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ORIGINAL RESEARCH |
From the Department of Obstetrics and Gynecology, University Hospital of Puerto Real, Puerto Real, Cadiz, Spain.
Address reprint requests to: José L. Bartha, St. Michael’s Hospital, Fetal Medicine Research Unit, Division of Obstetrics and Gynaecology, Southwell Street, Bristol BS2 8EG, United Kingdom; E-mail: j.bartha{at}bristol.ac.uk.
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ABSTRACT |
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METHODS: Sixty-six women in three groups were prospectively studied: 1) women diagnosed with fetal growth restriction (n = 22), 2) women with preeclampsia (n = 22), and 3) healthy pregnant women (controls) frequency matched for age, parity, and gestational age (n = 22). Primary outcomes were maternal serum concentrations of both ß-hCG and VEGF. Placental insufficiency was defined by a pulsatility index in umbilical artery greater than the 99th percentile for gestation.
RESULTS: Maternal serum concentrations of ß-hCG and VEGF were greater in women with preeclampsia than in controls (P = .001 and P = .002, respectively) and women with fetal growth restriction (P = .002 and P = .002, respectively). Concentrations did not differ between women with fetal growth restricted fetuses and controls. Correlation between ß-hCG and VEGF was not significant in any of the studied groups. Serum VEGF concentrations were significantly increased in a subgroup of 12 women with placental insufficiency (P = .04) and correlated with ß-hCG concentrations (r = .63, P = .02).
CONCLUSION: Both VEGF and ß-hCG maternal serum concentrations were increased in women with preeclampsia but normal in women with fetal growth restriction, although VEGF concentrations were increased in those cases with placental insufficiency. Maternal serum ß-hCG and VEGF concentrations did not correlate except in women with placental insufficiency.
Several studies have reported a significant association between unexplained elevations of maternal sernum human chorionic gonadotropin (hCG) concentrations during the second trimester of pregnancy and a range of subsequent adverse pregnancy outcomes, including fetal growth restriction (FGR) and preeclampsia.1–6 However, the underlying mechanism supporting this association remains unclear.
There are several links between hCG and vascular endothelial growth factor (VEGF). In nonpregnant women, hCG influences VEGF production. Exposure of human granulosa cells to hCG stimulates the expression of VEGF messenger ribonucleic acid to promote the vascularization of the corpus luteum.7 In a similar way, administration of hCG in women undergoing in vitro fertilization increases urinary VEGF concentrations.8
In pregnant women, profiles of longitudinal concentrations of serum hCG are positively correlated with profiles of VEGF in early pregnancy, and this correlation is independent of gestational age.9 Recently, this novel function for hCG as an angiogenic factor has been recognized.10
On the other hand, VEGF receptors have been found in human trophoblast, and it has been suggested that VEGF might also play a role in the growth and differentiation of cytotrophoblast at implantation.11 Therefore, VEGF might also indirectly stimulate hCG production. Thus, hCG and VEGF might have a reciprocal relationship in pregnancy.
The aim of the present study was to study the relationships between hCG and VEGF in either normal pregnant women during late pregnancy or in women with pregnancy complications.
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MATERIALS AND METHODS |
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Fetal growth restriction was defined first during gestation as a fetal abdominal circumference below the 10th percentile for gestation and second at delivery as a weight below the 10th percentile for gestation according to our own data adjusted to our local population.
Preeclampsia was considered in cases of previously healthy, normotensive women with a blood pressure recording of 140/90 mm Hg or higher on two or more occasions, at least at 4-hour intervals, developing after the 20th week of pregnancy, and with proteinuria. Proteinuria was defined as 300 or more mg protein per 24 hours, assessed by 24-hour urine collections.
Placental insufficiency was defined as a pulsatility index in umbilical artery greater than the 99th percentile for gestation.
Venous blood was collected by venepuncture in 10-mL silicone-coated Vacutainer blood-collecting tubes containing no additives between 7:00 AM and 9:00 AM in a fasting state. Blood was allowed to clot at room temperature and was then centrifuged for 20 minutes at 2000g. Aliquots of the serum were then stored at -80°C until they were required for the assay.
Total VEGF was determined with use of a commercially available chemiluminescent immunoassay (Research and Diagnostics Systems, R&D Systems Inc., Minneapolis, MN). It consists of a solid-phase enzyme-linked immunosorbent assay designed to measure VEGF165 concentrations. It contains Sf 21-expressed recombinant human VEGF165 and antibodies raised against the recombinant protein. The minimum detectable dose of VEGF ranged from 0.78 to 2.95 pg/mL. The intra- and interassay variation coefficients were 3.2% and 6.8%, respectively.
Beta-human chorionic gonadotropin was measured by a standard automated method (microparticle enzymoimmunoassay with the AxSYM System, Abbot Laboratories, Abbott Park, IL).
Statistical analysis was performed with SPSS software (SPSS Inc., Chicago, IL). We were looking for relevant correlations between hCG and VEGF. Therefore, we considered a correlation coefficient greater than 0.5 for calculating the sample size. We calculated that 20 were needed women in each group to show a significant correlation with an 
level of .05 and a power of 80% for a correlation coefficient of 0.6. We enrolled 22 women in each group to account for withdrawals. Distributions were checked with histogram and Kolmogorov–Smirnov test. Data are presented as mean and standard deviation for normally distributed variables. In cases of nonnormal variables, data are shown as median and interquartile range. Qualitative variables are expressed as numbers and percentages.
To compare groups, we first used one-way analysis of variance in cases of normal variables and Kruskal-Wallis test in cases of nonnormal variables. Post hoc analysis with Neuman-Keuls test was used to compare groups. Because both VEGF and ß-hCG were distributed in a nonnormal manner, Spearman correlation coefficient was used to study the relationships between these variables. To compare proportions (qualitative variables), the 
2 test and the Fisher exact test (when expected numbers were less than 5) were used. Statistical significance was set at the 95% level (P < .05).
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RESULTS |
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). There were seven cases (31.8%) of FGR in the preeclamptic group.
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). For ß-hCG, post hoc tests showed that these significant differences were between women with preeclampsia and controls (P = .001) and between preeclampsia and FGR (P = .002), but not between FGR and controls. For VEGF, the significant differences were also between preeclampsia and control (P = .002) and between preeclampsia and FGR (P = .002), but again there was no difference between women with FGR and controls.
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There were 12 cases of Doppler-defined placental insufficiency (seven in the group of FGR and five in the preeclamptic group). All these cases showed absence of end-diastolic flow or reverse flow in the umbilical artery Doppler. Three of the five patients with preeclampsia had also FGR. The gestational age of this group was 34.8 ± 3.6 weeks. Maternal serum VEGF concentrations were significantly greater in these patients than in the control group (P = 04) (Table 3). Similarly, serum ß-hCG concentrations were greater in these women, but the difference with the control group did not reach statistical significance. In this particular group of women, there was a significant correlation between the concentrations of ß-hCG and VEGF (r = .63, P = .02).
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DISCUSSION |
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Maternal concentrations of VEGF are also increased in women with preeclampsia, and VEGF has been suggested to have an important role in the pathophysiology of the disease12 and proposed as a marker for that condition.13
We found increased concentrations of both ß-hCG and VEGF in women with preeclampsia, but we did not find a significant link between them. This strongly suggests the presence of different pathophysiologic mechanisms for these two different secretory responses.
As with preeclampsia, elevations of midtrimester maternal serum ß-hCG have been found to be associated with an increased risk for low birth weight and small for gestational age infants.14 However, there are conflicting reports as to the predictive value of ß-hCG concentrations for FGR.1,15 Women with FGR and normal umbilical Doppler measurements (small for gestational age) have normal concentrations of serum ß-hCG, but they seem to be increased in cases of FGR due to Doppler-defined placental insufficiency.16 In fact, umbilical artery Doppler velocimetry might serve as a predictor of pregnancy outcome in the high-risk group characterized by unexplained elevated ß-HCG.17
At present, there is no strong evidence to suggest a role for VEGF in FGR. Actually, some studies have showed no differences in placental expression of VEGF between normal placentas and placentas from FGR fetuses.18,19 However, these studies defined FGR on the basis of low birth weight, and in one of them19 cases with placental insufficiency were specifically excluded. VEGF expression is upregulated by hypoxia,20 and it is known that FGR infants, particularly those associated with placental insufficiency, show signs of chronic hypoxia at delivery. Along this line, a strong correlation between VEGF staining in villous syncytiotrophoblasts and cord hematocrit, an indirect indicator of fetal oxygenation, has been found in cases of FGR infants.19 Therefore, it is logical to hypothesize that VEGF is increased in cases of placental hypoxia, as has been demonstrated in vitro.21 On the other hand, it has been hypothesized that decreased VEGF production as a primary factor could be involved in abnormal placental angiogenesis and be a cause of some cases of FGR.22 Thus, either increased or decreased expression of VEGF might theoretically be possible in cases of FGR, depending on whether these changes in VEGF expression are a cause or consequence of this condition.
We found no alterations in maternal serum VEGF concentrations and no significant correlation between VEGF and ß-hCG in pregnancies complicated by FGR. However, when cases with placental insufficiency were analyzed separately, we found both increased serum concentrations of VEGF and significant correlation with ß-hCG.
Our results suggest that in cases of Doppler-defined placental insufficiency, both secretory phenomena might be linked to the same stimulus. Whether or not placental hypoxia is the stimulus for that remains unknown, but in experiments in vitro, hypoxia differently influences the production of VEGF and the production of hCG. Hypoxia resulted in an increase in VEGF production but had inconsistent effects on hCG production.21 However, the present study was not designed to evaluate the relationships between hCG and VEGF in cases of placental insufficiency. This was a secondary outcome, and in this group both preeclamtic and FGR cases were mixed. Therefore, further studies are needed to either confirm or deny our findings.
In conclusion, maternal serum concentrations of both VEGF and hCG were increased in women with preeclampsia but normal in cases of FGR, although VEGF concentrations were increased in women with placental insufficiency. We found no significant correlation between maternal serum concentrations of hCG and VEGF in any of the studied groups, except in women with placental insufficiency associated with either preeclampsia or FGR, for which both secretory phenomena might be linked.
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Footnotes |
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Received April 1, 2003. Received in revised form June 11, 2003. Accepted July 10, 2003.
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REFERENCES |
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2. Aquilina J, Maplethorpe R, Ellis P, Harrington K. Correlation between second trimester maternal serum inhibon-A and human chorionic gonadotropin for the prediction of pre-eclampsia. Placenta 2000;21:487–92.[Medline]
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16. Bartha JL, Comino-Delgado R, Arrabal J, Escobar MA. Third-trimester maternal serum beta-HCG level and umbilical blood flow in fetal growth retardation. Int J Gynecol Obstet 1997;57:27–31.[Medline]
17. Yaron Y, Jaffa AJ, Har-Toov J, Lavi H, Legum C, Evans MI. Doppler velocimetry of the umbilical artery as a predictor of outcome in pregnancies characterized by elevated beta-subunit human chorionic gonadotropin. Fetal Diagn Ther 1997;12:353–5.[Medline]
18. Lash G, MacPherson A, Liu D, Smith D, Charnock-Jones S, Baker P. Abnormal fetal growth is not associated with altered chorionic villous expression of vascular endothelial growth factor mRNA. Mol Hum Reprod 2001;7: 1093–8.
19. Tse JY, Lao TT, Chan CC, Chiu PM, Cheung AN. Expression of vascular endothelial growth factor in third-trimester placentas is not increased in growth-restricted fetuses. J Soc Gynecol Invest 2001;8:77–82.[Medline]
20. Matsumoto LC, Bogic L, Brace RA, Cheung CY. Prolonged hypoxia upregulates vascular endothelial growth factor messenger RNA expression in ovine fetal membranes and placenta. Am J Obstet Gynecol 2002;186: 303–10.[Medline]
21. Taylor CM, Stevens H, Anthony FW, Wheeler T. Influence of hypoxia on vascular endothelial growth factor and chorionic gonadotrophin-derived cell lines: JEG, Jar and BeWo. Placenta 1997;18:451–8.[Medline]
22. Ashmed A, Perkins J. Angiogenesis and intrauterine growth restriction. Baillieres Clin Obstet Gynaecol 2000; 14:981–98.
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