The Role of Neuregulin-1 in Steatotic and Non-Steatotic Liver Transplantation from Brain-Dead Donors

Abstract

Background. Brain death (BD) and steatosis are key risk factors to predict adverse post-transplant outcomes. We investigated the role of Neuregulin-1 (NRG1) in rat steatotic and non-steatotic liver transplantation (LT) from brain death donors (DBD). Methods: NRG1 pathways were characterized after surgery. Results: NRG1 and p21-activated kinase 1 (PAK1) levels increased in steatotic and non-steatotic grafts from DBDs. The abolishment of NRG1 effects reduced PAK1. When the effect of either NRG1 nor PAK1 was inhibited, injury and regenerative failure were exacerbated. The benefits of the NRG-1-PAK1 axis in liver grafts from DBDs were associated with increased vascular endothelial growth factor-A (VEGFA) and insulin growth factor-1 (IGF1) levels, respectively. Indeed, VEGFA administration in non-steatotic livers and IGF1 treatment in steatotic grafts prevented damage and regenerative failure resulting from the inhibition of either NRG1 or PAK-1 activity in each type of liver. Exogenous NRG1 induced greater injury than BD induction. Conclusions: This study indicates the benefits of endogenous NRG1 in liver grafts from DBDs and underscores the specificity of the NRG1 signaling pathway depending on the type of liver: NRG1-PAK1-VEGFA in non-steatotic livers and NRG1-PAK1-IGF1 in steatotic livers. Exogenous NRG1 is not an appropriate strategy to apply to liver grafts from DBD.

NO–IL-6/10–IL-1β axis: a new pathway in steatotic and non-steatotic liver grafts from brain-dead donor rats

“…Ob Zucker rats showed severe and macrovesicular and microvesicular fatty infiltration in hepatocytes. In contrast, Ln Zucker rats show no evidence of steatosis ( 15 , 45 , 56 ). In addition, we also evaluated fibrosis presence or absence with Sirius Red staining in paraffin-embedded liver samples, as this staining is one of the best understood techniques able to selectively highlight collagen networks ( 57 ).…”

“…To determine hepatic edema, we weighed liver samples and then placed them in a drying oven at 55°C until they reached a constant weight. The increase in the wet-to-dry weight ratio represents hepatic edema ( 45 , 46 ).…”

“…Hepatic levels of lipid peroxidation were determined via measurements of malondialdehyde (MDA) formation by using the thiobarbiturate reaction ( 45 , 47 ). For this, frozen tissue samples were homogenized in 2 ml of Tris buffer at pH 7.…”

“…To function as an index of the level of accumulation of neutrophils, we determined hepatic levels of myeloperoxidase (MPO) photometrically using a substrate of 3,3′,5,5′-tetramethylbenzidine ( 45 , 47 ). For this, liver samples were homogenized in phosphate buffer (0.05 M KH 2 PO 4 , pH 6) with 0.5% hexadecyltrimethylammonium bromide (HTBA), then sonicated for 30 s at 20% power.…”

“…In liver samples obtained from all experimental group studies, we assessed the degree of liver injury also by staining paraffin-embedded liver sections with H&E and then using a method of counting points to determine a histological score ( 47 ). An experienced pathologist blinded to the allocation of animals into different experimental groups reviewed biopsy specimens and scored the histologic findings according to the following scale: grade 0, zero or minimal evidence of injury; grade 1, the injury is mild, showing cytoplasmic vacuolation and focal nuclear pyknosis; grade 2, the injury is moderate ranging to severe with extensive nuclear pyknosis, cytoplasmic hypereosinophilia, and the loss of intercellular borders; grade 3, severe necrosis showing disintegration of the hepatic cords, hemorrhage, and neutrophil infiltration; and finally grade 4, very severe necrosis exhibiting disintegration of the hepatic cords, hemorrhaging, and neutrophil infiltration ( 41 , 45 ).…”