Up-regulation of microRNA 506 leads to decreased Cl−/HCO3− anion exchanger 2 expression in biliary epithelium of patients with primary biliary cirrhosis

Abstract

Cl−/HCO3−anion exchanger 2 (AE2) participates in intracellular pH homeostasis and secretin-stimulated biliary bicarbonate secretion. AE2/SLC4A2 gene expression is reduced in liver and blood mononuclear cells from patients with primary biliary cirrhosis (PBC). Our previous findings of hepatic and immunological features mimicking PBC in Ae2-deficient mice strongly suggest that decreased AE2 expression might be involved in the pathogenesis of PBC. Here we tested the potential role of hsa-microRNA 506 (miR-506) – predicted as candidate to target AE2 mRNA – for the decreased expression of AE2 in PBC. Real-time qPCR showed that miR-506 expression is increased in PBC livers versus normal liver specimens. In situ hybridization in liver sections confirmed that miR-506 is upregulated in the intrahepatic bile ducts of PBC livers compared with normal and primary-sclerosing-cholangitis livers. Precursor-mediated overexpression of miR-506 in SV40-immortalized normal human cholangiocytes (H69 cells) led to decreased AE2 protein expression and activity, as indicated by immunoblotting and microfluorimetry, respectively. Moreover, miR-506 overexpression in 3D-cultured H69 cholangiocytes blocked the secretin-stimulated expansion of cystic structures developed under the three-dimensional conditions. Luciferase assays and site-directed mutagenesis demonstrated that miR-506 specifically may bind the 3’UTR region of AE2 mRNA and prevent protein translation. Finally, cultured PBC cholangiocytes showed decreased AE2 activity together with miR-506 overexpression compared to normal human cholangiocytes, and, transfection of PBC cholangiocytes with anti-miR-506 was able to improve their AE2 activity. Conclusion miR-506 is upregulated in cholangiocytes from PBC patients, binds the 3’UTR region of AE2 mRNA and prevents protein translation, leading to diminished AE2 activity and impaired biliary secretory functions. In view of the putative pathogenic role of decreased AE2 in PBC, miR-506 may constitute a potential therapeutic target for this disease.

Bile Acids and Deregulated Cholangiocyte Autophagy in Primary Biliary Cholangitis

“…Interestingly, cholangiocytes in PBC patients show suppressed anion exchanger (AE2; SLC4A2), which is possibly caused by upregulated micoRNA-506, which result in impaired biliary HCO 3 -formation [44] . This may cause impaired 'biliary HCO 3 -umbrella,' which protects cholangiocytes against the toxic effects of human hydrophobic bile acids [44] . GCDC is a major hydrophobic bile acid in cholestasis and impaired umbrella may result in GCDC-induced cholangiocyte damage [3,4] .…”

MicroRNAs in liver tissue engineering — New promises for failing organs

“…One interesting finding has been reported by Banales et al, who observed upregulation of miR-506 in primary biliary cirrhosis. The authors show that miR-506 regulates expression of the Cl − /HCO3 − anion exchanger 2 (AE2) and that suppression of miR-506 leads to improved AE2 function in primary biliary cirrhosis cholangiocytes [85]. In addition, Glaser et al reported that, after hepatic injury, secretin produced by cholangiocytes and S cells reduced miR125b and let7a levels, resulting in up-regulation of VEGF and NGF.…”

Epigenetics and Primary Biliary Cirrhosis: a Comprehensive Review and Implications for Autoimmunity

“…PBC patients have significantly decreased levels of AE2. Banales et al demonstrated that miR-506, upregulated in cholangiocytes from PBC patients, could bind the 3′ UTR of AE2 mRNA and prevent protein translation, leading to diminished AE2 activity and impaired biliary secretory functions [174]. Recently, Ananthanarayanan et al reported that miR-506 was also responsible for a decreased level of InsP 3 R3 [173], an intracellular Ca channel that is concentrated along the subapical portion of the endoplasmic reticulum where it regulates bicarbonate HCO 3 − and Cl− to support the functions of AE2.…”