Protective potential of the gallbladder in primary sclerosing cholangitis

Abstract

Background & aims: Gallbladder enlargement is common in patients with primary sclerosing cholangitis (PSC). The gallbladder may confer hepatoprotection against bile acid overload, through the sequestration and cholecystohepatic shunt of bile acids. The aim of this study was to assess the potential impact of the gallbladder on disease features and bile acid homeostasis in PSC.

Methods: Patients with PSC from a single tertiary center who underwent liver MRI with three-dimensional cholangiography and concomitant analyses of serum bile acids were included. Gallbladder volume was measured by MRI and a cut-off of 50 ml was used to define gallbladder enlargement. Bile acid profiles and PSC severity, as assessed by blood tests and MRI features, were compared among patients according to gallbladder size (enlarged vs. normal-sized) or presence (removed vs. conserved). The impact of cholecystectomy was also assessed in the Abcb4 knockout mouse model of PSC.

Results: Sixty-one patients with PSC, all treated with ursodeoxycholic acid (UDCA), were included. The gallbladder was enlarged in 30 patients, whereas 11 patients had been previously cholecystectomized. Patients with enlarged gallbladders had significantly lower alkaline phosphatase, a lower tauro-vs. glycoconjugate ratio and a higher UDCA vs. total bile acid ratio compared to those with normal-sized gallbladders. In addition, gallbladder volume negatively correlated with the hydrophobicity index of bile acids. Cholecystectomized patients displayed significantly higher aspartate aminotransferase and more severe bile duct strictures and dilatations compared to those with conserved gallbladder. In the Abcb4 knockout mice, cholecystectomy caused an increase in hepatic bile acid content and in circulating secondary bile acids, and an aggravation in cholangitis, inflammation and liver fibrosis.

Conclusion: Altogether, our findings indicate that the gallbladder fulfills protective functions in PSC.

Impact and implications: In patients with primary sclerosing cholangitis (PSC), gallbladder status impacts on bile acid homeostasis and disease features. We found evidence of lessened bile acid toxicity in patients with PSC and enlarged gallbladders and of increased disease severity in those who were previously cholecystectomized. In the Abcb4 knockout mouse model of PSC, cholecystectomy causes an aggravation of cholangitis and liver fibrosis. Overall, our results suggest that the gallbladder plays a protective role in PSC.

Keywords: ABC, ATP-binding cassette transporter; Abcb4 knockout mice; BA, bile acid; Bile acids; C4, 7α-hydroxy-4-cholesten-3-one; CFTR, cystic fibrosis transmembrane conductance regulator; CK19, cytokeratin 19; Cholecystectomy; FGF19, fibroblast growth factor 19; Gallbladder volume; HPLC-MS/MS, high-performance liquid chromatography coupled to tandem mass spectrometry; IBD, inflammatory bowel disease; MRC, magnetic resonance cholangiography; Magnetic resonance imaging; PSC, primary sclerosing cholangitis; UDCA, ursodeoxycholic acid; ULN, upper limit of normal.

Impact and implications: In patients with primary sclerosing cholangitis (PSC), gallbladder status impacts on bile acid homeostasis and disease features. We found evidence of lessened bile acid toxicity in patients with PSC and enlarged gallbladders and of increased disease severity in those who were previously cholecystectomized. In the Abcb4 knockout mouse model of PSC, cholecystectomy causes an aggravation of cholangitis and liver fibrosis. Overall, our results suggest that the gallbladder plays a protective role in PSC.

Fig. 1

Gallbladder enlargement and related cholestatic profile in patients with PSC. (A) Gallbladder volume was measured on MRI in patients with PSC and conserved gallbladders, who were subsequently separated into two groups with enlarged (≥50 ml, n = 30) or normal-sized (<50 ml, n = 20) gallbladders, herein illustrated by gallbladder volumes of 11 and 103 ml, respectively. (B-D) Graphs showing the distribution of gallbladder volume (B), serum alkaline phosphatase levels (C), tauroconjugates/total BA ratios, Tauro-/glycoconjugate ratios and UDCA/total BA ratios (D), in the two groups. Levels of significance are indicated in the figure (Mann-Whitney test). A p value of less than 0.05 was considered significant.

Fig. 2

Impact of cholecystectomy on PSC features in patients. Comparison of (A) aspartate aminotransferase, (B–C) cholangiographic features at MRI including representative images (B) and quantitative analyses (C), in patients with PSC whose gallbladders were conserved (n = 50) or previously removed (n = 11). Levels of significance are indicated in the figure (Mann-Whitney test for A; Chi-squared test for C). A p value of less than 0.05 was considered significant. CBD, common bile duct; LHD, left hepatic duct; RHD, right hepatic duct.

Fig. 3

Impact of cholecystectomy on phenotypic traits of PSC in Abcb4^-/- mice. Liver samples were collected from Sham-operated (n = 7) or cholecystectomized (n = 6) Abcb4^-/- mice, 5 weeks after surgery, for (A) histological analysis of H&E-stained tissue sections (left panel, cholangitis characterized by periductal inflammation [arrows] and onion-skin fibrosis [arrowheads]), according to an adaptation of Nakanuma score (cholangitis activity, fibrosis score and bile duct loss were evaluated using a 0 to 3 scale (right panel, hepatitis activity=0 in all samples, not shown); (B) analysis of ductular reaction, inflammation and fibrosis, assessed by CK-19, F4/80 and Sirius red staining, respectively; (C) RT-qPCR analysis of proinflammatory (Tnfα) and profibrotic factor (Col1a1, Acta2) expression. Data were normalized for Hprt1 and expressed relative to a pool of hepatic mRNA from WT mice. Levels of significance are indicated in the figure (Student’s t test). A p value of less than 0.05 was considered significant. Scale bars = 250 μm for H&E and F4/80; 500 μm for CK-19, 1 mm for Sirius red. RT-qPCR, quantitative reverse-transcription PCR; WT, wild-type.

Fig. 4

Impact of cholecystectomy on bile acid metabolism in Abcb4^-/- mice. Liver and plasma samples were collected from Sham-operated (n = 7) or cholecystectomized (n = 6) Abcb4^-/- mice, 5 weeks after surgery, for (A) RT-qPCR analysis of hepatic expression of genes involved in bile acid homeostasis (Data were normalized for Hprt1 and expressed relative to a pool of hepatic mRNA from WT mice); (B–C) HPLC-MS/MS analysis of total bile acids (B) and individual bile acid species (C) in liver and plasma. Levels of significance are indicated in the figure (Student’s t test). A p value of less than 0.05 was considered significant. RT-qPCR, quantitative reverse-transcription PCR; WT, wild-type.