EXPERIMENTAL ISCHEMIA AND REPERFUSION INJURY

N. Kuriyama¹, S. Duarte², T. Hamada³, R.W. Busuttil², A.J. Coito^4
1Surgery, Dumont-UCLA Transplant Ctr, Los Angeles/CA/UNITED STATES OF AMERICA, ²Surgery, Dumont-UCLA Transplant Ctr, Los Angeles/UNITED STATES OF AMERICA, ³, Hepatobiliary Pancreatic and Transplant Surgery, Tsu/JAPAN, ⁴Surgery, Dumont-UCLA Tranpslant Ctr, Los Angeles/CA/UNITED STATES OF AMERICA

Body: Introduction: Ischemia and reperfusion injury (IRI) remains a major problem in liver transplantation. Tenascin-C (TNC), an extracellular matrix (ECM) glycoprotein and adamage-associated molecular pattern (DAMP) molecule, is normally not expressed in adult tissues, with the exception of lymphoid tissues; however, it becomes upregulated during acute and chronicinflammation. While several in vitro assays have supported a key regulatory role for TNC in leukocyte migration, its precise function at sites of tissue injury remains unclear. Here, we test thehypothesis that TNC plays a critical role in the mechanism of hepatic IRI. Methods and results: TNC gene knockout mice (TNC-KO) and matched wild-type (WT) control littermates weresubmitted to partial warm ischemia in the left/medium hepatic lobes for 90 minutes, followed by reperfusion. TNC was virtually absent in naïve livers, and it was early upregulated in WT liversafter IRI; TNC deposition was mostly detected in the vascular areas of damaged control livers. Liver function, as evidenced by AST (1,902 ± 1,435 vs. 9,008 ± 1,774, p < 0.01), and byALT (2,067 ± 1,436 vs. 27,340 ± 6,834, p < 0.001) levels (U/L), was significantly improved in TNC-KO mice post-IRI, compared to WT controls. Histological damage was sheltered inTNC-KO livers, which showed significantly less signs of vascular congestion and necrosis post-IRI. Moreover, TUNEL staining illustrated reduced levels of apoptotic cells (47.2 ± 17.1 vs. 128.6± 8.9, p < 0.01) in the TNC-KO livers at 6h post-IRI, which were correlated with significantly reduced active caspase-3. While the inactive pro-caspase-3 was detected in TNC -/- and WTlivers post-IRI, the mature 17-kDa caspase-3 was predominantly detected in WT control livers at 6h post-IRI. In contrast, the 17-kDa active caspase-3 was profoundly depressed in TNC -/- livers (3-5fold; p<0.01) at 6h post-IRI, and virtually absent in naïve livers. Furthermore, TNC deficiency was associated with increased liver cell regeneration (PCNA positive cells: 64.5 ± 3.9vs. 18.4 ± 6.5, p < 0.0001; 24h) post-IRI. Others have previously shown that tenascin is capable of promoting leukocyte tethering and rolling, under flow, more efficiently than selectins.In our settings, lack of TNC was associated with a profound decrease in Ly6G+ neutrophil (6h: 6.8 ± 2.6 vs. 29.3 ± 11.2, p <0.05; 24h: 21.3 ± 8.4 vs. 64.7 ± 7.3, p <0.05) and in Mac-1+ macrophage (6h: 15.2 ± 8.9 vs. 49.1 ± 13.9, p < 0.05; 24h: 29.2 ± 13.7 vs. 85.9 ± 8.7, p < 0.05) infiltration post-IRI. Moreover, IL-6 (p <0.05) and CXCL2 (p < 0.05) expressions, which have been linked to liver IRI, were largely depressed in TNC-KO post-IRI. Conclusions: Our novel data, using tenascin-C deficient mice,support the view that TNC is an important player in hepatic IRI. We show that specifically targeting TNC profoundly ameliorated hepatic IRI. TNC deficiency exerted an anti-apoptotic function,improved liver regeneration, and depressed leukocyte recruitment and activation in liver IRI. Therefore, these findings offer a rationale for the development of therapeutic approaches based on newmechanistic concepts of hepatic IRI.

Disclosure: All authors have declared no conflicts of interest.