New technology enables researchers to better understand early placental development

A recent study done jointly by researchers at American and Israeli universities and hospitals has revealed crucial insights into the early development of specialized tissues that protect the fetus during human pregnancy, with the findings potentially helping in the prevention of dangerous pregnancy complications, like preeclampsia and preterm birth.

The research, which focused on the development of the maternal-fetal interface in the first half of pregnancy, reveals new knowledge of how a mother’s uterine arteries transform for healthy placental growth and function, and how foreign fetal cells manage to escape maternal immune response while invading the mother’s uterine wall. Even when in close proximity, the mother’s potent immune cells do not attack the fetus’s cells.

The study results, published in July in the peer-reviewed Nature journal, were made possible by the use of new technology, known as multiplexed Ion Beam Imaging by time-of-flight, or MIBI-TOF, in Dr. Mike Angelo’s lab at Stanford University. MIBI-TOF is a special instrument that enables scientists to detect dozens more genetic markers and proteins simultaneously than optical light while examining tissue samples.

“Mike invented this amazing microscope which really revolutionized pathology,” said Dr. Shirley Greenbaum, the study’s first author.

Following her medical studies and residency in obstetrics and gynecology at Hadassah Medical Center, Greenbaum did her postdoctoral studies at Stanford, beginning in 2017. In her clinical work, she treated patients with preeclampsia, a serious and potentially fatal complication for a mother and her fetus, which affects five-to-eight percent of pregnancies.

“Preeclampsia is such a puzzling disease and it is specific mostly to humans. There have been dozens of years of research in the field and we understand more, but we haven’t made much progress in terms of treatment. We basically deliver women who are diagnosed with preeclampsia early, even at the price of the baby’s prematurity, to save the mother and baby’s life,” Greenbaum said.

Six dynamic MIBI color overlays from different time points during the first half of the pregnancy exploring trophoblast invasion in early pregnancy villi and maternal decidua. (Courtesy of Stanford University Department of Pathology)

Among the difficulties in studying the causes of preeclampsia on the cellular level is the fact that it is not diagnosed until at least the 20th week of pregnancy, when it is too dangerous to take placental samples. In addition, since the condition is found almost exclusively in humans, there are no animal models to study in the lab.

Examination of placentas after preeclampsia births has shown that the arteries in the layer that borders the maternal and fetal sides of the placenta, called the decidua, maintain their smooth muscle and do not uncoil from tight spirals. For a healthy pregnancy, the arteries must change their structure and open up properly to provide oxygen and nutrients to the placenta and fetus.

Dr. Shirley Greenbaum (Orit Balicer-Tsur)

“The current thinking is that preeclampsia actually originates in the critical window of the early weeks of pregnancy because that’s where all the remodeling of maternal arteries takes place,” Greenbaum said.

To investigate this theory, the research team obtained 66 samples of non-medically indicated pregnancy terminations (between weeks six and 20) from the University of California in San Francisco. They used MIBI-TOF to examine around 500,000 cells and 588 arteries in intact deciduas from different stages of pregnancy. MIBI-TOF allowed the scientists to detect up to 40 markers simultaneously at the single-cell level — a whole new level of resolution.

They were able to see how in healthy pregnancies, the maternal arteries uncoiled properly over time as the placenta adhered to the uterine wall.

“These arteries, when they become dilated, can deliver a low-pressure, low-velocity flow of blood into the placenta. Then the fetal finger-like structures in the placenta called villi bask in the blood and take the nutrients and oxygen so that the baby can grow and the pregnancy can develop in the optimal way,” Greenbaum said.

The study’s second focus related to the normal “invasion of fetal cells” into the mother’s uterine wall tissue. The researchers saw, surprisingly, that the mother’s healthy and robust immune system did not attack the foreign fetal cells.

MIBI color overlay of 16 weeks gestation decidua sample. A thin and disrupted smooth muscle layer can be seen in blue, and invading fetal cells are denoted in magenta. Fetal cells can be observed within the artery lumen and surrounding it. (Courtesy of Stanford University Department of Pathology)

“That’s amazing because in any other situation, those foreign cells with different proteins and DNA would immediately evoke an immune response,” Greenbaum noted.

To understand this phenomenon, Greenbaum and the others needed to know what cell populations were present in the layer surrounding the pregnancy and what proteins they were expressing. They found that a whopping 50% of the cells in the maternal side of the decidua were immune cells.

“That’s striking. I mean, not many tissues in the body have such a high percentage of immune cells. Not only that, but here, they’re not attacking the pregnancy,” Greenbaum said.

MIBI-TOF (multiplexed ion beam imaging by time of flight) instrument at the Angelo Lab in Palo Alto, California. (Courtesy of Stanford University Department of Pathology)

Thanks to the advanced technology at the team’s disposal, they were able to not only determine the percentage of immune cells at every stage of the first half of pregnancy, but also to identify the specific subpopulations of the maternal and fetal cells in the area and the proteins they express to communicate with one another.

One finding was that some more tolerogenic (tolerant) immune cell populations increase with the advancement of the pregnancy, while others that are considered to be more aggressive decrease.

“This did not surprise me, but we found that some of those proteins expressed have a tolerogenic effect on the immune system in other cases. It was pretty amazing to see some similarities between tolerogenic proteins (such as PD-L1) in the maternal-fetal interface, also play a role in suppressing the immune response in cancer,” Greenbaum said.

“It’s unpleasant to compare between a fetus and cancer, but it is really interesting from the biological perspective,” she said.

Now that Greenbaum and her colleagues have mapped out the cells and proteins involved in the normal physiological process of healthy maternal-fetal interface development, it would be possible to make meaningful comparisons to what happens in preeclampsia. MIBI-TOF could make this possible.

“We would look again at placentas from preeclampsia births and hopefully we will spot differences in some of the [cell] populations and in the expression of some interesting proteins that we that we’ve captured. And then we’ll go from there,” Greenbaum said.