Cl − /HCO 3 − 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.