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Fertility study shows increased estrogen shortens window of implantation in mice

Findings may explain low fertility rates with IVF

Vanderbilt University Medical Center

VANDERBILT One of the most vexing causes of infertility is the failure of the embryo to implant in the uterus. It's estimated that roughly three-quarters of all embryos fail to implant properly. And in those women being treated for infertility, even when healthy embryos are transplanted the failure rate is high: up to 70 percent fail to dock successfully in the uterine lining.

The interplay of signals between embryo and uterus during implantation is complex, yet recent findings by a group of Vanderbilt researchers may provide clues to better understand it. The group, led by S.K. Dey, Ph.D., Dorothy Overall Wells Professor of Pediatrics and Cell & Developmental Biology, has found evidence in mice suggesting that estrogen plays a critical role in determining the window of uterine receptivity for embryo implantation.

A report of the researchers' work, which could have implications for improving human fertility, currently appears in the online version of the journal Proceedings of the National Academy of Sciences, USA.

The studies show that too much estrogen alters expression of implantation-related genes in the uterus, which can rapidly abolish uterine receptivity to the embryo. Moreover, the amount of estrogen sufficient to elicit these changes is very small.

"The most exciting aspect of this investigation is that a very narrow range of estrogen levels can alter embryo implantation and gene expression," said Dey, who is also head of the division of Reproductive and Developmental Biology. "We're talking about nanogram levels here, which I would consider to be almost physiological doses in the mouse."

For their studies, the scientists allowed mice to copulate, then removed the ovaries of female mice to halt normal production of estrogen and progesterone, both of which are required for implantation to occur. Daily doses of progesterone kept the mice in a state of "delayed implantation," where any developing embryo, or blastocyst, present would remain suspended in a kind of dormancy. It was at this stage that the researchers were able to parse estrogen's effect on the implantation process.

They found that a single injection of either 3, 10, or 25 nanograms of estrogen was capable of initiating implantation. However, injection of three nanograms of estrogen was enough to prolong uterine receptivity in the mice for up to four days. Increasing the estrogen dose to 10 nanograms rapidly closed the window of receptivity: within 24 hours, implantation no longer occurred. A second dose of estrogen, whether low or high, did not rescue receptivity after an initial high dose.

What's more remarkable, Dey says, is that the same sensitivity to progesterone doesn't seem to exist. "You can increase progesterone, say from one to four milligrams, and still, implantation would be fine," he said. "And a high progesterone level does not counteract this estrogen effect."

When the researchers looked at expression of a handful of key, implantation-specific genes at the site of blastocyst implantation, they found an abnormal pattern of activation in mice that received high doses of estrogen. Future studies may shed light on other gene expression changes, and on whether this effect is patchy or uniform in the uterus.

"It will be really interesting to see whether the gene expression pattern is changing from region to region within the uterus with changing estrogen levels," said Dey.

The applicability of these findings to humans is uncertain yet promising. Levels of estrogen in women undergoing in vitro fertilization rise due to the ovarian hyperstimulation required for retrieving multiple eggs. One prediction from their investigation, Dey says, is that exposure of the uterus to these increased estrogen levels may be contributing to high implantation failure rates.

Estrogen levels in women can be affected in other ways, as well, whether through the drugs or medications they take, the foods they eat, or certain environmental toxins they might encounter, such as some pesticides, which can behave as estrogens. And, there may be ethnic or population differences, based on genetics, which influence sensitivity to estrogen. Such factors could contribute toward implantation failures generally, even among those couples not participating in assisted reproduction programs.

Determining whether estrogen levels are as restrictive in humans as in mice will likely not be easy, says Dey.

"Studies in women would be very difficult, and may be impossible," he said. "But it might be done by taking small biopsies of the uterus during the menstrual cycle and carefully looking at gene expression with changing estrogen and progesterone levels. That way, we might get some idea. Alternatively, similar studies in sub-human primates may provide more meaningful information that could be relevant to humans".

The current studies were done in collaboration with Sanjoy K. Das, Ph.D., associate professor of Pediatrics, and Bibhash C. Paria, Ph.D., associate professor of Pediatrics, and were largely the work of former research associate Wen-ge Ma, now a graduate student at UCSF, and Haengseok Song, Ph.D., a former graduate student, now a post-doctoral fellow at Washington University in St. Louis.

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The studies were supported by grants from the NIH and the Mellon Foundation.

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