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Many drug and therapeutic modalities have emerged over the past few years. However, successful commercialization is dependent on their safety and efficacy evaluations. Several preclinical models are available for drug-screening and safety evaluations, including cellular- and molecular-level models, tissue and organoid models, and animal models. Organoids are three-dimensional cell cultures derived from primary tissues or stem cells that are structurally and functionally similar to the original organs and can self-renew, and they are used to establish various disease models. Human hepatobiliary organoids have been used to study the pathogenesis of diseases, such as hepatitis, liver fibrosis, hepatocellular carcinoma, primary sclerosing cholangitis and biliary tract cancer, as they retain the physiological and histological characteristics of the liver and bile ducts. Here, we review recent research progress in validating drug toxicity, drug screening and personalized therapy for hepatobiliary-related diseases using human hepatobiliary organoid models, discuss the challenges encountered in current research and evaluate the possible solutions.

Background & Aims: Primary sclerosing cholangitis (PSC) is characterized by chronic inflammation and progressive fibrosis of the biliary tree. The bile acid receptor TGR5 (GPBAR1) is found on biliary epithelial cells (BECs), where it promotes secretion, proliferation and tight junction integrity. Thus, we speculated that changes in TGR5-expression in BECs may contribute to PSC pathogenesis., Methods: TGR5-expression and -localization were analyzed in PSC livers and liver tissue, isolated bile ducts and BECs from Abcb4 -/- , Abcb4 -/- /Tgr5 Tg and ursodeoxycholic acid (UDCA)- or 24-norursodeoxycholic acid (norUDCA)-fed Abcb4 -/- mice. The effects of IL8/IL8 homologues on TGR5 mRNA and protein levels were studied. BEC gene expression was analyzed by single-cell transcriptomics (scRNA-seq) from distinct mouse models., Results: TGR5 mRNA expression and immunofluorescence staining intensity were reduced in BECs of PSC and Abcb4 -/- livers, in Abcb4 -/- extrahepatic bile ducts, but not in intrahepatic macrophages. No changes in TGR5 BEC fluorescence intensity were detected in liver tissue of other liver diseases, including primary biliary cholangitis. Incubation of BECs with IL8/IL8 homologues, but not with other cytokines, reduced TGR5 mRNA and protein levels. BECs from Abcb4 -/- mice had lower levels of phosphorylated Erk and higher expression levels of Icam1, Vcam1 and Tgfβ2. Overexpression of Tgr5 abolished the activated inflammatory phenotype characteristic of Abcb4 -/- BECs. NorUDCA-feeding restored TGR5-expression levels in BECs in Abcb4 -/- livers., Conclusions: Reduced TGR5 levels in BECs from patients with PSC and Abcb4 -/- mice promote development of a reactive BEC phenotype, aggravate biliary injury and thus contribute to the pathogenesis of sclerosing cholangitis. Restoration of biliary TGR5-expression levels represents a previously unknown mechanism of action of norUDCA., Lay Summary: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease-associated with progressive inflammation of the bile duct, leading to fibrosis and end-stage liver disease. Bile acid (BA) toxicity may contribute to the development and disease progression of PSC. TGR5 is a membrane-bound receptor for BAs, which is found on bile ducts and protects bile ducts from BA toxicity. In this study, we show that TGR5 levels were reduced in bile ducts from PSC livers and in bile ducts from a genetic mouse model of PSC. Our investigations indicate that lower levels of TGR5 in bile ducts may contribute to PSC development and progression. Furthermore, treatment with norUDCA, a drug currently being tested in a phase III trial for PSC, restored TGR5 levels in biliary epithelial cells., Competing Interests: Conflict of interest MR, LS, CK, KF, JS, KD, JH, EL, JV, SFG, GH, SJF, IE, DN, CDF, MGB, GG, SV, JPM, MV, KS, TL, KK, DH, TL, MH have nothing to disclose.VK has served as speaker for Falk Foundation, Albireo and Abbvie. JRH has served on the advisory board for Novartis and Orkla Health. JRH has received research support from Biogen unrelated to the current work. THK has received consulting/speaker reimbursement from Gilead and Intercept unrelated to the current work. CS has served as speaker for Falk Foundation. MT has served as speaker for Falk Foundation, Gilead, Intercept and MSD; he has advised for Albireo, BiomX, Boehringer Ingelheim, Falk Pharma GmbH, Genfit, Gilead, Intercept, Jannsen, MSD, Novartis, Phenex, Regulus and Shire. MT further received travel grants from Abbvie, Falk, Gilead and Intercept and research grants from Albireo, Cymabay, Falk, Gilead, Intercept, MSD and Takeda. MT is also co-inventor of patents on the medical use of NorUDCA filed by the Medical Universities of Graz and Vienna (WO2006119803, WO20099013334). PF received norUDCA, an unrestricted research grant, travel grants and advisory board fees from Dr. Falk Pharma GmbH. PF is listed as co-inventor of patents for the medical use of norUDCA filed by the Medical University of Graz (WO2006119803, WO20099013334). UB received grants for investigator-initiated studies by Dr Falk GmbH, Freiburg, and Intercept, San Diego., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

E7766 represents a novel class of macrocycle-bridged dinucleotides and is under clinical development for immuno-oncology. In this report, we identified mechanism of systemic clearance E7766 and investigated the hepatobiliary transporters involved in the disposition of E7766 and potential drug interactions of E7766 as a victim of organic anion-transporting polypeptide (OATP) inhibitors. In bile-duct cannulated rats and dogs, E7766 was mainly excreted unchanged in bile (>80%) and to a lesser extent in urine (<20%). Sandwich-cultured human hepatocytes (SCHHs), transfected cells, and vesicles were used to phenotype the hepatobiliary transporters involved in the clearance of E7766. SCHH data showed temperature-dependent uptake of E7766 followed by active biliary secretion. In vitro transport assays using transfected cells and membrane vesicles confirmed that E7766 was a substrate of OATP1B1, OATP1B3, and multidrug resistance-associated protein 2. Phenotyping studies suggested predominant contribution of OATP1B3 over OATP1B1 in the hepatic uptake of E7766. Studies in OATP1B1/1B3 humanized mice showed that plasma exposure of E7766 increased 4.5-fold when coadministered with Rifampicin. Physiologically based pharmacokinetic models built upon two independent bottom-up approaches predicted elevation of E7766 plasma exposure when administered with Rifampicin, a clinical OATP inhibitor. In conclusion, we demonstrate that OATP-mediated hepatic uptake is the major contributor to the clearance of E7766, and inhibition of OATP1B may increase its systemic exposure. Predominant contribution of OATP1B3 in the hepatic uptake of E7766 was observed, suggesting polymorphisms in OATP1B1 would be unlikely to cause variability in the exposure of E7766. SIGNIFICANCE STATEMENT: Understanding the clearance mechanisms of new chemical entities is critical to predicting human pharmacokinetics and drug interactions. A physiologically based pharmacokinetic model that incorporated parameters from mechanistic in vitro and in vivo experiments was used to predict pharmacokinetics and drug interactions of E7766, a novel dinucleotide drug. The findings highlighted here may shed a light on the pharmacokinetic profile and transporter-mediated drug interaction propensity of other dinucleotide drugs., (Copyright © 2021 by The Author(s).)

Background: We have previously shown that obeticholic acid (OCA) upregulates the biliary excretion of asymmetric dimethylarginine (ADMA), an inhibitor of iNOS regulating the activity of matrix metalloproteinases (MMPs). Here, the effects of OCA on MMP-2 and MMP-9 activity in liver, bile and serum were evaluated after hepatic ischemia/reperfusion (I/R) injury., Material and Methods: Male Wistar rats (n = 20) were orally administered 10 mg/kg/day of OCA (5 days) and subjected to a 60-min ischemia and 60-min reperfusion. Bile, serum and tissue were collected for MMP-2 and MMP-9 activity quantification. The MMP regulator tissue reversion-inducing cysteine rich protein with Kazal motifs (RECK), tissue inhibitor of metalloproteinases (TIMPs), iNOS and biliary levels of LDH, γGT, glucose and ADMA were quantified., Results: In the I/R group, OCA administration reduced MMP-2 and MMP-9 in liver, bile and serum. A downregulation of tissue RECK and TIMPs, observed under I/R, were recovered by OCA. Immunohistochemical staining of hepatic tissue demonstrated that RECK expression is mainly localized in both cholangiocytes and hepatocytes. Hepatic iNOS positively correlated with tissue MMP-2 and MMP-9 activity. Biliary levels of LDH, γGT and glucose were lower in I/R rats treated with OCA; in bile, MMP levels positively correlated with LDH and γGT., Conclusion: Thus, OCA administration confers protection to cholangiocytes via downregulation of biliary MMPs in livers submitted to I/R. This event is associated with hepatic RECK- and TIMP-mediated MMP decrease., Competing Interests: The authors declare that Luciano Adorini is a part-time consultant of Intercept Pharmaceuticals. The specific roles of this author are articulated in the ‘author contributions’ section. However, the Intercept Pharmaceuticals did not provide support in the form of salaries for authors and did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The commercial affiliation does not alter our adherence to PLOS ONE policies on sharing data and materials.

Background: Preconditioning of donor livers before organ retrieval may improve organ quality after transplantation. We investigated whether preconditioning with metformin reduces preservation injury and improves hepatobiliary function in rat donor livers during ex situ normothermic machine perfusion (NMP) and after orthotopic liver transplantation., Methods: Lewis rats were administered metformin via oral gavage, after which a donor hepatectomy was performed followed by a standardized cold storage period of 4 hours. Graft assessment was performed using NMP via double perfusion of the hepatic artery and portal vein. In an additional experiment, rat donor livers preconditioned with metformin were stored on ice for 4 hours and transplanted to confirm postoperative liver function and survival. Data were analyzed and compared with sham-fed controls., Results: Graft assessment using NMP confirmed that preconditioning significantly improved ATP production, markers for hepatobiliary function (total bile production, biliary bilirubin, and bicarbonate), and significantly lowered levels of lactate, glucose, and apoptosis. After orthotopic liver transplantation, metformin preconditioning significantly reduced transaminase levels., Conclusions: Preconditioning with metformin lowers hepatobiliary injury and improves hepatobiliary function in an in situ and ex situ model of rat donor liver transplantation.

Background and Purpose: Irinotecan-induced diarrhea (IID) results from intestinal damages by its active metabolite SN-38. Alleviation of these damages has focused on lowering luminal SN-38 concentrations. However, it is unclear if the enteric bioavailability of SN-38 is mostly dependent on luminal SN-38 concentrations., Experimental Approach: Irinotecan (50 mg/kg, i.p. once daily for 6 days) was administered to female wildtype FVB, Mdr1a (-/-), Mrp2 (-/-) and Bcrp1 (-/-) mice for pharmacokinetic (PK), toxicokinetic (TK) and biodistribution studies. Plasma PK/TK profiles and tissues drug distribution were determined after first or sixth daily doses, along with activities of blood and gut esterases and intestinal Ugts. Caco-2 cells and bile-cannulate mice were used to further investigate intestinal and biliary disposition of irinotecan and its metabolites., Key Results: Significant differences in IID severity were observed with the susceptible rank of Bcrp1(-/-) > wildtype FVB > Mdr1a(-/-) > Mrp2(-/-). This rank order did not correlate with biliary excretion rates of SN-38/SN-38G. Rather, the severity was best correlated (R = 0.805) with the intestinal ratio of C ss SN-38/SN-38G, a measure of gut Ugt activity. On the contrary, IID was poorly correlated with plasma AUC ratio of SN-38/SN-38G (R = 0.227). Increased intestinal esterase activities due to repeated dosing and gut efflux transporter functionality are the other key factors that determine SN-38 enteric exposures., Conclusion and Implications: Intestinal SN-38 exposure is mainly affected by intestinal Ugt activities and blood esterase activities, and strongly correlated with severity of IID. Modulating intestinal SN-38 concentration and gut Ugt expression should be the focus of future studies to alleviate IID., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objective: To determine if use of the oral cholecystagogue, Ensure Plus (EP), in hepatobiliary scintigraphy (HBS) leads to a similar distribution of normal and abnormal gallbladder ejection fractions (GBEFs) versus other historical secondary findings of chronic biliary disease in a similar patient population compared with the conventional cholecystokinin analog, sincalide. The HBS findings analyzed included the GBEF, small bowel transit time, gallbladder fill time, and reversal of the normal gallbladder versus small bowel transit. The secondary objectives were to determine whether patient outcomes were significantly different for EP and sincalide HBS study patients following cholecystectomy, namely, the surgical pathology and patient-reported biliary-type pain., Methods: We reviewed all HBS examinations over a retrospective 34-month period. Data from 446 patients who underwent sincalide or EP HBS with GBEF determination for evaluation of chronic symptoms concerning for biliary etiology met the inclusion criteria. The aforementioned HBS findings and postsurgical patient outcomes were obtained for each patient group., Results: Comparing HBS examinations performed with sincalide or EP, no significant differences were noted in the major HBS findings of similar patient populations. Outcomes for the sincalide and EP groups status post cholecystectomy were assessed to determine their sensitivity, specificity, positive predictive value, and negative predictive value. The outcomes assessed were the histopathology and patient-reported amelioration of biliary-type pain. No significant differences were noted., Conclusions: The sincalide and EP cholecystagogues did not lead to significantly different HBS findings for similar patient populations or postcholecystectomy outcomes.

The aim of the present study was to investigate the potential effects of isotretinoin on the biliary system in patients with acne vulgaris receiving isotretinoin therapy. This was a preliminary retrospective study involving 40 patients with severe acne vulgaris who attended the dermatology clinic and were administered different doses (20 or 30 mg/day) of isotretinoin. Serum levels of AST, ALT, ALP, GGT, total bilirubin, direct bilirubin, and indirect bilirubin at the beginning and at the first month of therapy were scanned, recorded, and statistically analyzed. Total and indirect bilirubin levels at the first month of treatment in 30 patients, receiving isotretinoin at a dose of 20 mg/day, were significantly lower compared to the baseline values (p = .02 and p = .03, respectively), whereas AST and GGT serum levels were significantly higher (p = .003 and p = .006 respectively). No significant reduction in total and indirect bilirubin levels was detectable at the first month of treatment in 10 patients receiving isotretinoin at a dose of 30 mg/day; however, AST, ALP, and GGT levels were significantly elevated in these patients (p = .023; p = .004; and p = .001, respectively). To our knowledge, there is no previous study investigating the effects of isotretinoin on the biliary system, and, therefore, the present study is a preliminary one. Our findings implicate that low dose (20 mg/day) isotretinoin therapy can potentially reduce total and indirect bilirubin levels. Long-term, large-scale, prospective studies with patients receiving different doses of isotretinoin may provide more reliable information regarding the bilirubin lowering effects of isotretinoin and optimum dosing for achieving this clinical effect., (© 2019 Wiley Periodicals, Inc.)

Ceftriaxone (CTRX) forms salts with calcium (Ca) in the gall bladder and bile duct, and induces the formation of gallstones. In this study, factors of CTRX-induced gallstone formation were extracted from the results of a retrospective survey using the Japanese Adverse Drug Event Report (JADER), and the causal relationship between the factors and gallstone formation was investigated. From JADER, 136 patients who developed 'gallstone-related disorder' with CTRX as a suspected drug were extracted. The incidence of gallstone-induced adverse effects was high in patients treated with CTRX at a dose exceeding the normal daily dose and in children younger than 10 years old, suggesting that CTRX at a high level is a factor for gallstone formation. Thus, after mixing CTRX and Ca 2+ at different concentrations under different pH condition, the number of particles in the solutions was measured using a Coulter counter. As a result, the number of minute particles significantly increased at all pH values when Ca 2+ and CTRX were mixed at a concentration of 10 mEq/L or higher and 1.5 g/L or higher, respectively. At pH 6.5 or 7.0, visible crystals were detected when 25 mEq/L of Ca 2+ and 2.0 g/L of CTRX were mixed. Based on these findings, attention should be sufficiently paid to the development of 'gallstone-related disorder' in pediatric patients and in patients treated with CTRX at a dose exceeding the normal dose. Furthermore, gallstone formation and growth may be promoted when CTRX and Ca 2+ coexist at high concentrations under low pH conditions., (Copyright © 2019 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Human biliary tree stem/progenitor cells (hBTSCs), reside in peribiliary glands, are mainly stimulated by primary sclerosing cholangitis (PSC) and cholangiocarcinoma. In these pathologies, hBTSCs displayed epithelial-to-mesenchymal transition (EMT), senescence characteristics, and impaired differentiation. Here, we investigated the effects of cholest-4,6-dien-3-one, an oxysterol involved in cholangiopathies, on hBTSCs biology. hBTSCs were isolated from donor organs, cultured in self-renewal control conditions, differentiated in mature cholangiocytes by specifically tailored medium, or exposed for 10 days to concentration of cholest-4,6-dien-3-one (0.14 mM). Viability, proliferation, senescence, EMT genes expression, telomerase activity, interleukin 6 (IL6) secretion, differentiation capacity, and HDAC6 gene expression were analyzed. Although the effect of cholest-4,6-dien-3-one was not detected on hBTSCs viability, we found a significant increase in cell proliferation, senescence, and IL6 secretion. Interestingly, cholest-4.6-dien-3-one impaired differentiation in mature cholangiocytes and, simultaneously, induced the EMT markers, significantly reduced the telomerase activity, and induced HDAC6 gene expression. Moreover, cholest-4,6-dien-3-one enhanced bone morphogenic protein 4 (Bmp-4) and sonic hedgehog (Shh) pathways in hBTSCs. The same pathways activated by human recombinant proteins induced the expression of EMT markers in hBTSCs. In conclusion, we demonstrated that chronic exposition of cholest-4,6-dien-3-one induced cell proliferation, EMT markers, and senescence in hBTSC, and also impaired the differentiation in mature cholangiocytes.

Estrogen-induced cholestasis is a common etiology of hepatic diseases in women with contraceptives administration, pregnancy or hormone replacement therapy. Farnesoid X receptor (FXR) is a member of nuclear receptor super family of ligand-activated transcription factors that is highly expressed in liver. FXR is acknowledged to contribute to the bile acid homeostasis, as well as the pathogenesis and progression of cholestasis. Specific targeting of FXR is an innovative approach for the treatment of cholestasis. The current study aimed to verify the anti-cholestasis effect of yangonin that is a natural product isolated from Kava via FXR signaling pathway in vivo and in vitro. The analyses of FXR gain- or loss-of-function were performed. Yangonin treatment ameliorates estrogen-induced cholestasis through increasing bile flow and biliary bile acid output. The mechanisms were an induction in the hepatic efflux transporters (Bsep and Mrp2) and an inhibition in hepatic uptake transporter (Ntcp) by yangonin. Likewise, yangonin through repressing Cyp7a1, Cyp8b1 and inducing Sult2a1 expression suppressed bile acid synthesis and promoted bile acid metabolism. Furthermore, yangonin improved estrogen-induced inflammatory cell infiltration and the inflammation gene expression. In vitro experiments further consolidated that yangonin alleviated estrogen-caused cholestasis via FXR activation. Noteworthily, the effects of yangonin were enhanced by FXR expression plasmids but abrogated by FXR siRNA. In conclusion, yangonin alleviates estrogen-induced cholestasis, due to FXR-mediated gene regulation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

In the liver, several approaches are used to investigate and predict the complex issue of drug-induced transporter inhibition. These approaches include in vitro assays and pharmacokinetic models that predict how inhibitors modify the systemic and liver concentrations of the victim drugs. Imaging is another approach that shows how inhibitors might alter liver concentrations stronger than systemic concentrations. In perfused rat livers associated with a gamma counter that measures liver concentrations continuously, we previously showed how fluxes across transporters generate the hepatocyte concentrations of two clinical imaging compounds, one with a low extraction ratio [gadobenate dimeglumine (BOPTA)] and one with a high extraction ratio [mebrofenin (MEB)]. BOPTA and MEB are transported by rat organic anion transporting polypeptide and multiple resistance-associated protein 2, which are both inhibited by rifampicin. The aim of the study is to measure how rifampicin modifies the hepatocyte concentrations and membrane clearances of BOPTA and MEB and to determine whether these compounds might be used to investigate transporter-mediated drug-drug interactions in clinical studies. We show that rifampicin coperfusion greatly decreases BOPTA hepatocyte concentrations, but increases those of MEB. Rifampicin strongly decreases BOPTA hepatic clearance. In contrast, rifampicin decreases moderately MEB hepatic clearance and blocks the biliary intrinsic clearance, increasing MEB hepatocyte concentrations. In conclusion, low concentrations prevent the quantification of BOPTA biliary intrinsic clearance, while MEB is a promising imaging probe substrate to evidence transporter-mediated drug-drug interactions when inhibitors act on influx and efflux transporters., (Copyright © 2019 by The American Society for Pharmacology and Experimental Therapeutics.)

Background: Using a meta-analysis of randomized controlled trials (RCTs), this study aimed to investigate the efficacy and safety of pemafibrate, a novel selective peroxisome proliferator-activated receptor α modulator, in patients with dyslipidemia., Methods: A search was performed using the MEDLINE, Cochrane Controlled Trials Registry, and ClinicalTrials.gov databases. We decided to employ RCTs to evaluate the effects of pemafibrate on lipid and glucose metabolism-related parameters in patients with dyslipidemia. For statistical analysis, standardized mean difference (SMD) or odds ratio (OR) and 95% confidence intervals (CIs) were calculated using the random effect model., Results: Our search yielded seven RCTs (with a total of 1623 patients) that satisfied the eligibility criteria of this study; hence, those studies were incorporated into this meta-analysis. The triglyceride concentration significantly decreased in the pemafibrate group (SMD, - 1.38; 95% CI, - 1.63 to - 1.12; P < 0.001) than in the placebo group, with a reduction effect similar to that exhibited by fenofibrate. Compared with the placebo group, the pemafibrate group also showed improvements in high-density and non-high-density lipoprotein cholesterol levels as well as in homeostasis model assessment for insulin resistance. Furthermore, the pemafibrate group showed a significant decrease in hepatobiliary enzyme activity compared with the placebo and fenofibrate groups; and, total adverse events (AEs) were significantly lower in the pemafibrate group than in the fenofibrate group (OR, 0.60; 95% CI, 0.49-0.73; P < 0.001). In contrast, the low-density lipoprotein cholesterol level was significantly higher in the pemafibrate group than in the placebo (P = 0.006) and fenofibrate (P < 0.001) groups., Conclusions: The lipid profile significantly improved in the pemafibrate group than in the placebo group. In addition to the pemafibrate group having an improved lipid profile, which was comparable with that of the fenofibrate group, the AEs were significantly lower than in the fenofibrate group and an improvement in hepatobiliary enzyme activity was also recognized. However, we believe that actual clinical data as well as long-term efficacy and safety need to be investigated in the future.

Acetaminophen (APAP) overdose is one of the world's leading causes of drug-induced hepatotoxicity. Although traditional methods such as histological imaging and biochemical assays have been successfully applied to evaluate the extent of APAP-induced liver damage, detailed effect of how APAP overdose affect the recovery of hepatobiliary metabolism and is not completely understood. In this work, we used intravital multiphoton microscopy to image and quantify hepatobiliary metabolism of the probe 6-carboxyfluorescein diacetate in APAP-overdose mice. We analyzed hepatobiliary metabolism for up to 7 days following the overdose and found that the excretion of the probe molecule was the most rapid on Day 1 following APAP overdose and slowed down on Days 2 and 3. On Day 7, probe excretion capability has exceeded that of the normal mice, suggesting that newly regenerated hepatocytes have higher metabolic capabilities. Our approach may be further developed applied to studying drug-induced hepatotoxicity in vivo., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Three-dimensional (3D) printing of decellularized extracellular matrix (dECM) hydrogels is a promising technique for regenerative engineering. 3D-printing enables the reproducible and precise patterning of multiple cells and biomaterials in 3D, while dECM has high organ-specific bioactivity. However, dECM hydrogels often display poor printability on their own and necessitate additives or support materials to enable true 3D structures. In this study, we used a sacrificial material, 3D-printed Pluronic F-127, to serve as a platform into which dECM hydrogel can be incorporated to create specifically designed structures made entirely up of dECM. The effects of 3D dECM are studied in the context of engineering the intrahepatic biliary tree, an often-understudied topic in liver tissue engineering. Encapsulating biliary epithelial cells (cholangiocytes) within liver dECM has been shown to lead to the formation of complex biliary trees in vitro. By varying several aspects of the dECM structures' geometry, such as width and angle, we show that we can guide the directional formation of biliary trees. This is confirmed by computational 3D image analysis of duct alignment. This system also enables fabrication of a true multi-layer dECM structure and the formation of 3D biliary trees into which other cell types can be seeded. For example, we show that hepatocyte spheroids can be easily incorporated within this system, and that the seeding sequence influences the resulting structures after seven days in culture. STATEMENT OF SIGNIFICANCE: The field of liver tissue engineering has progressed significantly within the past several years, however engineering the intrahepatic biliary tree has remained a significant challenge. In this study, we utilize the inherent bioactivity of decellularized extracellular matrix (dECM) hydrogels and 3D-printing of a sacrificial biomaterial to create spatially defined, 3D biliary trees. The creation of patterned, 3D dECM hydrogels in the past has only been possible with additives to the gel that may stifle its bioactivity, or with rigid and permanent support structures that may present issues upon implantation. Additionally, the biological effect of 3D spatially patterned liver dECM has not been demonstrated independent of the effects of dECM bioactivity alone. This study demonstrates that sacrificial materials can be used to create pure, multi-layer dECM structures, and that strut width and angle can be changed to influence the formation and alignment of biliary trees encapsulated within. Furthermore, this strategy allows co-culture of other cells such as hepatocytes. We demonstrate that not only does this system show promise for tissue engineering the intrahepatic biliary tree, but it also aids in the study of duct formation and cell-cell interactions., (Copyright © 2018. Published by Elsevier Ltd.)

Background: Diarrhea is a severe side effect of irinotecan, a pro-drug of SN-38 used for the treatment of many types of cancers. Pre-clinical and clinical studies showed that decreasing the colonic exposure of SN-38 can mitigate irinotecan-induced diarrhea., Objective: The purpose of this study is to evaluate the anti-diarrhea potential of Xiao-Chai-Hu-Tang (XCHT), a traditional Chinese herbal formula, against irinotecan-induced diarrhea by determining if and how XCHT alters the disposition of SN-38., Methods: LC-MS/MS was used to quantify the concentrations of irinotecan and its major metabolites (i.e., SN-38, SN-38G). An Intestinal perfusion model was used to determine the effect of XCHT on the biliary and intestinal secretions of irinotecan, SN-38, and SN-38G. Pharmacokinetic (PK) studies were performed to determine the impact of XCHT on the blood and fecal concentrations of irinotecan, SN-38, and SN-38G., Results: The results showed that XCHT significantly inhibits both biliary and intestinal excretions of irinotecan, SN-38, and SN-38G (range: 35% to 95%). PK studies revealed that the fecal concentrations of irinotecan and SN-38 were significantly decreased from 818.35 ± 120.2 to 411.74 ± 138.83 µg/g or from 423.95 ± 76.44 to 245.63 ± 56.72 µg/g (p<0.05) by XCHT, respectively, suggesting the colonic exposure of SN-38 is significantly decreased by XCHT. PK studies also showed that the plasma concentrations of irinotecan, SN-38, and SN-38G were not affected by XCHT., Conclusion: In conclusion, XCHT significantly decreased the exposure of SN-38 in the gut without affecting its plasma level, thereby possessing the potential of alleviating irinotecan-induced diarrhea without negatively impacting its therapeutic efficacy., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Bile acid imbalance causes progressive familial intrahepatic cholestasis type 2 (PFIC2) or type 3 (PFIC3), severe liver diseases associated with genetic defects in the biliary bile acid transporter bile salt export pump (BSEP; ABCB11) or phosphatidylcholine transporter multidrug resistance protein 3 (MDR3; ABCB4), respectively. Mdr2 -/- mice (a PFIC3 model) develop progressive cholangitis, ductular proliferation, periportal fibrosis, and hepatocellular carcinoma (HCC) because the nonmicelle-bound bile acids in the bile of these mice are toxic. We asked whether the highly hydrophilic bile acids generated by Bsep -/- mice could protect Mdr2 -/- mice from progressive liver damage. We generated double-KO (DKO: Bsep -/- and Mdr2 -/- ) mice. Their bile acid composition resembles that of Bsep -/- mice, with increased hydrophilic muricholic acids, tetrahydroxylated bile acids (THBAs), and reduced hydrophobic cholic acid. These mice lack the liver pathology of their Mdr2 -/- littermates. The livers of DKO mice have gene expression profiles very similar to Bsep -/- mice, with 4,410 of 6,134 gene expression changes associated with the Mdr2 -/- mutation being suppressed. Feeding with THBAs partially alleviates liver damage in the Mdr2 -/- mice. Hydrophilic changes to biliary bile acid composition, including introduction of THBA, can prevent the progressive liver pathology associated with the Mdr2 -/- (PFIC3) mutation., (Copyright © 2019 Wang et al.)

Pharmacology principles provide key understanding that underpins the clinical and research roles of nuclear medicine practitioners. Indeed, the scope of practice of the nuclear medicine technologist demands knowledge and understanding of indications, contraindications, warnings, precautions, proper use, drug interactions, and adverse reactions for each medication to be used. This article is the third in a series of articles that aims to enhance the understanding of pharmacologic principles relevant to nuclear medicine. This article will build on the introductory concepts, terminology, and principles of pharmacology explored in the first 2 articles in the series. Specifically, this article will focus on the pharmacologic principles associated with interventional medications encountered in renal and biliary imaging. A companion article (part 3B) will examine the less commonly used interventional medications in general nuclear medicine and prototype adjunctive medications. Future articles will address the pharmacology related to nuclear cardiology, the emergency crash cart, and contrast media associated with CT and MRI., (© 2018 by the Society of Nuclear Medicine and Molecular Imaging.)

Background: Primary sclerosing cholangitis (PSC) is a chronic, progressive cholestatic biliary disease associated with inflammatory bowel disease (IBD) with no known cure., Aim: To evaluate the effect of biological therapies on PSC progression in IBD patients., Methods: We performed a retrospective cohort study of 88 cases (75 unique patients with 12 patients treated >1 biologics) of IBD (48 ulcerative colitis, 24 Crohn's disease and 3 indeterminate colitis) with concomitant PSC who received biological therapy (42 infliximab, 19 adalimumab, 27 vedolizumab) between June 2002 and October 2017. Hepatic biochemistries were compared using the paired t-test (patients served as their own controls) ≤3 months before and 6-8 and 12-14 months after biological initiation. Radiographic information of biliary stenosis and liver fibrosis were obtained via abdominal ultrasound, abdominal magnetic resonance imaging and magnetic resonance elastography., Results: Use of adalimumab was associated with a significant decrease in alkaline phosphatase (ALP) after 6-8 months (P = 0.03; mean change -70 U/L, standard deviation [SD] 88 U/L) compared to vedolizumab (mean change +50 U/L, SD 142 U/L) or infliximab (mean change +37 U/L, SD 183 U/L) but the change was not significant after 12-14 months (P = 0.24). No significant decreases were observed with AST, ALT, total or direct bilirubin, elastography score or radiographic imaging of biliary tree dilation/strictures with any biological therapy after 6-8 or 12-14 months., Conclusions: Current evidence suggests that biological therapies used for the treatment of IBD are not effective treatments for PSC. Further study is needed to elucidate any potential beneficial effect of adalimumab on PSC., (© 2018 John Wiley & Sons Ltd.)

Background and Objectives: Prophylactic antibiotics are frequently administered for transarterial chemoembolization (TACE) of hepatocellular carcinoma (HCC). In patients without previous biliary instrumentation, infection risk from TACE is low. We hypothesized that there is a negligible rate of infection in these patients without prophylactic antibiotics., Methods: We reviewed consecutive patients undergoing TACE between 7/1/2013-6/15/2016. All patients had an intact Sphincter of Oddi, received no peri-procedural antibiotics, and had 30+ days follow-up. Level of arterial selection was recorded. Baseline Child-Pugh (CP) and Barcelona Clinic Liver Cancer (BCLC) scores were recorded. The primary outcome measure was the absence of clinical or imaging findings of hepatic abscess within 30 days., Results: A total of 171 patients underwent 235 TACE procedures. CP scores were A (n = 109), B (n = 47), and C (n = 15). BCLC scores were 0 (n = 1), A (n = 108), B (n = 47), and C (n = 15). TACE was performed segmentally (n = 208) or lobar (n = 27). Three patients died of non-infectious causes before 30 days. No hepatic abscesses developed in evaluable patients: 0/232 infusions., Conclusions: In patients with HCC and an intact Sphincter of Oddi, TACE was performed safely without prophylactic antibiotics. The majority of the patients were BCLC and CP A/B. Additional study of BCLC and CP C patients is warranted., (© 2018 Wiley Periodicals, Inc.)

Cholangiopathies are progressive disorders with largely unknown pathoetiology and limited treatment options. We aimed to develop a novel surgical technique with direct access to the bile ducts that would complement existing mouse models of cholestasis, biliary inflammation, and fibrosis and present a new route of administration for testing of potential treatment strategies. We developed a surgical technique to access the murine biliary tree by injection of different solvents through catheterization of the gall bladder with simultaneous clamping of the common bile duct. To demonstrate the applicability of the technique, we injected either phosphate-buffered saline or dimethyl sulfoxide in concentrations of 50 or 65% and compared these groups with sham-operated mice. The surgery was optimized to achieve a mortality rate close to 0. There were no significant changes in pain, activity level, or mortality from the day of the surgery until euthanization for any groups. Injection of phosphate-buffered saline or 50% dimethyl sulfoxide was generally well-tolerated, whereas 65% dimethyl sulfoxide led to higher weight loss, an increase of serum alanine transaminase, and histological portal inflammation. There were no signs of inflammation in the gut. We have developed a bile duct injection technique that is well-tolerated, easily reproducible, and that may complement existing models of cholangiopathies. Direct access to the bile ducts without causing harm to the hepatobiliary or intestinal tissue may be valuable in future studies of normal biliary physiology and different pathophysiological mechanisms of disease and to test novel therapeutic strategies. NEW & NOTEWORTHY To evaluate tolerability of the bile duct to injection of both polar and nonpolar compounds, we established a novel biliary injection technique. This technique is well-tolerated, easily reproducible, and with direct access to the bile ducts for studies of the murine biliary tree. The bile duct injection technique may complement existing animal models and be a valuable tool in future studies of normal biliary physiology or pathophysiology and to test novel therapeutic strategies.

Cystic fibrosis transmembrane conductance regulator (CFTR), the channel mutated in cystic fibrosis (CF), is expressed by the biliary epithelium (i.e., cholangiocytes) of the liver. Progressive clinical liver disease (CF-associated liver disease; CFLD) occurs in around 10% of CF patients and represents the third leading cause of death. Impaired secretion and inflammation contribute to CFLD; however, the lack of human-derived experimental models has hampered the understanding of CFLD pathophysiology and the search for a cure. We have investigated the cellular mechanisms altered in human CF cholangiocytes using induced pluripotent stem cells (iPSCs) derived from healthy controls and a ΔF508 CFTR patient. We have devised a novel protocol for the differentiation of human iPSC into polarized monolayers of cholangiocytes. Our results show that iPSC-cholangiocytes reproduced the polarity and the secretory function of the biliary epithelium. Protein kinase A/cAMP-mediated fluid secretion was impaired in ΔF508 cholangiocytes and negligibly improved by VX-770 and VX-809, two small molecule drugs used to correct and potentiate ΔF508 CFTR. Moreover, ΔF508 cholangiocytes showed increased phosphorylation of Src kinase and Toll-like receptor 4 and proinflammatory changes, including increased nuclear factor kappa-light-chain-enhancer of activated B cells activation, secretion of proinflammatory chemokines (i.e., monocyte chemotactic protein 1 and interleukin-8), as well as alterations of the F-actin cytoskeleton. Treatment with Src inhibitor (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyramidine) decreased the inflammatory changes and improved cytoskeletal defects. Inhibition of Src, along with administration of VX-770 and VX-809, successfully restored fluid secretion to normal levels., Conclusion: Our findings have strong translational potential and indicate that targeting Src kinase and decreasing inflammation may increase the efficacy of pharmacological therapies aimed at correcting the basic ΔF508 defect in CF liver patients. These studies also demonstrate the promise of applying iPSC technology in modeling human cholangiopathies. (Hepatology 2018;67:972-988)., (© 2017 by the American Association for the Study of Liver Diseases.)

Background: We previously showed that increased asymmetric dimethylarginine (ADMA) biliary excretion occurs during hepatic ischemia/reperfusion (I/R), prompting us to study the effects of the farnesoid X receptor (FXR) agonist obeticholic acid (OCA) on bile, serum and tissue levels of ADMA after I/R., Material and Methods: Male Wistar rats were orally administered 10mg/kg/day of OCA or vehicle for 5 days and were subjected to 60 min partial hepatic ischemia or sham-operated. After a 60 min reperfusion, serum, tissue and bile ADMA levels, liver mRNA and protein expression of ADMA transporters (CAT-1, CAT-2A, CAT-2B, OCT-1, MATE-1), and enzymes involved in ADMA synthesis (protein-arginine-N-methyltransferase-1, PRMT-1) and metabolism (dimethylarginine-dimethylaminohydrolase-1, DDAH-1) were measured., Results: OCA administration induced a further increase in biliary ADMA levels both in sham and I/R groups, with no significant changes in hepatic ADMA content. A reduction in CAT-1, CAT-2A or CAT-2B transcripts was found in OCA-treated sham-operated rats compared with vehicle. Conversely, OCA administration did not change CAT-1, CAT-2A or CAT-2B expression, already reduced by I/R. However, a marked decrease in OCT-1 and increase in MATE-1 expression was observed. A similar trend occurred with protein expression., Conclusion: The reduced mRNA expression of hepatic CAT transporters suggests that the increase in serum ADMA levels is probably due to decreased liver uptake of ADMA from the systemic circulation. Conversely, the mechanism involved in further increasing biliary ADMA levels in sham and I/R groups treated with OCA appears to be MATE-1-dependent.

Curcumin, an aromatic phytoextract from the turmeric (Curcuma longa) rhizome, has been used for centuries for a variety of purposes, not the least of which is medicinal. A growing body of evidence suggests that curcumin has a broad range of potentially therapeutic pharmacological properties, including anti-inflammatory, anti-fibrotic, and anti-neoplastic effects, among others. Clinical applications of curcumin have been hampered by quality control concerns and limited oral bioavailability, although novel formulations appear to have largely overcome these issues. Recent in vitro and in vivo studies have found that curcumin's cytoprotective and other biological activities may play a role in an array of benign and malignant hepatobiliary conditions, including but not limited to non-alcoholic fatty liver disease, cholestatic liver disease (e.g. primary sclerosing cholangitis), and cholangiocarcinoma. Here we provide an overview of fundamental principles, recent discoveries, and potential clinical hepatobiliary applications of this pleiotropic phytocompound.

Aim: Glucagon-like peptide 1 receptor agonists (GLP1-RA) have been associated with an increased risk of pancreatitis and pancreatic cancer. Prior meta-analyses of randomized controlled trials failed to show any significant increase of risk; however, those meta-analyses did not include the recently published cardiovascular outcome trials (CVOT) with GLP1-RA, which provide a substantial additional body of data. The aim of the present meta-analysis is to assess the effect of GLP1-RA on pancreatitis, pancreatic cancers and cholelithiasis., Materials and Methods: A Medline search for GLP-1 receptor agonists (exenatide, liraglutide, lixisenatide, albiglutide, dulaglutide or semaglutide) was performed, collecting all randomized clinical trials with a duration >11 weeks, enrolling patients with type 2 diabetes and comparing a GLP-1 receptor agonist with placebo or any other non-GLP-1 receptor agonist drug., Results: Of the 113 trials fulfilling inclusion criteria, 13 did not report information on pancreatitis, whereas 72 reported no events in all treatment groups. The incidence of pancreatitis and pancreatic cancer with GLP1-RA was not significantly different from that observed in comparator arms (MH-OR [95% CI] 0.93 [0.65-1.34], P = .71, and 0.94 [0.52-1.70], P = .84, respectively), whereas, a significantly increased risk of cholelithiasis (MH-OR [95% CI] 1.30 [1.01-1.68], P = .041) was detected., Conclusions: Presently available data confirm the safety of GLP-1 receptor agonists for pancreatitis. Conversely, therapy with those drugs is associated with an increased risk of cholelithiasis, which deserves further investigation., (© 2017 John Wiley & Sons Ltd.)

It has been known from previous studies that body fluids, such as cerebrospinal fluid, lung surfactant, and urine, have a strong impact on the bacterial killing of many anti-infective agents. However, the influence of human bile on the antimicrobial activity of antibiotics is widely unknown. Human bile was obtained and pooled from 11 patients undergoing cholecystectomy. After sterilization of the bile fluid by gamma irradiation, its effect on bacterial killing was investigated for linezolid (LZD) and tigecycline (TGC) against Enterococcus faecalis ATCC 29212. Further, ciprofloxacin (CIP), meropenem (MEM), and TGC were tested against Escherichia coli ATCC 25922. Time-kill curves were performed in pooled human bile and Mueller-Hinton broth (MHB) over 24 h. Bacterial counts (in CFU per milliliter after 24 h) of bile growth controls were approximately equal to MHB growth controls for E. coli and approximately 2-fold greater for E. faecalis , indicating a promotion of bacterial growth by bile for the latter strain. Bile reduced the antimicrobial activity of CIP, MEM, and TGC against E. coli as well as the activity of LZD and TGC against E. faecalis This effect was strongest for TGC against the two strains. Degradation of TGC in bile was identified as the most likely explanation. These findings may have important implications for the treatment of bacterial infections of the gallbladder and biliary tract and should be explored in more detail., (Copyright © 2017 American Society for Microbiology.)

Purpose/Aim: Oxidative stress plays an important role in the pathogenesis of acute pancreatitis (AP). We compared the therapeutic effects of Ukrain (NSC 631570) and N-acetylcysteine (NAC) in rats with AP., Materials and Methods: Forty male Sprague Dawley rats were divided into four groups: controls; AP; AP with NAC; and AP with Ukrain. AP was induced via the ligation of the bile-pancreatic duct; drugs were administered intraperitoneally (i.p.) 30 min and 12 h after AP induction. Twenty-four hours after AP induction, animals were sacrificed and the pancreas was excised. Levels of malondialdehyde (MDA) and nitric oxide (NO), and activity levels of tumor necrosis factor (TNF)-α, and myeloperoxidase (MPO) were measured in tissue samples. Total oxidant status (TOS), total antioxidant status (TAS), and total bilirubin, as well as activity levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), amylase and lipase were measured in serum samples. Pancreatic tissue histopathology was also evaluated., Results: Test drugs reduced levels of MDA, NO, TNF-α, total bilirubin, AST, ALT, TOS and MPO, amylase and lipase activities (P < 0.001), and increased TAS (P < 0.001). Rats treated with test drugs attenuated AP-induced morphologic changes and decreased pancreatic damage scores compared with the AP group (P < 0.05). Both test drugs attenuated pancreatic damage, but the therapeutic effect was more pronounced in rats that received Ukrain than in those receiving NAC., Conclusions: These results suggest that treatment with Ukrain or NAC can reduce pancreatic damage via anti-inflammatory and antioxidant effects.

The objective of this work was to investigate the mechanisms of hepatobiliary toxicity caused by thienopyrimidone MCHR1 antagonists using BMS-773174 as a tool molecule. Co-administration of the pan CYP inhibitor 1-aminobenzotriazole with BMS-773174 prevented hepatobiliary damage, and direct delivery of the diol metabolite BMS-769750 caused hepatobiliary toxicity, identifying the diol and possibly its downstream hydroxyacid (BMS-800754) metabolite as the toxic species. Rat liver gene expression revealed treatment-related changes in hepatic transporters and induction of oval cell-specific genes including deleted malignant tumor 1 (Dmbt1). The metabolites did not alter hepatic transporter activities, suggesting that transporter-mediated cholestasis was not involved. Because injury to biliary epithelium can result in adaptive hyperplasia, rat biliary epithelial cells (BECs) were isolated and exposed to the oxidative metabolites. BMS-769750 was cytotoxic to BECs, but not rat hepatocytes, suggesting a role of the diol in biliary epithelial injury. BMS-800754 was cytotoxic to rat hepatocytes therefore its contribution to hepatocyte injury in rats is a possibility. Induction of Dmbt1 in rat BECs was investigated because of its role in hepatic progenitor cell differentiation/proliferation during injury. Dmbt1 mRNA was induced by BMS-769750, but not BMS-800754 in BECs; this induction and cellular injury was confirmed with diol metabolites formed by other compounds with the same hepatobiliary liability. In conclusion, hepatobiliary injury by thienopyrimidinone MCHR1 antagonists was driven through a CYP-mediated bioactivation pathway. Induction of Dmbt1 mRNA coupled with cellular injury suggests that injury of biliary epithelium may be the first step toward an adaptive proliferative response causing BDH by these compounds., (© The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The role of endoplasmic reticulum stress and the unfolded protein response (UPR) in cholestatic liver disease and fibrosis is not fully unraveled. Tauroursodeoxycholic acid (TUDCA), a hydrophilic bile acid, has been shown to reduce endoplasmic reticulum (ER) stress and counteract apoptosis in different pathologies. We aimed to investigate the therapeutic potential of TUDCA in experimental secondary biliary liver fibrosis in mice, induced by common bile duct ligation. The kinetics of the hepatic UPR and apoptosis during the development of biliary fibrosis was studied by measuring markers at six different timepoints post-surgery by qPCR and Western blot. Next, we investigated the therapeutic potential of TUDCA, 10 mg/kg/day in drinking water, on liver damage (AST/ALT levels) and fibrosis (Sirius red-staining), in both a preventive and therapeutic setting. Common bile duct ligation resulted in the increased protein expression of CCAAT/enhancer-binding protein homologous protein (CHOP) at all timepoints, along with upregulation of pro-apoptotic caspase 3 and 12, tumor necrosis factor receptor superfamily, member 1A (TNFRsf1a) and Fas-Associated protein with Death Domain (FADD) expression. Treatment with TUDCA led to a significant reduction of liver fibrosis, accompanied by a slight reduction of liver damage, decreased hepatic protein expression of CHOP and reduced gene and protein expression of pro-apoptotic markers. These data indicate that TUDCA exerts a beneficial effect on liver fibrosis in a model of cholestatic liver disease, and suggest that this effect might, at least in part, be attributed to decreased hepatic UPR signaling and apoptotic cell death., Competing Interests: The authors declare no conflict of interest.

Notch signaling has been demonstrated to be involved in ductular reactions and fibrosis. Previous studies have shown that Huang Qi Decoction (HQD) can prevent the progression of cholestatic liver fibrosis (CLF). However, whether HQD affects the Notch signaling pathway is unclear. In this study, CLF was established by common bile duct ligation (BDL) in rats. At the end of the first week, the rats were randomly divided into a model group (i.e., BDL), an HQD group, and a sorafenib positive control group (SORA) and were treated for 3 weeks. Bile duct proliferation and liver fibrosis were determined by tissue staining. Activation of the Notch signaling pathway was evaluated by analyzing expressions of Notch-1, -2, -3, and -4, Jagged (JAG) 1, and Delta like (DLL)-1, -3, and -4. The results showed that HQD significantly reduced the deposition of collagen and the Hyp content of liver tissue and inhibited the activation of HSCs compared with the BDL group. In addition, HQD significantly decreased the protein and mRNA expressions of TGF-[Formula: see text]1 and [Formula: see text]-SMA. In contrast, HQD significantly enhanced expression of the Smad 7 protein. HQD also reduced biliary epithelial cell proliferation, and reduced the mRNA levels of CK7, CK8, CK18, SRY-related high mobility group-box gene (SOX) 9, epithelial cell adhesion molecule (EpCAM) and the positive areas of CK19 and OV6. In addition, the mRNA and protein expressions of Notch-3, -4, JAG1, and DLL-1, -3 were significantly reduced in the HQD compared to the BDL group. These results demonstrated that HQD may prevent biliary liver fibrosis through inhibition of the Notch signaling pathway, and it may be a potential treatment for cholestatic liver disease.

30-99% of administered nanoparticles will accumulate and sequester in the liver after administration into the body. This results in reduced delivery to the targeted diseased tissue and potentially leads to increased toxicity at the hepatic cellular level. This review article focuses on the inter- and intra-cellular interaction between nanoparticles and hepatic cells, the elimination mechanism of nanoparticles through the hepatobiliary system, and current strategies to manipulate liver sequestration. The ability to solve the "nanoparticle-liver" interaction is critical to the clinical translation of nanotechnology for diagnosing and treating cancer, diabetes, cardiovascular disorders, and other diseases., (Copyright © 2016. Published by Elsevier B.V.)

Transporter-mediated alterations in bile acid disposition may have significant toxicological implications. Current methods to predict interactions are limited by the interplay of multiple transporters, absence of protein in the experimental system, and inaccurate estimates of inhibitor concentrations. An integrated approach was developed to predict altered bile acid disposition due to inhibition of multiple transporters using the model bile acid taurocholate (TCA). TCA pharmacokinetic parameters were estimated by mechanistic modeling using sandwich-cultured human hepatocyte data with protein in the medium. Uptake, basolateral efflux, and biliary clearance estimates were 0.63, 0.034, and 0.074 mL/min/g liver, respectively. Cellular total TCA concentrations (Ct,Cells) were selected as the model output based on sensitivity analysis. Monte Carlo simulations of TCA Ct,Cells in the presence of model inhibitors (telmisartan and bosentan) were performed using inhibition constants for TCA transporters and inhibitor concentrations, including cellular total inhibitor concentrations ([I]t,cell) or unbound concentrations, and cytosolic total or unbound concentrations. For telmisartan, the model prediction was accurate with an average fold error (AFE) of 0.99-1.0 when unbound inhibitor concentration ([I]u) was used; accuracy dropped when total inhibitor concentration ([I]t) was used. For bosentan, AFE was 1.2-1.3 using either [I]u or [I]t This difference was evaluated by sensitivity analysis of the cellular unbound fraction of inhibitor (fu,cell,inhibitor), which revealed higher sensitivity of fu,cell,inhibitor for predicting TCA Ct,Cells when inhibitors exhibited larger ([I]t,cell/IC50) values. In conclusion, this study demonstrated the applicability of a framework to predict hepatocellular bile acid concentrations due to drug-mediated inhibition of transporters using mechanistic modeling and cytosolic or cellular unbound concentrations., (Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.)

In our previous work, we identified a natural toxin, biliatresone, from Dysphania glomulifera and D. littoralis, endemic plants associated with outbreaks of biliary atresia in Australian neonatal livestock. Biliatresone is a very rare isoflavonoid with an α-methylene ketone between two phenyls, 1,2-diaryl-2-propenone, along with methylenedioxy, dimethoxyl, and hydroxyl functional groups, that causes extrahepatic biliary toxicity in zebrafish. The toxic core of biliatresone is a methylene in the α-position relative to the ketone of 1,2-diaryl-2-propenone that serves as an electrophilic Michael acceptor. The α-methylene of biliatresone spontaneously conjugated with water and methanol (MeOH), respectively, via Michael addition in a reverse phase high-performance liquid chromatography (RP-HPLC) analysis. We here report the reactivity of biliatresone toward glutathione (GSH), several amino acids, and other thiol- or imidazole-containing biomolecules. LC-MS and HPLC analysis of the conjugation reaction showed the reactivity of biliatresone to be in the order histidine > N-acetyl-d-cysteine (D-NAC) = N-acetyl-l-cysteine (L-NAC) > histamine > glutathione ≥ cysteine ≫ glycine > glutamate > phenylalanine, while serine and adenine had no reactivity due to intramolecular hydrogen bonding in the protic solvents. The reactivity of ethyl vinyl ketone (EVK, 1-penten-3-one), an example of a highly reactive α,ß-unsaturated ketone, toward GSH gave a 6.7-fold lower reaction rate constant than that of biliatresone. The reaction rate constant of synthetic 1,2-diaryl-2-propen-1-one (DP), a core structure of the toxic molecule, was 10-fold and 1.5-fold weaker in potency compared to the reaction rate constants of biliatresone and EVK, respectively. These results demostrated that the methylenedioxy, dimethoxyl, and hydroxyl functional groups of biliatresone contribute to the stronger reactivity of the Michael acceptor α-methylene ketone toward nucleophiles compared to that of DP and EVK.

Primary biliary cholangitis (PBC), previously known as primary biliary cirrhosis, is a model autoimmune disease with chronic cholestasis characterized by the hallmark of anti-mitochondrial antibodies and treated with ursodeoxycholic acid (UDCA). However, approximately 20-40% of patients incompletely respond to UDCA and have an increased risk of disease progression. Although there have been significant advances in the immunobiology of PBC, these have yet to be translated into newer therapeutic modalities. Current approaches to controlling the immune response include broad immunosuppression with corticosteroids as well as targeted therapies directed against T and B cells. In contrast, ameliorating cholestasis is the focus of other therapies in development, including obeticholic acid. In this article the authors will discuss ongoing clinical trials and, in particular, the rationale for choosing agents that may effectively target the aberrant immune response.

Fasiglifam (TAK-875), a selective G-protein-coupled receptor 40 agonist, was developed for the treatment of type 2 diabetes mellitus; however, its development was terminated in phase III clinical trials because of liver safety concerns. Our preliminary study indicated that intravenous administration of 100 mg/kg of TAK-875 increased the serum total bile acid concentration by 3 to 4 times and total bilirubin levels by 1.5 to 2.6 times in rats. In the present study, we examined the inhibitory effects of TAK-875 on hepatobiliary transporters to explore the mechanisms underlying its hepatotoxicity. TAK-875 decreased the biliary excretion index and the in vitro biliary clearance of d₈-taurocholic acid in sandwich-cultured rat hepatocytes, suggesting that TAK-875 impaired biliary excretion of bile acids, possibly by inhibiting bile salt export pump (Bsep). TAK-875 inhibited the efflux transporter multidrug resistance-associated protein 2 (Mrp2) in rat hepatocytes using 5 (and 6)-carboxy-2',7'-dichlorofluorescein as a substrate. Inhibition of MRP2 was further confirmed by reduced transport of vinblastine in Madin-Darby canine kidney cells overexpressing MRP2 with IC₅₀ values of 2.41 μM. TAK-875 also inhibited the major bile acid uptake transporter Na(+)/taurocholate cotransporting polypeptide (Ntcp), which transports d₈-taurocholic acid into rat hepatocytes, with an IC₅₀ value of 10.9 μM. TAK-875 significantly inhibited atorvastatin uptake in organic anion transporter protein (OATP) 1B1 and OATP1B3 cells with IC₅₀ values of 2.28 and 3.98 μM, respectively. These results indicate that TAK-875 inhibited the efflux transporter MRP2/Mrp2 and uptake transporters Ntcp and OATP/Oatp, which may affect bile acid and bilirubin homeostasis, resulting in hyperbilirubinemia and cholestatic hepatotoxicity., (Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.)

Previous studies suggest an interdependent relationship between liver and intestine for cholesterol elimination from the body. We hypothesized that a combination of ursodiol (Urso) and ezetimibe (EZ) could increase biliary secretion and reduce cholesterol reabsorption, respectively, to promote cholesterol excretion. Treatment with Urso increased hepatic ABCG5 ABCG8 (G5G8) protein and both biliary and fecal sterols in a dose-dependent manner. To determine whether the drug combination (Urso-EZ) further increased cholesterol excretion, mice were treated with Urso alone or in combination with two doses of EZ. EZ produced an additive and dose-dependent increase in fecal neutral sterol (FNS) elimination in the presence of Urso. Finally, we sequentially treated wide-type and G5G8-deficient mice with Urso and Urso-EZ to determine the extent to which these effects were G5G8 dependent. Although biliary and FNS were invariably lower in G5G8 KO mice, the relative increase in FNS following treatment with Urso alone or the Urso-EZ combination was not affected by genotype. In conclusion, Urso increases G5G8, biliary cholesterol secretion, and FNS and acts additively with EZ to promote fecal sterol excretion. However, the stimulatory effect of these agents was not G5G8 dependent., (Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.)

The cause of Cystic fibrosis liver disease (CFLD), is unknown. It is well recognized that hepatic exposure to hydrophobic bile salts is associated with the development of liver disease. For this reason, we hypothesize that, CFTR dependent variations, in the hepatic handling of hydrophobic bile salts, are related to the development CFLD. To test our hypothesis we studied, in Cftr-/- and control mice, bile production, bile composition and liver pathology, in normal feeding condition and during cholate exposure, either acute (intravenous) or chronic (three weeks via the diet). In Cftr-/- and control mice the basal bile production was comparable. Intravenous taurocholate increased bile production to the same extent in Cftr-/- and control mice. However, chronic cholate exposure increased the bile flow significantly less in Cftr-/- mice than in controls, together with significantly higher biliary bile salt concentration in Cftr-/- mice. Prolonged cholate exposure, however, did not induce CFLD like pathology in Cftr-/- mice. Chronic cholate exposure did induce a significant increase in liver mass in controls that was absent in Cftr-/- mice. Chronic cholate administration induces a cystic fibrosis-specific hepatobiliary phenotype, including changes in bile composition. These changes could not be associated with CFLD like pathological changes in CF mouse livers. However, chronic cholate administration induces liver growth in controls that is absent in Cftr-/- mice. Our findings point to an impaired adaptive homeotrophic liver response to prolonged hydrophobic bile salt exposure in CF conditions.

NSC-743380 (1-[(3-chlorophenyl)-methyl]-1H-indole-3-carbinol) is in early stages of development as an anticancer agent. Two metabolites reflect sequential conversion of the carbinol functionality to a carboxaldehyde and the major metabolite, 1-[(3-chlorophenyl)-methyl]-1H-indole-3-carboxylic acid. In an exploratory toxicity study in rats, NSC-743380 induced elevations in liver-associated serum enzymes and biliary hyperplasia. Biliary hyperplasia was observed 2 days after dosing orally for 2 consecutive days at 100mg/kg/day. Notably, hepatotoxicity and biliary hyperplasia were observed after oral administration of the parent compound, but not when major metabolites were administered. The toxicities of a structurally similar but pharmacologically inactive molecule and a structurally diverse molecule with a similar efficacy profile in killing cancer cells in vitro were compared to NSC-743380 to explore scaffold versus target-mediated toxicity. Following two oral doses of 100mg/kg/day given once daily on two consecutive days, the structurally unrelated active compound produced hepatic toxicity similar to NSC-743380. The structurally similar inactive compound did not, but, lower exposures were achieved. The weight of evidence implies that the hepatotoxicity associated with NSC-743380 is related to the anticancer activity of the parent molecule. Furthermore, because biliary hyperplasia represents an unmanageable and non-monitorable adverse effect in clinical settings, this model may provide an opportunity for investigators to use a short-duration study design to explore biomarkers of biliary hyperplasia., (Published by Elsevier Inc.)

The dysregulation of phosphatidylinositol 3-kinase (PI3K)-dependent pathways is implicated in several human cancers making it an attractive target for small molecule PI3K inhibitors. A series of potent pyridyltriazine-containing inhibitors of class Ia PI3Ks were synthesized and a subset of compounds was evaluated in exploratory repeat-dose rat toxicology studies. Daily oral dosing of compound 1: in Sprague Dawley rats for four consecutive days was associated with hepatobiliary toxicity that included biliary epithelial hyperplasia and hypertrophy, periductular edema, biliary stasis, and acute peribiliary inflammatory infiltrates. These histological changes were associated with clinical pathology changes that included increased serum liver enzymes, total bile acids, and bilirubin. The predominant clearance pathway of 1: was shown in vitro and in a bile-duct cannulated rat (14)C-ADME study to be P450-mediated oxidative metabolism. An O-demethylated pyridine metabolite, M3: , was identified as a candidate proximal metabolite that caused the hepatotoxicity. Co-administration of the pan-P450 inhibitor 1-aminobenzotriazole with 1: to rats significantly reduced the formation of M3: and prevented liver toxicity, whereas direct administration of M3: reproduced the toxicity. Structural changes were introduced to 1: to make the methoxypyridine ring less susceptible to P450 oxidation (compound 2: ), and addition of a methyl group to the benzylic carbon (compound 3: ) improved the pharmacokinetic profile. These changes culminated in the successful design of a clinical candidate 3: (AMG 511) that was devoid of liver toxicity in a 14-day rat toxicity study. Herein, we describe how a metabolism-based structure-activity relationship analysis allowed for the successful identification of a PI3K inhibitor devoid of off-target toxicity., (© The Author 2014. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Analysis of exploratory behavior of the offspring of female rats with chronic alcohol-induced injury to the hepatobiliary system demonstrates reduced exploratory activity in the object recognition test. Results of the test with a novel object confirm these findings and suggest that the offspring of alcoholized animals showed increased anxiety at the visual contact with the novel object due to impaired memory expressed in the development of short-term memory.

Because apoB-containing lipoproteins are pro-atherogenic and their secretion by liver and intestine largely depends on microsomal triglyceride transfer protein (MTP) activity, MTP inhibition strategies are actively pursued. How decreasing the secretion of apoB-containing lipoproteins affects intracellular rerouting of cholesterol is unclear. Therefore, the aim of the present study was to determine the effects of reducing either systemic or liver-specific MTP activity on cholesterol metabolism and reverse cholesterol transport (RCT) using a pharmacological MTP inhibitor or a genetic model, respectively. Plasma total cholesterol and triglyceride levels were decreased in both MTP inhibitor-treated and liver-specific MTP knockout (L-Mttp(-/-)) mice (each P < 0.001). With both inhibition approaches, hepatic cholesterol as well as triglyceride content was consistently increased (each P < 0.001), while biliary cholesterol and bile acid secretion remained unchanged. A small but significant decrease in fecal bile acid excretion was observed in inhibitor-treated mice (P < 0.05), whereas fecal neutral sterol excretion was substantially increased by 75% (P < 0.001), conceivably due to decreased intestinal absorption. In contrast, in L-Mttp(-/-) mice both fecal neutral sterol and bile acid excretion remained unchanged. However, while total RCT increased in inhibitor-treated mice (P < 0.01), it surprisingly decreased in L-Mttp(-/-) mice (P < 0.05). These data demonstrate that: i) pharmacological MTP inhibition increases RCT, an effect that might provide additional clinical benefit of MTP inhibitors; and ii) decreasing hepatic MTP decreases RCT, pointing toward a potential contribution of hepatocyte-derived VLDLs to RCT.

Breast cancer resistance protein (BCRP) and multidrug resistance-associated protein 2 (MRP2) are members of the ATP binding cassette (ABC) transporter family located in the canalicular membrane of hepatocytes that mediate biliary excretion of many drugs and endogenous compounds. BCRP and MRP2 have overlapping substrate profiles. Predicting drug disposition in the setting of altered transport function has important clinical significance. This investigation was designed to establish an in vitro model system to evaluate the impact of impaired Mrp2 and Bcrp function on hepatobiliary drug disposition. To achieve Bcrp knockdown by RNA interference (RNAi), sandwich-cultured hepatocytes (SCH) from Mrp2-deficient (TR(-)) and wild-type (WT) rats were infected with adenoviral vectors to express shRNA targeting Bcrp (Ad-siBcrp) at multiplicity of infection (MOI) of 1-10. MOI of 5 was identified as optimal. At MOI of 5, viral infection as well as WT or TR(-) status was statistically significant predictors of the rosuvastatin (RSV) biliary excretion index (BEI), consistent with the known role of Bcrp and Mrp2 in the biliary excretion of RSV in vivo in rats. Relative to WT rat SCH, marginal mean BEI (%) of RSV in TR(-) rat SCH decreased by 28.6 (95% CI: 5.8-51.3). Ad-siBcrp decreased marginal mean BEI (%) of RSV by 13.3 (7.5-9.1) relative to SCH infected with adenoviral vectors expressing a nontargeting shRNA (Ad-siNT). The BEI of RSV was almost ablated in TR(-) rat SCH with Bcrp knockdown (5.9 ± 3.0%) compared to Ad-siNT-infected WT rat SCH (45.4 ± 6.6%). These results demonstrated the feasibility of Bcrp knockdown in TR(-) rat SCH as an in vitro system to assess the impact of impaired Bcrp and Mrp2 function. At MOI of 5, viral infection had minimal effects on RSV total accumulation, but significantly decreased marginal mean taurocholate total accumulation (pmol/mg of protein) and BEI (%) by 9.9 (7.0-12.8) and 7.5 (3.7-11.3), respectively, relative to noninfected SCH. These findings may be due to off-target effects on hepatic bile acid transporters, even though no changes in protein expression levels of the hepatic bile acid transporters were observed. This study established a strategy for optimization of the knockdown system, and demonstrated the potential use of RNAi in SCH as an in vitro tool to predict altered hepatobiliary drug disposition when canalicular transporters are impaired.

Unlabelled: Multidrug resistance transporter 3/ATP-binding cassette protein subfamily B4 (MDR3/ABCB4) is a critical determinant of biliary phosphatidylcholine (PC) secretion. Clinically, mutations and partial deficiencies in MDR3 result in cholestatic liver injury. Thus, MDR3 is a potential therapeutic target for cholestatic liver disease. Fenofibrate is a peroxisome proliferator-activated receptor (PPAR) α ligand that has antiinflammatory actions and regulates bile acid detoxification. Here we examined the mechanism by which fenofibrate regulates MDR3 gene expression. Fenofibrate significantly up-regulated MDR3 messenger RNA (mRNA) and protein expression in primary cultured human hepatocytes, and stimulated MDR3 promoter activity in HepG2 cells. In silico analysis of 5'-upstream region of human MDR3 gene revealed a number of PPARα response elements (PPRE). Electrophoretic mobility shift (EMSA) and chromatin immunoprecipitation (ChIP) assays demonstrated specific binding of PPARα to the human MDR3 promoter. Targeted mutagenesis of three novel PPREs reduced inducibility of the MDR3 promoter by fenofibrate. In collagen sandwich cultured rat hepatocytes, treatment with fenofibrate increased secretion of fluorescent PC into bile canaliculi., Conclusion: Fenofibrate transactivates MDR3 gene transcription by way of the binding of PPARα to three novel and functionally critical PPREs in the MDR3 promoter. Fenofibrate treatment further stimulates biliary phosphatidylcholine secretion in rat hepatocytes, thereby providing a functional correlate. We have established a molecular mechanism that may contribute to the beneficial use of fenofibrate therapy in human cholestatic liver disease., (© 2014 by the American Association for the Study of Liver Diseases.)

Background: Livers derived from donation after circulatory death (DCD) are increasingly accepted for transplantation. However, DCD livers suffer additional donor warm ischemia, leading to biliary injury and more biliary complications after transplantation. It is unknown whether oxygenated machine perfusion results in better preservation of biliary epithelium and the peribiliary vasculature. We compared oxygenated hypothermic machine perfusion (HMP) with static cold storage (SCS) in a porcine DCD model., Methods: After 30 min of cardiac arrest, livers were perfused in situ with HTK solution (4°C) and preserved for 4 h by either SCS (n = 9) or oxygenated HMP (10°C; n = 9), using pressure-controlled arterial and portal venous perfusion. To simulate transplantation, livers were reperfused ex vivo at 37°C with oxygenated autologous blood. Bile duct injury and function were determined by biochemical and molecular markers, and a systematic histological scoring system., Results: After reperfusion, arterial flow was higher in the HMP group, compared to SCS (251±28 vs 166±28 mL/min, respectively, after 1 hour of reperfusion; p = 0.003). Release of hepatocellular enzymes was significantly higher in the SCS group. Markers of biliary epithelial injury (biliary LDH, gamma-GT) and function (biliary pH and bicarbonate, and biliary transporter expression) were similar in the two groups. However, histology of bile ducts revealed significantly less arteriolonecrosis of the peribiliary vascular plexus in HMP preserved livers (>50% arteriolonecrosis was observed in 7 bile ducts of the SCS preserved livers versus only 1 bile duct of the HMP preserved livers; p = 0.024)., Conclusions: Oxygenated HMP prevents arteriolonecrosis of the peribiliary vascular plexus of the bile ducts of DCD pig livers and results in higher arterial flow after reperfusion. Together this may contribute to better perfusion of the bile ducts, providing a potential advantage in the post-ischemic recovery of bile ducts.

Experience of treatment of 17 patients, suffering cholangitis of various genesis, using antiseptic Decasan, is presented. Clinical efficacy of the preparation in complex treatment of cholangitis, confirmed by results of the bile bacteriological investigation, was noted.

Background and Aim: Functional disorders of the biliary tract involve gallbladder and sphincter of Oddi and cause pain and/or digestive troubles. In this context, in addition to pharmacological treatments, an important role is played by the use of sodium-sulphate and sulphate-bicarbonate mineral waters that, because of their composition into ions macro and trace elements, can stimulate the release or modulate the activity of some neurohumoral regulators of the digestive process. We want to do a study on the effects of hydropinotherapy with a sulphate-bicarbonate-calcium-magnesium mineral water in patients suffering from pain and other symptoms caused by biliary dyskinesias, biliary sand (without gallstones), or following a cholecystectomy (post-cholecystectomy syndromes)., Materials and Methods: We enrolled 43 patients suffering from those affections; all the patients did two cycles in one year of hydropinotherapy with Acqua Santa at Italy's Chianciano Spa; 20 of these patients did a third cycle of hydropinotherapy in the second year of the study. At the end of the second and of the third cycle we compared the frequency of eleven main symptoms in both groups and we also performed an longitudinal-observational study on the frequency of those symptoms before the beginning of the first cycle of the therapy and at the end of the second and of the third cycle. Statistical analyses were based on the use of Pearson's χ2 test., Results: The frequency of the symptoms observed at the end of second and third cicle of hydropinotherapy was significantly lower than that considered before starting therapy. The differences were statistically significant., Conclusions: The results of our research regarding the hydropinotherapy by sulphate-bicarbonate-calcium-magnesium mineral water show a significant improvement of symptoms in patients suffering from disturbances of biliary tract.

Efforts are put into developing toxicogenomics-based toxicity testing methods using in vitro human cell models for improving human risk assessment/replacing animal models. Human in vitro liver models include HepG2, HepaRG and primary human hepatocytes (PHH). Studies on comparability/applicability of these cell types mainly focus on assessing baseline biotransformation capacities/cytochrome P450-inducibility, but compound-induced gene expression profiles are at least as important. Therefore, we compared baseline and aflatoxin B1- and benzo(α)pyrene-induced gene expression profiles in HepG2, HepaRG and PHH (11-13 donors). At baseline, all liver models differ from each other with respect to whole genome gene expression levels. PHH show profound inter-individual differences, and are most similar to HepaRG. After compound exposure, induced gene expression profiles are more similar between cell models, especially for benzo(α)pyrene. Pathways involved in compound metabolism are induced in all 3 models, while others are more pronounced in a specific cell model. Examples are transcriptomic modifications of carbohydrate-related genes (HepaRG) and of receptor-related genes (PHH) after benzo(α)pyrene exposure, and of cell cycle-related genes (HepG2) after aflatoxin B1 exposure. PHH gene expression responses are the most heterogeneous. In conclusion, at base line level PHH are more similar to HepaRG than to HepG2, but for toxicogenomics applications both cell lines perform equally well in comparison to PHH., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Unlabelled: Biliary lipid secretion plays an important role in gallstone disease and reverse cholesterol transport (RCT). Using Sr-bI/Abcg5 double knockout mice (dko), the present study investigated the differential contribution of two of the most relevant transporters: adenosine triphosphate (ATP)-binding cassette subfamily G member 5 and 8 (ABCG5/G8) and scavenger receptor class B type I (SR-BI) to sterol metabolism and RCT. Plasma cholesterol levels increased in the following order, mainly due to differences in high density lipoprotein (HDL): Abcg5 ko < wild type < Sr-bI/Abcg5 dko < Sr-bI ko. Liver cholesterol content was elevated in Sr-bI ko only (P < 0.05). In Sr-bI/Abcg5 dko plasma plant sterols were highest, while hepatic plant sterols were lower compared with Abcg5 ko (P < 0.05). Under baseline conditions, biliary cholesterol secretion rates decreased in the following order: wild type > Sr-bI ko (-16%) > Abcg5 ko (-75%) > Sr-bI/Abcg5 dko (-94%), all at least P < 0.05, while biliary bile acid secretion did not differ between groups. However, under supraphysiological conditions, upon infusion with increasing amounts of the bile salt tauroursodeoxycholic acid, Abcg5 became fully rate-limiting for biliary cholesterol secretion. Additional in vivo macrophage-to-feces RCT studies demonstrated an almost 50% decrease in overall RCT in Sr-bI/Abcg5 dko compared with Abcg5 ko mice (P < 0.01)., Conclusion: These data demonstrate that (1) SR-BI contributes to ABCG5/G8-independent biliary cholesterol secretion under basal conditions; (2) biliary cholesterol mass secretion under maximal bile salt-stimulated conditions is fully dependent on ABCG5/G8; and (3) Sr-bI contributes to macrophage-to-feces RCT independent of Abcg5/g8., (Copyright © 2013 American Association for the Study of Liver Diseases.)