Clearing Estrogen's Bad Name

The summer of 2002 was a tense time for us. One announcement and all of our knowledge and experience as basic researchers was put to question.

On July 10th, Jacques Rossouw, the director of the Women's Health Initiative (WHI) clinical trial, which tracked the long term effects of hormone therapy (HT) on women, announced the trial was being halted three years ahead of schedule. The results showed an unacceptable proportion of women were harmed by the therapy. The data implied that HT did not protect women against heart disease or protect against memory loss and other neurodegenerative conditions such as dementia, and in fact increased their risk of stroke and breast cancer. At the time, some 14 million US women were taking HT to relieve postmenopausal symptoms or to lower their risk for osteoporosis. In the days and weeks that followed, women around the world would call their doctors in panic, and many would stop taking the medication altogether. But the news went against much of what we had been learning about estrogens in the laboratory.

Numerous studies had suggested that hormone treatment, using synthetic estrogens or a combination of synthetic estrogens and progestins, protected women from many diseases, whereas the absence of estrogen made women more vulnerable. Some of these studies were observational, retrospective, and meta-analytical studies. But a few were clinical trials where hormones or placebos were administered to groups of women. The WHI, however, with 20,000 participants, was the largest clinical trial to date. With numbers like that it was very difficult to argue that estrogen wasn't harmful.

But there was cause to argue. Only a few years earlier, my lab had used animal models to show that estradiol, the major and most potent estrogen secreted by the human ovary, was protective against damage caused by stroke. A graduate student of mine, Dena Dubal, had asked a deceptively simple question: Would estradiol influence the extent of injury in rats after we experimentally induced cerebrovascular stroke by blocking blood flow to a major vessel in the brain?

We were pretty astounded to find that the animals with very low levels of estradiol experienced approximately half the amount of injury compared to animals without estrogens. What's more, estradiol had to be present before we performed the stroke injury; otherwise treatment was totally ineffective in protecting the brain. We tried different doses of estradiol and were amazed to find that very low doses were as effective as higher doses. Although the experiment was performed in mice, it seemed to be in direct opposition to the WHI findings.

We weren't the only researchers perplexed, and a bit dismayed by the WHI results. Estrogens are primarily thought of as reproductive hormones, produced in abundance by the ovaries from the time a woman goes through puberty until she goes through the menopause. But in the past 10 years many investigators, including ourselves, performed studies that led to an understanding of estrogens as fascinating pleiotropic hormones which act on a plethora of physiological functions not directly related to reproduction, and therefore also important in male physiology. The field had come to appreciate that estrogens were important players in the immune system, in cardiovascular function, in bone formation and breakdown, in fat distribution and metabolism, and in learning and memory.

As basic researchers, we know that our models are imperfect. But our goals are to perform studies that cannot be performed in humans because they are too expensive, too complex, or because ethical reasons prevent them. We know that a gap exists between what happens in the mouse and what happens in the human. Just the same, I was not ready to chalk up the last 10 years of my life in estrogen research to a bad mouse model; at least not without a second look at the results.

My first reactions were ones I imagine many researchers would experience: Were our findings wrong? Had our work been discredited? Would I have more difficulty getting our work funded? As my lab started to tease apart the results of the trial in our weekly lab meetings, we began to realize that there were many questions the trial had not answered.

The community of estrogen researchers rallied together to probe more deeply. Why the discrepancies between this major clinical trial and so many of the previous studies in humans or animal models? Meetings and symposia were organized to brainstorm what these results meant for all of us and how this research fit into the larger story of estrogen function. My lab began to reanalyze and reassess our studies and their design to try to understand why our results were so at odds with the WHI.

On the molecular level, we could see how even low levels of estrogens influenced many genes that regulate cell survival and cell death. In particular, we found that low concentrations of estradiol protected the brain by suppressing apoptosis. Dena Dubal, along with another graduate student, Shane Rau, found that estradiol could change the expression of multiple genes and proteins, sometimes very subtly and in anatomically specific places. They found that estradiol helps maintain levels of bcl-2, a protein which reduces apoptosis after stroke. They also found that estradiol inhibits caspases, proteins that mediate cell death. Most importantly, stroke increased the expression of estrogen receptors to allow estrogen to protect the brain and enhance its repair. All of these changes tip the balance toward cell survival and against cell death.¹ It was clear that one of the reasons our results seemed to differ was because the WHI study had tracked women before stroke had occurred to capture their risk, but didn't follow women after their stroke to see if women taking estrogen had improved recoveries, as our models had suggested.

It is important to point out that men also produce estrogen, and are also protected after stroke when estrogens are present. However, it is not usually possible to treat men with estrogens because the hormone has feminizing effects that are not acceptable. This is why it is so important for us to investigate the mechanisms by which estrogens act. Our hope is that this understanding would lead toward the development of compounds that use the same protective mechanisms, but do not have the other effects of estrogens.

The mechanisms by which estrogens act is an area of incredibly intense study. A cytosolic estrogen receptor discovered in the 1960's was thought to act by transporting estrogens attached to the receptor to the nucleus where the hormone-receptor complex bound to DNA and acted as a transcription factor, altering gene expression (see graphic on below). For more than 30 years, this was the exclusive way that estrogens were thought to act: via a single receptor that acted by binding to the promoter region of DNA to increase or decrease transcription of estrogen responsive genes. In 1996 Jan Ake Gustafsson and Ken Korach discovered that there are actually two different estrogen receptor subtypes: one called estrogen receptor alpha (ERα) and a new form, ERβ.² More recently, my lab realized that estrogens may act without a receptor or by binding to a receptor which can then activate second messenger systems without ever traveling to the nucleus and binding to DNA.

Our laboratory has followed these studies with great interest because we wondered whether the protective actions of estrogens in the brain depend upon the classical pathway or the more novel mechanisms of actions. We designed an experiment to use either ERα- or ERβ-knock out mice and found that ERα plays a pivotal role in protecting the brain against cell death. When this receptor is knocked out, estradiol does not protect the brain. ERβ proved not to play a role in protection; when we knocked out ERβ, mice continued to be protected just like wild-type mice.

It appears that low physiological levels of estradiol therapy act predominantly through the classical estrogen mechanisms of action. It is interesting to contrast this with findings from James Simpkins' lab that much higher concentrations of some estrogen analogs act through the more recently discovered mechanisms which bypass direct estrogen receptor, ERα or ERβ activation. This is important because this information may allow us to develop drugs that target a specific receptor to allow better outcomes from stroke.

What did these studies mean in terms of the results of the WHI? Two aspects of the clinical study design are worth considering in this context. First, only synthetic hormones were used. Premarin and Prempro are composed of substances that attempt to mimic the endogenous hormones synthesized by the ovary, but they are not the same structures. This might account for differential actions of these substances compared to endogenous hormones. Second, the average age of women when they started in the WHI trial was 63 years old and most of them had not had hormone therapy previous to the trial. Since women typically undergo the transition to the postmenopausal state around the age of 51, it means that most of them had been estrogen deficient for about 12 years.

We decided to investigate what would happen if we delayed the time of hormone treatment in mice to match that of the WHI. Two of my postdoctoral fellows, Shotaro Suzuki and Candice Brown, found that if we did not treat mice immediately after "menopause," but waited for several weeks (the equivalent of several human years), that hormone treatment was totally ineffective. What was even more exciting is that we found that one way estradiol acts is by suppressing inflammation, thereby preventing cell death. But if we waited a few weeks before starting to treat the mice, we were unable to suppress inflammation.³ The ability of estradiol to protect the brain by suppressing inflammation also depends upon the presence of ERα, since delay in the initiation of treatment no longer induced an increase in this receptor the way immediate therapy did. Just as our earlier studies had suggested, the timing of treatment is a critical factor to consider.

Many other investigators have asked whether all kinds of estrogens, synthetic or endogenous, act the same way. The answer is a definite no. Different kinds of estrogens and different concentrations of hormone have different effects. And the hormonal milieu makes a big difference in how estrogens act as they can have completely opposite effects depending upon what other hormones are present and whether they have been there before or after estrogen exposure (click here for graphic). While the estrogen research community had started to scratch the surface of the complex actions of this hormone before the WHI trial, our collective knowledge would not have predicted the outcome then. Today, with the insights into human physiology that the WHI trial brought, we have a much clearer picture of the nuances of this hormone.

One of the most exciting areas in neuroscience today is that of neurogenesis. The dogma had been that all neurons were born during embryonic and early postnatal development. Whatever neurons we were endowed with was what we had to work with. If someone suffered from a neurological disease or suffered a brain injury, the only way we could recover brain function was to encourage the remaining neurons to function more efficiently. About 20 years ago, several investigators began to question this assumption. They found that under some circumstances, new neurons were born even in adulthood. This set off a chase for finding compounds that would stimulate the birth of new neurons.

We had learned that estrogens protected the brain after stroke by limiting damage caused by inflammation. Until now, however, we hadn't considered whether part of the improved post-stroke recovery could be cause by the birth of new neurons. Researchers had noted that estrogen was important in the developing brain of an embryo. In the fetal and early postnatal brain, estrogens are factors that mediate neurogenesis, synapse formation, and glial differentiation, to name a few effects. More recently our studies have shown that estrogens are potent neurogenic factors after stroke injury in the adult brain. We showed that this effect depends on both ERα and ERβ.⁴ Not only can estradiol limit the effects of inflammatory damage, but apparently it also promotes repair and birth of new neurons. At the present time, we are uncertain about how estradiol works to enhance neurogenesis and are studying the cellular and molecular mechanisms that underlie these important actions. We are most excited about these new findings since they hold promise for the use of estrogens in the long-term repair and recovery of brain function.

Clinical trials such as the WHI are impressive for the number of women they study. But because of prohibitive costs of performing such a large clinical trial, only a limited number of questions can be addressed. In the year following the announcement that the WHI trial was halted, 40% of women stopped taking hormone replacement therapy. By following up on these paradoxical results, the field realized that results should have been interpreted with more caution and that conclusions were more limited than they seemed at first.⁵ This powerful hormone, with its many different analogs, different receptors, and the multiple mechanisms of action, must be used carefully. Its functions depend upon dose, preparation, method of administration, the recipient's age, genetics and previous exposure to the hormone. Although all of these caveats apply to all hormones, estrogens should be expected to be among the most complex ones to understand: they exhibit a diurnal rhythm, a monthly rhythm that is determined by the menstrual cycle, and they act differently depending upon whether other reproductive hormones are present in high or low concentrations.

It was a unique time. Although the trial created controversy in the community, between those who thought estrogen harmed and those who were convinced it would help, both sides were deeply motivated to understand the full story. We learned that estrogens are not the panacea people thought they were, but neither are they always harmful. It made us interact with one another with an openness I hadn't seen before. A basic researcher can live her entire professional life without ever having her assumptions tested against the ultimate system of human physiology. For me, it was a rare opportunity.

Phyllis Wise is the Provost and Executive Vice President at the University of Washington in Seattle, where she also leads an active research group in the Department of Obstetrics and Gynecology.

This is my story as a long-time "drug user" of various HRT combinations over a period of 26 years. I started out on HRT 6 months into menopause. The hot flashes were vicious--roughly every 45 minutes--and were ruining my life. For the next 6 years I was switched from one hormone combo to another (all synthetics), the worst being premarin-provera. I could not tolerate any of them without having almost daily headaches. Even the patch caused a slight skin reaction. The last combo caused such bad migraines that I discontinued all hormones & within a week felt wonderful, but within a month or so the hot flashes came back in force. I toughed it out for 6 more years without hormones ( big mistake).Along with the flashes came heart problems such as palpitations and shortness of breath. The doctor put me on beta blockers, then calcium channel blockers with the effects being so bad (extreme fatigue and diarrhea) that I was barely functional. After 6 months of this I had to get off these drugs. I was then introduced to estradiol and prometrium, low dose, and this combo worked! I was not constantly tired, nauseous, and there were no more hot flashes.The other interesting thing was that my "heart problems" vanished. I can only conclude from my own research and experiences as a human guinea pig that I was so low on estrogen that this is what was the primary cause of the physical symptoms. I have now been on estradiol-prometrium for the last 9 years with no side effects. It is a god-send to feel good, have energy & a zest for life . I realize that this is anecdotal evidence,but you have to listen to your own body and do what you believe is best, BUT educate yourself first and don't take a doctor's word as the gospel truth. What they call the "standard of care" is not always good for everyone. And investigate bioidenticals as a source of treatment.

Relating to the previous two commentors, one cannot blame Big Pharma for the woes of the world. One must know two things about the people who work in industry: they really do wish to help people, and they are out to make a lot of money. Recognize, however, that each "they" pertains to two very different groups, first the scientists and second the marketers. Unfortunately, the upper-level management of large corporation is usually not made up of scientists and usually sides with the marketers to help the bottom-line and please the shareholders. Hence, with marketers in charge of disseminating "the truth," some real science can be ignored with a clear conscience.

Understanding this, there is only one way to beat this system: competition. Good old capitalism! When a new company creates more natural equivalents or includes more precise and correct hormone formulations to obtain better outcomes, then and only then will things change.

Knowing thy competition, stop sarcastically laughing or complaining, take control, and do something about it.

Whenever estrogen is taken in a pill form the level of estrogen goes up while the available level of testosterone and many other androgens are cut in half. Most people don't realize that any oral estrogen pill is also an anti-testosterone pill.

The women in the WHI were never told this information and were led to believe that they were being given HRT or replacement for the estradiol and every other hormone their ovaries no longer made.

We were all misled. We were led to believe that HRT failed us. Premarin and Prempro in no way represent the hormonal milieu of ovarian function. Perhaps that is why there was no hormone testing done in the WHI.

Informed consent laws are supposed to protect the participants from this kind of misinformation and manipulative research.

We can only hope that in the future the NIH grows a conscience and follows their informed consent laws and creates meaningful research.

At this point in time we should be figuring out the best levels of estradiol combined with other hormones rather than still proving its importance. Wyeth, the maker of Premarin and Prempro, by continuing to calling these hormone preparations HRT, HT or hormone therapy and MHT, menopausal hormone therapy, continues to stall any modernization of treatment.

Beth Rosenshein
Diamond Research Foundation
www.DiamondRF.org

personally I would not use anything synthetic in my body especially when I can have access to bio-identical hormones which are made up to replace quantities which are lower in my body, ie specific to my body not one size fits all... why is there no decent research in this area?? well, it's, most likely money as usual bec bio-identical hormones can't be patented (because they come from plants and not some synthetic molecule) so the pharmaceutical companies prefer giving us fake stuff rather than the real thing...too bad most women aren't informed altho most of my friends are, since I do my best to pass the word about which books to read and what kinds of questions to ask.....makes you wonder really. Does big pharma have any desire to create products which are good for us?? Hahaha of course not!