Liver graft washout is determinant for the efficient prevention of reperfusion injury: A role for the cytoskeleton and glycocalyx preservation

Antecedents: Hepatic ischemia reperfusion injury (I/R) contributes to the initial poor function or primary non-function after transplantation. This is due to the cold preservation, rewarming and reperfusion, respectively. We evaluated the benefits of using a new rinse solution against I/R. Experimental: Sprague-Dawley rats (180-200 g; n= 6 for each group), were classified as follows: Group 1(controls)= Livers preserved in UW solution (24 hours; 4 °C) were flushed with Ringer lactate solution(RLS) and subjected to 2 h-reperfusion at 37 °C using an ex-vivo model; Group 2 = Same as 1 but the liver grafts were flushed with the new rinse solution composed by CaCl2.2H2O, KH2PO4, NaH2PO4, MgSO4.7H2O, lactobionate and rafinose at pH=7.4; Group 3: same as 2 but with polyethyleneglycol-35 (PEG-35) addition at 1g /L. Group 4 = same as 3 but with PEG-35 addition at 5g/L. Determinations: Liver injury (AST/ALT); mitochondrial damage (GLDH); function (bile output and vascular resistance), oxidative stress (MDA), nitric oxide; as well as glycocalyx (syndecan, hyaluronic acid) and cytoskeleton (filament and globular actin fraction) degradation were assessed. Also metalloproteinases (MMP2 and MMP9) were determined. In addition, actin confocal microscopy studies were carried out. Results: PEG-35 at 1g/L and 5g/L reduced AST/ALT levels, ameliorated liver function when compared to Groups 1 and 2. This was accompanied by decreases in GLDH, MDA and increases in e-NOS activation. Interestingly, the PEG-35 benefits were closely associated with the inhibition of MMP9 and the stabilization of actin filament fraction (western blot and confocal microscopy).Glycocalyx was also protected (low syndecan-1 and hyaluronic acid levels). Conclusions: The most suitable liver graft protection was observed for PEG35 (5 g/L) (“graft post-conditioning”) that reinforced the stabilization of the liver graft cytoskeleton and glycocalyx.