Vascular Endothelial Growth Factor (VEGF) is a potent regulator of placental vascular function. one major band was observed at 180, 235, 130 and 130 kD, respectively. All of these bands were corresponding to their positive controls. Of these five proteins studied, only VEGFR-1 levels were increased (< 0.05; 1.7 fold) in PE placentas. The expression of VEGF and the four VEGF receptors was confirmed using immunohistochemistry. They were primarily present in syncytiotrophoblasts and endothelial cells of villous capillaries and large vessels. Thus, together with the previous reports that VEGFR-1 mediates trophoblast function and inhibits VEGF-induced angiogenesis and endothelium-dependent vasodilation, these data suggest that the increased VEGFR-1 expression may alter VEGF-mediated function on trophoblast and endothelial cells in PE placentas. < 0.05. Results The mRNA expression of total VEGF, EG-VEGF and the four VEGF receptors (VEGFR-1, VEGFR-2, NP-1, and NP-2) in human placentas was first confirmed using the RT-PCR analysis (Fig. 1). One band for each mRNA studied was observed at the estimated size as shown in Table 1 and these PCR products were confirmed by sequencing, indicating the specificity of each primer set. The mRNA levels of total VEGF, EG-VEGF and the four VEGF receptors quantified using real-time PCR are shown in Determine 2. The mRNA levels of total VEGF and VEGFR-1 were increased 2.8 and 2.7 fold (< 0.05) respectively in PE vs normal placentas. 39432-56-9 supplier No significant difference in mRNA levels of EG-VEGF and the other three VEGF receptors was observed between PE and normal placentas. Among these genes studied, VEGFR-1 was the most abundant (1/8 of -actin), followed by VEGFR-2 (1/19), NP1 (1/66), VEGF (1/186), EG-VEGF (1/473), and NP-2 (1/631) in normal placentas. Fig. 1 RT-PCR analysis for VEGF, EG-VEGF, VEGFR-1, VEGFR-2, NP-1, and NP-2 in human placentas. The total RNA samples (0.5 g/gene) from one normal placenta were used for PCR amplification. The PCR products were confirmed by sequencing and used as standards ... Fig. 2 Real-time PCR analysis of the mRNA levels of VEGF, EG-VEGF, VEGFR-1, VEGFR-2, NP-1, and NP-2 in human placentas from normal and PE pregnancies. For each gene, cDNA was amplified from total RNA (2 g/sample) of normal or PE placentas and 4 l ... To 39432-56-9 supplier determine whether expression of individual VEGF isoforms differed in PE vs normal placentas, mRNA levels of three major VEGF isoforms (VEGF121, 165, and 189) were quantified using semi-quantitative RT-PCR (Fig 3). The Spp1 overall mRNA levels of these three VEGF isoforms were increased (< 0.05) 1.8 fold in PE vs normal placentas. Compared with normal pregnancy, the placental mRNA levels of three VEGF isoforms in PE were elevated (< 0.05) 1.8, 1.9, and 1.7 fold, respectively for VEGF189, 165, and 121, as compared with normal placentas. Fig. 3 Semi-quantitative RT-PCR analysis for VEGF isoforms in human placentas from normal and PE pregnancies. The total RNA (2 g/sample) was used for generating cDNA and PCR products were run on 4% agarose gels. (A) A representative agarose gel. The ... Protein expression of VEGF and its four receptors in normal and PE placental tissues was determined by Western blot analysis (Fig. 4). The VEGF antibody detected two major bands approximately at 20 and 25 kD (Fig. 4A). The former was corresponding to the molecular mass of recombinant human VEGF165, while the latter is similar to the reported molecular mass of VEGF 189 (32). There was no significant difference in protein expression of both VEGF isoforms between 39432-56-9 supplier normal and PE placentas (Fig. 4B). For VEGFR-1, VEGFR-2, NP-1 and NP-2, one major band, corresponding to its respective control, was also identified at approximately 180, 235, 130, and 130 kD, respectively (Fig. 4A). VEGFR-1, but not the other three receptors (VEGFR-2, NP-1 and NP-2), protein.