Estrogen regulates vascular endothelial growth/permeability factor expression in 7,12-dimethylbenz(a)anthracene-induced rat mammary tumors.
Vascular endothelial growth/permeability factor (VEG/PF) is expressed in some normal tissues and at high levels in a wide range of tumors. This growth factor is believed to be a key mediator of angiogenesis. Recent reports have shown that VEG/PF mRNA in the normal rat uterus is stimulated by estradiol (E2). In this study, we investigated the expression of VEG/PF in the mammary gland and 7,12-dimethylbenz(a)anthracene (DMBA)-induced, hormone-dependent mammary tumor of the rat model, and also whether VEG/PF is regulated by E2. VEG/PF mRNA from tumor extracts was amplified by RT-PCR with VEG/PF primers and generated two main products which corresponded in size to those expected for VEG/PF 164 and 120. In some cases, a third product corresponding in size to that expected for VEG/PF 188 was also generated. No such PCR products were generated from equal amount of RNA from normal mammary tissue, rat brain, or liver. Using immunocytochemistry, VEG/PF expression was detected in the epithelial cells of the tumors. We developed an ELISA assay to measure VEG/PF protein concentrations and found a 4-fold difference between normal mammary glands (1.3 +/- 0.11 ng/mg protein) and tumors (4.44 +/- 0.66) (P < 0.01). E2 treatment (5 microg/rat, s.c.) of rats 24 h after ovariectomy, greatly enhanced the expression of RT-PCR products in tumors within 2 h, which reached a maximum at 6-8 h but declined by 48 h. VEG/PF concentrations were also increased 8-12 h after E2 injection. When rats were given two injections of aromatase inhibitor 4-hydroxyandrostenedione (4-OHA 10 mg/rat s.c.) 24 h apart, to reduce estrogen concentrations, a low level of RT-PCR products was maintained for at least 96 h. After a single injection of 4-OHA, RT-PCR products remained low until 36 h when an increase occurred corresponding with a rise in plasma E2 levels. Injection of E2 2 h after 4-OHA treatment, caused a rise in RT-PCR products in 6-8 h. However, there was no significant change in VEG/PF concentrations. An increase in VEG/PF protein concentrations followed the increase in mRNA levels by 4-6 h. Thus, it appears that E2 causes a rapid induction of VEG/PF expression in mammary tumors that is similar to that observed in the normal uterus. These findings suggest that one mechanism by which estrogen acts as a mammary tumor promotor is by stimulating VEG/PF, leading to increased tumor angiogenesis and/or permeability of the microvessels to allow tumor cell migration.
Nakamura
J
J
Department of Pharmacology and Experimental Therapeutics, School of Medicine, University of Maryland, Baltimore 21201, USA.
Savinov
A
A
Lu
Q
Q
Brodie
A
A
D015127:9,10-Dimethyl-1,2-benzanthracene; D000735:Androstenedione / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D016228:Endothelial Growth Factors / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D007150:Immunohistochemistry; D008222:Lymphokines / Q000235:genetics / Q000378:metabolism*; D008321:Mammary Glands, Animal / Q000378:metabolism; D015674:Mammary Neoplasms, Animal / Q000139:chemically induced / Q000378:metabolism*; D010052:Ovariectomy; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D013997:Time Factors; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D004791:Enzyme Inhibitors; D008222:Lymphokines; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D000735:Androstenedione; D004958:Estradiol; D015127:9,10-Dimethyl-1,2-benzanthracene; C014594:formestane