Comments for:
Abstract
Estrogen is an important vasoprotective molecule that causes the rapid dilation of blood vessels by activating endothelial nitric oxide synthase (eNOS) through an unknown mechanism. In studies of intact ovine endothelial cells, 17β-estradiol (E2) caused acute (five-minute) activation of eNOS that was unaffected by actinomycin D but was fully inhibited by concomitant acute treatment with specific estrogen receptor (ER) antagonists. Overexpression of the known transcription factor ERα led to marked enhancement of the acute response to E2, and this was blocked by ER antagonists, was specific to E2, and required the ERα hormone-binding domain. In addition, the acute response of eNOS to E2 was reconstituted in COS-7 cells cotransfected with wild-type ERα and eNOS, but not by transfection with eNOS alone. Furthermore, the inhibition of tyrosine kinases or mitogen-activated protein (MAP) kinase kinase prevented the activation of eNOS by E2, and E2 caused rapid ER-dependent activation of MAP kinase. These findings demonstrate that the short-term effects of estrogen central to cardiovascular physiology are mediated by ERα functioning in a novel, nongenomic manner to activate eNOS via MAP kinase–dependent mechanisms.
Authors
Zhong Chen, Ivan S. Yuhanna, Zoya Galcheva-Gargova, Richard H. Karas, Michael E. Mendelsohn, Philip W. Shaul
Re: Rapid nongenomic actions of estrogen
Submitter: Philip W. Shaul | Philip.Shaul@email.swmed.edu
University of Texas Southwestern Medical Center
Published July 12, 1999
Drs. Sudhir and Komesaroff raise a variety of interesting points regarding our recent report demonstrating a role for estrogen receptor-alpha (ERalpha) in the rapid activation of endothelial nitric oxide synthase (eNOS) in endothelial cells (1). First, they suggest that caution be taken in the extrapolation of our findings to human physiology because of the concentration of 17beta-estradiol that was tested (10–8M). We respectfully reiterate that we have previously demonstrated that eNOS activation readily occurs in this model system at concentrations of 10–10 M estradiol and above (2). Circulating estradiol levels in women range from about 3 x 10–10 to 4 x 10–10 M in the follicular phase to about 2 x 10–9 M at the time of ovulation, and levels to nearly 7 x 10–8 M may be achieved during pregnancy. Importantly, serum estradiol levels fall after menopause to values of approximately 2 x 10–11 to 7 x 10–11 M, which is below the threshold concentration observed in our model (2–4). As such, we would suggest that the dose-response observed in our studies is in close correlation with relevant estradiol levels in women.
Second, the comment is made that there are also endothelium-independent mechanisms mediating rapid, estrogen-induced vasodilation. We agree, and such mechanisms would include the opening of calcium-activated potassium channels in vascular smooth muscle through a nitric oxide– and cyclic guanosine monophosphate–dependent process, thus relaxing the smooth muscle directly (4–6). In addition, we agree that prostaglandin-mediated mechanisms may play a role. We have recently observed that estradiol causes rapid, receptor-mediated increases in prostacyclin production in endothelial cells (7). Multiple mechanisms are most likely involved in the acute effects of estrogen on vasomotor tone, and the relevance of different mechanisms probably varies between specific vascular beds.
Finally, Drs. Sudhir and Komesaroff note that membrane-associated estrogen receptors may play a role in rapid responses to estrogen in a variety of cell types. In support of this concept, we have recently reported that ERalpha protein is detectable in purified plasma membranes from endothelial cells, and that estradiol activates eNOS in isolated plasma membranes in a receptor-dependent manner (8). When all of these issues are considered, it is apparent that further studies are warranted in our model to determine the basis for nongenomic responses mediated by ERalpha in endothelial cells. However, we fully agree that additional investigation is also needed to delineate the mechanisms underlying the other rapid effects of estrogens, as well as androgens, on the vascular wall.
Philip W. Shaul, MD
Department of Pediatrics, University of Texas–Southwestern Medical Center
1. Chen, Z., et al. 1999. Estrogen receptor alpha mediates nongenomic activation of eNOS by estrogen. J. Clin. Invest. 103:401–406.
2. Lantin-Hermoso, R.L., et al. 1997. Estrogen acutely stimulates nitric oxide synthase activity in fetal pulmonary artery endothelium. Am. J. Physiol.273:L119–L126.
3. Yen, S.S.C., and Jaffe, R.B. 1991. Reproductive endocrinology: physiology, pathophysiology and clinical management. 3rd edition. W.B. Saunders. Philadelphia, PA.
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Rapid nongenomic actions of estrogen
Submitter: Krishnankutty Sudhir | k.sudhir@alfred.org.au
Baker Medical Research Institute and Alfred Hospital
Published July 12, 1999
Chen et al. have recently presented evidence that the rapid, nongenomic activation of endothelial nitric oxide synthase (eNOS) by estrogen is mediated by estrogen receptor-alpha, probably via tyrosine kinases or MAP kinase (1). They show that estradiol (10–8 M) induces a rapid increase in eNOS activity in endothelial cells, and that this increase is fully inhibited by tamoxifen and the nonselective estrogen receptor antagonist ICI 182,780. They also suggest that "the specific level of E2 that was studied is readily achievable during pregnancy." It is in fact unusual for such values to be attained; furthermore, estradiol values in pregnancy are about 100-fold greater than those in nonpregnant cycling women (2). Accordingly, their findings must be interpreted with caution if extrapolated to human physiology.
Nonetheless, this is an elegant study that provides a possible mechanism for the rapid, endothelium-dependent vasorelaxation in response to estradiol that has been observed in both women (3, 4) and men (5); it may also explain rapid attenuation of endothelin-1–induced coronary vasoconstriction in pigs (6). However, this may be just one, albeit important, part of the story. Additional evidence suggests that other mechanisms may also play a role in the rapid, nongenomic effects of estradiol in blood vessels. For example, estradiol induces rapid, endothelium-independent vasodilation in dog coronary arteries in vivo, an effect that is not blocked by the nonselective estrogen receptor antagonist ICI 182,780 and is undiminished by balloon denudation of the coronary artery (7). Furthermore, it appears likely that rapid estrogen actions are mediated, at least in part, by events triggered from the outer cell surfaces (8). Characterization of estrogen-binding membrane proteins shows that, at least in rabbit uterus, there are differences from classical cytosolic estrogen receptors, as shown by differences in inhibition by ICI 182,780, tamoxifen, and antibody binding (9), suggesting that the membrane binding site is structurally related to, but distinct from, intracellular estrogen receptors. Finally, recent evidence suggests that the rapid effects of estrogen on acetylcholine-induced vasodilatation in the cutaneous microvasculature (5) may occur via mechanisms independent of nitric oxide release, and may involve prostanoid pathways instead (10, 11), suggesting yet another possible mechanism for nongenomic actions of estrogen. Of interest, testosterone also induces rapid coronary vasodilation, an effect that is not inhibited in vitro by an androgen receptor antagonist (12) or blocked in vivo by ICI 182,780 (13), suggesting that it is not mediated via aromatization to estrogens.
In conclusion, we believe that the findings of Chen et al., important as they are, should be seen in the context of these other data. The nongenomic actions of estrogen are mediated by several distinct, possibly tissue-specific, pathways that, in some cases at least, may be common to both estrogens and androgens and independent of classical receptors.
Krishnankutty Sudhir, MD, PhD, FRACP, FACC, Associate Professor of Medicine
Paul A. Komesaroff, MD, PhD, FRACP, Associate Professor of Medicine
Hormones and the Vasculature Laboratory, Baker Medical Research Institute and Alfred Hospital, PO Box 6492, Melbourne 8008, Victoria, Australia. Phone: 011-613-9276-3263; Fax: 011-613-9276-2461.
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