Liver ischemia-reperfusion injury (IRI) remains a challenging problem in clinical orthotopic liver transplantation (OLT). Tissue inhibitor of metalloproteinases-3 (TIMP-3) is an extracellular matrix (ECM)-associated protein known to regulate metalloproteinases. Its expression has been linked to a variety of physiological and pathological functions, including regulation of inflammation. In this study we assessed the functional significance of TIMP-3 expression in a well-established mouse model of hepatic IRI. Methods: TIMP-3 deficient mice (TIMP-3 -/-) and matched wild-type (WT) control littermates were submitted to 60-min of partial warm ischemia followed by 6h of reperfusion. Results: TIMP-3 mRNA expression was detected in WT livers and absent in TIMP-3-/- livers. Compared to WT controls, TIMP-3-/- mice showed significantly higher AST (5,090±2,376 vs. 7.882±2895; p<0.05) and ALT (6,526±2548 vs. 12,493±4937; p<0.02) levels (IU/L) post-IRI. Overall, loss of TIMP-3 resulted in further lobular architecture disruption and severe necrosis after IRI. TIMP-3-/- livers showed significantly increased Mac-1 (32.4±11.2 vs. 57.9±13.4/HPF; p<0.005) and Ly6G (37.7±13.8 vs. 76.3±17.0/HPF; p<0.002) leukocyte infiltration and higher levels of pro-inflammatory cytokines, such as IL-1β (p<0.02), IL-6 (p<0.02) and TNF-α (p<0.01). Moreover, the inability of TIMP-3-/- mice to express TIMP-3 led to a significant loss of e-cadherin expression in the membranes of hepatocytes near the portal area. Indeed, while the full-length e-cadherin 120 kDa protein was readily detected in naïve (WT and TIMP-3-/-) livers and in WT livers post-IRI, it was significantly depressed in TIMP-3-/- livers after reperfusion (p<0.005). In contrast, the 35 kDa e-cadherin fragment was notably increased in TIMP-3-/- livers post-reperfusion (p<0.02), suggesting that TIMP-3 hampers e-cadherin shedding after liver IRI. Conclusion: Our results show for the first time that TIMP-3 deficiency results in enhanced leukocyte recruitment and liver damage after IRI. They also evidence a protective role for TIMP-3 in preventing the proteolytic cleavage of hepatic e-cadherin, an important mediator of intercellular adhesion and tissue integrity.

CITATION INFORMATION: Fujii T, Duarte S, Busuttil R, Coito A. Tissue Inhibitor of Metalloproteinase-3 (TIMP-3) Protects Against Loss of Hepatic Integrity After Ischemia-Reperfusion Injury. Am J Transplant. 2016;16 (suppl 3).

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