Adaptive Downregulation of Cl-/HCO3- Exchange Activity in Rat Hepatocytes under Experimental Obstructive Cholestasis

Abstract

In obstructive cholestasis, there is an integral adaptive response aimed to diminish the bile flow and minimize the injury of bile ducts caused by increased intraluminal pressure and harmful levels of bile salts and bilirrubin. Canalicular bicarbonate secretion, driven by the anion exchanger 2 (AE2), is an influential determinant of the canalicular bile salt-independent bile flow. In this work, we ascertained whether AE2 expression and/or activity is reduced in hepatocytes from rats with common bile duct ligation (BDL), as part of the adaptive response to cholestasis. After 4 days of BDL, we found that neither AE2 mRNA expression (measured by quantitative real-time PCR) nor total levels of AE2 protein (assessed by western blot) were modified in freshly isolated hepatocytes. However, BDL led to a decrease in the expression of AE2 protein in plasma membrane fraction as compared with SHAM control. Additionally, AE2 activity (J(OH)-, mmol/L/min), measured in primary cultured hepatocytes from BDL and SHAM rats, was decreased in the BDL group versus the control group (1.9 +/- 0.3 vs. 3.1 +/- 0.2, p<0.005). cAMP-stimulated AE2 activity, however, was not different between SHAM and BDL groups (3.7 +/- 0.3 vs. 3.5 +/- 0.3), suggesting that cAMP stimulated insertion into the canalicular membrane of AE2-containing intracellular vesicles, that had remained abnormally internalized after BDL. In conclusion, our results point to the existence of a novel adaptive mechanism in cholestasis aimed to reduce biliary pressure, in which AE2 internalization in hepatocytes might result in decreased canalicular HCO3- output and decreased bile flow.