Resumen de Papel de ae2 en la regulación del phi en colangiocitos de ratón
Background & Aims: Cl−/HCO3− anion exchanger 2 (AE2) is involved in intracellular pH (pHi) regulation and transepithelial acid-base transport, including secretin-stimulated biliary bicarbonate excretion. AE2 gene expression was found to be reduced in liver biopsies and blood mononuclear cells from patients with primary biliary cirrhosis (PBC), a disease characterized by chronic nonsuppurative cholangitis associated with antimitochondrial antibodies (AMA) and other autoimmune phenomena. AE2/Ae2 is the effector of bicarbonate secretion in human and rat cholangiocytes. Here we analyzed the role of Ae2 in the regulation of pHi in cholangiocytes isolated from normal and Ae2a,b¿deficient mice. Results: Initially, we isolated intrahepatic bile duct units (IBDUs) from both Ae2a,b+/+ and Ae2a,b¿/¿ mice which were then cultured on rat-tail collagen. By the 4th day in culture, IBDUs started to grow as cell clusters in a monolayer fashion. Cultures were released from fibroblast contamination by sequential dispase treatments during the first 3-4 cell-passages, when most cells tested as positive for cytokeratin-7 immunostaining. We used microfluorimetry to analyze pHi changes in response to acid or alkali loads and compared Ae2a,b¿/¿ cholangiocytes with the normal mouse cholangiocytes. Ae2a,b+/+ cholangiocytes exhibited a sodium-independent chloride/bicarbonate exchange activity which was absent in the Ae2a,b¿/¿ cholangiocytes, indicating that such an activity in normal cholangiocytes is mediated by Ae2. On the other hand, both Ae2a,b+/+ and Ae2a,b¿/¿ mouse cholangiocytes exhibit a chloride-independent bicarbonate secretion. Bile secretion agonists cAMP and ACh enhanced bicarbonate secretion in Ae2a,b+/+ but not in Ae2a,b¿/¿ cholangiocytes. Conclusions: Ae2a,b deficiency prevents genuine AE activity in mouse cholangiocytes. On the other hand, both Ae2a,b+/+ and Ae2a,b¿/¿ cholangiocytes possess the ability to acidify their pHi via a chloride-independent bicarbonate secretion mechanism.
