He, Huiling (1999) Expression of Vascular Endothelial Growth Factor-A and its Receptors in Uteroplacental Tissues of the Normotensive and Hypertensive Pregnant Rat. Research Master thesis, Victoria University.

Abstract

Hypertension, is a common clinical complication of pregnancy, often leading to preeclampsia and fetal intrauterine growth retardation. Many studies have shown that vascular endothelial growth factor A (VEGF-A) is a potent angiogenic factor, that is, it is responsible for the formation of new blood vessels from existing vessels. VEGF-A has been shown to be expressed in uteroplacental tissues, particularly during implantation. Levels of VEGF-A mRNA have also been shown to be significantly lower in placental tissue from pre-eclamptic women compared with control women (Cooper et al, 1996). The first aim of this project, was to establish the use of RT-PCR (reverse transcriptase-polymerase chain reaction), Northern blot analysis, immunohistochemistry and in situ hybridization, to measure the expression and localization of VEGF-A, vascular endothelial growth factor receptor 1 (VEGFR-1) and vascular endothelial growth factor receptor 2 (VEGFR-2) in normal rat uteroplacental tissues at early, mid and late gestation. The second aim of this project was to measure expression of VEGF-A, VEGFR-1 and VEGFR-2 in uteroplacental tissues of the normotensive (WKY) and the spontaneously hypertensive rat (SHR) at 7, 11 and 19 days of gestation. Spontaneously hypertensive rats have been previously established as an animal model for pre-eclampsia. The uterus containing the placenta and fetus was removed from normal Sprague Dawley, WKY and SHR rats over a range of gestational ages. After 11 days of gestation, the uterus and placenta were separated. Tissues were frozen immediately in liquid nitrogen and stored at -80oC, or fixed in 10% formaldehyde for immunohistochemistry. Total RNA was extracted using TRIZOL reagent and mRNA levels of VEGF-A and its receptors were examined using RT-PCR and Northern blot analysis. mRNA levels of VEGF-A and its receptors were compared with those of the housekeeping gene glyceraldehyde 3-phosphate dehydrogenase (G3PDH). We have successfully used RT-PCR to measure mRNA expression of VEGF-A, VEGFR-1 and VEGFR-2; Northern blot analysis to measure expression of VEGF-A, and immunohistochemistry to localize VEGF-A protein in rat uteroplacental tissues. We have shown that VEGF-A and its receptors were expressed in uteroplacental tissues of the normal rat. We found no major differences in the expression levels of VEGF-A and VEGFR-1 in uteroplacental tissues at early, mid and late gestation. We showed that VEGF-A164 was the predominant isoform found in uteroplacental tissues. Finally, we found no major differences in the expression levels of VEGF-A, VEGFR-1 and VEGFR-2 in uteroplacental tissues in normotensive rats (WKY) compared with hypertensive rats (SHR). We conclude that VEGF-A and its receptors, VEGFR-1 and VEGFR-2, were expressed in uteroplacental tissues of pregnant rats. Furthermore, mRNA levels of VEGF-A and its receptors were comparable in normotensive and hypertensive rats. These results suggest that VEGF-A and its receptors may have an important role in the development of the fetal and maternal portions of the placenta, but that they are not the primary factors involved in the aetiology of pre-eclampsia.

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