2010 - TTS International Congress


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Experimental Ischemia and Reperfusion Injury

111.1 - Development of Hepatic Ischemia and Reperfusion Injury is Independent of Myeloid Cell-Derived COX-2

Presenter: Sergio, Duarte, Los Angeles, United States
Authors: Duarte S., Kuriyama N., Ishikawa T., Busuttil R., Herschman H., Coito A.

DEVELOPMENT OF HEPATIC ISCHEMIA AND REPERFUSION INJURY IS INDEPENDENT OF MYELOID CELL-DERIVED COX-2 EXPERIMENTAL ISCHEMIA AND REPERFUSION INJURY

S. Duarte1, N. Kuriyama2, T. Ishikawa3, R.W. Busuttil1, H.R. Herschman3, A.J. Coito2
1Surgery, Dumont-UCLA Transplant Ctr, Los Angeles/UNITED STATES OF AMERICA, 2Surgery, Dumont-UCLA Tranpslant Ctr, Los Angeles/CA/UNITED STATES OF AMERICA, 3Molecular And Medical Pharmacology, David Geffen School of Medicine - UCLA, Los Angeles/CA/UNITED STATES OF AMERICA

Body: Cyclooxygenase-2 (COX-2) is a powerful inflammatory mediator linked to liver ischemia and reperfusion injury (IRI). Inhibition of COX-2 expression significantly ameliorates liver and brain IRI as we,and others, have demonstrated using COX-2 deficient mice and mice treated with COX-2 inhibitors. However, the use of COX-2 inhibitors present clinical limitations and, to improve their therapeuticapplicability, there is a need to identify the cell types in which COX-2 expression leads to tissue damage. Observations that leukocytes of myeloid origin, including macrophages, are major sources ofCOX-2 support a rationale to dissect the function of myeloid COX-2 in hepatic IRI. Methods and Results: To investigate the role of myeloid COX-2 in IRI, we usedCOX-2 floxed "knock-in" mice, in which exons 4 and 5 of mouse COX-2 are flanked by loxP sites. COX-2 floxed mice were crossed to LysM Cre mice to obtainmyeloid-specific COX-2 knock-out (COX-2–M/–M; COX-2flox/flox, LysMCre/+) and wild-type (WT; COX-2flox/flox, LysM+/+) mice. WhileLPS-activated WT macrophages are capable of expressing COX-2, cultured COX-2–M/–M macrophages fail to express COX-2 in response to pro-inflammatory stimuli. COX-2–M/–M mice and their WT control littermates were subjected to 90 min of 70% partial liver ischemia followed by 6h (n=5/group) and 24h (n=7/group) of reperfusion. COX-2 proteinexpression was significantly depressed in COX-2–M/–M livers (2.2 fold; p<0.02) at 24h post-IRI, as compared to WT livers; COX-2 was comparably upregulated inCOX-2–M/–M and WT livers after 6h of reperfusion. Somewhat unexpectedly, liver injury, as assessed by serum AST (6h: 13,280±2,843 vs. 14,225±1,374; 24h:7,140±1,655 vs. 7,385±2,305) and ALT (6h: 42,775±10,427 vs. 33,575±3,835; 24h: 11,094±4,553 vs. 11,222±2,360) levels (U/L), was similar in COX-2–M/–M and WT mice post-liver IRI. Moreover, both COX-2–M/–M and WT livers developed extensive necrosis and vascular congestion post-IRI. We havepreviously shown that total COX-2 depletion results in a significant reduction of pro-inflammatory cytokine release and neutrophil infiltration after liver IRI. In contrast, depletion of myeloidCOX–2 was ineffective in depressing leukocyte infiltration and activation. Indeed, COX-2 -M/-M livers showed increased TNF-αand IL-6 mRNA levels (TNF-α: 0.431±0.09 vs. 0.246±0.04, p<0.02; IL-6: 0.705±0.06 vs. 0.086±0.05, p<0.001) at6h post-IRI, suggesting a potentially mild ‘protective’ role for myeloid COX-2. COX-2 –M/-M and WT livers showed similar levels of Ly-6G neutrophil infiltration (6h:75±18 vs. 81±28; 24h: 157±39 vs. 185±30) post-IRI. Mac-1 macrophage infiltration in COX-2 –M/-M livers was also similar at 6h (78±12 vs.76±18) post-IRI, and only slightly reduced at 24h (117±15 vs. 170±38; p<0.05) post-IRI, as compared to controls. Conclusions: Our novelresults, using mice in which COX-2 has been ablated selectively in myeloid cells, clearly show that myeloid COX-2, including macrophage COX-2, is not responsible for hepatic IRI. Moreover, thesefindings provide evidence that COX-2 mediated liver IRI results from COX-2 being predominantly expressed during the first few hours post-reperfusion by sources other than myeloid cells. Thesefindings are of particular importance for the development of potential COX-2 targeted therapies in hepatic IRI.

Disclosure: All authors have declared no conflicts of interest.


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