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Biotransformation of plant secondary metabolites and their modulatory effects on drug-metabolizing enzymes
Šadibolová, Michaela ; Boušová, Iva (advisor) ; Soukup, Ondřej (referee) ; Anzenbacherová, Eva (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Training Workplace Department of Biochemical Sciences Doctoral Degree Program Xenobiochemistry and Pathobiochemistry Candidate Mgr. Michaela Šadibolová Supervisor Assoc. Prof. PharmDr. Iva Boušová, Ph.D. Advisor Title of Doctoral Thesis Biotransformation of plant secondary metabolites and their modulatory effects on drug-metabolizing enzymes Sesquiterpenes and prenylflavonoids represent two classes of plant secondary metabolites that form an inherent part of human diet. They exert many beneficial biological activities, and accordingly, they are used as active constituents of herbal products. However, as xenobiotics, they also interact with a battery of drug-metabolizing enzymes and can in turn modulate their activity and/or expression, which may lead to herb-drug interactions. Sesquiterpenes and prenylflavonoids mentioned in this dissertation have been under-researched in this aspect. Three acyclic sesquiterpenes (farnesol, cis-nerolidol, trans-nerolidol) and three cyclic sesquiterpenes (α-humulene, β-caryophyllene, β-caryophyllene oxide) activated gene transcription mediated by pregnane X receptor; however, they did not alter the mRNA or protein expression of their downstream targets, namely cytochrome P450 (CYP) 3A4 and CYP2C, neither did...
Biotransformation of plant secondary metabolites and their modulatory effects on drug-metabolizing enzymes
Šadibolová, Michaela ; Boušová, Iva (advisor) ; Soukup, Ondřej (referee) ; Anzenbacherová, Eva (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Training Workplace Department of Biochemical Sciences Doctoral Degree Program Xenobiochemistry and Pathobiochemistry Candidate Mgr. Michaela Šadibolová Supervisor Assoc. Prof. PharmDr. Iva Boušová, Ph.D. Advisor Title of Doctoral Thesis Biotransformation of plant secondary metabolites and their modulatory effects on drug-metabolizing enzymes Sesquiterpenes and prenylflavonoids represent two classes of plant secondary metabolites that form an inherent part of human diet. They exert many beneficial biological activities, and accordingly, they are used as active constituents of herbal products. However, as xenobiotics, they also interact with a battery of drug-metabolizing enzymes and can in turn modulate their activity and/or expression, which may lead to herb-drug interactions. Sesquiterpenes and prenylflavonoids mentioned in this dissertation have been under-researched in this aspect. Three acyclic sesquiterpenes (farnesol, cis-nerolidol, trans-nerolidol) and three cyclic sesquiterpenes (α-humulene, β-caryophyllene, β-caryophyllene oxide) activated gene transcription mediated by pregnane X receptor; however, they did not alter the mRNA or protein expression of their downstream targets, namely cytochrome P450 (CYP) 3A4 and CYP2C, neither did...
Expression and activities of antioxidant enzymes in the in vivo models of non-alcoholic fatty liver disease
Sochová, Gabriela ; Boušová, Iva (advisor) ; Szotáková, Barbora (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Gabriela Sochová Supervisor: doc. PharmDr. Iva Boušová, Ph.D. Advisor: Mgr. Gabriela Svobodová Title of diploma thesis: Expression and activities of antioxidant enzymes in the in vivo models of non-alcoholic fatty liver disease Non-alcoholic fatty liver disease is one of the most common liver diseases. This disease is caused by an imbalance between fat intake and fat degradation. Lipid accumulation can be caused by several factors (genetic predisposition, diet, metabolic diseases, etc.). If these factors act for a long enough period, the disease can lead to liver fibrosis or even cirrhosis. The aim of this diploma thesis was to compare the expression and activity of the antioxidant enzymes glutathione-S-transferase (GST), glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD), and catalase in two models of NAFLD (dietary model (K) - high fat, fructose, and cholesterol diet (FFC); chemical model - induced by monosodium glutamate (MSG)). Furthermore, the effect of LiPR31 and liraglutide on the expression and activity of antioxidant enzymes were investigated in these models. Expression and activity were determined in 20,000×g supernatant of mouse liver. RT-qPCR was used...
The efflux transport activity of P-glycoprotein - 9.2 fromHaemonchus contortus
Kohoutová, Kateřina ; Matoušková, Petra (advisor) ; Boušová, Iva (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Kateřina Kohoutová Supervisor: doc. Ing. Petra Matoušková, Ph.D. Title of diploma thesis: The efflux transport activity of P-glycoprotein-9.2 from Haemonchus contortus Haemonchosis is a globally widespread disease affecting small ruminants. The causative agent of the disease is the gastrointestinal nematode Haemonchus contortus, which feeds of the blood of its host. The treatment of haemonchosis with anthelmintics is complicated by the worldwide spread of resistance. One possible cause of the development of anthelmintic resistance is a decrease in anthelmintic concentration in the worm due to increased expression of P-glycoproteins from ABC transporter superfamily. The study of the transport activity of these transporters could help to understand the mechanism of resistance development and make the treatment of haemonchosis more effective. The aim of this thesis was to characterize the transport activity of P-glycoprotein-9.2 and its interaction with a selected inhibitor using a heterologous expression system. The adherent cell line HEK293 was transfected with a plasmid carrying the Pgp-9.2 gene of the Haemonchus contortus. Successful expression was verified by dot blot. Unfortunately,...
Interactions of antiretroviral drugs with membrane transporters
Martinec, Ondřej ; Červený, Lukáš (advisor) ; Boušová, Iva (referee) ; Chládek, Jaroslav (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate: Mgr. Ondřej Martinec Supervisor: Assoc. Prof. PharmDr. Lukáš Červený, Ph.D. Title of doctoral thesis: Interactions of antiretroviral drugs with membrane transporters Oral delivery is the most common, convenient, and economical form of drug administration. Absorption of orally administered drugs occurs mainly in the intestine. Intestinal absorption can be reduced by the activity of efflux drug ABC transporters, mainly p-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2), located in the apical membrane of the intestinal epithelium. HIV-infected patients are dependent on lifelong pharmacotherapy, which includes a combination of three or more antiretroviral drugs. Hepatitis C (HCV) is a common co-infection of HIV. In addition, the HIV-positive population is aging, which is associated with burden of other comorbidities. This results in an indication of polypharmacy and thus an increased risk of drug-drug interactions. Many antiretroviral drugs used are substrates, inhibitors and /or inducers of ABCB1, so they might quantitatively affect the intestinal absorption of co-administered drugs (ABCB1 substrates), thereby affecting the efficacy/safety of treatment. As part of this...
Metabolism and transport of 6-prenylnaringenin in vitro
Horní, Martin ; Boušová, Iva (advisor) ; Raisová Stuchlíková, Lucie (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Martin Horni Supervisor: doc. PharmDr. Boušová Iva, Ph.D. Consultant: Mgr. Michaela Šadibolová Title of diploma thesis: Metabolism and transport of 6-prenylnaringenin in vitro Prenylflavonoids constitute a group of plant secondary metabolites with various positive effects on human health. Hops (Humulus lupulus L.) represent a rich source of prenylflavonoids including 6-prenylnaringenin (6-PN). Recently, 6-PN has been found to possess various biological effects including analgesic, antiproliferative, and estrogen-modulating activity. Despite its potential to be used as a nutraceutical, the pharmacokinetic properties of 6-PN are still unknown. This thesis had therefore two objectives. The first objective was to determine the hepatic and intestinal metabolism of 6-PN in humans as well as its hepatic metabolism in rats and mice in vitro using precision-cut tissue slices (PCTS). Samples were analyzed using UHPLC-MS/MS. 6-PN was efficiently metabolized predominantly into phase II metabolites with four glucuronides and four sulfates identified in the studied PCTS. For the first time, a novel metabolite of 6-PN formed as a result of concurrent glucuronidation and sulfation has been identified, which...
Antiproliferative and anthelmintic effects of fern extracts
Turoňová, Petra ; Matoušková, Petra (advisor) ; Boušová, Iva (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Bichemical Sciences Candidate: Petra Turoňová Supervisor: doc. Ing. Petra Matoušková, PhD. Title of diploma thesis: Antiproliferative and anthelmintic effects of fern extracts Ferns belong to spore vascular plants. In this study, an antiproliferative and anthelmintic effects of selected fern extracts were investigated. The antiproliferative effect of the extracts was tested on the SW480 cell line. This cell line was originally obtained from primary adenocarcinoma of the colon. The anthelmintic effect was tested on L3 larvae Haemonchus contortus, the most common internal parasite of sheep and goats. Firstly, to determine the antiproliferative effect of the extracts, an initial screening was performed. Cytotoxicity was evaluated by measuring absorbance on a Tecan spectrophotometer. The four fern species that showed most promising antiproliferative activity were tested at three time intervals and four concetrations. H. contortus larvae L3 were exposed to fern extracts for seven days and the percentage of developed L4 larvae was detected by microscope. Athyrium distentifolium, Dryopteris aemula, Davallia canariensis and Polystichum aeculatum showed the highest antiproliferative and anthelmintic activity. In addition, Dryopteris...
The efflux transport activity of P-glycoprotein - 9.2 fromHaemonchus contortus
Kohoutová, Kateřina ; Matoušková, Petra (advisor) ; Boušová, Iva (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Kateřina Kohoutová Supervisor: doc. Ing. Petra Matoušková, Ph.D. Title of diploma thesis: The efflux transport activity of P-glycoprotein-9.2 from Haemonchus contortus Haemonchosis is a globally widespread disease affecting small ruminants. The causative agent of the disease is the gastrointestinal nematode Haemonchus contortus, which feeds of the blood of its host. The treatment of haemonchosis with anthelmintics is complicated by the worldwide spread of resistance. One possible cause of the development of anthelmintic resistance is a decrease in anthelmintic concentration in the worm due to increased expression of P-glycoproteins from ABC transporter superfamily. The study of the transport activity of these transporters could help to understand the mechanism of resistance development and make the treatment of haemonchosis more effective. The aim of this thesis was to characterize the transport activity of P-glycoprotein-9.2 and its interaction with a selected inhibitor using a heterologous expression system. The adherent cell line HEK293 was transfected with a plasmid carrying the Pgp-9.2 gene of the Haemonchus contortus. Successful expression was verified by dot blot. Unfortunately,...
Expression and activities of antioxidant enzymes in the in vivo models of non-alcoholic fatty liver disease
Sochová, Gabriela ; Boušová, Iva (advisor) ; Szotáková, Barbora (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Gabriela Sochová Supervisor: doc. PharmDr. Iva Boušová, Ph.D. Advisor: Mgr. Gabriela Svobodová Title of diploma thesis: Expression and activities of antioxidant enzymes in the in vivo models of non-alcoholic fatty liver disease Non-alcoholic fatty liver disease is one of the most common liver diseases. This disease is caused by an imbalance between fat intake and fat degradation. Lipid accumulation can be caused by several factors (genetic predisposition, diet, metabolic diseases, etc.). If these factors act for a long enough period, the disease can lead to liver fibrosis or even cirrhosis. The aim of this diploma thesis was to compare the expression and activity of the antioxidant enzymes glutathione-S-transferase (GST), glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD), and catalase in two models of NAFLD (dietary model (K) - high fat, fructose, and cholesterol diet (FFC); chemical model - induced by monosodium glutamate (MSG)). Furthermore, the effect of LiPR31 and liraglutide on the expression and activity of antioxidant enzymes were investigated in these models. Expression and activity were determined in 20,000×g supernatant of mouse liver. RT-qPCR was used...
Effect of xanthohumol and 6-prenylnaringenin in the in vitro model of non-alcoholic fatty liver disease
Velecká, Eva ; Boušová, Iva (advisor) ; Zárybnický, Tomáš (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Eva Velecká Supervisor: doc. PharmDr. Iva Boušová, Ph.D. Consultant: Mgr. Michaela Šadibolová Title of diploma thesis: Effect of xanthohumol and 6-prenylnaringenin in the in vitro model of non-alcoholic fatty liver disease Non-alcoholic fatty liver disease (NAFLD) is a chronic disease characterized by the accumulation of triglycerides (TAG) in hepatocytes encompassing a wide spectrum of pathological states ranging from simple steatosis to non-alcoholic steatohepatitis (NASH) and eventually progressing to cirrhosis and hepatocellular carcinoma. Xanthohumol (XH), a prenylflavonoid found in common hop (Humulus lupulus), is currently being studied as a potential drug for NAFLD therapy due to its described anti- steatotic, anti-inflammatory, anti-fibrotic, hypolipidemic and antioxidant effects. The aim of this thesis was to study the potential hepatoprotective effect of XH and 6-prenylnaringenin (6PN), a minor prenylflavonoid also found in common hop. For this purpose, a murine in vitro NAFLD model was used in which steatosis was induced by a lipogenic medium containing palmitic and oleic acid, fructose, citrate, and insulin in precision-cut liver slices (PCLS). Steatotic and non-steatotic PCLS...

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