Effect of Ischemia–Reperfusion on Na⁺,K⁺-ATPase Expression in Human Liver Tissue Allograft: Image Analysis by Confocal Laser Scanning Microscopy

Abstract

We have analyzed the effect of ischemia-reperfusion on expression of hepatic Na+,K+-ATPase on bile canalicular (BCM) and basolateral membranes (BLM) in human liver allografts using confocal laser scanning microscopy imaging. Na+, K+-ATPase, an integral membrane enzyme, plays a key role in the physiology and structure of hepatocytes, where it maintains the electrochemical gradients for Na+ and K+ across the cell membrane. The concentrations of these ions as well as their gradients regulate the active transport across the plasma membrane for bile acid and water from sinusoidal to canalicular membranes. In addition, Na+,K+-ATPase is also involved in cellular structure because of its close relationship with submembrane microfilaments and its implication in tight junction assembly. Therefore, Na+,K+-ATPase appears as an indicator of tissue viability and hepatic functionality during liver transplantation. Its localization and its function in BCM are still controversial. As in previous studies, we found an enzyme expression in both BLM and BCM. We show that ischemia induced a decrease in Na+,K+-ATPase expression only in BCM. This result could be explained by the differences in biochemical membrane environment between basolateral and bile canalicular Na+,K+-ATPase. Membrane lipid fluidity, which is more elevated in BLM than in BCM, could protect the enzyme during ischemia. After reperfusion, Na+,K+-ATPase expression was strongly decreased in both BCM and BLM. This alteration following reperfusion is probably due to multiple factors: direct alteration of the enzyme catalytic subunit and modification of its environment and membrane lipid fluidity by free radicals and changes in ATP levels and ionic distribution. This important decrease in Na+,K+-ATPase expression of both BLM and BCM could disturb not only hepatic secretory function but also cellular volume and structure during the postoperative period.

Proteomic Analysis of Ischemia-Reperfusion Injury upon Human Liver Transplantation Reveals the Protective Role of IQGAP1

“…After 1 h of reperfusion, F-actin concentrates only at the hepatocyte junctions, surrounding BC, which present disorganized and dilated structures (R60, inset, arrow). These observations are in agreement with previous morphological studies reporting Factin remodeling in human transplanted liver during the course of IRI (5,11). Under these circumstances, the alteration of F-actin structure correlates with the loss of hepatocyte junctions and BC integrity and significantly impacts hepatocyte bile secretion and function (5).…”

“…Treatment of rat liver with compounds that alter actin polymerization, such as cytochalasin B and phalloidin, leads to the alteration of BC structure (canaliculi dilatation and loss of microvilli) as well as contractile properties (8,9). In addition, a major actin cytoskeleton alteration around the hepatocyte cell membrane has been reported in human liver in response to IRI and has also been related to an impairment of BC contraction, tight junction permeability, and subsequent maintenance of a normal bile flow during the postoperative period (10,11). An intact actin network is therefore required for BC integrity to ensure proper bile secretion, i.e.…”

Selective intrarenal human A1 adenosine receptor overexpression reduces acute liver and kidney injury after hepatic ischemia reperfusion in mice

“…37,38 For example, F-actin disruption is a well known stimulus of apoptosis in several cell lines. 39 We saw severe disruption of F-actin in liver and kidney proximal tubules after liver IR in EGFP lentivirus-injected mice.…”

Monoclonal antibody 16D10 to the COOH-terminal domain of the feto-acinar pancreatic protein targets pancreatic neoplastic tissues

“…Image Processing Images were processed as described previously (27). For each primary antibody, the staining was calculated as the ratio between the total fluorescence of the area (total specific fluorescence) and the surface of this area [mean specific fluorescence (MSF)].…”