ORIGINAL RESEARCH
Plasma Placenta Growth Factor Levels in Midtrimester Pregnancies May Lee Tjoa, MSc, John M. G. van Vugt, MD, PhD, Monique A. M. Mulders, Ruud B. H. Schutgens, PhD, Cees B. M. Oudejans, MD, PhD, and Inge J. van Wijk, PhD OBJECTIVE: Previous studies have shown decreased levels of placenta growth factor in serum of pregnant women with preeclampsia. The aim of this study was to investigate whether levels of placenta growth factor are decreased before the clinical onset of preeclampsia, and whether placenta growth factor levels are decreased in pregnancies complicated by intrauterine growth restriction. METHODS: From an ongoing longitudinal study, 101 plasma samples were collected from 72 pregnant women at weeks 11–21 of gestation. Placenta growth factor levels were determined retrospectively in plasma using an enzyme-linked immunosorbent assay. Correlations between plasma concentrations of placenta growth factor and pregnancy outcome were evaluated. RESULTS: Plasma samples of 72 patients were analyzed. Forty-four patients had no pregnancy complications, 18 developed preeclampsia, and 10 women had pregnancies complicated by intrauterine growth restriction. Between week 17 and week 21 of pregnancy, a significantly lower level of placenta growth factor was found in plasma of patients who later developed preeclampsia (n ⴝ 10), compared with control pregnancies (n ⴝ 25, P ⴝ .004). In women with a growth-restricted baby at birth (n ⴝ 5), levels of placenta growth factor were also low. CONCLUSIONS: Our results show that plasma placenta growth factor levels are decreased before preeclampsia is clinically evident. The data suggest that placenta growth factor may be useful to determine the relative risk of developing preeclampsia and intrauterine growth restriction. (Obstet Gynecol 2001;98:600 –7. © 2001 by the American College of Obstetricians and Gynecologists.)
From the Molecular Biology Laboratory, Department of Clinical Chemistry (MLT, MAMM, RBHS, CBMO, IJvW) and the Department of Obstetrics and Gynecology, Vrije Universiteit Medical Centre, Amsterdam, The Netherlands (JMGvV). This study was supported by a grant from the Dutch Praeventiefonds/ZON (#28-3022). The authors thank C. Mulder, MSc, A.A. Verstraeten, PhD, and E.R.A. Peters-Muller, MSc, for statistical analysis of the data, and the doctors from the Department of Obstetrics for their support. We thank R&D systems for providing ELISA Placenta Growth Factor kits.
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Preeclampsia is one of the leading causes of maternal and fetal morbidity and mortality. It is characterized by hypertension, proteinuria, and edema, and deliveries are often premature. Intrauterine growth restriction is frequently— but not necessarily—associated with preeclampsia. Less than 10% of all cases of preeclampsia occur before 32 completed weeks of gestation.1 Removal of the placenta is still the only cure for preeclampsia. Inappropriate development of the placenta is considered to be one of the primary causes of preeclampsia, although the exact pathogenesis remains unclear.2 Adequate placental function, required for successful pregnancy, depends on appropriate trophoblast invasion. During early pregnancy, extravillous trophoblast cells invade the maternal space, nestling within the wall of the maternal spiral arteries.3 The endovascular trophoblast cells adopt an endothelial phenotype, which is thought to lead to the modification of the spiral arteries into high-flow, low-resistance vessels.4 Shallow trophoblast invasion by the extravillous trophoblasts is the primary placentation defect in pregnancies complicated by intrauterine growth restriction, with or without preeclampsia. It has been shown that in these pregnancies the modification of the spiral arteries is restricted to the superficial portion of the decidua.5–7 It is also known that hypoxia regulates the trophoblast differentiation pathway. The first weeks of embryogenesis occur under relative hypoxic conditions, but as pregnancy progresses and true blood flow is established within the fetoplacental unit, the oxygen tension rises.8 In preeclampsia and (mild) intrauterine growth restriction, placental hypoxia continues to occur to the extent that the triggering of trophoblast differentiation and invasion fails to happen. The end result of this failure is the inability of the uterine vasculature to accommodate the fetus to an increase in blood flow with increasing gestational age. Adequate placentation also relies on the establishment and maintenance of a fetoplacental vascular network. The best-known angiogenic growth factor studied in the human placenta is vascular endothelial growth fac-
VOL. 98, NO. 4, OCTOBER 2001 © 2001 by The American College of Obstetricians and Gynecologists. Published by Elsevier Science Inc.
0029-7844/01/$20.00 PII S0029-7844(01)01497-1
Table 1. Clinical Characteristics of the Patient Population of Pregnant Women (Gestational Age, Weeks 11–21) for Plasma Placental Growth Factor Analysis
Maternal age at delivery (y) Gestational age at delivery (d) Number of primigravidae Highest diastolic blood pressure during pregnancy (mmHg) Placental weight (g) Birth weight (g)
Normal pregnancy (n ⫽ 44)
Preeclampsia (n ⫽ 18)
34 (⫾4) 275 (⫾11) 14 75 (⫾6)
34 (⫾4) 268 (⫾11) 5 100 (⫾9)*
700 (⫾122) 3565 (⫾435)
620 (⫾160) 3100 (⫾660)‡
Intrauterine growth restriction (n ⫽ 10) 33 (⫾5) 270 (⫾7) 4 75 (⫾6) 515 (⫾90)† 2585 (⫾135)*
The P values listed are calculated in comparison with the normal pregnancy population. * P ⬍ .001. † P ⬍ .01. ‡ P ⬍ .05.
tor.9 –11 Vascular endothelial growth factor stimulates endothelial cell growth and migration and angiogenesis during embryonic development. Recently, an additional member of this family of angiogenic factors, placenta growth factor, was identified.12 Placenta growth factor shares 53% homology with vascular endothelial growth factor at the amino acid level. Placenta growth factor and its receptor are expressed predominantly by trophoblast cells, which implies a potential autocrine role of placenta growth factor in regulating trophoblast function.13,14 However, the direct effects of placenta growth factor on various stages of angiogenesis have yet to be established.15 Unlike vascular endothelial growth factor, expression of placenta growth factor is downregulated by hypoxia.16 Placenta growth factor levels, as determined in serum, rise during the first and second trimesters of pregnancy, peaking during the early third trimester, before declining sharply.17 Reduced levels of circulating placenta growth factor are found in the maternal serum of women with preeclamptic pregnancies compared with normal pregnancies.17,18 These studies were conducted using serum samples from women with clinically manifested preeclampsia (in weeks 30 – 40 of gestation). We hypothesize that placenta growth factor is involved in the early process of placentation, and that levels of placenta growth factor are already reduced before 20 weeks’ gestation in women who will later develop preeclampsia and/or deliver a growth-restricted baby, compared with uncomplicated pregnancies. Analyses of placenta growth factor concentrations were conducted with first- and second-trimester plasma samples and correlated with clinical outcome. MATERIALS AND METHODS The protocol for this study was approved by the Medical Ethics Committee of the Vrije Universiteit Medical Centre, and procedures were in accordance with the ethical standard for human experimentation established by the
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Declaration of Helsinki of 1975, revised 1983. After we obtained written informed consent from study participants, we collected 25–30 mL of heparin blood from 72 pregnant women from our ongoing longitudinal study, once or twice during the first and second trimesters of pregnancy. The gestational age varied from week 11 to week 21 of pregnancy. All pregnancies were carefully monitored, and all laboratory and clinical data were stored in a database. The most important clinical characteristics of the patient population are listed in Table 1. Preeclampsia was defined as a blood pressure of more than 140 mmHg systolic or more than 90 mmHg diastolic in a woman who was normotensive before 20 weeks’ gestation, together with new-onset proteinuria, which is defined as more than 300 mg in a 24-hour specimen, according to the National High Blood Pressure Education Program Working Group on High Blood Pressure in Pregnancy.19 Cases complicated by pregnancy-induced hypertension, diabetes mellitus, renal disease, gestational diabetes, fetuses with congenital anomalies, intrauterine fetal death, and multifetal gestations were excluded from the study. All normotensive women were healthy throughout pregnancy and were admitted for delivery. Intrauterine growth restriction of neonates was defined as a birth weight under the 10th percentile for gestational age, according to Kloosterman.20 Birth weight and placental weight were determined immediately after delivery. For this study, the group with intrauterine growth restriction included only cases not complicated by preeclampsia. All blood samples were processed on the day of withdrawal, which has shown to be crucial for obtaining reliable results from the enzyme-linked immunosorbent assays (ELISAs). The heparin blood samples were subjected to density centrifugation on a multilayer Percoll gradient to separate blood cells for isolation of circulating trophoblast cells as described previously.21 Note that blood is diluted 2:1 in Hanks’ balanced salt solution
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(HBSS; Invitrogen Life Technologies, Carlsbad, CA) before loading on the gradient. A 2-mL plasma sample was taken from the top of the gradient after centrifugation and stored at ⫺80C until further use in the ELISA. For placenta growth factor analyses, 101 samples were processed. The distribution was as follows: 1) in weeks 11–14: 21 women total, 11 of which were normal, eight developed preeclampsia, and two delivered a growthrestricted baby; 2) in weeks 14 –17: 42 women total, 26 of which were normal, nine developed preeclampsia, and seven delivered a growth-restricted baby; and 3) in weeks 17–21 of gestation: 40 women total, 25 of which were normal, 10 developed preeclampsia, and five delivered a growth-restricted baby. Plasma levels of placenta growth factor were determined by means of an ELISA (R&D Systems, Abingdon, United Kingdom). According to the manufacturer, both recombinant and natural human placenta growth factor–1 are accurately measured with a minimum of 7 ng/L and a maximum of 1000 ng/L. Instructions were followed as provided by the manufacturer. For each sample, 100 L of plasma was used, and all samples were assayed in duplicate. The coefficient of interassay variation was 11%. For evaluation of the clinical characteristics of the patient population, test results are expressed as mean ⫾ standard deviation. One-way analysis of variance (ANOVA) followed by the Tukey test for multiple comparison was used when appropriate. If distribution was skewed, the Kruskal-Wallis test followed by the MannWhitney U test was used. Multiple regression analysis was performed for the three groups with placenta growth factor levels as dependent variable. A single sample per woman was included (the sample with the most advanced gestational age). Analysis of variance with a covariant (ANCOVA) using gestational age as covariant was used to test the differences in placenta growth factor levels between the three patient groups (normal, preeclampsia, and intrauterine growth restriction) at different gestational intervals. However, when placenta growth levels appeared to be independent of gestational age in a particular gestational period, the Kruskal-Wallis test was used, followed by the Mann-Whitney U test. Two consecutive blood samples were obtained from 29 patients; eight had developed preeclampsia, and four had developed intrauterine growth restriction. Blood samples were obtained at successive gestational ages. The increase in placenta growth factor levels was measured by calculating the slope of the line between the two points. The Mann-Whitney U test was used to compare the increases in the placenta growth factor levels in the three patient groups.
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To analyze correlations between placenta growth factor levels and placenta weight or birth weight, the Pearson’s correlation coefficient was calculated. A P value of less than .05 was considered statistically significant. All analyses were performed using the Statistical Package for Social Sciences, v 9.0 (SPSS Inc., Chicago, IL). RESULTS In Table 1, the clinical characteristics of the patient populations are shown. No difference was found in the average maternal age and the average gestational age at delivery in the three groups (ie, pregnancies without complications [controls], pregnancies with preeclampsia, and pregnancies with an intrauterine growth–restricted baby at birth). The highest diastolic blood pressure was found to be significantly different between the preeclampsia group and the other two groups (P ⬍ .001). Placenta weight at birth was significantly different between normal pregnancies and pregnancies complicated by intrauterine growth restriction (P ⫽ .006). The placenta weights did not differ significantly between the normal and preeclampsia groups (P ⫽ .185) or between the preeclampsia and intrauterine growth restriction groups (P ⫽ .242). The Mann-Whitney U test was used to evaluate the differences in birth weights between the three groups. Birth weights were significantly lower in the intrauterine growth–restricted group when compared with both the normal group (P ⬍ .001) and the preeclampsia group (P ⫽ .021). The birth weights of babies born after a preeclamptic pregnancy were also significantly lower than those in the normal group (P ⫽ .012). Although the majority of our patient population was composed of secundigravidas, a considerable number of primigravidas were included (Table 1). No differences were found in outcome of the placenta growth factor analyses between nulli- and multiparous patients. Before 11 weeks of pregnancy, levels of immunodetectable placenta growth factor were generally below detection levels (less than 15 ng/L), and therefore were not included. In Figure 1, placenta growth factor levels are shown for all 72 subjects between 11 and 21 weeks of pregnancy. Multiple regression analysis, with the normal group defined as baseline, showed that placenta growth factor levels in the preeclampsia group and in the intrauterine growth–restricted group were both significantly lower than in the normal group (P ⫽.011 for both). Regression lines are included in Figure 1. Between 11 and 14 weeks of pregnancy, the levels of placenta growth factor in plasma are relatively low (varying from 20 ng/L to 50 ng/L). Starting at 14 weeks’ gestation, the placenta growth factor levels rise gradu-
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Figure 1. Cross-sectional representation of plasma placenta growth factor levels during the first and second trimester of pregnancy (gestational age: weeks 11–21). Regression lines are included. Levels were determined in 72 pregnant women, 44 of whom had a normal pregnancy outcome (open circles, solid line), 18 developed preeclampsia (closed squares, broken line), and 10 developed intrauterine growth restriction (gray triangles, dotted line). Tjoa. Placenta Growth Factor. Obstet Gynecol 2001.
ally, and placenta growth factor levels were subsequently analyzed in two separate periods of gestation: weeks 14 –17 and weeks 17–21. As the placenta growth factor levels rose with increasing gestational age, we performed the ANCOVA test. There was a significant increase in
placenta growth factor levels with gestational age (P ⬍ .001) in the first time period studied. However, we could not demonstrate a difference in placenta growth factor concentrations among the three groups studied (P ⫽ .135) in this first gestational interval (Table 2 and Figure 2). In contrast, the second period, between 17 and 21 weeks’ gestation, a statistically significant difference was demonstrated among the three groups (P ⫽ .004), with placenta growth factor dependent on gestational age (P ⫽ .049). Further analysis showed that there is a significant difference in placenta growth factor levels between the normal and preeclampsia groups (P ⫽ .007). When comparing the placenta growth factor levels between the normal and the intrauterine growth restriction groups, the expression levels were significantly less (P ⫽ .026). However, in both cases, placenta growth factor was apparently independent of gestational age in this time interval (P ⫽ .118 and P ⫽ .075, respectively). For this reason, and because of the small number of patients included in these groups, we also evaluated the differences in placenta growth factor in weeks 17–21 using the Kruskal-Wallis test, followed by the Mann-Whitney U test. The difference between the normal and the preeclampsia group remained statistically significant (P ⫽ .004). However, the difference between the women with an intrauterine growth–restricted baby at birth and the normal group decreased (P ⫽ .057) (Table 3 and Figure 3). A correlation between placental weight at delivery and placenta growth factor levels measured between 17 and 21 weeks’ gestation was not observed for either normal pregnancies or preeclampsia (Pearson’s correlation coefficient ⫽ ⫺0.13 and ⫺0.24; P ⫽ .59 and P ⫽ .60, respectively). Similarly, no correlation was found between birth weight and placenta growth factor levels measured between 17 and 21 weeks’ gestation in these groups (Pearson’s correlation coefficient ⫽ 0.26 and 0.29; P ⫽ .19 and P ⫽ .40, respectively). Analysis of the increase in placenta growth factor levels in individual patients was performed when two consecutive blood samples were obtained. No differences in increase were found between the normal and preeclampsia groups (P ⫽ .63), the normal and intrauter-
Table 2. Placenta Growth Factor Concentration in Plasma of Pregnant Women in Weeks 14 –17 of Gestation
Normal Preeclampsia IUGR
Mean gestational age in weeks ⫹ days, (range) 15 ⫹ 4 (14–17) 15 ⫹ 5 (14 ⫹ 4–17) 15 ⫹ 4 (14–17)
Number
Mean PlGF concentration (ng/L)
Standard deviation (ng/L)
Range (ng/L)
26 9 7
68.8 46.6 46.6
28.2 31.3 25.7
23.9–117.6 17.7–99.5 23.2–98.1
PlGF ⫽ placenta growth factor; IUGR ⫽ intrauterine growth restriction.
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Figure 2. Placenta growth factor levels in weeks 14 –17 of gestation. A single sample was included per patient. Open circles, healthy patients (n ⫽ 26); closed squares, patients with preeclampsia (n ⫽ 9); and open triangles, patients with a pregnancy with intrauterine growth restriction (n ⫽ 7). We found no significant difference between normal and preeclampsia pregnancies or between normal and intrauterine growth-restricted pregnancies. Tjoa. Placenta Growth Factor. Obstet Gynecol 2001.
ine growth–restricted groups (P ⫽ .999), and the preeclampsia and intrauterine growth–restricted groups (P ⫽ .68). However, the placenta growth factor levels in the first blood samples were generally lower in preeclampsia patients than in normal patients, as well as in all consecutive blood samples. This is in agreement with the aforementioned analyses demonstrating a statistically significant difference in placenta growth factor levels measured in weeks 17–21 of gestation in normal pregnancies compared with pregnancies destined to develop preeclampsia. DISCUSSION In women with overt preeclampsia as analyzed beyond 30 weeks of gestation, Torry et al17 and Reuvekamp et al18 detected decreased levels of circulating placenta
growth factor. Both studies argue that the selective deficit of this angiogenic growth factor plays an important role in placentation during the first trimester of pregnancy. Placental dysfunction is considered to be one of the primary causes of preeclampsia. In a large number of pregnancies complicated by intrauterine growth restriction, trophoblast invasion and modification of uteroplacental spiral arteries is also impaired.5,7 We hypothesized that defective trophoblast invasion forms the primary defect in the etiology of both preeclampsia and intrauterine growth restriction,5–7 and that placenta growth factor concentrations might be aberrant in these patients before manifestation of clinical symptoms. Indeed, as shown in the present study, decreased plasma levels of placenta growth factor were found in the second trimester (weeks 17–21) in patients who developed pre-
Table 3. Placenta Growth Factor Concentration in Plasma of Pregnant Women in Weeks 17–21 of Gestation
Normal Preeclampsia IUGR
Mean gestational age in weeks ⫹ days (range) 18 ⫹ 5 (17 ⫹ 1–20 ⫹ 3) 18 ⫹ 6 (17 ⫹ 3–20) 19 (17 ⫹ 2–20 ⫹ 4)
Number
Mean PlGF concentration (ng/L)
Standard deviation (ng/L)
Range (ng/L)
25 10 5
124.1 83.8* 83.5†
39.8 12.3 22.5
54.7–200.2 64.4–106.8 44.3–99.8
PlGF ⫽ placenta growth factor; IUGR ⫽ intrauterine growth restriction. * P ⫽ .004. † P ⫽ .057.
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Figure 3. Placenta growth factor levels in weeks 17–21 of gestation. A single sample was included per patient. Open circles, healthy patients (n ⫽ 25); closed squares, patients with preeclampsia (n ⫽ 10); and open triangles, patients with a pregnancy with intrauterine growth restriction (n ⫽ 5). We found a significant difference between normal and preeclampsia pregnancies (**, P ⫽ .004). There is a tendency toward a statistically significant difference between normal and intrauterine growth–restricted pregnancies (*, P ⫽ .057). Tjoa. Placenta Growth Factor. Obstet Gynecol 2001.
eclampsia in the third trimester of pregnancy and in women who delivered a growth-restricted baby. The regression lines in Figure 1 show that the increase in placenta growth factor levels occurs in all three studied groups. However, the concentration of placenta growth factor in plasma remains lower in the preeclampsia and intrauterine growth restriction groups. These differences are not found in the first period (weeks 14 –17) of gestation, whereas the differences become statistically significant in the second period (weeks 17–21) of gestation for preeclampsia. The data from Torry et al17 and Reuvekamp et al18 show that in third trimester pregnancies these differences in levels of circulating placenta growth factor will be even more pronounced. The intrauterine growth restriction group shows similar placenta growth factor levels to the preeclampsia group, although the difference is not statistically significant. To be able to draw any firm conclusions, the number of patients in this group needs to be increased. The data presented in this study were obtained using plasma samples; preliminary experiments show that com-
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parable results are obtained using serum samples (data not shown). In a preliminary study we similarly analyzed placenta growth factor concentrations in plasma of women carrying twins. The plasma placenta growth factor concentrations are considerably higher in these women when compared with singleton pregnancies (data not shown). These additional data confirm the validity of the assay as an indicator of placental function, as placenta growth factor is predominantly synthesized by trophoblast cells and the placental mass is increased in twin pregnancies. However, we showed that there is no statistically significant correlation between the placental weight and placenta growth factor or between the birth weight and placenta growth factor for all three groups. Therefore, the decreased placenta growth factor levels in preeclampsia— characterized in this study by normal placental weight—rather indicate that levels of placenta growth factor might be representative for placental function rather than mass. Aberrant expression of placenta growth factor in early pregnancy indicates a potential role for this growth factor in placental function. The study by Desai et al22 describes a
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potential function of placenta growth factor in trophoblast cells through activation of the stress-activated protein kinase pathways. Activating the stress-activated protein kinase responses has been shown to regulate apoptosis. This raises the possibility that placenta growth factor might mediate trophoblast survival, by protecting against apoptosis.22 Interestingly, in placentas of both patients with preeclampsia and in pregnancies where there is intrauterine growth restriction, increased levels of apoptotic trophoblast cells have been found.23,24 Possibly in both pregnancy complications, decreased levels of placenta growth factor contribute to increased trophoblast apoptosis, which results in placental dysfunction. The majority of complications analyzed in our study were mild. No obvious correlation was found between the severity of the pregnancy complication (as measured by the levels of hypertension, proteinuria, and the gestational age at the onset of the disease) and placenta growth factor levels in the circulation. It will be interesting to determine cellular placenta growth factor concentrations in term placentas of these populations and to determine plasma placenta growth factor levels in severe and earlyonset cases of preeclampsia and intrauterine growth restriction. Contradicting data concerning placenta growth factor levels in pregnancy16,17 are probably due to the existence of three alternative transcripts: placenta growth factor 1, 2, and 3.25 In most studies—including this one—no distinction is made between these three subtypes. Detailed analysis of the expression profiles and specific function of all three subtypes separately, in early and term placental tissue and in serum/plasma samples throughout pregnancy, would be helpful in elucidating the role of placenta growth factor in pregnancy. Despite the differences found between the three studied groups, there is an overlap of placenta growth factor concentrations between study and control populations. Therefore, the use of placenta growth factor as an early marker, to identify individual patients destined to develop preeclampsia or intrauterine growth restriction, requires further evaluation (eg, of the three different subtypes generated by alternative splicing).
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phenotype as they differentiate. J Clin Invest 1997;99: 2139 –51. 5. Khong TY, De Wolf F, Robertson WB, Brosens I. Inadequate maternal vascular response to placentation in pregnancies complicated by preeclampsia and by small-forgestational age infants. Br J Obstet Gynaecol 1986;93: 1049 –59. 6. Zhou Y, Damsky CH, Fisher SJ. Pre-eclampsia is associated with failure of human cytotrophoblasts to mimic a vascular adhesion phenotype. J Clin Invest 1997;99: 2152– 64. 7. Sheppard BL, Bonnar J. Uteroplacental hemostasis in intrauterine fetal growth retardation. Semin Thromb Hemost 1999;25:443– 6. 8. Zhou Y, Genbacev O, Damsky CH, Fisher SJ. Oxygen regulates human cytotrophoblast differentiation and invasion: Implication for endovascular invasion in normal pregnancy and pre-eclampsia. J Reprod Immunol 1998;39: 197–213. 9. Ferrara N. Molecular and biological properties of vascular endothelial growth factor. J Mol Med 1999;77:527– 43. 10. Smith SK, He Y, Clark DE, Charnock-Jones DS. Angiogenic growth factor expression in placenta. Semin Perinatol 2000;24:82– 6. 11. Ahmed A, Dunk C, Ahmad S, Khaliq A. Regulation of placental vascular endothelial growth factor (VEGF) and placenta growth factor and soluble Flt-1 by oxygen–a review. Placenta 2000;21:S16 –S24. 12. Maglione D, Guerriero V, Viglietto G, Delli-Bovi P, Persico MG. Isolation of a human placenta cDNA coding for a protein related to the vascular permeability factor. Proc Natl Acad Sci USA 1991;88:9267–71. 13. Khaliq A, Li XF, Shams M, Sisi P, Acevedo CA, Whittle MJ, et al. Localisation of placenta growth factor (PlGF) in human term placenta. Growth Factors 1996;13:243–50. 14. Clark DE, Smith SK, Licence D, Evans AL, CharnockJones DS. Comparison of expression patterns for placenta growth factor, vascular endothelial growth factor (VEGF), VEGF-B and VEGF-C in the human placenta throughout gestation. J Endocrinol 1998;159:459 – 67. 15. Torry DS, Ahn H, Barnes EL, Torry RJ. Placenta growth factor: Potential role in pregnancy. Am J Reprod Immunol 1999;41:79 – 85. 16. Khaliq A, Dunk C, Jiang J, Shams M, Li XF, Acevedo C, et al. Hypoxia down-regulates placenta growth factor, whereas fetal growth restriction up-regulates placenta growth factor expression: Molecular evidence for “placental hyperoxia” in intrauterine growth restriction. Lab Invest 1999;79:151–70. 17. Torry DS, Wang HS, Wang TH, Caudle MR Torry RJ. Pre-eclampsia is associated with reduced levels of placenta growth factor. Am J Obstet Gynecol 1999;179:1539 – 44. 18. Reuvekamp A, Velsing-Aarts FV, Poulina IEJ, Capello JJ, Duits AJ. Selective deficit of angiogenic growth factors
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characterises pregnancies complicated by pre-eclampsia. Br J Obstet Gynaecol 1999;106:1019 –22. 19. National High Blood Pressure Education Program Working Group on High Blood Pressure in Pregnancy. Report of the National High Blood Pressure Education Program Working Group on High Blood Pressure in Pregnancy. Am J Obstet Gynecol 2000;183:S1–S22. 20. Kloosterman GJ. On intrauterine growth: The significance of prenatal care. Int J Gynaecol Obstet 1970;8:895–912. 21. van Wijk IJ, van Vugt JMG, Mulders MAM, Ko¨nst AAM, Weima SM, Oudejans CBM. Enrichment of fetal trophoblast cells from the maternal peripheral blood followed by detection of fetal deoxyribonucleic acid with a nested X/Y polymerase chain reaction. Am J Obstet Gynecol 1996; 174:871– 6. 22. Desai J, Holt-Shore V, Torry RJ, Caudle MR, Torry DS. Signal transduction and biological function of placenta growth factor in primary human trophoblast. Biol Reprod 1999;60:887–92.
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23. Genbacev O, DiFederico E, McMaster M, Fisher SJ. Invasive cytotrophoblast apoptosis in pre-eclampsia. Hum Reprod 1999;14:59 – 66. 24. Axt R, Kordina AC, Meyberg R, Reitnauer K, Mink D, Schmidt W. Immunohistochemical evaluation of apoptosis in placentae from normal and intrauterine growth restricted pregnancies. Clin Exp Obstet Gynecol 1999;26: 195– 8. 25. Cao Y, Ji W-R, Qi P, Rosin A, Cao Y. Placenta growth factor: Identification and characterization of a novel isoform generated by RNA alternative splicing. Biochem Biophys Res Comm 1997;235:493– 8. Address reprint requests to: Inge J. van Wijk, PhD, Molecular Biology Laboratory, Department of Clinical Chemistry, Vrije Universiteit Medical Centre, de Boelelaan 1117, 1081 HV Amsterdam, The Netherlands; E-mail:
[email protected]. Received December 27, 2000. Received in revised form May 30, 2001. Accepted June 7, 2001.
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