Ann Thorac Surg 1998;65:719-723
© 1998 The Society of Thoracic Surgeons
Original Articles: Cardiovascular
Granulocyte Elastase Release and Pulmonary Hemodynamics in Patients With Atrial Septal Defect
Hidenori Gohra, MD,
Yoshihiko Fujimura, MD,
Hiroshi Ito, MD,
Kimikazu Hamano, MD,
Tomoe Katoh, MD,
Nobuya Zempo, MD,
Kensuke Esato, MD
First Department of Surgery, Yamaguchi University School of Medicine, Yamaguchi, Japan
Accepted for publication September 15, 1997.
Dr Gohra, First Department of Surgery, Yamaguchi University School of Medicine, 1144 Kogushi, Ube, Yamaguchi 755, Japan.
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Abstract
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Background. In patients with increased pulmonary artery pressure, the pulmonary vascular endothelium is morphologically and functionally abnormal and may be vulnerable to neutrophil-mediated injury induced by cardiopulmonary bypass (CPB). We investigated the relation between levels of granulocyte elastase (GEL), interleukin-6, or interleukin-8 after CPB and preoperative pulmonary hemodynamics or changes in pulmonary function after the operation.
Methods. We measured plasma levels of GEL, interleukin-6, and interleukin-8 before and after CPB in patients who underwent closure of an atrial septal defect. Preoperative and postoperative respiratory index were evaluated. Preoperative pulmonary hemodynamics were determined within 1 month before the operation.
Results. The level of GEL rose significantly after CPB from baseline (164.8 ± 81.3 versus 819.4 ± 320.3 µg/L; p < 0.01). Levels of interleukin-6 and interleukin-8 showed no significant changes after CPB. Peak level of GEL was significantly correlated with preoperative systolic pulmonary artery pressure (r = 0.76; p = 0.017), mean pulmonary artery pressure (r = 0.75; p = 0.021) and pulmonary-to-systemic arterial pressure ratio (r = 0.77; p = 0.016), but not with the hemodynamic variables for pulmonary blood flow or pulmonary resistance. Moreover, the value of (postoperative respiratory index - preoperative respiratory index)/preoperative respiratory index was positively correlated with the peak level of GEL (r = 0.72; p = 0.030).
Conclusions. The increase in GEL level after CPB is proportional to the increase in preoperative pulmonary artery pressure, which may cause the accordant pulmonary vascular damage.
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Introduction
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Cardiopulmonary bypass (CPB) is the most widely used technique for cardiac operations. After CPB, a degree of pulmonary dysfunction develops in many patients [1][2][3]. An activation of inflammatory mediators and neutrophil that is initiated by the exposure of blood to artificial surface during CPB and reperfusion may be responsible for the lung injury [3][4][5]. It has been demonstrated that the level of granulocyte elastase (GEL) released from neutrophils is elevated during CPB [6][7], and GEL has been implicated in neutrophil-mediated tissue injury [8]. The endothelium is the interface between a tissue and activated neutrophils, and may be the primary target in neutrophil-mediated tissue injury. Interleukin-8 (IL-8) and interleukin-6 (IL-6) are chemoattractants for neutrophils and stimulate degranulation of neutrophils, thereby releasing GEL [9][10].
It has been shown that the pulmonary vascular endothelium is morphologically and functionally abnormal in patients with congenital heart defects and pulmonary hypertension [11]. In patients with increased pulmonary artery pressure, pulmonary blood flow, or pulmonary vascular resistance, pulmonary vascular endothelium might be vulnerable to the injury caused by CPB, and high levels of circulating GEL, IL-6, and IL-8 might be shown after CPB.
In the present study, we measured the levels of GEL, IL-6, and IL-8 before and after CPB in adult patients undergoing closure of an atrial septal defect (ASD). We evaluated the relation between these levels and preoperative pulmonary hemodynamic characteristics or the changes in pulmonary function after the operation.
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Patients and Methods
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Patients
Nine patients (5 men and 4 women) undergoing closure of an ASD with CPB were studied. The present study was approved by the ethical committee of our institution. Information about the study was given to each patient, and written consent was obtained according to the guidelines of the ethical committee. Age ranged from 19 to 60 years, with a mean age of 48.0 ± 13.1 years. Cardiac catheterization was performed within 1 month before the operation, and the following values were determined or calculated by standard formulas: systolic pulmonary artery pressure, mean pulmonary artery pressure, pulmonary blood flow, total pulmonary resistance, pulmonary vascular resistance, pulmonary-to-systemic arterial pressure ratio, pulmonary-to-systemic arterial blood flow ratio, pulmonary-to-systemic arterial resistance ratio, and left-to-right shunt ratio. Pulmonary blood flow was determined by the direct Fick method and was used for calculating total pulmonary resistance and pulmonary vascular resistance.
Surgical Management and Cardiopulmonary Bypass
Anesthesia was induced by midazolam, and intermittent positive-pressure ventilation with nitrous oxygen was facilitated by muscle relaxants. Fentanyl was used for maintenance. The operation was performed through a median sternotomy. After heparin (300 IU/kg) was given, the superior and inferior venae cavae and ascending aorta were cannulated to institute CPB. The CPB consisted of a Biopump (Bio-Console; Bio-Medicus Inc, Eden Prairie, MN) and a membrane oxygenator (Capiox-E; Terumo Co, Tokyo, Japan). The circuit was primed with 2.0 to 2.2 L of Ringer’s lactate and colloid solution (albumin or plasma protein fraction). Betamethasone (4 mg/kg) was added to the priming solution. The hematocrit value was 18% to 22% during CPB. A flow rate of 2.4 L · min-1 · m-2 was used to maintain a systemic perfusion pressure of 60 to 80 mm Hg. Multidose cold crystalloid cardioplegia was used for myocardial protection. Five of 9 patients underwent direct closure of the ASD, and 4 patients underwent patch closure of the ASD. The duration and temperature of CPB are shown in Table 1.
Blood Sampling
Samples of arterial blood were obtained via a catheter placed in the radial artery for the measurement of GEL, IL-6, and IL-8 at the following times: (1) before the operation, (2) immediately after cessation of CPB, (3) 24 hours after the operation, (4) 3 days after the operation, and (5) 6 days after the operation. Blood samples were collected into vacuum tubes containing ethylenediaminetetraacetic acid, and were immediately centrifuged (600 g for 10 minutes) at 4°C. Plasma was stored at -70°C until cytokine levels were assayed. Concentration of GEL was measured by enzyme-linked immunosorbent assay using an enzyme immunoassay kit (Merck, Darmstadt, Germany) according to the manufacturer’s instructions. Enzyme-linked immunoassays for IL-6 and IL-8 were performed using an enzyme-linked immunosorbent assay kit (Toray-Fuji Bionics Inc, Tokyo, Japan).
Immediately after induction of anesthesia and after the operation, arterial blood gas analysis was carried out with an inspiratory oxygen concentration of 100% to calculate respiratory index (RI) according to the following formula (inspired oxygen fraction = 1.0, 1 atm, 37°C):
where AaDO2 is the arterial-alveolar difference of oxygen tension, PaO2 is the oxygen tension in the systemic arterial blood, and PaCO2 is the carbon dioxide tension in the systemic arterial blood. The changes in the pulmonary function was evaluated with the value calculated by the following formula:
Statistical Analysis
Data are expressed as the mean ± the standard deviation. Differences between data at different time points were analyzed using repeated-measures analysis of variance and then the Tukey test. Pearson’s correlation tests were performed to compare levels of GEL, IL-6, or IL-8 and measurements obtained by cardiac catheterization, CPB time, aortic cross-clamp time, or RI. Values were considered statistically significant if the p value was less than 0.05.
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Results
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Pulmonary Hemodynamic Characteristics
Preoperative pulmonary hemodynamics are summarized in Table 2. There was 1 patient included in the present study who had pulmonary hypertension (ie, mean pulmonary artery pressure >25 mm Hg).
Changes in Levels of Granulocyte Elastase, Interleukin-6, and Interleukin-8
In all 9 patients, GEL level rose to a peak value immediately after cessation of CPB. The GEL level was significantly higher immediately after cessation of CPB compared with that before the operation (164.8 ± 81.3 versus 819.4 ± 320.3 µg/L; p < 0.01), remained elevated at 24 hours after the operation (448.6 ± 138.9 µg/L; p < 0.05), and returned to the baseline value after 3 days postoperatively (Fig 1). Levels of IL-6 and IL-8 also rose to their peak immediately after cessation of CPB (from 17.2 ± 12.8 to 29.4 ± 17.5 pg/mL, from 3.1 ± 2.6 to 8.8 ± 8.8 pg/mL, respectively) and decreased thereafter. However, no significant changes were found in the levels of IL-6 or IL-8 throughout the period of the present study.
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Changes in the level of granulocyte elastase. The granulocyte elastase level rose significantly after cessation of cardiopulmonary bypass (CPB), remained higher at 24 hours after the operation compared with before the operation, and returned to the baseline after postoperative day 3 (POD).
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Correlation Between Granulocyte Elastase, Interleukin-6, or Interleukin-8 Levels and Cardiopulmonary Bypass Time or Aortic Cross-Clamp Time
The peak levels of GEL and IL-6 were not significantly correlated with CPB time or aortic cross-clamp time. The peak level of IL-8 was positively correlated with CPB time (r = 0.70; p = 0.041) but not with aortic cross-clamp time. There was no significant correlation between the peak level of GEL and that of IL-6 or IL-8.
Correlation Between the Peak Level of Granulocyte Elastase, Interleukin-6, Interleukin-8 and Pulmonary Hemodynamic Variables
The preoperative levels of GEL, IL-6, and IL-8 showed no significant correlation with any of the hemodynamic variables (ie, systolic pulmonary artery pressure, mean pulmonary artery pressure, pulmonary capillary wedge pressure, pulmonary blood flow, total pulmonary resistance, pulmonary vascular resistance, pulmonary-to-systemic arterial pressure ratio, pulmonary-to-systemic arterial blood flow ratio, pulmonary-to-systemic arterial resistance ratio, or left-to-right shunt ratio).
The peak level of GEL was significantly correlated with systolic pulmonary artery pressure (r = 0.76; p = 0.017) (Fig 2), mean pulmonary artery pressure (r = 0.75; p = 0.021) (Fig 3), and pulmonary-to-systemic arterial pressure ratio (r = 0.77; p = 0.016) (Fig 4), but not with pulmonary blood flow, total pulmonary resistance, pulmonary vascular resistance, pulmonary-to-systemic arterial blood flow ratio, pulmonary-to-systemic arterial resistance ratio, or left-to-right shunt ratio (Table 3). The peak levels of IL-6 and IL-8 were not significantly correlated with any of the hemodynamic variables.
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Scattergram showing relation between the peak level of granulocyte elastase (GEL) and preoperative systolic pulmonary artery pressure (sPAP). Regression analysis revealed a significant positive correlation between the peak level of GEL and sPAP ( ; r = 0.76; p = 0.017).
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Scattergram showing relation between the peak level of granulocyte elastase (GEL) and preoperative mean pulmonary artery pressure (mPAP). Regression analysis revealed a significant positive correlation between the peak level of GEL and mPAP ( ; r = 0.75; p = 0.021).
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Scattergram showing relation between the peak level of granulocyte elastase (GEL) and preoperative pulmonary-to-systemic arterial pressure ratio (Pp/Ps). Regression analysis revealed a significant positive correlation between the peak level of GEL and Pp/Ps. (  ; r = 0.77; p = 0.016).
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Changes in Respiratory Index
The value of (
, which could represent the changes in the pulmonary function, was positively correlated with the peak level of GEL (r = 0.72; p = 0.030) (Fig 5).
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Scattergram showing relation between the changes in respiratory index (RI) and the peak level of granulocyte elastase (GEL). Regression analysis revealed a significant positive correlation between the changes in RI and the peak level of GEL. ( ; r = 0.72; p = 0.030).
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Comment
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In the present study, plasma levels of GEL, IL-6, and IL-8 were determined before and after CPB in 9 patients with ASD. We analyzed the correlation between the peak level of GEL, IL-6, or IL-8 and the pulmonary hemodynamic variables obtained preoperatively or the changes in pulmonary function (ie, RI). The level of GEL rose significantly after CPB, but IL-6 and IL-8 levels did not. The peak level of GEL demonstrated positive correlation with the pulmonary arterial pressure, but not with the pulmonary blood flow or pulmonary resistance. The changes in RI were correlated with the peak level of GEL.
Granulocyte elastase is a serine protease stored within the azurophilic granules of the neutrophil and released upon degranulation or disintegration of the cells. Interleukin-6 and IL-8 play an important role in the release of GEL as neutrophil chemotactic factors [9][10]. Granulocyte elastase-mediated pulmonary endothelial and epithelial cell injury has been demonstrated in vitro [8] and in vivo [12]. Granulocyte elastase released from adherent neutrophils can impair the functional integrity of endothelium by degrading collagen, elastin, proteoglycan, and fibronectin, which form the endothelial extracellular matrix and basement membrane [13]. Although we did not confirm that GEL, IL-6, and IL-8 measured in the present study originated from the lung, pulmonary circulation may be a major source of GEL, IL-6, and IL-8 based on the observations that neutrophils are sequestered in the lung after release of the aortic cross-clamp [14] and that neutrophil-mediated lung injury occurs after CPB [15].
In patients with congenital heart defects and an increased pulmonary arterial pressure, the pulmonary vascular endothelial integrity is abnormal [11][16] and thus the levels of GEL, IL-6, and IL-8 might be increased after CPB. To test this theory, we studied patients with ASD because CPB time and aortic cross-clamp time are relatively short; therefore, changes in the levels of GEL, IL-6, and IL-8 might reflect the pulmonary endothelial damage with only a small effect of CPB and aortic cross-clamping itself.
Frering and associates [17] have shown that plasma concentration of IL-6 and IL-8 rose significantly from baseline after CPB in patients who were undergoing valve replacements. Finn and colleagues [5] reported that IL-8 release was elevated significantly after CPB and was correlated with the length of CPB in patients with congenital heart disease. In the present study, the IL-8 level was correlated with CPB time, but IL-6 and IL-8 levels did not change significantly after CPB. This might be because the duration of CPB was relatively short and severe inflammatory response did not develop, and because the betamethasone suppressed the increase in IL-6 and IL-8 levels.
On the other hand, the GEL level was elevated significantly after CPB. The peak level of GEL was not correlated with the level of IL-6 or IL-8, which stimulate GEL release, although Finn and colleagues [5] have shown that GEL release was correlated with IL-8 concentration. These findings suggest that some factor other than neutrophil chemotactic factors (ie, IL-6 and IL-8) caused the degranulation of neutrophils and the release of GEL. Ultrastructural or functional damage of pulmonary vascular endothelium caused by increased pulmonary vascular pressure may be involved in the release of GEL.
The increase in pulmonary artery pressure is associated with an increase in the number of mitochondria and in the volume density of ribosome and rough endoplasmic reticulum in pulmonary vascular endothelium [11][16][18], resulting in increased endothelial metabolic function. Moreover, as an adaptation of the heightened metabolic activity, alterations in endothelial surface appearance such as an increased areal proportion of microvilli and a change in cell shape develop, by which the surface area of cytoplasmic membrane in contact with the lumen is preserved [11]. The altered endothelial surface characteristics associated with an increase in pulmonary artery pressure may lead to abnormal interactions with neutrophils, causing them to degranulate and release GEL, especially when they are activated by CPB. Our finding that the peak level of GEL was positively correlated with the hemodynamic variables for pulmonary artery pressure (ie, systolic pulmonary artery pressure, mean pulmonary artery pressure, and pulmonary-to-systemic arterial pressure ratio) supports this suggestion. The data also suggest that the extent of endothelial injury is related to the degree of increase in pulmonary artery pressure, even when the pulmonary artery pressure does not reach the level of pulmonary hypertension. It is unlikely that neutrophils are activated by altered endothelium before CPB, because the preoperative level of GEL is not correlated with pulmonary artery pressure. We can only speculate that CPB causes the abnormal interaction of altered endothelium with neutrophils through high levels of adhesion molecules on the endothelium, CD11b on neutrophils, or complement activity. Rabinovitch and coworkers [11] have shown that changes in pulmonary vascular endothelial surface are inconsistent in patients with increased pulmonary blood flow or with intimal hyperplasia that contributes to an increase in pulmonary vascular resistance. Their findings could explain our finding that the peak level of GEL was not correlated with the hemodynamic variables representing pulmonary blood flow (ie, pulmonary blood flow, pulmonary-to-systemic arterial blood flow ratio, and left-to-right shunt ratio) or those representing pulmonary vascular resistance (ie, total pulmonary resistance, pulmonary vascular resistance, and pulmonary-to-systemic arterial resistance ratio).
Respiratory index reflects intrapulmonary shunting [19] and is used as an indicator of oxygen transport in the lung [20]. In the present study, the changes in RI were correlated with the peak level of GEL, indicating that severe lung injury occurred in patients with a high level of GEL.
In summary, the results of the present study have shown that the peak level of GEL is correlated with the preoperative pulmonary artery pressure and that the changes in the pulmonary function after the operation are correlated with the peak level of GEL. These results indicate that the increase in the GEL level after CPB is proportional to the increase in the preoperative pulmonary artery pressure, which may cause the accordant pulmonary vascular damage that develops after CPB.
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Abstract
Introduction
