β-Defensin 1 Is Prominent in the Liver and Induced During Cholestasis by Bilirubin and Bile Acids via Farnesoid X Receptor and Constitutive Androstane Receptor.

Thomas Klag,Jan Wehkamp,Nisar P Malek,Thomas S Weiss,Dirk Ehmann,Brigitte Vollmar,Christoph P Berg,Kerstin Abshagen,Maria Thomas,Eduard F Stange,Rania Dayoub,Wolfgang E Thasler,Ulrich M Zanger,Lioba Courth,Daniela Mailänder-Sanchez

doi:10.3389/fimmu.2018.01735

Abstract

Knowledge about innate antimicrobial defense of the liver is limited. We investigated hepatic expression and regulation of antimicrobial peptides with focus on the human beta defensin-1 (hBD-1). Radial diffusion assay was used to analyze antimicrobial activity of liver tissue. Different defensins including hBD-1 and its activator thioredoxin-1 (TXN) were analyzed in healthy and cholestatic liver samples by qPCR and immunostaining. Regulation of hBD-1 expression was studied in vitro and in vivo using bile duct-ligated mice. Regulation of hBD-1 via bilirubin and bile acids (BAs) was studied using siRNA. We found strong antimicrobial activity of liver tissue against Escherichia coli. As a potential mediator of this antimicrobial activity we detected high expression of hBD-1 and TXN in hepatocytes, whereas other defensins were minimally expressed. Using a specific antibody for the reduced, antimicrobially active form of hBD-1 we found hBD-1 in co-localization with TXN within hepatocytes. hBD-1 was upregulated in cholestasis in a graded fashion. In cholestatic mice hepatic AMP expression (Defb-1 and Hamp) was enhanced. Bilirubin and BAs were able to induce hBD-1 in hepatic cell cultures in vitro. Treatment with siRNA and/or agonists demonstrated that the farnesoid X receptor (FXR) mediates basal expression of hBD-1, whereas both constitutive androstane receptor (CAR) and FXR seem to be responsible for the induction of hBD-1 by bilirubin. hBD-1 is prominently expressed in hepatocytes. It is induced during cholestasis through bilirubin and BAs, mediated by CAR and especially FXR. Reduction by TXN activates hBD-1 to a potential key player in innate antimicrobial defense of the liver.

Highlights

There is growing evidence that bacterial translocation from the gut may contribute to progression of chronic liver damage leading to liver fibrosis through increased inflammation [1,2,3,4]
We found a strong inhibition of E. coli growth, using cationic protein fractions of liver homogenates, where Human beta defensin-1 (hBD-1) is cumulating, as shown by immunoblot analysis (Figure 1)
We performed a screening study to investigate the basal expression of several defensins in primary cultured human hepatocytes (PHH) (Figure S1 in Supplementary Material) and again hBD-1 mRNA was prominent while other β-defensins were only marginally expressed or absent

Summary

Introduction

There is growing evidence that bacterial translocation from the gut may contribute to progression of chronic liver damage leading to liver fibrosis through increased inflammation [1,2,3,4]. Once established, liver cirrhosis may lead to severe infectious complications like bacterial peritonitis and sepsis [7]. First line of defense against these commensals and pathogenic microorganisms is the epithelium with its mucus barrier, consisting of mucins and antimicrobial peptides (AMPs) like alpha-defensins from Paneth cells in the small intestine and β-defensins in the colon, inhibiting bacterial translocation to portal blood [8]. Human beta defensin-1 (hBD-1) is ubiquitous in most epithelia and in the colon, for example, it is regulated via nuclear receptors (NRs) like PPARγ [9]. We investigated hepatic expression and regulation of antimicrobial peptides with focus on the human beta defensin-1 (hBD-1)

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