Intrauterine growth restriction (IUGR) is a complicated placental vascular disease resulting in low birth weight, preterm delivery, and increased perinatal morbidity and mortality 1234. IUGR may be caused by various fetal, maternal, and placental factors 1235. Angiogenesis, defined as the development of new vascular structures, is a placental factor playing an important role in the development of IUGR 23467.

Angiogenesis involves the branching of new microvessels from pre-existing larger blood vessels. It is an important factor in normal embryogenesis and in physiological processes such as ovulation and the menstrual cycle 8910. Angiogenesis plays a role in the development of the villous vasculature and the formation of terminal villi in the human placenta. Placental vascular growth begins early in pregnancy and continues throughout gestation 911. The villous vasculature increases in number rather than vessel type from the 21st day of development until the end of the first trimester. From the 26th week of gestation until term, villous vascular growth changes from branching to non-branching angiogenesis due to the formation of mature intermediate villi that specialize in gas exchange. Specific angiogenic and inhibitory factors regulate these processes 489. IUGR occurs as a result of the failure of elongation, branching, and dilation of the capillary loops and of terminal villous formation 4.

Vascular endothelial growth factor (VEGF), basic-fibroblast growth factor (b-FGF), and endothelial nitric oxide synthase (eNOS; type III nitric oxide enzyme) have been identified as positive regulators of angiogenesis 8. They are strongly expressed during embryonic and fetal development, especially in the first trimester 1610121314.

VEGF, one of the first angiogenic factors identified, is widely believed to be the most important regulator of both normal and pathological angiogenesis 15. It plays an essential role in the formation of new blood vessels 812. In pregnancy, VEGF participates in the proliferation, migration, and metabolic activity of trophoblasts 34612. It is expressed by human villous and extravillous trophoblasts, and conclusive evidence indicates that it regulates trophoblast function by stimulating release of nitric oxide 4. b-FGF acts as a modulator of tissue differentiation and placental angiogenesis 361013, and eNOS has an important role in the regulation of placental blood flow 14.

In this study, we investigated the relationship between placental angiogenesis and the expression of VEGF-A, b-FGF, and eNOS in normal-term pregnancy and IUGR placentas.

Clinical and pathological data, including maternal, neonatal, and placental parameters are shown in Table 1. The mean gestational age of the normal pregnancies and those complicated by IUGR was 38.9 ± 5.9 weeks and 36.3 ± 22.9 weeks, respectively. Birth weight, placental weight, placental diameter, umbilical cord length, and APGAR scores were demonstrated to be significantly different between the two groups.

The clinical and morphological features commonly associated with IUGR observed in 55 placentas are recorded in Table 2. A histological comparison of normal term pregnancy and IUGR placental tissues showed widespread infarct areas and increased syncytial knots or Tenney-Parker changes in the IUGR placentas (Fig. 1A, B).

IUGR is an important cause of perinatal morbidity and mortality. In developed countries, the incidence of IUGR is 3%, whereas in developing countries, it reaches 15-20%. It is one of the most commonly recognized abnormalities of the fetal condition and is a compounding factor in 26% or more of stillbirths 145. It may also have long-term health implications for adults 17. For these reasons, the accurate prediction, diagnosis, and appropriate management of pregnancies complicated with IUGR are important. In accordance with the clinical definition of IUGR, we observed significantly reduced fetal birth weight, amniotic fluid, and placental size and weight in our IUGR group.

The placenta, which normally has a rich vasculature, plays an important role in the development of IUGR. The most common cause of IUGR is placenta ischemia in which insufficient placental function results from deteriorated uteroplacental perfusion 35718. The clinical features of ischemic placental disease are revealed throughout the second half of pregnancy, but the pathophysiological processes initiating the disease originate in the first half 17. The process of promoting the development of neovessels may be activated by chemo-cytokines in some pathological situations such as ischemia 12. Structural evidence suggests that placental oxygenation is important in controlling fetoplacental angiogenesis and hence, villous differentiation 9. Insufficient uteroplacental perfusion leading to abnormal angiogenesis may result in the pathophysiology of IUGR 34.

Changes appearing during placental development can be used as early markers of pathologies that may occur later in pregnancy. Placental ischemia is one such pathology, and it can be investigated in relation to various angiogenic mediators 17. A hypoxic stimulus may lead to an excessive proliferation of villous capillaries and connective tissue via growth factors such as VEGF and FGF 19. Abnormal vasculogenesis, angiogenesis, and pseudovasculogenesis is correlated with the impaired placental and fetal development seen in complicated pregnancies such as IUGR 3 Angiogenesis may be regulated by oxygen status and by the production of angiogenic growth factors and their natural receptors and antagonists by vascular endothelial cells, pericytes, and trophoblasts. The resulting changes in fetal vasculature are associated with altered patterns of villous growth 18. The results from our study support the literature reporting that an alteration in placental development accompanying deteriorated angiogenesis occurs in IUGR 4111718. Contrary to our findings, Lyall et al. 20 demonstrated a reduction in villous placenta VEGF expression in placental villous tissue from pregnancies complicated by IUGR and preeclampsia.

It is thought that the angiogenic factors VEGF, b-FGF, and eNOS have important roles in villous proliferation, trophoblast function, and angiogenesis as characterized by the formation of new vascular structures in the villous stroma 14. VEGF is thought to exert a dual role in the placenta, acting on both angiogenesis and trophoblast function during placental development. The roles of VEGF, b-FGF and eNOS in placental angiogenesis may be altered in conditions such as IUGR 414. The increased expression of VEGF-A, b-FGF, and eNOS that we have found in IUGR placentas may promote increased endothelial cell proliferation and migration and pathological angiogenesis 17. It is likely that normal placental angiogenesis depends on the regulation of vascular development by a complex relationships among these factors and that they play an important role in the development of IUGR.

It is necessary to explain the regulatory mechanism of placental vascular development in order to elucidate the pathogenesis of IUGR and the associated placental vascular insufficiency. The observed, increased expression of VEGF-A, b-FGF, and eNOS suggests that abnormal angiogenic activity, caused by insufficient uteroplacental perfusion, results in the pathophysiology of IUGR.

IUGR: Intrauterine growth restriction; VEGF: Vascular endothelial growth factor; VEGF-A: Vascular endothelial growth factor-A; b-FGF: Basic-fibroblast growth factor; eNOS: Endothelial nitric oxide synthase; H&E: Hematoxylin and eosin; B-SA: Biotin-streptavidin; DAB: 3,3-diaminobenzidine tetrahydrochloride; CVSC: Chorionic villous stromal cells; VSMC: Vascular smooth muscle cells; VVEC: Villous vascular endothelial cells; CT: Cytotrophoblasts; ST: Syncytiotrophoblasts; EVT: Extra villous trophoblasts.

The authors declare that they have no competing interests.

FB conducted the design of the study, performed microscopic evaluation, and drafted the manuscript. AB participated in the design of the study and performed the selection of appropriate cases and data collection and helped to draft the manuscript. BDG and NOK participated in the design of the study and immunohistochemical evaluation. MIH and MH and SOO conceived of the study, and participated in its design and coordination and helped to draft the manuscript. EA participated in the design of the study and performed statistical analysis. All authors read and approved the final manuscript.