high fat diet group was sequenced from high to low as liver > lung > spleen > small intestine > kidney > larger intestine > stomach. The oatp2b1 protein expression in the lung tissue was sequenced from high to low as the overstrain group > the normal group > the high fat diet group (P > 0.05). The oatp2b1 protein expression in the spleen tissue was sequenced from high to low as the high fat diet group > the normal group > the overstrain group (P > 0.05). The oatp2b1 protein expression in the kidney tissue was sequenced from high to low as the normal group > the overstrain group > the high fat diet group (P > 0.05). The oatp2b1 protein expression in the liver tissue was sequenced from high to low as the normal group > the high fat diet group > the overstrain group (P > 0.05). Of them, the oatp2b1 protein expressed extremely less in the stomach, large intestine, and small intestine. The oatp2b1 protein expression in the normal group was sequenced from high to low as lung >spleen > liver, kidney > stomach, larger intestine, and small intestine. The oatp2b1 protein expression in the overstrain group was sequenced from high to low as lung > spleen > kidney > liver > stomach, larger intestine, and small intestine. The oatp2b1 protein expression in the high fat diet group was sequenced from high to low as spleen > lung > kidney > liver > stomach, larger intestine, and small intestine. However, there was no statistical significance among the three groups by pair-wise comparison (P > 0.05).

CONCLUSIONSKidney and liver might play important roles in the transportation and transformation of damp under the state of Pi deficiency syndrome. Oatp2b1 may be one of the material bases involved in the transportation and transformation of damp turbidity. Pi's function of governing transportation and transformation of damp might not only include the functions of the gastrointestinal tract, but also include partial liver and kidney functions.

Animals ; Diet, High-Fat ; Disease Models, Animal ; Fatigue ; diagnosis ; metabolism ; Kidney ; metabolism ; Liver ; metabolism ; Male ; Medicine, Chinese Traditional ; Organic Anion Transporters ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo analyze the clinical features and SLC25A13 gene mutations of a child with citrin deficiency complicated with purpura, convulsive seizures and methioninemia.

METHODSThe patient was subjected to physical examination and routine laboratory tests. Blood amino acids and acylcarnitines, and urine organic acids and galactose were analyzed respectively with tandem mass spectrometry and gas chromatographic mass spectrometry. SLC25A13 gene mutation screening was conducted by high resolution melt (HRM) analysis.

RESULTSThe petechiae on the patient's face and platelet count (27×10(9)/L, reference range 100×10(9)/L-300×10(9)/L) supported the diagnosis of immunologic thrombocytopenic purpura (ITP). Laboratory tests found that the patient have abnormal coagulation, cardiac enzyme, liver function and liver enzymes dysfunction. Tandem mass spectrometry also found methionine to be increased (286 μmol/L, reference ranges 8-35 μmol/L). The patient did not manifest any galactosemia, citrullinemia and tyrosinemia. Analysis of SLC25A13 gene mutation found that the patient has carried IVS16ins3kb, in addition with abnormal HRM result for exon 6. Direct sequencing of exon 6 revealed a novel mutation c.495delA. The same mutation was not detected in 100 unrelated healthy controls. Further analysis of her family has confirmed that the c.495delA mutation has derived from her farther, and that the IVS16ins3kb was derived from her mother.

CONCLUSIONThe clinical features and metabolic spectrum of citrin deficiency can be variable. The poor prognosis and severity of clinical symptoms of the patient may be attributed to the novel c.495delA mutation.

Amino Acid Metabolism, Inborn Errors ; genetics ; pathology ; Calcium-Binding Proteins ; deficiency ; genetics ; DNA Mutational Analysis ; methods ; Female ; Glycine N-Methyltransferase ; deficiency ; genetics ; Humans ; Infant ; Mitochondrial Membrane Transport Proteins ; genetics ; Organic Anion Transporters ; deficiency ; genetics ; Pedigree ; Purpura ; genetics ; pathology ; Seizures ; genetics ; pathology

AIM: The present study was undertaken to characterize the effects of Wuling San on urate excretion and renal function, and explore its possible mechanisms of action in hyperuricemic mice. METHODS: Mice were administered with 250 mg·kg(-1) potassium oxonate by gavage once daily (10 animals/group) for seven consecutive days to develop a hyperuricemia model. Different doses of Wuling powder were orally initiated on the day 1 h after oxonate was given, separately. Allopurinol was used as a positive control. Serum and urine levels of uric acid and creatinine, and fractional excretion of uric acid (FEUA) were measured in hyperuricemic mice treated with Wuling San and allopurinol. Simultaneously, renal mRNA and protein levels of urate transporter 1 (mURAT1), glucose transporter 9 (mGLUT9), organic anion transporter 1 (mOAT1), as well as organic cation/carnitine transporters mOCT1, mOCT2 and mOCTN2, were assayed by semi-quantitative RT-PCR and Western blot methods, respectively. RESULTS AND CONCLUSION Compared to the hyperuricemia control group, Wuling San significantly reduced serum uric acid and creatinine levels, increased 24 h urate and creatinine excretion, and FEUA in hyperuricemic mice, exhibiting its ability to enhance urate excretion and improve kidney function. Wuling San was found to down-regulate mRNA and protein levels of mURAT1 and mGLUT9, as well as up-regulate mOAT1 in the kidney of hyperuricemic mice. Moreover, Wuling San up-regulated renal mRNA and protein levels of mOCT1, mOCT2 and mOCTN2, leading to kidney protection in this model.
Animals ; Drugs, Chinese Herbal ; administration & dosage ; Glucose Transport Proteins, Facilitative ; genetics ; metabolism ; Humans ; Hyperuricemia ; drug therapy ; genetics ; metabolism ; Kidney ; drug effects ; metabolism ; Male ; Mice ; Organic Anion Transport Protein 1 ; genetics ; metabolism ; Organic Anion Transporters ; genetics ; metabolism ; Up-Regulation ; drug effects ; Uric Acid ; metabolism

Liver is regarded as one of the most important organs for drug clearance in the body, which mediates both the metabolism and biliary excretion of drugs. Transporters are a class of functional membrane proteins and control the movement of substances into or out of cells. Transporters, which are extensively expressed in the liver, play important roles in the drug hepatic disposition by regulating the uptake of drugs from blood into hepatocytes or the efflux of drugs and their metabolites into bile. In this review, the localization, functions and substrate selectivity of the major transporters in the liver will be summarized, and the impacts of these transporters on drug hepatic disposition, the potential drug-drug interactions as well as their genetic polymorphisms will also be reviewed.
ATP Binding Cassette Transporter, Sub-Family G, Member 2 ; ATP-Binding Cassette Transporters ; genetics ; metabolism ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Bile ; metabolism ; Biological Transport ; Drug Interactions ; Humans ; Liver ; metabolism ; Membrane Transport Proteins ; genetics ; metabolism ; Metabolic Clearance Rate ; Multidrug Resistance-Associated Proteins ; genetics ; metabolism ; Neoplasm Proteins ; genetics ; metabolism ; Organic Anion Transporters ; genetics ; metabolism ; Organic Anion Transporters, Sodium-Dependent ; metabolism ; Organic Anion Transporters, Sodium-Independent ; genetics ; metabolism ; Organic Cation Transport Proteins ; genetics ; metabolism ; Pharmacokinetics ; Polymorphism, Genetic ; Symporters ; metabolism

OBJECTIVETo investigate the diagnostic value of histopathological changes in the liver of patients with neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD).

METHODSLiver specimens from 10 cases of NICCD were evaluated by hematoxylin-eosin stain, histochemistry and immunohistochemistry (EnVision method). SLC25A13 mutation analysis was performed to correlate with histopathology.

RESULTSMost specimens showed varying degrees of fat deposition in hepatocytes, necrotic inflammation, cholestasis and fibrosis (so-called tetralogy). The combination of the above four histological changes was highly characteristic for NICCD. With the progression of the disease, hepatic fibrosis deteriorated and ultimately led to cirrhosis.

CONCLUSIONSNICCD should be suspected in the presence of cholestasis during infancy. A liver biopsy must be performed to rule out other liver diseases. The tetralogy of the hepatic histopathological changes has a highly diagnostic value for NICCD, which is also practical for accurately assessing the degree of inflammation and fibrosis, and similarly the progression of hepatic cirrhosis.

Biopsy ; Calcium-Binding Proteins ; deficiency ; genetics ; metabolism ; Cholestasis, Intrahepatic ; etiology ; genetics ; pathology ; Disease Progression ; Female ; Hepatocytes ; pathology ; Humans ; Infant ; Liver ; pathology ; Liver Cirrhosis ; pathology ; Male ; Mitochondrial Membrane Transport Proteins ; genetics ; Mutation ; Organic Anion Transporters ; deficiency ; genetics ; metabolism

OBJECTIVETo study organic anion transporting polypeptide (OATP) superfamily member 4a1 (oatp4a1) mRNA expression in the Pi deficiency model rats, thus exploring its mechanism for transporting and transforming the dampness.

METHODSSix SD rats of SPF grade were used to prepare over-fatigue impairing Pi model. Another 12 SD rats were randomly divided into the blank control group and the high fat diets group, 6 in each. The special binding tube was used for the over-fatigue impairing Pi model group on the odd day, 3 h each time. Then the rats were forced to swim in the cold water (10 degrees C +/- 1 degrees C) for 7 min on the even day, for 2 successive weeks. Rats in the model group and the blank control group were granulated feed for 12 weeks, while rats in the high fat group were fed with high fat diet for 12 weeks. All rats were free to take food and drink water. The mRNA and protein expressions of oatp4al were detected in the Fei, Pi, Gan, Shen, Wei, Xiaochang, and Dachang using Real-time fluorescent quantitative polymerase chain reaction (FQ-PCR) and Western blot.

RESULTSRats in the model group were idled together with lousy defecate and coarse skin. They ate and drank less, and lost body weight (P<0.05). They were consistent with clinical manifestations of Pi deficiency syndrome, indicating that the over-fatigue impairing Pi animal model was successfully established. Rats in the high fat group started to have poor appetite and languish spirit, move lazily and addict to sleep, have coarse, dark, and colorless hair 9 weeks later, indicating phlegm dampness syndrome. Compared with the blank control group, the average body weight increased in the high fat group at the 9th week (P<0.05). The oatp4a1 mRNA expressed in the Fei, Pi, Gan, Shen, Wei, Xiaochang, and Dachang. There was no statistical difference in the oatp4al mRNA expression among all tissues (P>0.05). The oatp4al mRNA expressions were higher in the Fei and Shen of the high fat group than in the Gan (P<0.05).

CONCLUSIONSoatp4al might be one of the basic substances in the transportation and transformation of phlegm dampness. Of them, Fei, Shen, and Dachang might play important roles in the transportation and transformation of phlegm dampness.

Animals ; Antiporters ; metabolism ; Diet, High-Fat ; Eye Proteins ; metabolism ; Fatigue ; metabolism ; Male ; Organic Anion Transporters ; metabolism ; RNA, Messenger ; genetics ; Rats ; Yin Deficiency ; metabolism