The expression of vascular endothelial growth factor isoforms in cultured human endometrial stromal cells and its regulation by 17beta- oestradiol

Angiogenesis or formation of new blood vessels is required for regeneration of the endometrium after its breakdown during each menstruation. Vascular endothelial growth factor (VEGF), a family of recently discovered angiogenic factors, may be involved in the repair and growth of the endometrium. In this study reverse transcription-polymerase chain reaction (PCR) was used to confirm the presence of VEGF mRNA and restriction enzyme digestion to confirm the identity of PCR products generated from different VEGF isoforms in cultured human endometrial stromal cells. The shortest isoform, VEGF 121, was the most abundant in quiescent stromal cells. It was about one-and-a-half times that of VEGF 165. The longest isoform, VEGF 206, was not detected; only a relatively weak signal for VEGF 189 was detectable. The mRNA for VEGF increased 2-fold after stimulation by 17beta-oestradiol (10 nM) for 30 min. A further increase to 3-fold above baseline occurred after 2 h incubation and remained steady at 6 h incubation, but decreased to 2-fold of baseline after 15 h. There was no differential stimulation of mRNA for VEGF isoforms: the ratio of VEGF 121 to 165 remained constant at 3:2 during the course of the incubation, with the exception at 15 h incubation when the ratio was 2:1. The VEGF protein, determined by specific enzyme immunoassay, increased from undetectable at baseline to 79.8 +/- 18.9 pg/ml (n = 4, mean +/- SD, 9.6 cm2/well/ml) after 2 h, with a further significant increase to 249.5 +/- 27.3 pg/ml after 15 h and 695.0 +/- 41.4 pg/well after 39 h. At 15 h incubation, the specific oestradiol antagonist ICI 182,780 (1 microM) significantly reduced VEGF secretion by 25% from 249.5 +/- 27.3 to 189.0 +/- 26.6 pg/ml. Thus, VEGF showed specific patterns of isoform expression in the human endometrial stromal cells; oestradiol (10 microM) stimulated, but not differentially, the mRNA for VEGF isoforms.