8B,C). Moreover, Bcl-2, a known inhibitor of cell death, was almost absent in the TIMP-1−/− livers at 48 hours post-IRI (0.13 ± 0.08 versus 0.69 ± 0.19; P < 0.05) (Fig. 8A). Finally, phosphorylation of mTOR inhibitor Akt, a 57-kD protein-serine/threonine kinase with prosurvival-associated functions,22 was depressed in TIMP-1−/− livers (0.10 ± 0.07 versus 0.44 ± 0.30; P < 0.05) at 48 hours post-IRI (Fig. 8A). At 7

days post-IRI, Bcl-2 was still reduced (≈0.6-fold; P < 0.05) in TIMP-1−/− livers compared to controls. Hence, these results support a major protective role for TIMP-1 expression in hepatic IRI. The understanding of the functions of TIMPs in liver IRI has the potential to contribute to the development of novel therapeutic approaches to prevent hepatic IRI and, consequently, to improve the outcome of liver transplantation. In this study we investigated the functional significance of TIMP-1 expression in a well-established 90-minute mouse model of partial liver warm IRI.4 Interactions between ECM components and cell adhesion receptors regulate leukocyte functions; therefore, it is not unanticipated that enzymatic degradation of ECM can alter leukocyte behaviors.23 Indeed, cells employ proteolytic enzymes, particularly MMPs, to control the ECM turnover, to release growth factors, and to migrate across ECM.24 There is a growing body of evidence supporting key functions

for MMP expression Barasertib datasheet in the pathogenesis of liver diseases.3, 25, 26 In this regard, we have previously shown that MMP-9 regulates leukocyte recruitment and contributes to hepatic IRI.4 Although TIMP-1 can inhibit a broad range of MMPs, it is particularly potent for MMP-9.27 However, compared to MMP-9, the role of its natural inhibitor, TIMP-1, is virtually unknown in liver IRI. TIMP-1 expression is very low in naive livers and it is induced after liver IRI; however,

it is still insufficient to prevent an elevated MMP activity in liver IRI.11 In the present study we show that TIMP-1 deficiency resulted in further exaggerated up-regulation of MMP-9 activity Adenosine triphosphate and, more strikingly, it led to a poor survival rate after reperfusion. This is particularly interesting in that the model of partial liver IRI is nonlethal.14 Indeed, all TIMP-1+/+ mice survived hepatic IRI despite the significant damage detected in the livers after reperfusion; in contrast, only three out of eight TIMP-1−/− mice survived more than 4 days after liver IRI. In general, TIMP-1−/− mice showed additional impairment of liver function and more severe lesions, which likely led to their death between the second and fourth day postreperfusion. Although infiltrating leukocytes are recognized as mediators of hepatic IRI,3, 28 the mechanisms involved in their recruitment to sites of inflammatory stimulation in liver are still far from being understood. TIMP-1−/− livers showed massive leukocyte accumulation post-IRI.