New Insights in Fatty Liver Preservation: A Role for Carbonic Anhydrase II.

Abstract

Purpose: During liver graft preservation, the switch to anaerobic metabolism provokes accumulation of lactic acid and leads to lower pH. Studies on the relevance of acidic pH are controversial. It can either activate proteases and induce cell death or protect cell against apoptosis. Carbonic anhydrases (CA) are ubiquitous metalloenzymes involved in many physiological and pathological processes, including pH and CO[/sub]2[/sub] homeostasis. However, the role of CA in liver graft preservation has not been investigated. The aim of this work was to evaluate the effects of CAII by its addition to the IGL-1 preservation solution or when it was inhibited by acetazolamide (AZ). A close CAII relationship with well known protective factors (AKT, MAP kinases) and apoptosis prevention was established Experimental: Steatotic livers from obese male Zucker rats (9 weeks aged) were classified as follows: Group1: Livers preserved for 24h (4°C) in IGL-1 solution and then subjected to “ex vivo” perfusion (2h at 37°C); Group2: same as Group1 but IGL-1 supplemented with CAII (10 μg/ml) and Group 3: Same as Group 1 but livers were pretreated with AZ (i.v. 30mg/Kg) before liver preservation. Liver injury (transaminases) and function (bile production) were measured. Akt, GSK3β, Caspase 3, Caspase 9 and MAPKs (p38, ERK and JNK) were also determined by western blot techniques. Results and Discussion: Fatty livers preserved in IGL-1+CAII showed lower injury and better function when compared to IGL-1 alone. IGL-1+CAII solution induced a significant phosphorylation of Akt and significant decreases in MAPKs (ERK, p38 and JNK) after 2h-reperfusion. This was consistent with a major decrease of liver apoptosis parameters (caspase 3, caspase 9, and GSK3β). Surprisingly, the AZ pretreatment protected efficiently steatotic liver grafts, as revealed by lower transaminases, higher bile production and decreases in MAPK levels. AZ showed differential protective mechanism of liver as it fails to activate Akt. Also, it reduced vascular resistance which was not observed with CAII addition. Conclusion: Data reported here, demonstrated that CAII is a potential pharmacological target for preserving liver grafts against cold ischemia reperfusion injury.