Thoracic Surgery Directors Association Award. Cobalt chloride pretreatment attenuates myocardial apoptosis after hypothermic circulatory arrest.
BACKGROUND
Deep hypothermic circulatory arrest (DHCA) causes myocyte injury as a consequence of ischemia and reperfusion. Previous studies have shown that hypoxia or hypoxia-mimetic agents (cobalt chloride [CoCl2] or deferoxamine [DFX]) limit myocyte necrosis by upregulating the transcription factor hypoxia-inducible factor. However, it remains unknown whether these agents attenuate myocardial apoptosis after DHCA. This study tested the hypotheses (1) that hypoxia, DFX, or CoCl2 preconditioning attenuates myocardial apoptosis during DHCA; and (2) that the protective mechanism involves the altered expression of apoptosis regulatory proteins pAkt (antiapoptotic), Bcl-2 (antiapoptotic), and Bax (proapoptotic).


METHODS
Anesthetized neonatal piglets were randomly assigned to four groups (n = 6 in a group): control (NaCl injection); hypoxia (pO2 of 30 to 40 mm Hg for 3 hours); DFX injection; or CoCl2 injection. Twenty-four hours later, the animals underwent cardiopulmonary bypass (CPB) and 110 minutes of DHCA. One week after CPB, percentage of apoptotic myocytes (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling [TUNEL] assay) and expression of the pAKT, Bcl-2, and Bax were assessed by Western blot.


RESULTS
Although preconditioning with hypoxia and DFX failed to show a protective benefit, CoCl2 pretreatment significantly attenuated myocardial apoptosis (9.3% +/- 4.1%) versus controls (33.8% +/- 9.7%, p = 0.042). That was associated with increased myocardial pAkt expression (0.19 +/- 0.006 in CoCl2 versus 0.12 +/- 0.008 in control, p < 0.001). The expression of Bcl-2 was also significantly higher in the CoCl2 group (0.15 +/- 0.02) versus control (0.11 +/- 0.01, p = 0.007), whereas Bax expression was lower (0.34 +/- 0.04 versus 0.54 +/- 0.03 for control, p < 0.001).


CONCLUSIONS
Preconditioning with CoCl2 before prolonged DHCA in neonatal piglets attenuates myocardial apoptosis by mechanisms involving phosphorylation of Akt, upregulation of the antiapoptotic protein Bcl-2, and decreased expression of the proapoptotic protein Bax.
Kerendi
Faraz
F
Division of Cardiothoracic Surgery, Carlyle Fraser Heart Center, Emory University School of Medicine, Atlanta, Georgia, USA.
Kirshbom
Paul M
PM
Halkos
Michael E
ME
Wang
Ning-Ping
NP
Kin
Hajime
H
Jiang
Rong
R
Zhao
Zhi-Qing
ZQ
Kanter
Kirk R
KR
Guyton
Robert A
RA
Vinten-Johansen
Jakob
J
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis* / Q000235:genetics; D001363:Awards and Prizes; D015153:Blotting, Western; D002315:Cardiopulmonary Bypass / Q000009:adverse effects; D050792:Circulatory Arrest, Deep Hypothermia Induced / Q000009:adverse effects*; D003035:Cobalt / Q000008:administration & dosage / Q000627:therapeutic use*; D053938:DNA Fragmentation; D003676:Deferoxamine / Q000008:administration & dosage / Q000627:therapeutic use; D005786:Gene Expression Regulation / Q000187:drug effects; D019254:Genes, bcl-2; D006321:Heart / Q000187:drug effects*; D000860:Hypoxia / Q000150:complications; D051793:Hypoxia-Inducible Factor 1 / Q000096:biosynthesis / Q000235:genetics; D020287:In Situ Nick-End Labeling; D019157:Ischemic Preconditioning, Myocardial / Q000379:methods*; D009206:Myocardium / Q000378:metabolism / Q000473:pathology*; D010766:Phosphorylation; D011229:Preanesthetic Medication; D011499:Protein Processing, Post-Translational; D051057:Proto-Oncogene Proteins c-akt / Q000096:biosynthesis / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics; D011897:Random Allocation; D034421:Sus scrofa; D013903:Thoracic Surgery; D014481:United States; D051028:bcl-2-Associated X Protein / Q000096:biosynthesis / Q000235:genetics
D051017:Apoptosis Regulatory Proteins; D051793:Hypoxia-Inducible Factor 1; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D003035:Cobalt; D051057:Proto-Oncogene Proteins c-akt; C018021:cobaltous chloride; D003676:Deferoxamine