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Recent functional studies have suggested that, in addition to simple diffusion, carrier-mediated transport may play an important role in foetal unconjugated bilirubin (UCB) uptake by the placenta. We have investigated the role of organic anion-transporting polypeptides (OATPs) in UCB transport by the placenta-maternal liver tandem. RNA was obtained from human liver (hL), human placenta (hPl) at term, and purified (> 80%) cytokeratin-7-positive mononucleated human trophoblast cells (hTCs). By analytical reverse transcription (RT)-PCR, agarose gel electrophoresis separation and sequencing, the mRNA of OATP-A ( SLC21A3 ) and OATP-8 ( SLC21A8 ) was identified in hL, hPl and hTCs, whereas that of OATP-C ( SLC21A6 ) was detectable only in hL. Real-time quantitative RT-PCR revealed that in hL the abundance of mRNA was OATP-8 > OATP-C >> OATP-A, whereas in hPl and hTCs this was OATP-8 >> OATP-A >> OATP-C. Expression levels for these OATPs were hL >> hTCs > hPl. Injection of mRNA of OATP-A, OATP-C or OATP-8 or RNA from hL, hPl or hTCs into Xenopus laevis oocytes conferred on them the ability to take up [(3)H]17 beta-D-glucuronosyl oestradiol ([(3)H]E(2)17 beta G) and [(3)H]UCB, although in the case of OATP-A mRNA, the induced uptake of [(3)H]UCB was very low. Cis -inhibition of [(3)H]E(2)17 beta G and [(3)H]UCB uptake by both unlabelled E(2)17 beta G and UCB was found in all cases. The affinity and efficiency of [(3)H]UCB transport was OATP-C > OATP-8. Kinetic parameters for [(3)H]UCB uptake induced by RNA from hTCs resembled most closely those of OATP-8. In conclusion, our results suggest that OATP-8 may play a major role in the carrier-mediated uptake of foetal UCB by the placental trophoblast, whereas both OATP-8 and OATP-C may substantially contribute to UCB uptake by adult hepatocytes.

Unconjugated bilirubin (UCB) is currently believed to cross the placenta only by passive diffusion. To assess whether carrier-mediated transport might be involved, the uptake of [(3)H]-UCB by basal (bTPM) and apical (aTPM) plasma membrane vesicles from human placental trophoblast at term was investigated. In both types of vesicles, the uptake of [(3)H]-UCB into an osmotically sensitive space was temperature-dependent, independent of the presence of Na(+), and not affected by changes in membrane potential. The uptake of [(3)H]-UCB by aTPM, but not bTPM, was activated by ATP hydrolysis and inhibited by vanadate. Thus, the exact contribution of both inside out and right-side out bTPM to UCB uptake could not be distinguished, while only inverted aTPM were expected to carry out ATP-dependent UCB uptake. In bTPM and aTPM, uptake of free (unbound) [(3)H]-UCB (B(f)) consisted of a dominant, saturable, presumably carrier-mediated process and a diffusional component that became predominant only at B(f) near or above aqueous solubility limit for UCB (70 nM ). For bTPM, K(m)=7.2 nM; V(max)=9.8 pmol/20s/mg protein; and diffusion coefficient (K(D))=0.14 ml/20s/mg protein. For aTPM in the presence of 9.5m M ATP, K(m)=18 n M; V(max)=131 pmol/20s/mg protein; and K(D)=0.47 ml/20s/mg protein. The uptake of [(3)H]-UCB by bTPM was cis-inhibited by estrone-3-sulfate and estradiol-17 beta-glucuronide and trans-stimulated by unlabelled UCB and bromosulphopthalein. ATP-dependent UCB uptake by aTPM was cis-inhibited by doxorubicin, cholic acid, methotrexate and pronenecid. These findings suggest the presence of distinct transporters in the two domains of human placental trophoblast that could cooperate to transfer UCB from the foetus to the maternal circulation.

This study was undertaken to determine the effects of long-distance running on serum bilirubin fractions. Thirteen male ultramarathon runners participating in a 100-km race volunteered for the study. Venous blood samples were obtained shortly before and immediately after the race. Decreased serum haptoglobin levels (-66%) indicated the presence of hemolysis. After accounting for plasma-volume loss, significant post-race increases were found for creatine kinase (+20-fold), creatine kinase-MB (+252%), alanine aminotransferase (+42%), aspartate aminotransferase (+193%), gammaglutamyl transpeptidase (+56%), and glutamate dehydrogenase (+58%) serum activities, suggesting that running causes alterations of both muscle and liver tissues. Serum concentration of total bilirubin was significantly elevated (+106%) following the race, with changes corresponding to both unconjugated (+96%) and conjugated esters (+283%) of the pigment and significant increases in the ratio of esterified to total bilirubin. Our data show that long-distance running causes increases in the different serum bilirubin fractions which can be accounted for both hemolysis and hepatic disturbances.

The effects of subclinical fasciolosis at various stages of its development on bilirubin metabolism were investigated in rats. Studies were carried out at 3 (initial lesions), 6 (parenchymal degeneration), and 11 weeks (biliary localization of the parasite) after infection with an oral dose of 20 metacercariae of Fasciola hepatica. Both bile flow and bile acid secretion were significantly reduced by Weeks 3 and 6 postinfection, while a significantly increased bile flow was found in the 11-week group. Serum concentration of unconjugated bilirubin was elevated at 3-6 weeks postinfection and that of conjugated bilirubin and the esterified/total bilirubin ratio increased in all infected rats. Biliary excretion of bilirubin mono- and diconjugates was maximally increased by 3-6 weeks, remaining elevated at 11 weeks. No significant change in the biliary diconjugate/monoconjugate ratio was found in any of the groups. UDP-glucuronosyltransferase activity was reduced by Weeks 3 and 6. This was accompanied by a significantly lowered liver cytochrome P450 concentration and an elevation in hepatic heme oxygenase activity. Blood hemoglobin concentration was significantly reduced and reticulocyte count increased by 6 and 11 weeks. Our results indicate that enhanced bilirubin production, cholestasis, and a reduced conjugating activity could be responsible for the changes in plasma and bile pigment composition during the acute stage of fasciolosis. Hemolysis and a high bilirubin excretion are maintained when the parasite migrates into the biliary ducts.

Disturbances of bilirubin metabolism such as jaundice or pigment gallstone formation occur during total parenteral nutrition (TPN). We have studied the effects of TPN on bile flow and bile acid secretion and on the hepatobiliary transport of bilirubin in rats. Animals on parenteral nutrition for 5 days received 4.8 g of amino acids and 6.9 g of glucose daily. Controls were orally fed animals. Bile flow and bile acid secretion were not significantly modified by TPN. Serum bile acid and alkaline phosphatase levels were significantly increased in TPN animals when compared with the controls (+98% and +38%, respectively), which points to a relative cholestasis in the TPN rats. The biliary excretion of bilirubin monoconjugates and bilirubin diconjugates was significantly increased (+72% and +78%, respectively). This provides evidence for enhanced production of the pigment. Serum concentration of total bilirubin was enhanced in the TPN rats (+240%). The esterified/total bilirubin ratio in serum increased, whereas the bilirubin diconjugates/bilirubin monoconjugates ratio decreased. These facts, together with the minor reduction of hepatic bilirubin UDP glucuronosyltransferase activity (-12%), suggest that hyperbilirubinemia would be a consequence of both cholestasis and increased bilirubin production. The alterations reported here could contribute to the explanation of hyperbilirubinemia and pigment gallstone formation in patients maintained on parenteral nutrition.