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 report...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 CYP1 A2, CYP3 A4, UGT1 A6, UGT1 A9, and UGT2 B15, 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 CYP1 A-and CYP3 A-mediated metabolism.展开更多
Ginkgolic acids(GAs), primarily found in the leaves, nuts, and testa of ginkgo biloba, have been identified with suspected allergenic, genotoxic and cytotoxic properties. However, little information is available about...Ginkgolic acids(GAs), primarily found in the leaves, nuts, and testa of ginkgo biloba, have been identified with suspected allergenic, genotoxic and cytotoxic properties. However, little information is available about GAs toxicity in kidneys and the underlying mechanism has not been thoroughly elucidated so far. Instead of GAs extract, the renal cytotoxicity of GA(15 : 1), which was isolated from the testa of Ginkgo biloba, was assessed in vitro by using MDCK cells. The action of GA(15 : 1) on cell viability was evaluated by the MTT and neutral red uptake assays. Compared with the control, the cytotoxicity of GA(15 : 1) on MDCK cells displayed a time-and dose-dependent manner, suggesting the cells mitochondria and lysosomes were damaged. It was confirmed that GA(15 : 1) resulted in the loss of cells mitochondrial trans-membrane potential(ΔΨm). In propidium iodide(PI) staining analysis, GA(15 : 1) induced cell cycle arrest at the G0/G1 and G2/M phases, influencing on the DNA synthesis and cell mitosis. Characteristics of necrotic cell death were observed in MDCK cells at the experimental conditions, as a result of DNA agarose gel electrophoresis and morphological observation of MDCK cells. In conclusion, these findings might provide useful information for a better understanding of the GA(15 : 1) induced renal toxicity.展开更多
NTCP is specifically expressed on the basolateral membrane of hepatocytes, participating in the enterohepatic circulation of bile salts, especially conjugated bile salts, to maintain bile salts homeostasis. In additio...NTCP is specifically expressed on the basolateral membrane of hepatocytes, participating in the enterohepatic circulation of bile salts, especially conjugated bile salts, to maintain bile salts homeostasis. In addition, recent studies have found that NTCP is a functional receptor of HBV and HDV. Therefore, it is important to study the interaction between drugs and NTCP and identify the inhibitors/substrates of NTCP. In the present study, a LLC-PK1 cell model stably expressing human NTCP was established, which was simple and suitable for high throughput screening, and utilized to screen and verify the potential inhibitors of NTCP from 102 herbal medicinal ingredients. The results showed that ginkgolic acid(GA)(13 : 0), GA(15 : 1), GA(17 : 1), erythrosine B, silibinin, and emodin have inhibitory effects on NTCP uptake of TCNa in a concentration-dependent manner. Among them, GA(13 : 0) and GA(15 : 1) exhibited the stronger inhibitory effects, with IC_(50) values being less than 8.3 and 13.5 mmol·L^(-1), respectively, than the classical inhibitor, cyclosporin A(CsA)(IC_(50) = 20.33 mmol·L^(-1)). Further research demonstrated that GA(13 : 0), GA(15 : 1), GA(17 : 1), silibinin, and emodin were not substrates of NTCP. These findings might contribute to a better understanding of the disposition of the herbal ingredients in vivo, especially in biliary excretion.展开更多
Stellera chamaejasme L.is a traditional Chinese medicine with a long history to treat stubborn skin ulcer,and it also has antiviral and antitumor effects.Neochamaejasmine B(NCB),Neochamaejasmine A(NCA)and Chamaechromo...Stellera chamaejasme L.is a traditional Chinese medicine with a long history to treat stubborn skin ulcer,and it also has antiviral and antitumor effects.Neochamaejasmine B(NCB),Neochamaejasmine A(NCA)and Chamaechromone(CMC)are the major components in dried roots of Stellera chamaejasme L..Our studies suggested that NCB,NCA and CMC are inhibitors of Organic anion transporter 1(OAT1).OAT1 is encoded by solute carrier family 22 member 6 gene(SLC22 A6)in humans and plays a critical role in the organic anion drug uptake and excretion in the kidney.Lamivudine is the typical substrate of OAT1 and is frequently used in combination with other antiviral drugs in clinical antiviral treatments.The aim of this study is to investigate the interaction and its mechanism between these bi-flavone components in Stellera chamaejasme L.and lamivudine via OAT1 both in vitro and in vivo.In vitro,the uptake studies in Madin-Darby canine kidney(MDCK)cells overexpressing OAT1 suggested that NCB inhibited the uptake of 6-CFL and lamivudine.Similar results were obtained for NCA and CMC.NCB was a noncompetitive and competitive inhibitor interaction with OAT1.IC50 values of NCB,NCA and CMC for inhibiting OAT1-mediated lamivudine transport were 2.46,8.35 and 0.61μmol·L^–1,respectively.In vivo,the pharmacokinetic results of lamivudine in rats showed that the mean area under the plasma concentration-time curve(AUC0-∞)and maximal plasma concentration(Cmax)of lamivudine after co-administration is increased 2.94-fold and 1.87-fold,respectively,compared to lamivudine administration alone.The results of interactions between lamivudine and these bi-flavone components in Stellera chamaejasme L.extracts via OAT1 in vivo are consistent with studies in vitro.The inhibition of OAT1-mediated uptake of lamivudine by NCB,NCA and CMC is the possible mechanism for Stellera chamaejasme L.extracts improving the oral bioavailability of lamivudine in rats.展开更多
基金supported by the National Key Project of China(No.2017YFC0908600)the National Natural Science Foundation of China(No.81173120)the National Natural Science Foundation of Zhejiang Province(No.LQ15H310003)
文摘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 CYP1 A2, CYP3 A4, UGT1 A6, UGT1 A9, and UGT2 B15, 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 CYP1 A-and CYP3 A-mediated metabolism.
基金supported by National Natural Science Foundation of China(No.81173120)International Science&Technology Cooperation Program of China(No.2014DFE30050)
文摘Ginkgolic acids(GAs), primarily found in the leaves, nuts, and testa of ginkgo biloba, have been identified with suspected allergenic, genotoxic and cytotoxic properties. However, little information is available about GAs toxicity in kidneys and the underlying mechanism has not been thoroughly elucidated so far. Instead of GAs extract, the renal cytotoxicity of GA(15 : 1), which was isolated from the testa of Ginkgo biloba, was assessed in vitro by using MDCK cells. The action of GA(15 : 1) on cell viability was evaluated by the MTT and neutral red uptake assays. Compared with the control, the cytotoxicity of GA(15 : 1) on MDCK cells displayed a time-and dose-dependent manner, suggesting the cells mitochondria and lysosomes were damaged. It was confirmed that GA(15 : 1) resulted in the loss of cells mitochondrial trans-membrane potential(ΔΨm). In propidium iodide(PI) staining analysis, GA(15 : 1) induced cell cycle arrest at the G0/G1 and G2/M phases, influencing on the DNA synthesis and cell mitosis. Characteristics of necrotic cell death were observed in MDCK cells at the experimental conditions, as a result of DNA agarose gel electrophoresis and morphological observation of MDCK cells. In conclusion, these findings might provide useful information for a better understanding of the GA(15 : 1) induced renal toxicity.
基金supported by National Nature Scientific Foundation of China(No.8117312)International Science&Technology Cooperation Program of China(No.2014DFE30050)+1 种基金Program for Zhejiang Leading Team of S&T Innovation Team(No.2011R50014)Fundamental Research Funds for the Central Universities of China Ministry of Education(2016XZZX001-08)
文摘NTCP is specifically expressed on the basolateral membrane of hepatocytes, participating in the enterohepatic circulation of bile salts, especially conjugated bile salts, to maintain bile salts homeostasis. In addition, recent studies have found that NTCP is a functional receptor of HBV and HDV. Therefore, it is important to study the interaction between drugs and NTCP and identify the inhibitors/substrates of NTCP. In the present study, a LLC-PK1 cell model stably expressing human NTCP was established, which was simple and suitable for high throughput screening, and utilized to screen and verify the potential inhibitors of NTCP from 102 herbal medicinal ingredients. The results showed that ginkgolic acid(GA)(13 : 0), GA(15 : 1), GA(17 : 1), erythrosine B, silibinin, and emodin have inhibitory effects on NTCP uptake of TCNa in a concentration-dependent manner. Among them, GA(13 : 0) and GA(15 : 1) exhibited the stronger inhibitory effects, with IC_(50) values being less than 8.3 and 13.5 mmol·L^(-1), respectively, than the classical inhibitor, cyclosporin A(CsA)(IC_(50) = 20.33 mmol·L^(-1)). Further research demonstrated that GA(13 : 0), GA(15 : 1), GA(17 : 1), silibinin, and emodin were not substrates of NTCP. These findings might contribute to a better understanding of the disposition of the herbal ingredients in vivo, especially in biliary excretion.
基金supported by the National Key R&D Program of China(Nos.2017YFE0102200,2017YFC0908600)Zhejiang Provincial Natural Science Foundation of China(LY15H310004,LY16H280014)Natural Science Foundation of China(No.81573502)
文摘Stellera chamaejasme L.is a traditional Chinese medicine with a long history to treat stubborn skin ulcer,and it also has antiviral and antitumor effects.Neochamaejasmine B(NCB),Neochamaejasmine A(NCA)and Chamaechromone(CMC)are the major components in dried roots of Stellera chamaejasme L..Our studies suggested that NCB,NCA and CMC are inhibitors of Organic anion transporter 1(OAT1).OAT1 is encoded by solute carrier family 22 member 6 gene(SLC22 A6)in humans and plays a critical role in the organic anion drug uptake and excretion in the kidney.Lamivudine is the typical substrate of OAT1 and is frequently used in combination with other antiviral drugs in clinical antiviral treatments.The aim of this study is to investigate the interaction and its mechanism between these bi-flavone components in Stellera chamaejasme L.and lamivudine via OAT1 both in vitro and in vivo.In vitro,the uptake studies in Madin-Darby canine kidney(MDCK)cells overexpressing OAT1 suggested that NCB inhibited the uptake of 6-CFL and lamivudine.Similar results were obtained for NCA and CMC.NCB was a noncompetitive and competitive inhibitor interaction with OAT1.IC50 values of NCB,NCA and CMC for inhibiting OAT1-mediated lamivudine transport were 2.46,8.35 and 0.61μmol·L^–1,respectively.In vivo,the pharmacokinetic results of lamivudine in rats showed that the mean area under the plasma concentration-time curve(AUC0-∞)and maximal plasma concentration(Cmax)of lamivudine after co-administration is increased 2.94-fold and 1.87-fold,respectively,compared to lamivudine administration alone.The results of interactions between lamivudine and these bi-flavone components in Stellera chamaejasme L.extracts via OAT1 in vivo are consistent with studies in vitro.The inhibition of OAT1-mediated uptake of lamivudine by NCB,NCA and CMC is the possible mechanism for Stellera chamaejasme L.extracts improving the oral bioavailability of lamivudine in rats.
