Effect of Chrysin and Natural Coumarins on UGT1A1 and 1A6 Activities in Rat and Human Hepatocytes in Primary Culture

Abstract

Flavonoids and coumarins are naturally occurring compounds that are widely distributed in vegetables and have a broad pharmacological activity. Inducibility of UDP-glucuronosyltransferases (UGTs) by xenobiotics is well documented and can be considered beneficial for health. In particular, UGT1A1-dependent bilirubin conjugation plays a critical role in the detoxification of neurotoxic bilirubin and phenobarbital-mediated UGT1A1 induction therapy is commonly used in the treatment of unconjugated hyperbilirubinemic diseases such as Crigler-Najjar type II disease. In the present study, the effects of the flavone chrysin and six natural coumarins isolated from various Rutaceous plants on UGT1A6-dependent p-nitrophenol and/or UGT1A1-dependent bilirubin glucuronoconjugation activities were evaluated in cultured rat and human hepatocytes and compared to those of the prototypical UGT1A inducers β-naphthoflavone, phenobarbital and clofibric acid. After 3 days of treatment at a concentration of 25 μM, the pyranocoumarins avicennin and cis-avicennol, and the furocoumarins bergapten and imperatorin, increased by 2-fold UGT1A1-dependent activity, equivalent to the increases obtained with chrysin at 25 μM, whereas in the presence of the simple coumarins such as coumarin or umbelliferone, UGT1A1-dependent activity was not modified. In terms of structural requirements for UGT1A1 induction, the present study suggests that the B-ring (phenyl) for chrysin and the furan or pyran rings for coumarins are essential for the biological activity.

Abbreviations

BCA:bicinchoninic acid

Bili:biliribin

BSA:bovine serum albumin

CLO:clofibric acid

FCS:foetal calf serum

HBSS:Hank’s buffered salt solution

HEPES:4-(2-hydroxyethyl)piperazine-1-ethane sulfonic acid

NF:β-naphthoflavone

PB:Phenobarbital

PBS:phosphate buffered saline

PNP:p-nitrophenol

UDPGA:Uridine 5′-diphosphoglucuronic acid

UGTs:UD-glucuronosyltransferases

Coumarins - Rutaceae - chrysin - avicennin - cis-avicennol - imperatorin - bergapten - umbelliferone - UGT1A1 - UGT1A6 - hepatocytes

References

• 1 Tukey R H. Human UDP-glucuronosylltransferases: metabolism, expression and disease.  Annu Rev Pharmacol Toxicol. 2000;  40 581-616.
• 2 Tephly T R, Burchell B. UDP-glucuronosyltransferases: a family of detoxifying enzymes.  Trends Pharmacol Sci. 1990;  11 276-9.
• 3 Embola C W, Sohn O S, Fiala E S, Weisburger J H. Induction of UDP-glucuronosyltransferase 1 (UDP-GT1) gene complex by green tea in male F344 rats.  Food Chem Toxicol. 2002;  40 841-4.
• 4 Van der Logt E MJ, Roelofs H MJ, Nagengast F M, Peters W HM. Induction of rat hepatic and intestinal UDP-glucuronyltransferases by naturally occuring dietary anticarcinogens.  Carcinogenesis. 2003;  24 1651-6.
• 5 Walle U K, Walle T. Induction of human UDP-glucuronosyltransferase UGT1A1 by flavonoids - structural requirements.  Drug Metab Dispos. 2002;  30 564-9.
• 6 Moon Y J, Wang X, Morris M E. Dietary flavonoids: effect on xenobiotic and carcinogen metabolism.  Toxicol In Vitro. 2006;  20 187-210.
• 7 Rosselli S, Maggio A, Bellone G, Formisano C, Basile A, Cicala C. et al . Antibacterial and anticoagulant activities of coumarins isolated from the flowers of Magydaris tomentosa .  Planta Med. 2007;  73 103-9.
• 8 Hoult J RS, Paya M. Pharmacological and biochemical actions of simple coumarins: Natural products with therapeutic potentiel.  Gen Pharmacol. 1996;  27 713-22.
• 9 Roux D, Muyard F, Girard C, Colombain M, Tillequin F, Waterman P G. et al . Phebaclavin I, a novel 3-prenylated coumarin, and trichoclin acetate, a new natural furocoumarin, from the aerial parts of Phebalium aff. tuberculosum (Rutaceae).  Nat Prod Res. 2006;  20 279-83.
• 10 Girard C, Roux D, Muyard F, Colombain M, Bévalot F, Tillequin F. et al . Three new 3-substituted coumarins from the twigs of Rhadinothamnus rudis ssp. amblycarpus (Phebalium rude ssp. amblycarpum) (Rutaceae).  Z Naturforsch [B]. 2005;  60 561-4.
• 11 Girard C, Muyard F, Colombain M, Tillequin F, Waterman P G, Bévalot F. Coumarins from Phebalium aff. brachycalyx .  Biochem Syst Ecol. 2005;  33 1195-201.
• 12 Chlouchi A, Girard C, Bévalot F, Tillequin F, Waterman P G, Muyard F. Coumarins and furoquinoline alkaloids from Philotheca deserti var. deserti (Rutaceae).  Biochem Syst Ecol. 2006;  34 71-4.
• 13 Carmichael J, DeGraff W G, Gazdar A F, Minna J D, Mitchell J B. Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of chemosensitivity testing.  Cancer Res. 1987;  47 936-42.
• 14 Binda D, Lasserre-Bigot D, Bonet A, Thomassin M, Come M P, Guinchard C. et al . Time course of cytochromes P450 decline during rat hepatocyte isolation and culture: effect of L-NAME.  Toxicol In Vitro. 2003;  17 59-67.
• 15 David P, Viollon C, Alexandre E, Azimzadeh A, Nicod L, Wolf P. et al . Metabolic capacities in cultured human hepatocytes obtained by a new isolating procedure from non-wedge small liver biopsies.  Hum Exp Toxicol. 1998;  17 544-53.
• 16 Chesné C, Guyomard C, Fautrel A, Poulain M G, Frémond B, De Jong H. et al . Viability and function in primary culture of adult hepatocytes from various animal species and human beings after cryopreservation.  Hepatology. 1993;  18 406-14.
• 17 Richert L, Binda D, Hamilton G, Viollon-Abadie C, Alexandre E, Bigot-Lasserre D. et al . Evaluation of the effect of culture configuration on morphology, survival time, antioxidant status and metabolic capacities of cultured rat hepatocytes.  Toxicol In Vitro. 2002;  16 89-99.
• 18 Viollon-Abadie C, Lasserre D, Debruyne E, Nicod L, Carmichael N, Richert L. Phenobarbital, β-naphtoflavone, clofibrate, and pregnenolone-16α-carbonitrile do not affect hepatic thyroid hormone UDP-glucuronosyltransferase activity, and thyroid gland function in mice.  Toxicol Appl Pharmacol. 1999;  155 1-12.
• 19 Heirwegh K PM, Van Der Vijver M, Fevery J. Assay and properties of digitonin-activated bilirubin uridine diphosphate glucuronosyltransferase from rat liver.  Biochem J. 1972;  144 57-61.
• 20 Smith P K, Krohn R I, Hermanson G T, Malia A K, Gartner F H, Provenzano M I. et al . Measurement of protein using bicinchoninic acid.  Anal Biochem. 1985;  150 76-85.
• 21 Viollon-Abadie C, Bigot-Lasserre D, Nicod L, Carmichael N, Richert L. Effects of model inducers on thyroxine UDP-glucuronosyltransferase activity in vitro rat and mouse hepatocyte cultures.  Toxicol In Vitro. 2000;  14 504-12.
• 22 Galijatovic A, Otake Y, Walle U K, Walle T. Induction of UDP-glucuronyltransferase by the flavonoids chrysin and quercetin in Caco-2 cells-potentiel role in carcinogen bioinactivation.  Pharm Res. 2001;  18 374-9.
• 23 Walle U K, Walle T. Induction of human UDP-glucuronosyltransferase UGT1A1 by flavonoids - structural requirements.  Drug Metab Dispos. 2002;  30 564-9.
• 24 Smith C M, Graham R A, Krol W L, Silver I S, Negishi M, Wang H. et al . Differential UGT1A1 induction by chrysin in primary human hepatocytes and HepG2 cells.  J Pharmacol Exp Ther. 2005;  315 1256-64.
• 25 Richert L, Lamboley C, Viollon-Abadie C, Grass P, Hartmann N, Laurent S. et al . Effects of clofibric acid on mRNA expression profiles in primary cultures of rat, mouse and human hepatocytes.  Toxicol Appl Pharmacol. 2003;  191 130-46.
• 26 Smith C M, Faucette S R, Wang H, Lecluyse E L. Modulation of UDP-glucuronosyltransferase 1A1 in primary human hepatocytes by prototypical inducers.  J Biochem Mol Toxicol. 2005;  19 96-108.
• 27 Soars M G, Petullo D M, Eckstein J A, Kasper S C, Wrighton S A. An assessment of UDP-glucuronosyltransferase induction using primary human hepatocytes.  Drug Metab Dispos. 2004;  32 140-8.
• 28 Ritter J K, Kessler F K, Thompson M T, Grove A D, Auyeung D J, Fisher R A. Expression and inducibility in the human bilirubin UGT-Glucuronosyltransferase UGT1A1 in liver and cultured primary hepatocytes: evidence for both genetic and environmental influences.  Hepatology. 1999;  30 476-84.