- Email: [email protected]
Maternal serum and umbilical cord blood leptin concentrations with fetal growth restriction
Maternal Serum and Umbilical Cord Blood Leptin Concentrations With Fetal Growth Restriction Marcella Pighetti, MD, Giovanni A. Tommaselli, MD, Antonio D’Elia, MD, Costantino Di Carlo, MD, Angela Mariano, MD, Angela Di Carlo, MD, and Carmine Nappi, MD OBJECTIVE: To ascertain whether fetal growth restriction is associated with alterations of leptin concentrations in umbilical cord blood and maternal serum. METHODS: Maternal serum and umbilical cord blood leptin concentrations were determined by immunoradiometric assay at term in 43 women with uncomplicated singleton pregnancies (group A) and in 27 women with singleton pregnancies complicated by fetal growth restriction (group B), all with normal pregravid body mass index (BMI). RESULTS: Maternal serum leptin concentrations were significantly higher in group B compared with group A (45.0 ng/mL [range 34.2–54.9] versus 29.0 ng/mL [range 24.7– 33.3]; P < .01). Umbilical cord blood leptin levels were significantly lower in group B compared with group A (8.4 ng/mL [range 3.6 –13.2] versus 13.1 ng/mL [9.7–16.5]; P < .01). Maternal serum leptin levels were not significantly correlated with maternal BMI or with neonatal birth weight in either group. Umbilical cord blood leptin concentrations were significantly correlated with neonatal birth weight in both groups. CONCLUSION: Growth restricted fetuses at term show umbilical cord blood leptin concentrations significantly lower than those in normal fetuses, suggesting that fetal adipose tissue is a major source of leptin. Maternal serum leptin concentrations are higher in the presence of a growth restricted fetus. This increase might be due to an intrinsic placental mechanism, by which small placentas produce more leptin as a compensatory mechanism, or to early hypoxia. (Obstet Gynecol 2003;102:535– 43. © 2003 by The American College of Obstetricians and Gynecologists.)
Leptin is a 167-amino-acid protein mainly produced by adipocytes. It is thought to act on the hypothalamic centers, regulating energy expenditure and signalling the amount of body fat mass.1 Indeed, its serum levels are positively correlated with fat mass, percentage body fat, and body mass index (BMI).2 Leptin also exerts a number of metabolic functions, including an action on the From the Departments of Obstetrics and Gynecology and Clinical Pathology, University of Naples “Federico II,” Naples, Italy.
reproductive axis: ob/ob mice, mutants that lack circulating leptin, are infertile, and the administration of recombinant human leptin to these animals restores fertility.3,4 During pregnancy, maternal serum leptin levels progressively increase, peaking during the second trimester and then plateauing to a level at term three to four times higher than in nonpregnant women.5 This increase is only in part due to the weight gain experienced by pregnant women, because leptin levels are no longer significantly related to BMI during gestation.6 Furthermore, it has been demonstrated that leptin is produced by the placenta7 and that placental weight is significantly related to leptin levels in the umbilical cord blood.8 Thus, placental leptin might contribute to the increase of maternal serum leptin concentration, even though it is not known to what extent. Leptin levels in umbilical cord blood (ie, in the fetus), significantly increase starting by 34 weeks of pregnancy.9 A significant correlation exists between umbilical cord blood leptin concentration and neonatal ponderal index.10,11 This suggests that leptin might have a role in the regulation of fetal growth. Indeed, several authors evaluated umbilical cord blood leptin levels in pregnancies complicated by fetal growth restriction and found contrasting results. The aim of this study was to evaluate serum and umbilical cord blood leptin concentrations in normal and growth restricted pregnancies and correlate them with maternal BMI and birth weight of the newborns. MATERIAL AND METHODS Fifty women with uncomplicated singleton pregnancies and 30 women with singleton pregnancies complicated by asymmetric fetal growth restriction, all with normal pregravid BMI (20 –27 kg/m2), were consecutively enrolled from the outpatient clinic of our department. No subject was affected by diabetes or hypertension, and no patient developed complications during pregnancy. The diagnosis of fetal growth restriction was made when the
VOL. 102, NO. 3, SEPTEMBER 2003 © 2003 by The American College of Obstetricians and Gynecologists. Published by Elsevier.
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abdominal circumference by ultrasound measurement was below the tenth percentile of reference values for fetuses of similar age in Italy.12 Fetal growth restriction diagnosis was confirmed at birth if birth weight was below the tenth percentile according to Italian standards for birth weight and gestational age.13 The study was approved by our institutional review board. All patients gave their informed and written consent for participation in the study. All ultrasonographic measurements were performed by the same operator (MP) with the use of a Toshiba Powervision 6000 with a 5-MHz transabdominal convex probe (Toshiba Medical Systems, Rome, Italy). A maximum of 7 days elapsed between the final ultrasonography and births. Maternal blood samples were drawn from the antecubital vein upon hospitalization in our obstetric ward for parturition and centrifuged. All women were fasting and not in labor. Arterial umbilical cord blood samples were collected at the time of parturition and centrifuged. All sera were stored at ⫺20C until determination of leptin levels. Serum leptin levels were determined in duplicate with a human leptin immunoradiometric assay (Diagnostic Systems Laboraties Inc., Webster, TX) with a sensitivity of 0.1 ng/mL, an intraassay coefficient of variation of 2.6 – 4.9%, and an interassay coefficient of variation of 3.7– 6.6%. Statistical analysis of the data was performed on an IBM-compatible personal computer with the Statistical Package for Social Science 8.0 (SPSS Inc., Chicago, IL). Data distribution was assessed with the Shapiro-Wilk test. Maternal serum and umbilical cord blood leptin levels were nonparametric variables, and a Mann-Whitney U test was used to assess differences between groups. The other variables displayed a normal distribution, and an analysis of variance followed by Scheffe` procedure for post hoc comparison of means was used to identify differences between groups and between different times within the same group. Correlations between maternal serum and umbilical cord blood leptin levels were regressed on maternal BMI and neonatal birth weight for both normal pregnancies and pregnancies complicated by fetal growth restriction and were evaluated with the Spearman R test. Significance was set at P ⬍ .05. RESULTS Seven women with uncomplicated pregnancies and three women with fetal growth restricted pregnancies did not deliver in our department and were considered drop-outs. Thus, the study refers to 43 women with normal pregnancy (group A) and 27 women with fetal growth restriction (group B).
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Figure 1. Maternal serum leptin concentrations in normal pregnancies (group A) and pregnancies complicated by fetal growth restriction (group B). Horizontal lines represent median. *P ⬍ .05. Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
No statistical difference was observed between group A and group B in age (25.3 ⫾ 4.5 years in group A versus 27.1 ⫾ 3.2 years in group B), parity (1.4 ⫾ 0.3 in group A versus 1.2 ⫾ 0.4 in group B), gestational age at parturition (38.6 ⫾ 0.6 weeks in group A versus 37.2 ⫾ 1.1 weeks in group B), and BMI (24.7 ⫾ 0.9 kg/m2 in group A versus 24.5 ⫾ 0.9 kg/m2 in group B). Mean birth weight of newborns from group B was significantly lower compared with newborns from group A (1384 ⫾ 360 g in group B versus 3300 ⫾ 417 g in group A; P ⬍ .001). There was a similar number of female and male infants in both groups (group A: 17 male [40%], 26 female [60%]; group B: 12 male [44%], 15 female [56%]). Mean placental weight was significantly higher in group A compared with group B (620 ⫾ 215 g versus 385 ⫾ 130 g; P ⬍ .01). Maternal serum leptin concentrations were significantly higher in group B compared with group A (45.0 ng/mL [range 34.2–54.9] versus 29.0 ng/mL [range 24.7– 33.3]; P ⬍ .01) (Figure 1). Umbilical cord blood leptin levels were significantly lower in group B compared with group A (8.4 ng/mL [range 3.6 –13.2] versus 13.1 ng/mL [range 9.7–16.5]; P ⬍ .01) (Figure 2). No significant difference was observed in umbilical cord blood leptin levels between male and female fetuses (data not shown). Umbilical cord blood leptin concentrations were significantly correlated with neonatal birth weight in both groups (group A: R ⫽ .448, P ⬍ .01; group B: R ⫽ .876, P ⬍ .01) (Figure 3) but not with maternal BMI (group A: R ⫽ ⫺.240, P ⫽ nonsignificant; group B: R ⫽ ⫺.350, P ⫽ nonsignificant) (Figure 4). Maternal serum leptin lev-
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Figure 2. Umbilical cord blood leptin concentrations in normal pregnancies (group A) and pregnancies complicated by fetal growth restriction (group B). Horizontal lines represent median. *P ⬍ .05. Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
els were not significantly correlated with maternal BMI (group A: R ⫽ .298; group B: R ⫽ .300; P ⫽ nonsignificant) or with neonatal birth weight (group A: R ⫽ .287; group B: R ⫽ .291; P ⫽ nonsignificant) (Figures 5, 6). All women diagnosed as having a fetal growth restricted pregnancy delivered a small for gestational age infant. DISCUSSION In this study, we demonstrated that leptin concentrations in the umbilical cord blood of growth restricted newborns are significantly lower compared with normal for age infants. This result is in agreement with previous studies.9,14 –16 Because fetuses affected by growth restriction and small for gestational age newborns have a significantly reduced fat tissue accumulation,17 it has been hypothesized that lower leptin levels in umbilical cord blood found in these fetuses were determined by the limited amount of fetal fat tissue. Indeed, large for gestational age newborns have higher umbilical cord blood leptin levels compared with appropriate for gestational age (AGA) and small for gestational age newborns.11,14,18 However, determination of leptin concentrations in umbilical cord blood from growth restricted fetuses has shown discrepancies, with studies reporting lower14,15,18,19 but also higher20 leptin levels compared with normal fetuses. Indeed, Jaquet et al9 found that, at term, serum leptin concentrations were significantly lower in newborns with fetal growth restriction than in those with normal growth, whereas no differences were found before 34 weeks’ gestation. Similarly, Cetin et al16 found that growth restricted fetuses show umbilical cord blood
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leptin levels significantly lower than do AGA fetuses after 34 weeks. These authors also found no significant differences between fetal growth restriction and AGA when adjusting for fetal weight (leptin per kg). Both these studies seem to indicate that, after 34 weeks’ gestation, leptin synthesis and secretion are correlated with the development and growth of adipose tissue. Conversely, some studies report higher umbilical cord blood leptin levels in growth restricted fetuses.20 This might be owing to the role played by fetal oxygenation and acid– base balance, because it has been observed that growth restricted fetuses with signs of severe fetal distress have significantly higher leptin concetrations per kilogram of fetal weight than do AGA fetuses.16 Our fetal growth restriction population showed no sign of fetal distress and lower leptin levels than did the AGA controls. It is therefore possible that umbilical cord blood leptin levels depend more on adipose tissue and on fetal oxygenation status than on other antenatal development factors. More studies are needed to evaluate the role of fetal well-being on leptin secretion and leptin secretion patterns in different types of fetal growth restriction. It has been demostrated that the placenta is a major source of leptin during pregnancy.21 Because maternal serum leptin levels progressively rise during the first two trimesters of pregnancy, it has been hypothesized that placental leptin might substantially contribute to the determination of these concentrations.21 It is not yet clear why during the third trimester, when estrogenic and progestinic stimuli on leptin secretion are at their peak and placental weight and size at their maximum, maternal serum leptin concentrations remain unmodified. The finding of higher maternal serum leptin levels in fetal growth restriction-complicated pregnancies is difficult to explain. Leptin inhibits neuropeptide Y secretion in the hypothalamus in mice22 and, although there is no evidence,23 it is hypothesized that the same relationship exists in humans. It has been demonstrated that hypoxia induces an increase of leptin secretion by choriocarcinoma cells. Dotsch et al24 demonstrated that, in preeclampsia, elevated placental leptin expression is accompanied by lower placental neuropeptide Y messenger ribonucleic acid levels. Because neuropeptide Y is a potent vasoconstrictor,24 it is tempting to hypothesize that the placental neuropeptide Y–leptin axis counteracts the vasoconstriction associated with preeclamptic changes in the placenta. It might be that in fetal growth restriction-complicated pregnancies the first sign of hypoxia is an increase of placental leptin production, mirrored by elevated maternal serum leptin levels.25 Further
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Figure 3. Relationship between umbilical cord blood leptin levels and neonatal birth weight in normal pregnancies (A) (Spearman R ⫽ .448; P ⬍ .001) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .876; P ⬍ .001). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 4. Relationship between umbilical cord blood leptin levels and maternal body mass index in normal pregnancies (A) (Spearman R ⫽ ⫺.240; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ ⫺.350; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 5. Relationship between maternal serum leptin levels and maternal body mass index in normal pregnancies (A) (Spearman R ⫽ .287; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .291; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 6. Relationship between maternal serum leptin levels and neonatal birth weight in normal pregnancies (A) (Spearman R ⫽ .298; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .3; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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studies evaluating growth restricted fetuses with and without hypoxia evaluated by uterine and placental blood flow are needed, to assess whether maternal leptin serum levels can be used as a marker of fetal hypoxia. Furthermore, an inverse correlation between placental weight and maternal serum leptin concentrations has been demonstrated in normal pregnancies.8 Thus, in the presence of low-weight placentas, such as those found in cases of fetal growth restriction, leptin secretion, and therefore maternal serum concentrations, should be increased. Lea et al15 demonstrated that placentas from pregnancies complicated by fetal growth restriction express lower amounts of leptin compared with normal placentas. A possible explanation might be that the mean gestational age of the patients in Lea et al’s study was 33.2 ⫾ 1.88 weeks, whereas in our study the women with fetal growth restriction were almost all at term. This might have influenced the amount of placental leptin produced. Alternatively, the mechanisms involved in the compensation of growth restricted fetuses might not yet be established by 33 weeks’ gestation but only at term. In this study we confirmed that growth restricted fetuses at term show umbilical cord blood leptin concentrations significantly lower than those in normal fetuses, indicating that fetal adipose tissue is a major source of leptin. This conclusion is confirmed by the positive correlation between umbilical cord blood leptin concentrations and neonatal birth weight found in both groups. Furthermore, we also demonstrated that mothers with fetal growth restricted fetuses have higher serum leptin concentrations compared with mothers with uncomplicated pregnancies. This increase might be due to an intrinsic placental mechanism, by which small placentas produce more leptin as a compensatory mechanism, or to early hypoxia. Further research is needed to ascertain the role of hypoxia in growth restricted fetuses in determining placental leptin secretion and the intraplacental function of leptin in these situations.
REFERENCES 1. Zhang Y, Proenca R, Maffei M, Barone M, Leopold L, Friedman JM. Positional cloning of the mouse obese gene and its human homologue. Nature 1994;372:425–32. 2. Considine RV, Sinha MK, Heiman ML, et al. Serum immunoreactive leptin concentrations in normal-weight and obese women. N Engl J Med 1996;334:292–5. 3. Barash IA, Cheung CC, Weigle DS, Ren H, Kabigting EB, Kuijper JL, et al. Leptin is a metabolic signal to the reproductive system. Endocrinology 1996;137:3144–7.
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4. Chehab FF, Lim ME, Lu R. Correction of sterility defect in homozigous obese female mice by treatment with the human recombinant leptin. Nat Genet 1996;12:318–20. 5. Hardie L, Trayhurn P, Abramovich D, Fowler P. Circulating leptin in women: A longitudinal study in the menstrual cycle and during pregnancy. Clin Endocrinol 1997; 47:101–6. 6. McCarthy JF, Misra DN, Roberts JM. Maternal plasma leptin is increased in preeclampsia and positively correlates with fetal cord concentration. Am J Obstet Gynecol 1999; 180:731–6. 7. Masuzaki H, Ogawa Y, Isse N, Satoh N, Okazaki T, Shigemoto M, et al. Human obese gene expression: Adipocyte-specific expression and regional differences in the adipose tissue. Diabetes 1995;44:855–8. 8. Schubring C, Kiess W, Englaro P, Rascher W, Dotsch J, Hanitsch S, et al. Levels of leptin in maternal serum amniotic fluid, and arterial and venous cord blood: Relation to neonatal and placental weight. J Clin Endocrinol Metab 1997;82:1480–3. 9. Jaquet D, Leger J, Levy-Marchal C, Czernochow P. Ontogeny of leptin in human fetuses and newborns: Effect of intrauterine growth retardation on serum leptin concentrations. J Clin Endocrinol Metab 1998;83:1243–6. 10. Ong KK, Ahmed ML, Sheriff A, Woods KA, Watts A, Golding J, et al. Cord blood leptin is associated with size at birth and predicts infancy weight gain in humans. J Clin Endocrinol Metab 1999;84:1145–8. 11. Yang SW, Kim SJ. The relationship of the levels of leptin, insulin-like growth factor-I and insulin in cord blood with birth size, ponderal index, and gender difference. J Pediatr Endocrinol Metab 2000;13:289–96. 12. Todros T, Ferrazzi E, Giroli C, Nicolini U, Parodi L, Pavoni M, et al. Fitting growth curves to head and abdomen measurements of the fetus: A multicentric study. J Clin Ultrasound 1987;15:95–105. 13. Parazzini F, Cortinovis I, Bortolus R, Fedele L. Standard di peso alla nascita in Italia. [Birth weight standards in Italy] Ann Ost Gin Med Perinat 1991;CXII:203– 46. 14. Lepercq J, Challier JC, Guerre-Millo M, Cauzac M, Vidal H, Hauguel-de Mouzon S. Prenatal leptin production: Evidence that fetal adipose tissue produces leptin. J Clin Endocrinol Metab 2001;86:2409–13. 15. Lea RG, Howe D, Hannah LT, Bonneau O, Hunter L, Hoggard N. Placental leptin in normal, diabetic and fetal growth-retardated pregnancies. Mol Hum Reprod 2000;6: 763–9. 16. Cetin I, Morpurgo PS, Radaelli T, Taricco E, Cortellazzi D, Bellotti M, et al. Fetal plasma leptin concentrations: Relationship with different intrauterine growth patterns from 19 weeks to term. Pediatr Res 2000;48:646–51. 17. Widdowson EM, Southgate DAT, Hey EN. Nutrition and metabolism of the fetus and the newborn. In: Nutrition of the fetus and infant: Visser HKA, ed. The Hague, The Netherlands: Martinus-Nijhoff, 1979:169 –77.
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18. Varvarigou A, Mantzoros CS, Beratis NG. Cord blood leptin concentration in relation to intrauterine growth. Clin Endocrinol 1999;50:177–83. 19. Koistinen HA, Koivisto VA, Andersson S, Karonen S-L, Kontula K, Oksanen L, et al. Leptin concentration in cord blood correlates with intrauterine growth. J Clin Endocrinol Metab 1997;82:3328–30. 20. Shekawat PS, Garland JS, Shivpuri C, Mick GJ, Sasidharan P, Pelz CJ, et al. Neonatal cord blood leptin: Its relationship to birth weight, body mass index, maternal diabetes, and steroids. Pediatr Res 1998;43:338–43. 21. Masuzaki H, Ogawa Y, Sagawa N, Hosoda K, Matsumoto T, Mise H, et al. Nonadipose tissue production of leptin: Leptin as a novel placenta-derived hormone in humans. Nat Med 1997;3:1029–33. 22. Stephens TW, Basinski M, Bristow PK, Bue-Valleskey JM, Burgett SG, Craft L, et al. The role of neuropeptide Y in the antiobesity action of the obese gene product. Nature 1995;377:530–2. 23. Dotsch J, Nusken KD, Knerr I, Kirschbaum M, Repp R, Rascher W. Leptin and neuropeptide Y gene expression in
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human placenta: Ontogeny and evidence for similarities to hypothalamic regulation. J Clin Endocrinol Metab 1999; 84:2755–8. 24. Dotsch J, Adelmann M, Englaro P, Dotsch A, Hanze J, Blum WF. Relation of leptin and neuropeptide Y in human blood and cerebrospinal fluid. J Neurol Sci 1997;151: 185–8. 25. Mise H, Sagawa N, Matsumoto T, Yura S, Nanno H, Itoh H, et al. Augmentated placental production of leptin in preeclampsia: Possible involvement of placenta hypoxia. J Clin Endocrinol Metab 1998;83:3225–9. Address reprint requests to: Giovanni A. Tommaselli, MD, University of Naples “Federico II,” Department of Obstetrics and Gynecology, Via S. Pansini 5, 80131, Naples, Italy; E-mail: [email protected].
Received June 13, 2002. Received in revised form November 23, 2002. Accepted November 27, 2002.
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Leptin is a 167-amino-acid protein mainly produced by adipocytes. It is thought to act on the hypothalamic centers, regulating energy expenditure and signalling the amount of body fat mass.1 Indeed, its serum levels are positively correlated with fat mass, percentage body fat, and body mass index (BMI).2 Leptin also exerts a number of metabolic functions, including an action on the From the Departments of Obstetrics and Gynecology and Clinical Pathology, University of Naples “Federico II,” Naples, Italy.
reproductive axis: ob/ob mice, mutants that lack circulating leptin, are infertile, and the administration of recombinant human leptin to these animals restores fertility.3,4 During pregnancy, maternal serum leptin levels progressively increase, peaking during the second trimester and then plateauing to a level at term three to four times higher than in nonpregnant women.5 This increase is only in part due to the weight gain experienced by pregnant women, because leptin levels are no longer significantly related to BMI during gestation.6 Furthermore, it has been demonstrated that leptin is produced by the placenta7 and that placental weight is significantly related to leptin levels in the umbilical cord blood.8 Thus, placental leptin might contribute to the increase of maternal serum leptin concentration, even though it is not known to what extent. Leptin levels in umbilical cord blood (ie, in the fetus), significantly increase starting by 34 weeks of pregnancy.9 A significant correlation exists between umbilical cord blood leptin concentration and neonatal ponderal index.10,11 This suggests that leptin might have a role in the regulation of fetal growth. Indeed, several authors evaluated umbilical cord blood leptin levels in pregnancies complicated by fetal growth restriction and found contrasting results. The aim of this study was to evaluate serum and umbilical cord blood leptin concentrations in normal and growth restricted pregnancies and correlate them with maternal BMI and birth weight of the newborns. MATERIAL AND METHODS Fifty women with uncomplicated singleton pregnancies and 30 women with singleton pregnancies complicated by asymmetric fetal growth restriction, all with normal pregravid BMI (20 –27 kg/m2), were consecutively enrolled from the outpatient clinic of our department. No subject was affected by diabetes or hypertension, and no patient developed complications during pregnancy. The diagnosis of fetal growth restriction was made when the
VOL. 102, NO. 3, SEPTEMBER 2003 © 2003 by The American College of Obstetricians and Gynecologists. Published by Elsevier.
0029-7844/03/$30.00 doi:10.1016/S0029-7844(03)00668-9
535
abdominal circumference by ultrasound measurement was below the tenth percentile of reference values for fetuses of similar age in Italy.12 Fetal growth restriction diagnosis was confirmed at birth if birth weight was below the tenth percentile according to Italian standards for birth weight and gestational age.13 The study was approved by our institutional review board. All patients gave their informed and written consent for participation in the study. All ultrasonographic measurements were performed by the same operator (MP) with the use of a Toshiba Powervision 6000 with a 5-MHz transabdominal convex probe (Toshiba Medical Systems, Rome, Italy). A maximum of 7 days elapsed between the final ultrasonography and births. Maternal blood samples were drawn from the antecubital vein upon hospitalization in our obstetric ward for parturition and centrifuged. All women were fasting and not in labor. Arterial umbilical cord blood samples were collected at the time of parturition and centrifuged. All sera were stored at ⫺20C until determination of leptin levels. Serum leptin levels were determined in duplicate with a human leptin immunoradiometric assay (Diagnostic Systems Laboraties Inc., Webster, TX) with a sensitivity of 0.1 ng/mL, an intraassay coefficient of variation of 2.6 – 4.9%, and an interassay coefficient of variation of 3.7– 6.6%. Statistical analysis of the data was performed on an IBM-compatible personal computer with the Statistical Package for Social Science 8.0 (SPSS Inc., Chicago, IL). Data distribution was assessed with the Shapiro-Wilk test. Maternal serum and umbilical cord blood leptin levels were nonparametric variables, and a Mann-Whitney U test was used to assess differences between groups. The other variables displayed a normal distribution, and an analysis of variance followed by Scheffe` procedure for post hoc comparison of means was used to identify differences between groups and between different times within the same group. Correlations between maternal serum and umbilical cord blood leptin levels were regressed on maternal BMI and neonatal birth weight for both normal pregnancies and pregnancies complicated by fetal growth restriction and were evaluated with the Spearman R test. Significance was set at P ⬍ .05. RESULTS Seven women with uncomplicated pregnancies and three women with fetal growth restricted pregnancies did not deliver in our department and were considered drop-outs. Thus, the study refers to 43 women with normal pregnancy (group A) and 27 women with fetal growth restriction (group B).
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Figure 1. Maternal serum leptin concentrations in normal pregnancies (group A) and pregnancies complicated by fetal growth restriction (group B). Horizontal lines represent median. *P ⬍ .05. Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
No statistical difference was observed between group A and group B in age (25.3 ⫾ 4.5 years in group A versus 27.1 ⫾ 3.2 years in group B), parity (1.4 ⫾ 0.3 in group A versus 1.2 ⫾ 0.4 in group B), gestational age at parturition (38.6 ⫾ 0.6 weeks in group A versus 37.2 ⫾ 1.1 weeks in group B), and BMI (24.7 ⫾ 0.9 kg/m2 in group A versus 24.5 ⫾ 0.9 kg/m2 in group B). Mean birth weight of newborns from group B was significantly lower compared with newborns from group A (1384 ⫾ 360 g in group B versus 3300 ⫾ 417 g in group A; P ⬍ .001). There was a similar number of female and male infants in both groups (group A: 17 male [40%], 26 female [60%]; group B: 12 male [44%], 15 female [56%]). Mean placental weight was significantly higher in group A compared with group B (620 ⫾ 215 g versus 385 ⫾ 130 g; P ⬍ .01). Maternal serum leptin concentrations were significantly higher in group B compared with group A (45.0 ng/mL [range 34.2–54.9] versus 29.0 ng/mL [range 24.7– 33.3]; P ⬍ .01) (Figure 1). Umbilical cord blood leptin levels were significantly lower in group B compared with group A (8.4 ng/mL [range 3.6 –13.2] versus 13.1 ng/mL [range 9.7–16.5]; P ⬍ .01) (Figure 2). No significant difference was observed in umbilical cord blood leptin levels between male and female fetuses (data not shown). Umbilical cord blood leptin concentrations were significantly correlated with neonatal birth weight in both groups (group A: R ⫽ .448, P ⬍ .01; group B: R ⫽ .876, P ⬍ .01) (Figure 3) but not with maternal BMI (group A: R ⫽ ⫺.240, P ⫽ nonsignificant; group B: R ⫽ ⫺.350, P ⫽ nonsignificant) (Figure 4). Maternal serum leptin lev-
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Figure 2. Umbilical cord blood leptin concentrations in normal pregnancies (group A) and pregnancies complicated by fetal growth restriction (group B). Horizontal lines represent median. *P ⬍ .05. Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
els were not significantly correlated with maternal BMI (group A: R ⫽ .298; group B: R ⫽ .300; P ⫽ nonsignificant) or with neonatal birth weight (group A: R ⫽ .287; group B: R ⫽ .291; P ⫽ nonsignificant) (Figures 5, 6). All women diagnosed as having a fetal growth restricted pregnancy delivered a small for gestational age infant. DISCUSSION In this study, we demonstrated that leptin concentrations in the umbilical cord blood of growth restricted newborns are significantly lower compared with normal for age infants. This result is in agreement with previous studies.9,14 –16 Because fetuses affected by growth restriction and small for gestational age newborns have a significantly reduced fat tissue accumulation,17 it has been hypothesized that lower leptin levels in umbilical cord blood found in these fetuses were determined by the limited amount of fetal fat tissue. Indeed, large for gestational age newborns have higher umbilical cord blood leptin levels compared with appropriate for gestational age (AGA) and small for gestational age newborns.11,14,18 However, determination of leptin concentrations in umbilical cord blood from growth restricted fetuses has shown discrepancies, with studies reporting lower14,15,18,19 but also higher20 leptin levels compared with normal fetuses. Indeed, Jaquet et al9 found that, at term, serum leptin concentrations were significantly lower in newborns with fetal growth restriction than in those with normal growth, whereas no differences were found before 34 weeks’ gestation. Similarly, Cetin et al16 found that growth restricted fetuses show umbilical cord blood
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leptin levels significantly lower than do AGA fetuses after 34 weeks. These authors also found no significant differences between fetal growth restriction and AGA when adjusting for fetal weight (leptin per kg). Both these studies seem to indicate that, after 34 weeks’ gestation, leptin synthesis and secretion are correlated with the development and growth of adipose tissue. Conversely, some studies report higher umbilical cord blood leptin levels in growth restricted fetuses.20 This might be owing to the role played by fetal oxygenation and acid– base balance, because it has been observed that growth restricted fetuses with signs of severe fetal distress have significantly higher leptin concetrations per kilogram of fetal weight than do AGA fetuses.16 Our fetal growth restriction population showed no sign of fetal distress and lower leptin levels than did the AGA controls. It is therefore possible that umbilical cord blood leptin levels depend more on adipose tissue and on fetal oxygenation status than on other antenatal development factors. More studies are needed to evaluate the role of fetal well-being on leptin secretion and leptin secretion patterns in different types of fetal growth restriction. It has been demostrated that the placenta is a major source of leptin during pregnancy.21 Because maternal serum leptin levels progressively rise during the first two trimesters of pregnancy, it has been hypothesized that placental leptin might substantially contribute to the determination of these concentrations.21 It is not yet clear why during the third trimester, when estrogenic and progestinic stimuli on leptin secretion are at their peak and placental weight and size at their maximum, maternal serum leptin concentrations remain unmodified. The finding of higher maternal serum leptin levels in fetal growth restriction-complicated pregnancies is difficult to explain. Leptin inhibits neuropeptide Y secretion in the hypothalamus in mice22 and, although there is no evidence,23 it is hypothesized that the same relationship exists in humans. It has been demonstrated that hypoxia induces an increase of leptin secretion by choriocarcinoma cells. Dotsch et al24 demonstrated that, in preeclampsia, elevated placental leptin expression is accompanied by lower placental neuropeptide Y messenger ribonucleic acid levels. Because neuropeptide Y is a potent vasoconstrictor,24 it is tempting to hypothesize that the placental neuropeptide Y–leptin axis counteracts the vasoconstriction associated with preeclamptic changes in the placenta. It might be that in fetal growth restriction-complicated pregnancies the first sign of hypoxia is an increase of placental leptin production, mirrored by elevated maternal serum leptin levels.25 Further
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Figure 3. Relationship between umbilical cord blood leptin levels and neonatal birth weight in normal pregnancies (A) (Spearman R ⫽ .448; P ⬍ .001) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .876; P ⬍ .001). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 4. Relationship between umbilical cord blood leptin levels and maternal body mass index in normal pregnancies (A) (Spearman R ⫽ ⫺.240; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ ⫺.350; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 5. Relationship between maternal serum leptin levels and maternal body mass index in normal pregnancies (A) (Spearman R ⫽ .287; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .291; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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Figure 6. Relationship between maternal serum leptin levels and neonatal birth weight in normal pregnancies (A) (Spearman R ⫽ .298; P ⫽ nonsignificant) and pregnancies complicated by fetal growth restriction (B) (Spearman R ⫽ .3; P ⫽ nonsignificant). Pighetti. Leptin in Fetal Growth Restricted Pregnancies. Obstet Gynecol 2003.
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studies evaluating growth restricted fetuses with and without hypoxia evaluated by uterine and placental blood flow are needed, to assess whether maternal leptin serum levels can be used as a marker of fetal hypoxia. Furthermore, an inverse correlation between placental weight and maternal serum leptin concentrations has been demonstrated in normal pregnancies.8 Thus, in the presence of low-weight placentas, such as those found in cases of fetal growth restriction, leptin secretion, and therefore maternal serum concentrations, should be increased. Lea et al15 demonstrated that placentas from pregnancies complicated by fetal growth restriction express lower amounts of leptin compared with normal placentas. A possible explanation might be that the mean gestational age of the patients in Lea et al’s study was 33.2 ⫾ 1.88 weeks, whereas in our study the women with fetal growth restriction were almost all at term. This might have influenced the amount of placental leptin produced. Alternatively, the mechanisms involved in the compensation of growth restricted fetuses might not yet be established by 33 weeks’ gestation but only at term. In this study we confirmed that growth restricted fetuses at term show umbilical cord blood leptin concentrations significantly lower than those in normal fetuses, indicating that fetal adipose tissue is a major source of leptin. This conclusion is confirmed by the positive correlation between umbilical cord blood leptin concentrations and neonatal birth weight found in both groups. Furthermore, we also demonstrated that mothers with fetal growth restricted fetuses have higher serum leptin concentrations compared with mothers with uncomplicated pregnancies. This increase might be due to an intrinsic placental mechanism, by which small placentas produce more leptin as a compensatory mechanism, or to early hypoxia. Further research is needed to ascertain the role of hypoxia in growth restricted fetuses in determining placental leptin secretion and the intraplacental function of leptin in these situations.
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human placenta: Ontogeny and evidence for similarities to hypothalamic regulation. J Clin Endocrinol Metab 1999; 84:2755–8. 24. Dotsch J, Adelmann M, Englaro P, Dotsch A, Hanze J, Blum WF. Relation of leptin and neuropeptide Y in human blood and cerebrospinal fluid. J Neurol Sci 1997;151: 185–8. 25. Mise H, Sagawa N, Matsumoto T, Yura S, Nanno H, Itoh H, et al. Augmentated placental production of leptin in preeclampsia: Possible involvement of placenta hypoxia. J Clin Endocrinol Metab 1998;83:3225–9. Address reprint requests to: Giovanni A. Tommaselli, MD, University of Naples “Federico II,” Department of Obstetrics and Gynecology, Via S. Pansini 5, 80131, Naples, Italy; E-mail: [email protected].
Received June 13, 2002. Received in revised form November 23, 2002. Accepted November 27, 2002.
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