Delayed parenchymal (hepatocellular) killing occurs in liver grafts that begins after about 4 h and can lead to graft dysfunction and failure. The mechanisms of delayed parenchymal cell injury develops remain poorly understood. Our preliminary experiments indicate that the onset of the mitochondrial permeability transition (MPT) plays a key role in delayed parenchymal cell killing. Accordingly our specific aims are to 1) elucidate the role and mechanisms of mitochondrial inner membrane permeabilization in delayed hepatocellular injury to liver grafts;2) determine the relationship of the MPT with microcirculatory disturbances, leukocyte adhesion and inflammation to liver grafts;3) characterize how c-Jun N-terminal kinase isoforms (JNK1 and JNK2) contribute to MPT onset and liver graft failure and 4) to test the hypothesis that activation of the HIF-1/prolyl hydroxylase oxygen-sensing signal cascade blocks the MPT and minimizes storage/reperfusion injury to liver grafts. Our studies utilize the emerging technology of intravital multiphoton microscopy to visualize parameter- indicating fluorophores inside individual hepatocytes and their organelles in liver grafts of living rats and mice. Specifically, we will determine whether mitochondrial dysfunction in liver grafts is due to MPT onset and if dysfunction leads to necrosis and apoptosis;how enhanced ROS formation contributes to MPT onset;and if mitochondrial calcium dysregulation contributes to MPT onset. Additionally, we will evaluate competing hypotheses that reperfusion causes microcirculatory disturbances and inflammation that promote oxidative stress and MPT onset or that MPT-dependent cell death activates inflammation and microcirculatory disturbances. During the previous period of support, we showed that JNK activation occurs after liver transplantation and that pharmacological inhibition of JNK decreases graft injury and improves graft survival. Two JNK isoforms, JNK1 and JNK2, are expressed in liver. We will compare injury and survival of JNK1 and JNK2 deficient liver grafts and determine the effects of JNK isoform deficiency on MPT induction, microcirculatory disturbances and the inflammatory response in transplanted liver grafts. Based on preliminary experiments, we expect to show that JNK2 promotes MPT onset and hepatocellular injury to liver grafts. In a final series of experiments, we will assess the effects of ethyl-3,4-dihydroxybenzoate (EDHB), a prolyl hydroxylase inhibitor that activates the oxygen sensing cascade, on liver graft injury and characterize mechanisms of protection, including MPT inhibition, decreased oxidative stress, decreased nonparenchymal cell injury and increased expression of heme oxygenase-1 (HO-1). These experiments will lead to a fuller understanding of the pathogenesis of liver graft injury from storage/reperfusion injury relevant to improving the function and survival of marginal donor livers. Transaminases increase well beyond the upper limit of normal in virtually every human liver transplantation, indicating storage/reperfusion to every liver graft. Thus, new information from this project has the potential of improving the outcome of all human liver transplantations. Delayed parenchymal (hepatocellular) killing occurs in virtually every human liver transplantation and when severe is a principal cause of initial poor function and graft failure requiring retransplanation. This project will address mechanisms underlying delayed hepatocellular killing and test the hypothesis that onset of the mitochondrial permeability transition has a key role. The information gained from the proposed experiments will be useful for improving clinical outcomes of not only so-called marginal donor livers but of virtually every liver graft.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK037034-25
Application #
7858210
Study Section
Surgery, Anesthesiology and Trauma Study Section (SAT)
Program Officer
Doo, Edward
Project Start
1986-05-01
Project End
2013-04-30
Budget Start
2010-05-01
Budget End
2011-04-30
Support Year
25
Fiscal Year
2010
Total Cost
$598,772
Indirect Cost
Name
Medical University of South Carolina
Department
Biochemistry
Type
Schools of Medicine
DUNS #
183710748
City
Charleston
State
SC
Country
United States
Zip Code
29425
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Hendriks, Giel; Derr, Remco S; Misovic, Branislav et al. (2016) The Extended ToxTracker Assay Discriminates Between Induction of DNA Damage, Oxidative Stress, and Protein Misfolding. Toxicol Sci 150:190-203
Rehman, Hasibur; Liu, Qinlong; Krishnasamy, Yasodha et al. (2016) The mitochondria-targeted antioxidant MitoQ attenuates liver fibrosis in mice. Int J Physiol Pathophysiol Pharmacol 8:14-27
Maldonado, Eduardo N; DeHart, David N; Patnaik, Jyoti et al. (2016) ATP/ADP Turnover and Import of Glycolytic ATP into Mitochondria in Cancer Cells Is Independent of the Adenine Nucleotide Translocator. J Biol Chem 291:19642-50
Hu, Jiangting; Kholmukhamedov, Andaleb; Lindsey, Christopher C et al. (2016) Translocation of iron from lysosomes to mitochondria during acetaminophen-induced hepatocellular injury: Protection by starch-desferal and minocycline. Free Radic Biol Med 97:418-426
Hu, Jiangting; Ramshesh, Venkat K; McGill, Mitchell R et al. (2016) Low Dose Acetaminophen Induces Reversible Mitochondrial Dysfunction Associated with Transient c-Jun N-Terminal Kinase Activation in Mouse Liver. Toxicol Sci 150:204-15
Krishnasamy, Yasodha; Ramshesh, Venkat K; Gooz, Monika et al. (2016) Ethanol and High Cholesterol Diet Causes Severe Steatohepatitis and Early Liver Fibrosis in Mice. PLoS One 11:e0163342
Belosludtsev, Konstantin N; Belosludtseva, Natalia V; Agafonov, Alexey V et al. (2015) Effect of surface-potential modulators on the opening of lipid pores in liposomal and mitochondrial inner membranes induced by palmitate and calcium ions. Biochim Biophys Acta 1848:2200-5
Liu, Qinlong; Krishnasamy, Yasodha; Rehman, Hasibur et al. (2015) Disrupted Renal Mitochondrial Homeostasis after Liver Transplantation in Rats. PLoS One 10:e0140906
Azarashvili, T; Krestinina, O; Baburina, Yu et al. (2015) Combined effect of G3139 and TSPO ligands on Ca(2+)-induced permeability transition in rat brain mitochondria. Arch Biochem Biophys 587:70-7

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