Natural resistance to liver cold ischemia-reperfusion injury associated with the hibernation phenotype

Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
AJP Gastrointestinal and Liver Physiology (Impact Factor: 3.8). 04/2005; 288(3):G473-80. DOI: 10.1152/ajpgi.00223.2004
Source: PubMed


The success of liver grafts is currently limited by the length of time organs are cold preserved before transplant. Novel insights to improve viability of cold-stored organs may emerge from studies with animals that naturally experience low body temperatures (T(b)) for extended periods. In this study, we tested whether livers from hibernating ground squirrels tolerate cold ischemia-warm reperfusion (cold I/R) for longer times and with better quality than livers from rats or summer squirrels. Hibernators were used when torpid (T(b) < 10 degrees C) or aroused (T(b) = 37 degrees C). Livers were stored at 4 degrees C in University of Wisconsin solution for 0-72 h and then reperfused with 37 degrees C buffer in vitro. Lactate dehydrogenase (LDH) release after 60 min was increased 37-fold in rat livers after 72 h cold I/R but only 10-fold in summer livers and approximately three- to sixfold in torpid and aroused hibernator livers, despite twofold higher total LDH content in livers from hibernators compared with rats or summer squirrels. Reperfusion for up to 240 min had the least effect on LDH release in livers from hibernators and the greatest effect in rats. Compared with rats or summer squirrels, livers from hibernators after 72 h cold I/R showed better maintenance of mitochondrial respiration, bile production, and sinusoidal lining cell viability, as well as lower vascular resistance and Kupffer cell phagocytosis. These results demonstrate that the hibernation phenotype in ground squirrels confers superior resistance to liver cold I/R injury compared with rats and summer squirrels. Because hibernation-induced protection is not dependent on animals being in the torpid state, the mechanisms responsible for this effect may provide new strategies for liver preservation in humans.

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Available from: Martin J Mangino, Oct 04, 2015
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    • "Similarly, the decreases in the fold change of circulating levels of DBHB during hemorrhage in both AGS groups concurs with a low demand for glucose noted previously in IBA and hibernating ground squirrels. Metabolomic fingerprints of IBA- and EU-AGS were consistent with reports from other species showing a metabolic switch during the hibernation season [5]–[7], [14], [15], [45]–[48]. Surprisingly, both groups of animals avoided anaerobic glucose metabolism during I/R, consistent with resistance from I/R injury. "
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    ABSTRACT: Background Hemorrhagic shock (HS) following trauma is a leading cause of death among persons under the age of 40. During HS the body undergoes systemic warm ischemia followed by reperfusion during medical intervention. Ischemia/reperfusion (I/R) results in a disruption of cellular metabolic processes that ultimately lead to tissue and organ dysfunction or failure. Resistance to I/R injury is a characteristic of hibernating mammals. The present study sought to identify circulating metabolites in the rat as biomarkers for metabolic alterations associated with poor outcome after HS. Arctic ground squirrels (AGS), a hibernating species that resists I/R injury independent of decreased body temperature (warm I/R), was used as a negative control. Methodology/principal findings Male Sprague-Dawley rats and AGS were subject to HS by withdrawing blood to a mean arterial pressure (MAP) of 35 mmHg and maintaining the low MAP for 20 min before reperfusing with Ringers. The animals’ temperature was maintained at 37±0.5°C for the duration of the experiment. Plasma samples were taken immediately before hemorrhage and three hours after reperfusion. Hydrophilic and lipid metabolites from plasma were then analyzed via 1H–NMR from unprocessed plasma and lipid extracts, respectively. Rats, susceptible to I/R injury, had a qualitative shift in their hydrophilic metabolic fingerprint including differential activation of glucose and anaerobic metabolism and had alterations in several metabolites during I/R indicative of metabolic adjustments and organ damage. In contrast, I/R injury resistant AGS, regardless of season or body temperature, maintained a stable metabolic homeostasis revealed by a qualitative 1H–NMR metabolic profile with few changes in quantified metabolites during HS-induced global I/R. Conclusions/significance An increase in circulating metabolites indicative of anaerobic metabolism and activation of glycolytic pathways is associated with poor prognosis after HS in rats. These same biomarkers are absent in AGS after HS with warm I/R.
    PLoS ONE 09/2014; 9(9):e107493. DOI:10.1371/journal.pone.0107493 · 3.23 Impact Factor
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    • "It seems reasonable to suggest that moesin expression is an adaptation to hypothermia, which is consistent with our observations of the moesin defect in cold stored livers from non-hibernating mammals. In fact, livers retrieved from actively hibernating ground squirrels were determined to be resistant to cold storage injury, relative to cold stored livers retrieved from either non-hibernating summer active ground squirrels or from other non-hibernating rodent species like rats [14]. It is not known from these studies whether hibernating ground squirrels also have higher expression levels of moesin in the liver, like they do in their intestine, or if they express a more heavily phosphorylated moesin protein in the liver that would provide for more structural functionality. "
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    ABSTRACT: The objective of this study was to determine how expression and functionality of the cytoskeletal linker protein moesin is involved in hepatic hypothermic preservation injury. Mouse livers were cold stored in University of Wisconsin (UW) solution and reperfused on an Isolated Perfused Liver (IPL) device for one hour. Human hepatocytes (HepG2) and human or murine Sinusoidal Endothelial Cells (SECs) were cold stored and rewarmed to induce hypothermic preservation injury. The cells were transfected with: wild type moesin, an siRNA duplex specific for moesin, and the moesin mutants T558D and T558A. Tissue and cell moesin expression and its binding to actin were determined by western blot. Liver IPL functional outcomes deteriorated proportional to the length of cold storage, which correlated with moesin disassociation from the actin cytoskeleton. Cell viability (LDH and WST-8) in the cell models progressively declined with increasing preservation time, which also correlated with moesin disassociation. Transfection of a moesin containing plasmid or an siRNA duplex specific for moesin into HepG2 cells resulted in increased and decreased moesin expression, respectively. Overexpression of moesin protected while moesin knock-down potentiated preservation injury in the HepG2 cell model. Hepatocytes expressing the T558A (inactive) and T558D (active) moesin binding mutants demonstrated significantly more and less preservation injury, respectively. Cold storage time dependently caused hepatocyte detachment from the matrix and cell death, which was prevented by the T558D active moesin mutation. In conclusion, moesin is causally involved in hypothermic liver cell preservation injury through control of its active binding molecular functionality.
    Cryobiology 05/2014; 69(1). DOI:10.1016/j.cryobiol.2014.04.017 · 1.59 Impact Factor
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    • "Bouts of torpor are interspersed by short arousal periods, during which metabolism increases and body temperature returns to euthermia [2], [6], [7]. Key changes in physiological parameters are thought to lead to an increased resistance to ischemia/reperfusion [8], [9] allowing hibernating mammals to survive periods of torpor and arousal without signs of organ injury. Therefore, hibernating animals have been used in various studies as a model to investigate the effects of low body temperature and hypoxia on organs, in attempts to unravel the adaptations that allow these animals to cope with the physiological extreme conditions of torpor [5]. "
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    ABSTRACT: Hibernation is an energy-conserving behavior in winter characterized by two phases: torpor and arousal. During torpor, markedly reduced metabolic activity results in inactivity and decreased body temperature. Arousal periods intersperse the torpor bouts and feature increased metabolism and euthermic body temperature. Alterations in physiological parameters, such as suppression of hemostasis, are thought to allow hibernators to survive periods of torpor and arousal without organ injury. While the state of torpor is potentially procoagulant, due to low blood flow, increased viscosity, immobility, hypoxia, and low body temperature, organ injury due to thromboembolism is absent. To investigate platelet dynamics during hibernation, we measured platelet count and function during and after natural torpor, pharmacologically induced torpor and forced hypothermia. Splenectomies were performed to unravel potential storage sites of platelets during torpor. Here we show that decreasing body temperature drives thrombocytopenia during torpor in hamster with maintained functionality of circulating platelets. Interestingly, hamster platelets during torpor do not express P-selectin, but expression is induced by treatment with ADP. Platelet count rapidly restores during arousal and rewarming. Platelet dynamics in hibernation are not affected by splenectomy before or during torpor. Reversible thrombocytopenia was also induced by forced hypothermia in both hibernating (hamster) and non-hibernating (rat and mouse) species without changing platelet function. Pharmacological torpor induced by injection of 5'-AMP in mice did not induce thrombocytopenia, possibly because 5'-AMP inhibits platelet function. The rapidness of changes in the numbers of circulating platelets, as well as marginal changes in immature platelet fractions upon arousal, strongly suggest that storage-and-release underlies the reversible thrombocytopenia during natural torpor. Possibly, margination of platelets, dependent on intrinsic platelet functionality, governs clearance of circulating platelets during torpor.
    PLoS ONE 04/2014; 9(4):e93218. DOI:10.1371/journal.pone.0093218 · 3.23 Impact Factor
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