Estrogen inhibition of a key receptor that helps regulate blood pressure may be a new mechanism by which premenopausal women are protected from cardiovascular disease, researchers reported.
Findings from in vitro investigations published in the November issue of the journal Endocrinology suggest that estrogen signaling through the estrogen receptor protects the circulatory system by inhibiting the detrimental effects of the steroid hormone aldosterone signaling through the mineralocorticoid receptor.
Aldosterone activation of the mineralocorticoid receptor (MR) in the kidneys regulates sodium and fluid balance. MR also is expressed in the vasculature and in many other tissues. The steroid hormone has been shown to contribute to cardiovascular disease in both animal models and humans, researcher Iris Jaffe, MD, PhD, and colleagues from Tufts University, Boston, wrote.
It is well known that premenopausal women have increased protection against myocardial infarction and cardiovascular mortality compared to men of the same age, and endogenous estrogen is widely believed to mediate this protective effect. But the mechanisms for this have not been well understood, Jaffe explained in an interview.
“In clinical trials when we give estrogen back to postmenopausal women, we don’t always see the same benefits,” she told MedPage Today. “If we have a better understanding of how (endogenous) estrogen benefits women we might be able to mimic that.”
Mineralocorticoid Damages Blood Vessels
Jaffe has been studying the mineralocorticoid receptor in her lab at Tufts’ Sackler School of Graduate Biomedical Sciences for close to a decade.
“The mineralocorticoid receptor is a well known regulator of blood pressure, but in the last decade it has also become clear that mineralocorticoid receptors also damage blood vessels and promote heart attacks and strokes,” she said.
Study co-author Richard Karas, MD, PhD, studies the effects of estrogen on the cardiovascular system, and Jaffe said whenever mineralocorticoid receptor was found to have a detrimental effect on the vasculature of cells in her lab, Karas’ lab would find that estrogen prevented the same effect.
“We got to thinking that maybe these two receptors work together, and that is what led to this study,” Jaffe said.
The researchers hypothesized that estrogen (E2) signaling through the estrogen receptor (ER) may protect the vasculature by inhibiting the detrimental effects of aldosterone (aldo) signaling through the MR.
In a series of in vitro studies they demonstrated that E2-activated ER inhibited MR-mediated gene transcription from the mouse mammary tumor virus reporter in human embryonic kidney-293 cells. In contrast, aldo-activated MR did not affect ER-mediated gene transcription.
“The ER N terminus (amino acids 1–253) containing part of the DNA-binding domain is sufficient to inhibit MR genomic function, although point mutations reveal that DNA binding, ligand-independent activation, and rapid nongenomic ER signaling are not required for this effect,” the researchers wrote. “Furthermore, ER and MR are part of a complex in cell lysates, with amino acids 1–233 of the ERα N terminus being sufficient to complex with the MR.”
The ability of ER to inhibit MR-mediated gene transcription was also found to correlate with the ability of ER segments to both localize to the nucleus and complex with the MR. In cultured vascular endothelial cells expressing ER, E2 inhibited aldosterone induction of the vascular MR target gene intercellular adhesion molecule-1 (ICAM-1).
ICAM-1 induction by endothelial MR is known to promote vascular inflammation that could contribute to the mechanism of aldo-induced atherosclerosis. E2 also inhibits aldo induction of ICAM-1 protein and prevents aldo-enhanced leukocyte adhesion to endothelial cells.
‘Studies Support New Model for Cardio Protection’
The in vitro studies support a new model in which E2-activated ER in endothelial cells forms a complex with mineralocorticoid receptor in the nucleus to modulate mineralocorticoid regulation of the proinflammatory gene ICAM-1, the researchers noted.
“Although further studies are needed to clarify the detailed molecular mechanism(s) by which ER abrogates MR-mediated gene regulation, our findings are consistent with several possible mechanisms,” they wrote. “First, the data support that the mechanism requires ERα to form a complex with MR in the nucleus, either through direct binding or indirectly via other proteins, and that this interaction blocks MR function as a transcriptional factor.”
However, even though the ER segment 1–253 was identified as the smallest portion able to inhibit MR transcriptional activity, the smaller segment 1–233 was able to complex with MR.
“The explanation for this discrepancy may come from our finding that nuclear localization of 1–233 was much weaker than 1–253, suggesting nuclear localization of ER may also be critical for its interference with MR-mediated transcriptional activity,” the researchers wrote. They added that “this mechanism appears to be distinct from the well-characterized dimerization interaction of the retinoid-X-receptor with non-steroid nuclear receptors including the thyroid, retinoid and vitamin D receptors because this does not require DNA binding by the ER.”
Jaffe and colleagues noted that once the ER forms a complex with MR in the nucleus, “multiple potential mechanisms are consistent with the data.”
“ER may prevent MR from interacting with a co-factor that is required for aldo-induced transcriptional activation but is not needed for ER transcriptional activation,” they wrote. “This is consistent with the finding that MR does not mutually inhibit transcriptional activation of ER. Recently posttranslational modification of the MR by phosphorylation has been shown to modulate MR transcriptional function in specific renal cell types. Thus, another possibility could be that complexing with ER in the nucleus facilitates MR modifications that alter its transcriptional function.”
The researchers cited several potentially important study limitations including the use of a transformed human embryonic kidney cell line (HEK293) with overexpressed ER and MR proteins and a transfected supercoiled plasmid containing the MMTV, MR-responsive element as a transcriptional reporter for their initial studies exploring interactions between ER and MR transcriptional function.
“This system, although highly artificial, was chosen because these HEK293 cells lack endogenous ER, MR and GR, and hence, the expression of receptors and ER mutants could be carefully controlled to specifically and independently examine transcriptional function and the ER and the MR,” they wrote.
The lack of known vascular MR target gene DNA binding sites and an adequate MR antibody for chromatin immunoprecipitation prevented the use of more modern methods to confirm endogenous MR binding.
In Vivo Studies Needed to Confirm Findings
The researchers also noted that since ER expression is rapidly down-regulated in cultured cells, the ER in the studies was exogenously expressed.
“Future studies in whole vessels or in vivo will be needed to interrogate this novel mechanism in the setting of physiological levels or MR and ER,” they wrote.
Jaffe said she hopes next to study the estrogen receptor-mineralocorticoid receptor connection in a mouse model.
“If one of the benefits of estrogen is that it is preventing the detrimental effects of mineralocorticoid, we might be able to reproduce that effect in men and postmenopausal women by giving them mineralocorticoid antagonists,” she said.
Several anti-mineralocorticoid diuretics have been approved and are used to treat chronic heart failure, hypertension and a condition known as hyperaldosteronism.
“These drugs act on the mineralocorticoid receptor in the kidneys to lower blood pressure and decrease mortality in patients with heart failure, but I don’t think anyone has really thought about their blood vessel protective effect,” Jaffe said.
- Estrogen inhibition of a key receptor that helps regulate blood pressure may be a new mechanism by which premenopausal women are protected from cardiovascular disease.
- Note that the in vitro studies support a new model in which estrogen-activated estrogen receptor in endothelial cells forms a complex with mineralocorticoid receptor in the nucleus to modulate mineralocorticoid regulation of the proinflammatory gene ICAM-1.