Dysregulation of serum bile acids and FGF19 in alcoholic hepatitis

Abstract

Background & aims: The degree of cholestasis is an important disease driver in alcoholic hepatitis, a severe clinical condition that needs new biomarkers and targeted therapies. We aimed to identify the largely unknown mechanisms and biomarkers linked to cholestasis in alcoholic hepatitis.

Methods: Herein, we analyzed a well characterized cohort of patients with alcoholic hepatitis and correlated clinical and histological parameters and outcomes with serum bile acids and fibroblast growth factor 19 (FGF19), a major regulator of bile acid synthesis.

Results: We found that total and conjugated bile acids were significantly increased in patients with alcoholic hepatitis compared with controls. Serum FGF19 levels were strongly increased and gene expression of FGF19 was induced in biliary epithelial cells and ductular cells of patients with alcoholic hepatitis. De novo bile acid synthesis (CYP7A1 gene expression and C4 serum levels) was significantly decreased in patients with alcoholic hepatitis. Importantly, total and conjugated bile acids correlated positively with FGF19 and with disease severity (model for end-stage liver disease score). FGF19 correlated best with conjugated cholic acid, and model for end-stage liver disease score best with taurine-conjugated chenodeoxycholic acid. Univariate analysis demonstrated significant associations between FGF19 and bilirubin as well as gamma glutamyl transferase, and negative correlations between FGF19 and fibrosis stage as well as polymorphonuclear leukocyte infiltration, in all patients with alcoholic hepatitis.

Conclusion: Serum FGF19 and bile acids are significantly increased in patients with alcoholic hepatitis, while de novo bile acid synthesis is suppressed. Modulation of bile acid metabolism or signaling could represent a promising target for treatment of alcoholic hepatitis in humans.

Lay summary: Understanding the underlying mechanisms that drive alcoholic hepatitis is important for the development of new biomarkers and targeted therapies. Herein, we describe a molecule that is increased in patients with alcoholic hepatitis. Modulating the molecular pathway of this molecule might lead to promising targets for the treatment of alcoholic hepatitis.

Keywords: Bile acids; FGF19; Microbiome.

Figure 1. Patients with alcoholic hepatitis have significantly higher serum levels of total and conjugated bile acids than controls and patients with alcohol use disorder

Sera of 15 patients with alcoholic hepatitis, 9 patients with alcohol use disorder and 9 control subjects were analyzed. (A) Total bile acids. (B) Conjugated bile acids. (C) Taurine-conjugated bile acids. (D) Glycine-conjugated bile acids. (E) Unconjugated bile acids. (F) Bile acid composition. Only bile acids with contribution of > 2% in healthy subjects are depicted. One-Way ANOVA with Bonferroni post-test. ***P < 0.001. BA, bile acid; CA, cholic acid; CDCA, chenodeoxycholic acid; DCA, deoxycholic acid; G, glycine; HDCA, hyodeoxycholic acid; T, taurine; UDCA, ursodeoxycholic acid; LCA, lithocholic acid.

Figure 2. Patients with alcoholic hepatitis show significantly elevated serum levels of FGF19 and significantly suppressed de novo bile acid synthesis relative to controls and patients with alcohol use disorder

Sera of 15 patients with alcoholic hepatitis, 8–9 patients with alcohol use disorder and 9 control subjects were analyzed. (A) FGF19 concentrations. (B) C4 levels, a marker for de novo bile acid synthesis. One-Way ANOVA with Bonferroni post-test. *P < 0.05, **P < 0.01, ***P < 0.001. C4, 7-alpha-hydroxy-4-cholesten-3-one; FGF19, fibroblast growth factor 19.

Figure 3. Severe forms of alcoholic hepatitis are associated with marked hepatic FGF19 production, down-regulated hepatic bile acid uptake and synthesis, as well as induced bile acid efflux

Hepatic gene expression was quantitated in 12 patients with early alcoholic steatohepatitis, 11 patients with non-severe alcoholic hepatitis, 18 patients with severe alcoholic hepatitis (responders and non-responders), 10 with “terminal” alcoholic hepatitis (with liver failure and early transplantation), and 10 controls with non-diseased livers. (A) FGF19. (B) Bile acid uptake transporter NTCP (SLC10A1). (C) CYP7A1. (D) CYP8B1. (E) BSEP (ABCB11) important for bile acid excretion into bile canaliculi. (F – G) Transporters for bile acid efflux MRP4 (ABCC4; F) and MRP3 (ABCC3; G). One-Way ANOVA with Bonferroni post-test. *P < 0.05, **P < 0.01, ***P < 0.001. ABCB11, ATP-binding cassette, sub-family B member 11; ABCC3/4, ATP-binding cassette, sub-family C member 3/4; BSEP, bile salt export pump; CYP7A1, cytochrome P450 family 7 subfamily A member 1; CYP8B1, cytochrome P450 family 8 subfamily B member 1; FGF19, fibroblast growth factor 19; MRP3/4, multidrug resistance-associated protein 3/4; NTCP, Na+-taurocholate co-transporting polypeptide; SLC10A1, solute carrier family 10 member 1; TPM, transcripts per million.

Figure 4. FGF19 immunohistochemistry in liver biopsies from patients with alcoholic hepatitis

(A – C) Alcoholic hepatitis. (A) Intense staining in a large bile duct and moderate staining in bile ductules (magnification ×200). (B) Moderate staining in bile ductules (magnification ×400). (C) Intense staining in non-parenchymal cells in the sinusoids (magnification ×400). (D) Gallbladder specimen as positive control (magnification ×200). (E) Non-neoplastic liver tissue adjacent to resected HCC as negative control (magnification ×200). (F) Large caliber bile ducts in normal liver tissue adjacent to resected HCC stained positive for FGF19 (magnification ×100). FGF19, fibroblast growth factor 19; HCC, hepatocellular carcinoma.

Figure 5. Association of MELD score and FGF19 with several variables or clinical parameters

(A) MELD score and T-CDCA serum levels are similarly elevated in alcoholic hepatitis, and have a strong positive correlation. MELD scores and T-CDCA serum concentrations were analyzed from all patients in our cohorts with determinable MELD scores and T-CDCA serum concentrations (n=25). (B) Serum FGF19 levels were significantly higher in alcoholic hepatitis patients with above average serum bilirubin than in alcoholic hepatitis patients with below average serum bilirubin (mean was 16 mg/dl, n=128). (C) Serum FGF19 is significantly higher in alcoholic hepatitis patients with above average serum GGT (mean was 495 mg/dl, n= 61). (D) Serum FGF19 increases distinctly when the contribution of conjugated bile acids to the total bile acid content approaches 100%. FGF19 values from 15 patients with alcoholic hepatitis, 8 patients with alcohol use disorder and 9 control subjects, and their respective % conjugated bile acids were analyzed. FGF19, fibroblast growth factor 19; GGT, gamma-glutamyl-transferase; MELD, Model for End-stage Liver Disease; T-CDCA, taurine-conjugated chenodesoxycholic acid. **P < 0.01.