Myocardial cytochrome oxidase activity increases with age and hypoxemia in patients with congenital heart disease

Abstract

Background: Myocardial tolerance to ischemia is influenced by age and preoperative cyanosis through unknown mechanisms and significantly affects postoperative outcomes. Cytochrome c oxidase (CcOx), the terminal enzyme of the mitochondrial electron transport chain, may play a role in the susceptibility to ischemic-reperfusion (IR) injury. Our study aimed at investigating changes in human myocardial CcOx activity based on age and preoperative oxygen saturation to understand its role in transition from neonatal to mature myocardium and hypoxic conditions.

Methods: The right atrial appendage from patients undergoing first time surgical repair/palliation of congenital heart defects was analyzed for steady state CcOx activity by oxidation of ferrocytochrome c via spectrophotometry and steady state CcOx subunit I protein content by protein immunoblotting. Student's t-test compared CcOx activity and protein levels between patients with preoperative hypoxia and normoxia. Multiple linear regression analysis was used to assess the effects of age and preoperative arterial oxygen saturations (SaO₂) on CcOx protein activity and protein content.

Results: Thirty-two patients with a median (interquartile range) age of 83 days (8-174) and preoperative oxygen saturation 98% (85-100%) were enrolled. Independent of age, preoperative SaO₂ ⩽90% was associated with significantly greater CcOx steady state activity (p=0.004). Additionally, older age itself was associated with increased CcOx steady state activity (p=0.022); the combination of preoperative SaO₂ and age account for 33% of the variation in CcOx steady state activity (R²=0.332). There was no increase in the CcOx subunit I protein content with either age or preoperative hypoxia.

Conclusions: In patients with congenital heart disease, an increase in CcOx steady state activity is seen with increasing age. Hypoxia leads to upregulation of CcOx steady state activity without an increase in the amount of enzyme protein itself. Higher CcOx activity in older and cyanotic patients may indicate CcOx-dependent reactive oxygen species as the mechanism for IR injury.

Keywords: cyanotic congenital heart disease; cytochrome oxidase; ischemia reperfusion; myocardial protection; neonatal cardiac surgery.

Figure 1

Distribution of patients into ‘Preoperative Hypoxia’ vs. ‘Preoperative Normoxia’ groups by anatomical diagnosis and age. Six patients with anatomical disease defined as cyanotic heart defects crossed over to the ‘Normoxic Group’ for oxygen saturations >90%.

Figure 2

Effect of preoperative hypoxia on cytochrome c oxidase steady state activity; stratified by ‘Preoperative Hypoxia’ vs. ‘Preoperative Normoxia’ groups.

Figure 3

Cytochrome c oxidase steady state activity is increased in the‘Preoperative Hypoxia’ group compared to the ‘Preoperative Normoxia’ group.

Figure 4

Effect of age on cytochrome c oxidase steady state activity. Age in months is plotted on the Log¹⁰ scale and has a positive relationship with CcOx activity. Age along with preoperative SaO₂ as evaluated by multiple linear regression provides a moderate R² value of 0.332, accounting for 33% of the variation in CcOx activity.

Figure 5

Lack of correlation between cytochrome c oxidase activity and cytochrome c oxidase subunit I protein content.

Figure 6

Effect of preoperative hypoxia on cytochrome c oxidase subunit I protein content, ‘Preoperative Hypoxia’ vs. ‘Preoperative Normoxia’ groups.