Up-regulation of Bcl-2 through hyperbaric pressure transfection of TGF-β1 ameliorates ischemia-reperfusion injury in rat cardiac allografts

Grünenfelder, Jürg; Miniati, Douglas N; Murata, Seiichiro; Falk, Volkmar; Hoyt, E.Grant; Robbins, Robert C

Next »The Journal of Heart and Lung Transplantation
Volume 21, Issue 2 , Pages 244-250, February 2002

Up-regulation of Bcl-2 through hyperbaric pressure transfection of TGF-β1 ameliorates ischemia-reperfusion injury in rat cardiac allografts

Presented at the ISHLT 21^st Annual Meeting and Scientific Sessions, April 25–28, 2001, Vancouver BC, Canada.

Jürg Grünenfelder, MD
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA
- Reprint requests and correspondence: Jürg Grünenfelder, MD, Clinic for Cardiovascular Surgery, University Hospital Zürich, Rämistrasse 100, 8091 Zürich, Switzerland. Telephone: 0041 1 255-3801. Fax: 0041 1 255-4369
Douglas N Miniati, MD
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA
Seiichiro Murata, MD, PhD
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA
Volkmar Falk, MD, PhD
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA
E.Grant Hoyt
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA
Robert C Robbins, MD
- Department of Cardiothoracic Surgery, Falk Cardiovascular Research Building, Stanford University School of Medicine, Stanford, California, USA

Received 13 July 2001; accepted 29 August 2001.

Abstract

Background: Oxidative stress after ischemia-reperfusion of cardiac allografts leads to activation of cardiomyocytes and production of cytokines. Bcl-2, an inhibitor of the apoptotic pathway, also has strong antioxidant properties. Ischemia-reperfusion injury after transplantation leads to decreased bcl-2 and increased tumor necrosis factor (TNF)-α levels. Transforming growth factor (TGF)-β₁ is known to attenuate ischemia-reperfusion injury and inhibits apoptosis of myofibroblasts. We hypothesize that TGF-β₁, prevents bcl-2 cleavage and increased TNF-α production.

Methods: Rat PVG donor hearts were heterotopically transplanted into ACI recipients. Donor hearts were procured and assigned to groups: (1) intracoronary TGF-β₁ (200 ng/ml) perfusion and pressure at 78 psi for 45 minutes (n = 4); (2) intracoronary TGF-β₁ perfusion and incubation for 45 minutes without pressure (n = 4), (3) saline perfusion and incubation for 45 minutes without pressure (n = 4). Hearts were procured 4 hours after transplantation and analyzed by reverse transcriptase-polymerase chain reaction for bcl-2 mRNA expression, ELISA for TNF-α, and for myeloperoxidase activity (MPO).

Results: Bcl-2 decreased in untreated animals (bcl-2:G3PDH ratio = 0.85 ± 0.73 vs 1.16 ± 0.11, not significant [NS]), whereas TNF-α increased to 669.99 ± 127.09 vs 276.84 ± 73.65 pg/mg total protein in controls (p < 0.003). In TGF-β₁ pressure-treated hearts, bcl-2 was up-regulated (2.49 ± 0.6 vs 1.16 ± 0.11, controls, p < 0.005), whereas TNF-α was unchanged (396.1 ± 100.38 vs 276.84 ± 73.65 pg/mg, NS). Hearts treated with TGF-β₁ and pressure showed significant up-regulation of bcl-2 compared with hearts treated with TGF-β₁ without pressure (2.49 ± 0.6 vs 1.17 ± 0.6, p < 0.02). MPO showed no differences.

Conclusions: Bcl-2 is down-regulated and TNF-α up-regulated in this model of ischemia-reperfusion injury. Furthermore, TGF-β₁ is linked to this process and ameliorates reperfusion injury by up-regulating bcl-2 and inhibiting TNF-α. Therapeutic overexpression of myocardial TGF-β₁ may be clinically useful to control ischemia-reperfusion injury associated with cardiac transplantation.