High-Mobility Group Box-1 as a Novel Therapeutic Target for Reducing Pulmonary Ischemia-Reperfusion Injury in CLAWN Miniature Swine

Abstract

Purpose: Strategies that reduce ischemia-reperfusion injury (IRI) have the potential to expand the use of extended-criteria or donation after cardiac donors and improve both the short and long-term outcomes of transplantation. Because high-mobility group box 1 (HMGB1) has been reported to be a mediator of inflammation and cell injury, blocking of HMGB1 might represent a novel therapeutic strategy against IRI. However, there is limited data to characterize the role of HMGB1 on pulmonary IRI especially in large animals. The aim of this study was to determine the role of HMGB1 on IRI and to evaluate the cytoprotective effects of anti-HMGB1 antibodies as a translational research using our established, in situ porcine lung warm IRI model. Methods: Warm ischemia was induced in 12 CLAWN miniature swine by clamping the left main bronchus, pulmonary artery and veins for 90 minutes. In the control group (n=5), the animals were followed without any additional treatment. In the HMGB1 Ab group (n=4), 1mg/kg of anti-HMGB1 Ab was intravenously injected just before reperfusion. In the isotype Ab group (n=3), control IgY Ab was injected as the same fashion as the HMGB1 Ab group. Lung function and structure were serially accessed via lung biopsy, chest X-ray and blood gas analysis. Serum cytokine levels such as IL-1β and HMGB1 were measured in order to characterize the inflammatory response to IRI and evaluate the effect of the treatment on that response. Results: Serum levels of HMGB1 increased after IRI, peaking 2 days after reperfusion (14 ± 6 ng/ml) in the control group. Anti-HMGB1 Ab treatment resulted in significant reduction of HMGB1 (0.9 ± 0.5 ng/ml). Two hours after reperfusion, the PaO2 of the HMGB1 Ab group was 551 ± 19 mmHg, more than double of the control group (266 ± 60 mmHg). Lung biopsy in the control group showed significant damage to alveolar endothelial cells with intra-alveolar edema, hemorrhage, and fibrin exudates at POD2. In contrast, treatment with anti-HMGB1 Ab resulted in marked improvement of histological findings and less PCNA-positive infiltrating cells and TUNEL-positive apoptotic cells. Systemic IL-1β levels increased after IRI in both the control and HMGB1 groups, peaking 1 day after reperfusion, though there was no difference in the concentration between two groups (control vs. HMGB1 Ab group; 63 ± 26 vs. 62 ± 14 pg/ml). Furthermore, treatment of isotype control IgY Ab showed no effect on improving gas oxygenation or inhibiting lung damage after IRI. Conclusions: In this study, we demonstrated that HMGB1 played a major role in the progression of lung IRI and that reducing systemic HMGB1 by anti-HMGB1 Ab alone could reduce IRI independently of systemic IL-1β concentration using a clinically relevant large animal model. Targeting HMGB1 might represent a novel therapeutic strategy in clinical IRI associated with lung transplantation.