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J Thorac Cardiovasc Surg 2007;133:919-926
© 2007 The American Association for Thoracic Surgery

Cardiopulmonary Support and Physiology

Effect of profound hypothermia during circulatory arrest on neurologic injury and apoptotic repressor protein Bcl-2 expression in an acute porcine model

^a Department of Cardiac Surgery, University of Ioannina, School of Medicine, Ioannina, Greece
^b Department of Anatomy-Histology-Embryology, University of Ioannina, School of Medicine, Ioannina, Greece
^c Department of Pathology, University of Ioannina, School of Medicine, Ioannina, Greece
^d NHLI, Imperial College, University of London, London, United Kingdom
^e Cardiac Surgery Department, St Luke’s Roosevelt Hospital, Columbia University, New York, NY.

Read at the Eighty-sixth Annual Meeting of The American Association for Thoracic Surgery, Philadelphia, Pa, April 29-May 3, 2006.

Received for publication April 19, 2006; revisions received September 25, 2006; accepted for publication October 9, 2006.

^_* Address for reprints: E.O. Johnson, MD, Department of Anatomy-Histology-Embryology, University of Ioannina, School of Medicine, Ioannina 45110 Greece. (Email: oananiadou{at}yahoo.co.uk; ejohnson{at}cc.uoi.gr).

Objectives: We reported that the neocortex and hippocampus are selectively vulnerable to injury in an acute porcine model of hypothermic circulatory arrest at 18°C. We hypothesize that further cooling to 10°C could reduce neurologic injury in these regions. To further elucidate the mechanisms of neurologic injury and protection, we assessed the expression of the anti-apoptotic protein Bcl-2.

Methods: Twelve piglets underwent 75 minutes of hypothermic circulatory arrest at 18°C (n = 6) and 10°C (n = 6). After gradual rewarming and reperfusion, animals were put to death and brains were perfusion-fixed and cryopreserved. Regional patterns of neuronal apoptosis after hypothermic circulatory arrest were characterized by in situ DNA fragmentation with terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling (TUNEL) histochemistry. Bcl-2 protein expression was characterized with immunohistochemistry. Statistical comparisons were made by t test, analysis of variance, and Mann–Whitney U test, as appropriate.

Results: Concentrations of TUNEL(+) cells were significantly lower after profound hypothermia at 10°C compared with 18°C hypothermia in the sensory and motor neocortex and hippocampus (t test, P < .0001; P < .006; P < .006, respectively). Positive Bcl-2 immunostaining was observed only in the motor and sensory neocortex and hippocampus after 18°C hypothermic circulatory arrest. Profound cooling to 10°C resulted in a significant increase in Bcl-2 immunostaining in the motor and sensory cortex as compared with 18°C (Mann–Whitney U test, P < .05).

Conclusions: Deep hypothermia at 10°C protects the neocortex and hippocampus from insult during hypothermic circulatory arrest as suggested by significantly reduced TUNEL(+) staining in these areas. Although a concomitant increase in Bcl-2 expression was observed in the neocortex at 10°C, it remains unclear whether profound hypothermia deters from neuronal injury by activation of the anti-apoptotic protein Bcl-2.

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Discussion
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