Bilirubin UDP-glucuronyltransferase Research Articles

Bilirubin, the Gold Within

When heme-containing proteins such as hemoglobin and cytochrome P450 are degraded, their prosthetic groups cannot be metabolized and have to be excreted. Elimination of heme could be considered straightforward detoxification, but a large body of evidence shows that this pathway plays a role in various physiological processes as well. Article see p 2556 The first step in the catabolism of heme is cleavage of the porphyrin ring by heme oxygenase (HO); this reaction yields biliverdin, carbon monoxide, and iron. Two HO isoforms mediate this process, constitutively expressed HO-2 and inducible HO-1. Many cell types express HO, with high expression present in cells of the reticuloendothelial system involved in the degradation of erythrocytes.1 The first step in heme catabolism has attracted considerable interest because HO activity is involved in the regulation of processes such as arthrosclerosis, inflammation, and diabetes mellitus.1 This regulation is mediated by the products of HO action on heme, with the potent signaling molecule carbon monoxide likely playing a major role.1 The other product of HO action on heme is the green and water-soluble compound biliverdin. In mammals, levels of biliverdin are low because it is quickly converted to the hydrophobic yellow compound bilirubin by ubiquitously expressed biliverdin reductase. However, other animal species, such as fish,2 do not completely metabolize all biliverdin to bilirubin and excrete significant amounts of the green pigment. Because bilirubin is poorly water soluble, mammals require efficient conjugation with glucuronic acid by UDP glucuronyltransferase before bilirubin glucuronides can be excreted into bile. This aspect of heme metabolism has always puzzled scientists. Why is bilirubin the sole end product of heme degradation in mammals? Bilirubin is toxic, and the absence of bilirubin UDP glucuronyltransferase (UGT1A1) in patients with Crigler-Najjar syndrome causes high unconjugated bilirubin levels and subsequent severe brain damage. In …

895. Ex Vivo Gene Therapy of Metabolic Liver Disorders Using Uncultured and Lentivirally Transduced Hepatocytes

Background: At the present time, orthotopic liver transplantation is the only effective therapy for several congenital and acquired hepatic disorders. Because of a chronic organ shortage, and of the necessity of heavy and lifelong immunosuppression after transplantation, gene therapy represents a promising alternative. We have previously described an ex vivo gene therapy protocol that allows very high rates of transduction of human hepatocytes (up to 98%), in which freshly isolated hepatocytes are transduced in suspension and immediately transplanted. This protocol is easily performed, rapid, does not required primary culture of hepatocytes, and preserves full cell engraftment potential and in vivo cell functionality. In this study, we used the Gunn rat, which is the model for Crigler-Najjar syndrome type I, a defect in bilirubin UDP-glucuronyltransferase (BGT), to evaluate our approach.

170. Ex Vivo Gene Therapy of Metabolic Liver Disorders Using Uncultured and Lentivirally Transduced Hepatocytes

Top of pageAbstract At the present time, orthotopic liver transplantation is the only effective therapy for several congenital and acquired hepatic disorders. Because of a chronic organ shortage, and of the necessity of heavy and lifelong immunosuppression after transplantation, gene therapy represents a promising alternative. Ex vivo liver gene therapy approach carries several advantages: (i) it allows for an elective targeting of the hepatocytes thereby avoiding, without the need for highly tissue-specific expression systems, that the transgene be expressed in professional antigen-presenting cells, (ii) it avoids most of the potential biohazards linked to the in vivo approach such as vector dissemination and integration in inappropriate target cells. We have previously described a new liver-directed ex vivo gene therapy protocol that allows very high rates of transduction of human hepatocytes (up to 98%), in which freshly isolated hepatocytes are transduced while kept in suspension and immediately transplanted. This protocol is easily performed, rapid (<5 hours) and preserves full cell engraftment potential and in vivo cell functionality. Two prototypic metabolic liver diseases were used as models to evaluate our approach: Hemophilia B, a defect in blood coagulation Factor IX and Crigler-Najjar syndrome type I (CNI), a defect in bilirubin UDPglucuronyltransferase (BGT1). Stable and persistent expression of human Factor IX (hFIX), up to 200 days and 10% of physiologic levels, was observed in C57BL/6 receiving hepatocytes that have been transduced with vectors coding for hFIX under control of the liver-specific transthyretin promoter. Therefore, these results prompt further studies in mice and dog model of hemophilia B. To correct BGT1 deficiency in Gunn rat, which is the model for CNI, we constructed vectors coding for human BGT1 under control of the E1-alpha ubiquitous promoter. In rats receiving 6 millions transduced hepatocytes, serum bilirubin levels decreased by 20–30%, a decline equivalent to that found in previously reported studies using normal hepatocytes, and was sustained for up to 150 days. In contrast, in rats receiving untransduced hepatocytes, serum didn't decline but rather increased. Noteworthy, these results also suggested an absence of immune response against the transduced hepatocytes, as expected by selective targeting of the hepatocytes using an ex vivo approach, even though transgene expression was driven by a ubiquitous promoter. Recent clinical trials have shown that freshly isolated and uncultured allogeneic hepatocyte transplantation has resulted in therapeutic benefits in patients with metabolic liver disorders. Altogether, these data open new perspectives for the treatment of metabolic liver disorders using an ex vivo approach.

Long-term correction of bilirubin UDPglucuronyltransferase deficiency in rats by in utero lentiviral gene transfer

Bilirubin is glucuronidated by bilirubin UDP-glucuronyltransferase (UGT1A1) before biliary excretion. Because bilirubin is toxic, patients with Crigler–Najjar type I (CN), who have no UGT1A1 activity, suffer severe brain damage early in childhood. The Gunn rat is the model for CN type 1. Gunn rat fetuses were injected with 107 transducing units of UGT1A1 lentiviral vector at the end of the third trimester on embryonic day 19. Serum bilirubin of injected Gunn rats was lowered by 45% compared to untreated controls. This decrease was highly significant (P < 106) and was sustained for more than a year. In treated Gunn rats, bilirubin glucuronides were present in bile and UGT1A1 protein was detected in tissue. Liver, intestine, stomach, pancreas, and other organs were transduced and mostly contained 1% or less vector copies per genome. Tissue distribution was variable among experimental animals but high transduction levels were seen in pancreas and intestine in most animals. Immunohistochemistry of these organs revealed transduction of pancreatic acinar cells and intestinal epithelium. Injection of a lentiviral UGT1A1 vector into third-trimester Gunn rat fetuses corrects the metabolic deficiency and mediates a reduction of serum bilirubin levels that would be therapeutic in humans.

1183. Long Term Correction of Bilirubin UDP Glucuronyltransferase Deficiency in Rats by In Utero Lentiviral Gene Transfer

1183. Long Term Correction of Bilirubin UDP Glucuronyltransferase Deficiency in Rats by In Utero Lentiviral Gene Transfer

790 Acetaminophen overdose: relationship of glucuronyltransferase promoter variants to subsequent acute liver failure

Induction of hepatic microsomal bilirubin UDP_glucuronyltransferase and CYP4A1 P450 by 3,4,3′,4′-tetrachlorobiphenyl (TCB) and 3,4,5,3′,4′-pentachlorobiphenyl (PenCB) was studied in guinea pigs and rats. Both enzyme activities in guinea pigs were increased 4-fold or more over the control by PenCB treatment, while by treatment with a less toxic congener, TCB, these were increased to a lesser extent. On the contrary, the rat enzymes were significantly decreased by treatment with these co-planar polychlorinated biphenyls. These results suggest that the alterations of both bilirubin UDP-glucuronyltransferase and CYP4A P450 are the characteristic indices for the toxicites of polychlorinated hydrocarbons.

151 Long term correction of bilirubin UDP glucuronyltransferase deficiency in rats by in utero lentiviral gene transfer

151 Long term correction of bilirubin UDP glucuronyltransferase deficiency in rats by in utero lentiviral gene transfer

Gilbert's syndrome is not associated with HELLP syndrome.

The HELLP syndrome has been associated with postpartum unconjugated hyperbilirubinaemia. Several types of disorders cause unconjugated hyperbilirubinaemia, Gilbert's syndrome being the most common. In Caucasians a genetic defect in the TATA box of the promotor region of the gene encoding for bilirubin UDP-glucuronyltransferase is tightly associated with Gilbert's syndrome. This defect was assessed by polymerase chain reaction in 237 women with the HELLP syndrome in their obstetric history and 236 controls. Fifteen percent of the cases and 10% of the controls had a homozygous genetic defect (chi2 = 2.9; P = 0.23). No evidence was found that Gilbert's syndrome is associated with the HELLP syndrome.

Gilbert's syndrome is not associated with HELLP syndrome

The HELLP syndrome has been associated with postpartum unconjugated hyperbilirubinaemia. Several types of disorders cause unconjugated hyperbilirubinaemia, Gilbert's syndrome being the most common. In Caucasians a genetic defect in the TATA box of the promotor region of the gene encoding for bilirubin UDP-glucuronyltransferase is tightly associated with Gilbert's syndrome. This defect was assessed by polymerase chain reaction in 237 women with the HELLP syndrome in their obstetric history and 236 controls. Fifteen percent of the cases and 10% of the controls had a homozygous genetic defect (χ 2 = 2.9; P = 0.23). No evidence was found that Gilbert's syndrome is associated with the HELLP syndrome.

Association of human liver bilirubin UDP-glucuronyltransferase activity with a polymorphism in the promoter region of the UGT1A1 gene

Background/Aims: Gilbert's syndrome is a benign form of a deficiency in bilirubin glucuronidation. It is associated with a homozygous polymorphism, A(TA) 7TAA instead of A(TA) 6TAA, in the TATA-box of the promoter region of the bilirubin UDP-glucuronyltransferase gene. In this study the correlation between this promoter region polymorphism and in vitro human liver bilirubin UDP-glucuronyltransferase enzyme activity was investigated. Methods: Liver samples from organ transplant donors ( n=39) and two known Gilbert's syndrome patients were used for measuring bilirubin UDP-glucuronyltransferase enzyme activity and for isolation of DNA followed by detection of the promoter region polymorphism by polymerase chain reaction. Genotypes were assigned as follows; 6/6: homozygous for the A(TA) 6TAA-allele, 7/7: homozygous for the A(TA) 7TAA-allele, and 6/7: heterozygous with one of each alleles. Results: Seventeen out of 39 subjects (44%) had the homozygous 6/6 genotype, 18 subjects (46%) had the heterozygous 6/7 genotype, whereas four individuals (10%) and the two individuals with Gilbert's syndrome had the 7/7 genotype correlated with Gilber's syndrome. This resulted in an allele frequency of 0.33 for the A(TA) 7TAA-allele. The median bilirubin UDP-glucuronyltransferase enzyme activity of the 17 subjects with the 6/6 genotype (1565 nmol/g liver/h) was significantly higher than the activity of the 18 subjects with the 6/7 genotype (985 nmol/g liver/h; p<0.05) and the six individuals with the 7/7 genotype (749 nmol/g liver/h; p<0.005). No significant differences in enzyme activity were found between the 6/7 and the 7/7 genotype groups. Conclusions: The results indicate a close association between the promoter region genotype and the expression of hepatic bilirubin UDP-glucuronyltransferase enzyme activity. Subjects who have a 7/7 genotype have the lowest enzyme activity, whereas subjects with the heterozygous 6/7 genotype have an intermediate enzyme activity.

Dose-related increase in liver heme catabolism during rabbit aflatoxicosis

Aflatoxin B1 (AFB1) has been reported to decrease microsomal hepatic cytochrome P450 (P450) content and increase both total plasma bilirubin concentration and liver heme oxygenase activity. The purposes of this study were to determine whether liver hemoproteins contents and heme catabolizing enzymes were affected by the mycotoxin and whether these alterations were linked to hyperbilirubinemia. Male New Zealand rabbits were divided into three groups of five animals, each receiving for 5 days either arabic gum as vehicle or AFB1 at a daily oral dose of 0.05 or 0.10 mg/kg. These treatments affected neither cytochrome b5 content nor NADPH-cytochrome reductase activity. A linear dose-dependent decrease in cytochrome P450 content and increases in both heme oxygenase and biliverdin reductase activities were observed. Bilirubin UDP-glucuronyltransferase activity was dramatically decreased at both doses, whereas cholestasis occurred only at 0.10 mg/kg. An exponential dose-dependent increase in plasma bilirubin concentration was also observed. Both the simultaneous exponential increase in bilirubinemia associated to a reduced bilirubin UDP-glucuronyltransferase activity and the absence of cholestasis at 0.05 mg/kg, suggested that the hyperbilirubinemia is more probably related to an increased heme catabolism than to an altered bile duct permeability.

Induction of bilirubin UDP-glucuronyltransferase and CYP4A1 P450 by co-planar PCBs: Different responsiveness of guinea pigs and rats

Induction of hepatic microsomal bilirubin UDP_glucuronyltransferase and CYP4A1 P450 by 3,4,3′,4′-tetrachlorobiphenyl (TCB) and 3,4,5,3′,4′-pentachlorobiphenyl (PenCB) was studied in guinea pigs and rats. Both enzyme activities in guinea pigs were increased 4-fold or more over the control by PenCB treatment, while by treatment with a less toxic congener, TCB, these were increased to a lesser extent. On the contrary, the rat enzymes were significantly decreased by treatment with these co-planar polychlorinated biphenyls. These results suggest that the alterations of both bilirubin UDP-glucuronyltransferase and CYP4A P450 are the characteristic indices for the toxicites of polychlorinated hydrocarbons.

Glucuronidation of thyroid hormone in rat liver: effects of in vivo treatment with microsomal enzyme inducers and in vitro assay conditions

We investigated the effects of in vivo treatment with different microsomal enzyme inducers, including clofibrate (CLOF), hexachlorobenzene (HCB), 3-methylcholanthrene (MC), 3,3',4,4'-tetrachlorobiphenyl (TCB), and 2,3,7,8-tetrachloro-p-dioxin, as well as of in vitro addition of the detergent Brij 56 on the glucuronidation of T4, T3, and rT3 by UDP-glucuronyltransferase (UGT) activities of rat liver microsomes. The results were compared with measurements of UGT activities for bilirubin, p-nitrophenol (PNP), and androsterone. In general, glucuronidation rates were 5-fold or more higher with rT3 than with T4 or T3 as substrate. In liver microsomes from untreated rats, T4 UGT activity was stimulated by Brij 56 to a maximum of about 2-fold at 0.025% detergent. Treatment of Wistar rats for 4 days with CLOF (200 mg/kg BW.day) resulted in significant increases in UGT activities for T4 (to 154%), rT3 (to 155%), and bilirubin (to 194%), in particular if assayed in the presence of 0.025% Brij 56, but had little effect on the UGT activities for T3, PNP, and androsterone. The CLOF-induced increases in T4 and rT3 UGT activities were not observed in Gunn rats, which have a complete lack of bilirubin UGT activity and greatly impaired PNP UGT activity. Treatment of Wistar rats with a single injection of MC (50 mg/kg BW), TCB (50 mg/kg BW), or 2,3,7,8-tetrachloro-p-dioxin (6.25 micrograms/kg BW) resulted, after 4 days, in 6.3- to 7.3-fold increases in T4 UGT activity and 15.1- to 16.7-fold increases in rT3 UGT activity if determined in the absence of Brij 56, whereas T4 UGT activity was only increased by 33-68% when assayed in the presence of Brij 56. T3 glucuronidation was not affected (with Brij 56) or was increased by only 33-68% (without Brij 56) after treatment with these MC-type inducers. PNP UGT activity was induced 3.6- to 4.3-fold, whereas bilirubin and androsterone UGT activities were changed little by these treatments. Similar findings regarding T4, rT3, PNP, and bilirubin UGT activities were obtained after chronic treatment of WAG rats with HCB, another MC-type inducer. However, WAG rats lack androsterone UGT and show low T3 UGT activity, which was increased about 2.3-fold by HCB treatment. On the basis of these and previous findings it is concluded that at least three UGT isoenzymes are involved in the glucuronidation of thyroid hormone.

Multiple UDP-glucuronyltransferases for the glucuronidation of thyroid hormone with preference for 3,3',5'-triiodothyronine (reverse T 3)

We have studied the glucuronidation of the thyroid hormones T 4, T 3 and rT 3 by liver microsomes of Wistar, Gunn and WAG rats. Gunn rats have a defect in the gene coding for bilirubin and phenol UDP-glucuronyltransferase (UGT) isoenzymes; WAG rats have a genetic defect in androsterone UGT. In normal Wistar rats UGT activity was ≈5-fold higher for rT 3 than for T 4 or T 3. UGT activities for T 4 and rT 3, but not for T 3, were impaired in Gunn rats. Conversely, UGT activity for T 3, but not for T 4 or rT 3, was impaired in WAG rats. Thus, in rat liver rT 3 is glucuronidated much more rapidly than T 4 and T 3. Our results support the view that T 4 and rT 3 are glucuronidated by bilirubin and phenol UGTs and T 3 by androsterone UGT.

Induction of Bilirubin UDP-Glucuronyltransferase and P-450 4A Subfamily in Guinea Pigs by Treatment with Coplanar PCB (Proceedings of the 18th Symposium on Toxicology, and Environmental Health)

Induction of Bilirubin UDP-Glucuronyltransferase and P-450 4A Subfamily in Guinea Pigs by Treatment with Coplanar PCB (Proceedings of the 18th Symposium on Toxicology, and Environmental Health)

Glucuronidation of thyroid hormone in rat liver: effects of in vivo treatment with microsomal enzyme inducers and in vitro assay conditions.

We investigated the effects of in vivo treatment with different microsomal enzyme inducers, including clofibrate (CLOF), hexachlorobenzene (HCB), 3-methylcholanthrene (MC), 3,3',4,4'-tetrachlorobiphenyl (TCB), and 2,3,7,8-tetrachloro-p-dioxin, as well as of in vitro addition of the detergent Brij 56 on the glucuronidation of T4, T3, and rT3 by UDP-glucuronyltransferase (UGT) activities of rat liver microsomes. The results were compared with measurements of UGT activities for bilirubin, p-nitrophenol (PNP), and androsterone. In general, glucuronidation rates were 5-fold or more higher with rT3 than with T4 or T3 as substrate. In liver microsomes from untreated rats, T4 UGT activity was stimulated by Brij 56 to a maximum of about 2-fold at 0.025% detergent. Treatment of Wistar rats for 4 days with CLOF (200 mg/kg BW.day) resulted in significant increases in UGT activities for T4 (to 154%), rT3 (to 155%), and bilirubin (to 194%), in particular if assayed in the presence of 0.025% Brij 56, but had little effect on the UGT activities for T3, PNP, and androsterone. The CLOF-induced increases in T4 and rT3 UGT activities were not observed in Gunn rats, which have a complete lack of bilirubin UGT activity and greatly impaired PNP UGT activity. Treatment of Wistar rats with a single injection of MC (50 mg/kg BW), TCB (50 mg/kg BW), or 2,3,7,8-tetrachloro-p-dioxin (6.25 micrograms/kg BW) resulted, after 4 days, in 6.3- to 7.3-fold increases in T4 UGT activity and 15.1- to 16.7-fold increases in rT3 UGT activity if determined in the absence of Brij 56, whereas T4 UGT activity was only increased by 33-68% when assayed in the presence of Brij 56. T3 glucuronidation was not affected (with Brij 56) or was increased by only 33-68% (without Brij 56) after treatment with these MC-type inducers. PNP UGT activity was induced 3.6- to 4.3-fold, whereas bilirubin and androsterone UGT activities were changed little by these treatments. Similar findings regarding T4, rT3, PNP, and bilirubin UGT activities were obtained after chronic treatment of WAG rats with HCB, another MC-type inducer. However, WAG rats lack androsterone UGT and show low T3 UGT activity, which was increased about 2.3-fold by HCB treatment. On the basis of these and previous findings it is concluded that at least three UGT isoenzymes are involved in the glucuronidation of thyroid hormone.(ABSTRACT TRUNCATED AT 400 WORDS)

Analysis of the interaction uridin 5'-diphosphoglucuronic acid with intestinal bilirubin udp-glucuronyltransferase

1. 1. Bilirubin UDP-glucuronyltransferase Michaelis-Menten kinetic parameters for UDP-glucuronic acid were studied using native and digitonin activated microsomes obtained from rat intestinal mucosa. 2. 2. The intestinal enzyme showed a lower apparent V max compared with the hepatic enzyme in both native and activated microsomes; digitonin pretreatment enhanced V max 4 times in the former enzyme and 2 times in the latter. 3. 3. The affinity of UDP-glucuronic acid for the intestinal enzyme was about 2 times lower than that for the hepatic enzyme and it was not substantially modified by detergent neither in the intestine nor in the liver. 4. 4. The lipid analysis of intestinal and hepatic microsomes showed that the former present a higher content of cholesterol and a lower phosphatidylcholine/sphingomyelin ratio than the latter, accordingly the estimation of membrane fluidity using the fluorescence anisotropy of l,6-diphenyl-1,3,5-hexatriene indicated that intestinal microsomes are more “rigid” than the hepatic ones. 5. 5. These characteristics would provoke a restrictive milieu surrounding the enzyme that modifies its kinetic properties thus limiting its participation in the metabolism of bilirubin.

Failure of expression of the phenobarbital-induced enhancement of UDP-glycosyltransferases in native, sealed endoplasmic reticulum vesicles from rat liver.

Conflicting data have been published regarding the effects of phenobarbital treatment on bilirubin UDP-glucuronyltransferase activity in native liver microsomes. Recent evidence suggests that the bilirubin UDP-glycosyltransferase system faces the interior of microsomal vesicles, and that expression of its activities in sealed microsomes may be rate-limited by transport of UDP sugars across the membrane. These observations raise the possibility that the reported variability in the effects of phenobarbital may reflect differences in integrity of the membrane in microsomal preparations. We examined the effect of phenobarbital on bilirubin UDP-glucosyltransferase and the UDP-glucuronyltransferase activities towards bilirubin, 4-nitrophenol, and 1-naphthol using native rat liver microsomes with verified vesicle integrity. Phenobarbital-induced microsomes in which the membrane permeability barrier was eliminated by pretreatment with detergent displayed markedly higher UDP-glycosyltransferase activities towards all tested substrates compared with activities in similarly disrupted microsomes from untreated rats. In contrast, none of the transferase activities tested were significantly enhanced by phenobarbital treatment when the enzymic activities were assayed in sealed microsomes. Addition to the enzyme assay mixture of UDPGlcNAc, a presumed physiological activator of the UDP-glucuronyltransferases, failed to expose the enhanced UDP-glucuronyltransferase concentration in phenobarbital-induced sealed microsomes. Our findings are consistent with the idea that transport of UDP sugar across the membrane may be rate-limiting for expression of UDP-glycosyltransferase activities in sealed microsomes. Quantitative assessment of membrane integrity is an essential prerequisite in experiments designed to study the regulation of the microsomal UDP-glycosyltransferase system.

Kinetic properties of bilirubin UDP-glucuronyltransferase in squirrel monkeys exhibiting fasting hyperbilirubinemia

1. 1. Bolivian squirrel monkeys (BoSM),unlike Brazilian squirrel monkeys (BrSM), exhibit a marked fasting hyperbilirubinemia (FH) and serve as animal models for Gilbert's syndrome type I. 2. 2. Compared to BrSM, BoSM possess a higher apparent udpga k m (0.51 vs 0.29 mM) and lower V m (0.36 vs 0.48 nmol BR conjugated/min per mg microsomal protein) for hepatic bilirubin (BR) UDP-glucuronyltransferase (BR UDPG-T). 3. 3. Lineweaver-Burk plots are linear and obey Michaelis-Menten kinetics when UDP-acetylglucosamine is used as activator and UDPGA substrate concentrations are within the physiologic range present in the liver during the fed and fasted state (0.10-0.71 mM); above these concentrations, there is a discontinuity of kinetic plots as noted in other species. 4. 4. There is no effect of fasting on the K m of BR conjugation (i.e. sum of mono- and diglucuronides) in either monkey; however, fasting is associated with lower V m, values (15–20%) in each subspecies. 5. 5. By calculating the potential BR flux (nmol BR conjugated/min per kg) using known hepatic UDPGA concentrations, liver weights and in vitro K m and V m, a markedly lower BR flux is observed in BoSM (58.4 nmol/min per kg) than in BrSM (91.6nmol/min per kg). 6. 6. Significantly higher apparent UDPGAK m and lower V m of BR UDPG-T for conjugation of BR to BR monoglucuronide appears responsible in part for the four- to five-fold elevations in unconjugated BR in the liver and plasma in the fasted BoSM.

Proportion of conjugated bilirubin in bile in relation to hepatic bilirubin UDP-glucuronyltransferase activity

To elucidate the diagnostic relevance of biliary conjugated bilirubin, biliary bilirubin from normal volunteers (NV), patients with Gilbert's syndrome (GS) and Crigler-Najjar syndrome type II (C-N II), and from various rat strains was fractionated. Biliary bilirubin diglucuronide (BDG) was present at lower levels, and bilirubin monoglucuronide (BMG) and unconjugated bilirubin were present at higher levels in GS and C-N II compared with NV, which is consistent with decreased hepatic bilirubin UDP-glucuronyltransferase activity (BGTA). The level of biliary BDG was higher in Wistar-Kyoto rats and lower in heterozygous (Jj) Gunn rats than in SD and Wistar rats. The hepatic BGTA level in heterozygous (Jj) Gunn rats was decreased to 60% of that in Wistar rats, in accordance with decreased biliary BDG. On the other hand, BGTA in Wistar-Kyoto rats whose biliary BDG level was high, was not different from that of Wistar and SD rats. Thus, a correlation between BGTA and biliary bilirubin fractions may not exist on some occasions.