Brain injury with systemic inflammation in newborns with congenital heart disease undergoing heart surgery

Abstract

The potential role of systemic inflammation on brain injury in newborns with congenital heart disease (CHD) was assessed by measuring levels of central nervous system (CNS)-derived proteins in serum prior to and following cardiac surgery. A total of 23 newborns (gestational age, 39±1 weeks) with a diagnosis of CHD that required cardiac surgery with cardiopulmonary bypass (CPB) were enrolled in the current study. Serum samples were collected immediately prior to surgery and 2, 24 and 48 h following CPB, and serum levels of phosphorylated neurofilament-heavy subunit (pNF-H), neuron-specific enolase (NSE) and S100B were analyzed. Systemic inflammation was assessed by measuring serum concentrations of complement C5a and complement sC5b9, and the following cytokines: Interleukin (IL)-1β, IL-6, IL-8, IL-10, IL12p70, interferon γ and tumor necrosis factor (TNF)-α. Analysis of cord blood from normal term deliveries (n=26) provided surrogate normative values for newborns. pNF-H and S100B were 2.4- to 2.8-fold higher (P<0.0001) in patient sera than in cord blood prior to surgery and remained elevated following CPB. Pre-surgical serum pNF-H and S100B levels directly correlated with interleukin (IL)-12p70 (ρ=0.442, P<0.05). pNF-H was inversely correlated with arterial pO₂ prior to surgery (ρ=-0.493, P=0.01) and directly correlated with arterial pCO₂ post-CPB (ρ=0.426, P<0.05), suggesting that tissue hypoxia and inflammation contribute to blood brain barrier (BBB) dysfunction and neuronal injury. Serum IL12p70, IL-6, IL-8, IL-10 and TNF-α levels were significantly higher in patients than in normal cord blood and levels of these cytokines increased following CPB (P<0.001). Activation of complement was observed in all patients prior to surgery, and serum C5a and sC5b9 remained elevated up to 48 h post-surgery. Furthermore, they were correlated (P<0.05) with low arterial pO₂, high pCO₂ and elevated arterial pressure in the postoperative period. Length of mechanical ventilation was associated directly with post-surgery serum IL-12p70 and IL-8 concentrations (P<0.05). Elevated serum concentrations of pNF-H and S100B in neonates with CHD suggest BBB dysfunction and CNS injury, with concurrent hypoxemia and an activated inflammatory response potentiating this effect.

Keywords: S100B; cardiopulmonary bypass; cytokines; neonates; neuron specific enolase; phosphorylated neurofilament heavy chain.

Figure 1.

Immunoblot analysis of pNF-H in patient sera and mouse brain tissue. Molecular weight is indicated as kilodaltons (kDa). NF-H, neurofilament heavy chain subunit; NF-M, medium molecular weight subunit of neurofilament as identified in mouse brain.

Figure 2.

Concentration of CNS biomarkers in patient sera taken immediately prior to surgery (pre-) and at 2, 24 and 48 h post-CPB. Scatter plot represents individual patient data. Box plot: box boundaries indicates 25th and 75th percentile; line marks the median; whiskers indicate 10th and 90th percentiles. Statistical significance was determined by RM-ANOVA followed by Bonferroni post-hoc analysis for pairwise comparisons. *P<0.01; **P<0.001; ***P<0.0001. CPB, NSE, neuron-specific enolase; S100B, B subunit of calcium-binding protein S100; pNF-H, phosphorylated neurofilament heavy chain; repeated measures-ANOVA, analysis of variance; CPB, cardiac surgery with cardiopulmonary bypass.

Figure 3.

Concentration of cytokines in patient sera taken immediately prior to surgery (pre-) and 2, 24 and 48 h post-CPB. Scatter plot represents individual patient data. Box plot: box boundaries indicates 25th and 75th percentile; line marks the median; whiskers indicate 10th and 90th percentiles. Statistical significance was determined by RM-ANOVA followed by Bonferroni post-hoc analysis for pairwise comparisons. *P<0.01; **P<0.001; ***P<0.0001. CPB, cardiac surgery with cardiopulmonary bypass; RM-ANOVA, analysis of variance; IL, interleukin; INF, interferon; TNF, tumor necrosis factor.

Figure 4.

Concentration of complement in patient sera taken immediately prior to surgery and at 2, 24 and 48 h post-CPB. Scatter plot represents individual patient data. Box plot: box boundaries indicates 25th and 75th percentile; line marks the median; whiskers indicate 10th and 90th percentiles. Statistical significance was determined by RM-ANOVA followed by Bonferroni post-hoc analysis for pairwise comparisons. *P<0.01; ***P<0.0001.

Figure 5.

Spearman correlation analysis between brain biomarkers, pNF-H, S100B and cytokine IL-12p70 in patient sera obtained prior to surgery demonstrated similar correlation coefficients of ρ=0.442 and P<0.05. The straight line represents the best-fit linear regression between pNF-H and IL-12p70. Triangles or circles represent individual patient serum S100B or pNF-H values, respectively, plotted vs. serum IL-12p70. IL, interleukin; pNF-H, phosphorylated neurofilament-heavy subunit.