PEDIATRICS Vol. 113 No. 5 May 2004, pp. 1348-1351
Placenta Growth Factor Elevation in the Cord Blood of Premature Neonates Predicts Poor Pulmonary Outcome
Po-Nien Tsao, MD*,
Shu-Chen Wei, MD
,
Yi-Ning Su, MD
,
Chien-Nan Lee, MD, MPH||,
Hung-Chieh Chou, MD*,
Wu-Shiun Hsieh, MD* and
Fon-Jou Hsieh, MD||
* Department of Pediatrics, National Taiwan University Hospital, National Taiwan University College of Medicine, Taipei, Taiwan
Department of Internal Medicine, National Taiwan University College of Medicine, Taipei, Taiwan
Department of Medical Genetics, National Taiwan University College of Medicine, Taipei, Taiwan
|| Department of Obstetrics and Gynecology, National Taiwan University Hospital, National Taiwan University College of Medicine
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ABSTRACT
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Objective. To determine whether an elevated placenta growth factor (PlGF) level in cord blood is associated with increased risk for preterm infants to develop bronchopulmonary dysplasia (BPD).
Methods. Sixty-three preterm infants who were born at 34 weeks' gestation or earlier were enrolled. Two infants who died before 28 days' postnatal age could not be assigned a BPD status and were excluded. PlGF levels in cord blood were measured using enzyme-linked immunosorbent assay. Mann-Whitney rank sum test, Spearman correlation coefficients, and multivariable linear or logistic regression analyses were used for statistical analysis.
Results. The BPD group had a higher PlGF level, lower gestational age, lower birth weight (BW), higher incidence of endotracheal tube intubation, and longer duration of intubation. The PlGF levels in cord blood correlated negatively with gestational age and BW. However, multivariable logistic regression analyses revealed that only elevated cord blood PlGF levels and BW were associated with BPD after adjusting for all contributing factors. Furthermore, an increased PlGF level in cord blood was significantly correlated with the clinical severity of BPD, as measured by duration of intubation. At 17 mg/dL, the specificity of cord blood PlGF level in predicting BPD was 95%, the sensitivity was 53%, the positive predictive value was 83%, and the negative predictive value was 82%.
Conclusions. Measuring cord blood PlGF level at birth might be a biological marker for predicting the occurrence of BPD and allowing early therapeutic intervention.
Key Words: bronchopulmonary dysplasia • cord blood • placenta growth factor
Abbreviations: BPD, bronchopulmonary dysplasia • PlGF, placenta growth factor • VEGF, vascular endothelial growth factor • GA, gestational age • RDS, respiratory distress syndrome • BW, birth weight
With the widespread use of antenatal steroids, exogenous surfactant therapy, and improvements in neonatal care, the survival rate of very low birth weight infants has increased, but bronchopulmonary dysplasia (BPD) remains 1 of the major complications in premature infants who need prolonged ventilator support. The incidence of BPD ranges from 7.5% to 20% in infants who are born before 34 weeks.1,2
The cause of BPD includes immaturity, prolonged oxygen therapy, barotrauma, volume trauma, infection, and antioxidant/oxidant imbalance.3,4 In premature infants with BPD, the pathologic findings include alveolar hypoplasia, vascular arrest, adaptive dysmorphic changes, and variable interstitial proliferation.5,6 However, only a few candidate biological makers might be able to predict which infants are at greater risk for developing BPD.7,8 Consequently, early therapeutic intervention is difficult.
Placenta growth factor (PlGF), a member of the vascular endothelial growth factor (VEGF) family, is a 132–amino acid, 50-kDa dimeric glycoprotein. Present in normal tissues, especially the placenta, thyroid, and lungs, it is an important mediator of angiogenesis and hematopoiesis.9–13 In our previous study, we demonstrated that PlGF overexpression transgenic mice have enlarged airspace, similar to the pathologic findings of infants with BPD.14 The aim of this study was to determine whether PlGF levels in the cord blood could predict increased risk for subsequent BPD in preterm infants.
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METHODS
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Study Group
This study was performed at the National Taiwan University Hospital with the approval of the internal review board. All infants who were born at 34 weeks' gestation or earlier were enrolled in this study. Infants were excluded when there was evidence of prenatal maternal infection or infection within the first 3 days of life. We determined gestational age (GA) by the last menstrual date or prenatal ultrasound. Weekly prenatal steroids were routinely used when preterm labor occurred from 24 to 34 weeks of gestation. Respiratory distress syndrome (RDS) was defined as acute respiratory failure at birth with characteristic chest radiograph changes in the absence of sepsis, pneumonia, or other causes of respiratory distress. Exogenous surfactant was administered within 2 hours after birth to infants who had RDS and remained ventilator dependent and required a fraction of inspired oxygen >0.4 to maintain pulse oximeter saturation >90%. Infants, ventilated, requiring <40% oxygen and ventilator rate <18/min, were considered by the clinical management team to be ready for extubation. BPD was defined as the need for supplemental oxygen or mechanical ventilation at 28 days' postnatal age, in association with radiologic changes consistent with BPD. Demographic information and perinatal history were obtained from medical records.
Collection of Cord Blood and PlGF Measurement
Cord blood was collected in heparinized syringes on delivery and centrifuged within 15 minutes of collection. The plasma was kept at –70°C until analysis by a technician who was blinded to the patients' condition. The level of PlGF in the cord blood was assayed by a standardized sandwich enzyme-linked immunosorbent assay method (R&D Systems, Minneapolis, MN) in duplicate according to the manufacturer's protocol.
Data Analysis
Comparisons between unpaired groups were performed by Mann-Whitney rank sum test. The relationships between PlGF and GA, birth weight (BW), and ventilator days were analyzed by Spearman correlation coefficients. Multivariable analyses were conducted using logistic and linear regression. P < .05 was considered statistically significant.
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RESULTS
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Sixty-three neonates were included in this study. Two infants who died before 28 days' postnatal age were excluded because they could not be assigned a BPD status. Eighteen (30%) infants developed BPD. The mean GA in this series was 30.0 ± 2.8 weeks; the mean BW was 1.3 ± 0.5 kg. The BPD group had a lower GA, lower BW, higher incidence of endotracheal tube intubation, antenatal steroid use, and longer duration of intubation (Table 1). In addition, the BPD group had a higher PlGF level in cord blood (Fig 1). PlGF levels were below the sensitivity level (<7 pg/mL) of our assay in 1 (6%) BPD infant and 17 (40%) non-BPD infants (P = .008).
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Fig. 1. Cord blood levels of PlGF were significantly increased in infants with BPD compared with unaffected infants.
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The PlGF levels in cord blood correlated negatively with GA (r = –0.633, P < .001; Fig 2) and BW (r = –0.270, P = .035). However, using multivariable logistic regression with BPD as the dependent variable revealed that only PlGF level (P = .011) and BW (P = .024) were independently correlated (Table 2). In addition, infants with RDS had a higher PlGF level (P = .018). Only 6 premature infants with BPD could be matched by GA (28–34 weeks) with infants without BPD. These infants also demonstrated higher cord blood PlGF levels (12.3 mg/dL [8.8–33.4] vs 8.1 mg/dL [7.0–23.8]). However, the difference was not significant because of the small sample size.
Infants with a lower GA (P < .001), lower BW (P < .001), higher PlGF levels (P = .029), or RDS (P < .001) had a higher incidence of endotracheal intubation. After logistic regression analysis with endotracheal tube intubation as the dependent variable, only RDS (P = .035; odds ratio: = 7.5) and GA (P = .035; odds ratio: 0.6) were significantly correlated.
More and more evidence shows that duration of oxygen therapy is a less accurate surrogate to predict long-term pulmonary outcome because the criteria for the use of oxygen is rarely defined.15,16 Therefore, we used the duration of ventilation as another pulmonary outcome. The duration of intubation negatively correlated with GA (P < .001) and BW (P < .001) but positively correlated with the PlGF levels in cord blood (P < .001; Fig 3). Multivariable linear regression with intubation days as the dependent variable revealed that only PlGF level (P = .033) was independently correlated (Table 3).
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Fig. 3. Cord blood levels of PlGF showed significant linear correlation with the clinical severity of BPD as measured by duration of intubation.
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Additional analysis was performed using the alternative definition of BPD as supplemental oxygen or mechanical ventilation at 36 weeks' postmenstrual age (BPD36 weeks).17 Nine (15%) of 61 infants developed BPD36 weeks. The BPD36 weeks group also had a lower GA (27 weeks [24–34] vs 31 weeks [24–34]; P = .005), lower BW (828 g [638–1070] vs 1360 g [620–2328]; P < .001), longer duration of intubation (45 days [0–109] vs 0 days [0–45]; P = .001), and higher PlGF levels (15.1 mg/dL [8.8–474] vs 9.4 mg/dL [7–27.3]; P = .013).
A receiver operating characteristics curve was used to determine the best cutoff point of cord blood PlGF to discriminate between infants with and without BPD. The specificity of cord blood PlGF >17 pg/mL for BPD was 95%, the sensitivity was 53%, the positive predictive value was 83%, and the negative predictive value was 82%.
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DISCUSSION
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In this study, we demonstrated that PlGF is elevated in cord blood from preterm infants who subsequently developed BPD. Moreover, the concentrations of PlGF seemed to be closely related to the number of days on ventilator. In addition, the PlGF levels in cord blood correlated significantly and negatively with both GA and BW.
The VEGF family includes PlGF. It binds to the VEGF-1 receptor but not to the VEGF-2 receptor; the latter is thought to mediate the angiogenic and proliferative effects of VEGF. PlGF mRNA is present in normal tissues such as the placenta, thyroid, and lungs.10 Previously, we demonstrated that PlGF mRNA was detected in lungs during saccular division. It was downregulated rapidly after alveolarization was completed in mice.14 However, little is known about PlGF expression in human lungs during the last trimester of pregnancy. In this study, we demonstrated that PlGF levels in human cord blood declined with increasing GA, suggesting that PlGF may play a role during human lung development as a synergistic agent with VEGF to promote vasculogenesis.9 It should be downregulated after lung maturation.
In our previous report, we found that overexpression of PlGF in mice caused air space enlargement similar to BPD. Furthermore, we demonstrated that exogenous PlGF inhibited proliferation and promoted death of mouse type II pneumocytes in vitro.14 On the basis of these data, PlGF may exert a key influence on pulmonary remodeling by regulating type II pneumocyte proliferation or death. In this study, we showed that PlGF levels in cord blood were significantly elevated in the BPD group. Although PlGF levels were significantly and negatively correlated with GA, which is smaller in infants with BPD, PlGF still was a significant risk factor of BPD after multivariable logistic regression analysis. Furthermore, the PlGF levels in cord blood were independently correlated with the severity of BPD, as represented by the duration of intubation. These data revealed that PlGF might play a role, via its action on type II pneumocytes, in the pathogenesis of BPD.
The mechanisms that lead to increase PlGF in cord blood of infants with BPD remain unknown. Some suggested that there may be a genetic susceptibility to BPD.18–20 In our previous report, we found that maternal PlGF concentration was elevated in Down syndrome pregnancies during the early second trimester.21 We postulate that the additional copy of chromosome 21 might in some way upregulate the expression of PlGF gene or downregulate its degradation. In this study, we demonstrated that elevated cord blood PlGF level was an independent risk factor to develop BPD; it preceded clinical evidence of BPD. Overproduction of PlGF during alveolarization, which may have a genetic predisposition, might be responsible for the subsequently impaired saccular division and BPD.
Recently, 2 reports showed evidence that measuring bombesin-like peptide and keratinocyte growth factor concentrations during the early postnatal course could predict BPD. Cullen et al7 found that elevated urine bombesin-like peptide levels at 1 to 4 days after birth in preterm infants who were born before 28 weeks' gestation increased the risk of developing BPD. Danan et al8 showed that keratinocyte growth factor concentration within 5 days after birth was significantly higher in survivors without BPD than in those with BPD. However, using these 2 markers to predict BPD will be possible until 4 to 5 days after birth. Here, we demonstrated that using cord blood PlGF levels might predict BPD at birth.
In conclusion, the PlGF levels in cord blood correlated significantly and negatively with GA and BW. The cord blood PlGF elevation preceded BPD and was an independent risk factor. High PlGF levels in cord blood were significantly correlated with the clinical severity of BPD, indicating that PlGF might play a role in the pathogenesis of BPD. The measurement of cord blood PlGF level at birth might be a biological marker to permit early intervention and might provide a new therapeutic target to prevent BPD.
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ACKNOWLEDGMENTS
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We thank Hwai-I Yang for the excellent statistical analysis.
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FOOTNOTES
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Received for publication May 23, 2003; Accepted Sep 19, 2003.
Reprint requests to (F.J.H.) Department of Obstetrics and Gynecology, College of Medicine, National Taiwan University Hospital, No. 7, Chung-Shan South Road, Taipei, Taiwan. E-mail: fjhsieh{at}ha.mc.ntu.edu.tw
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PEDIATRICS (ISSN 1098-4275). ©2004 by the American Academy of Pediatrics
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