Mom’s immune system and microbiome may help predict premature birth

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Every Monday, Jennifer Degl
leads a group through the halls of the neonatal intensive care unit at Maria
Fareri Children’s Hospital in Valhalla, N.Y. The volunteers offer support to
the parents of babies born early and struggling to survive.

Seven
years ago, Degl, a high school science teacher in Putnam County, was one of those
anxious parents. Her daughter, Joy, was born at 23 weeks gestation, weighing
just over a pound. The baby spent her first four months in that NICU. Degl
wasn’t allowed to hold Joy or change her diaper for a month.

Although
already a mom to three boys, Degl was completely unprepared for the experience.

“People
call [the NICU] a roller coaster for a reason,” she says. Joy would gain half
an ounce and do well enough for Degl to start the 45-minute drive home to spend
time with her sons, only to be called back because the baby was having trouble
breathing and needed a breathing tube. “There is no smooth NICU ride,” Degl
says.

Like Joy, roughly 10 percent of children worldwide — an estimated 15 million babies — are born prematurely, or before 37 weeks gestation, each year. In developed countries, surviving an early birth has become more likely, thanks to the availability of intensive medical care. More than 98 percent of U.S. preemies survive infancy, according to a study published in the American Journal of Obstetrics and Gynecology in 2016, though as many as 44 percent of the youngest preemies don’t make it. Survival is least likely in nations with the fewest resources. Worldwide, complications associated with preterm birth are the leading cause of death in children younger than 5 years old.

But
survival is just step one. Many preemies face breathing problems, infections
and other complications that can cause issues well after infancy. Children born
prematurely can experience developmental delays and have a higher risk of
learning disorders such as attention-deficit/hyperactivity disorder. Many
require physical, speech and other types of therapy well into childhood. In the
United States, health issues related to prematurity collectively cost more than
$26 billion a year.

Joy
has needed physical therapy, feeding therapy and occupational therapy. She
still receives speech therapy twice a week, and because of scar tissue in her
lungs from the NICU ventilators, she wheezes when she exerts herself. A simple
cold is dangerous — she’s had pneumonia eight times.

Mom holding Joy
Jennifer Degl, holding her fourth baby, Joy, in the NICU, works to help other parents cope when their premature babies must stay in the hospital. Courtesy of J. Degl

For
decades, researchers and clinicians have sought ways to predict and prevent
preterm birth with little progress to show for it. “It’s extremely
frustrating,” says neonatologist Sylvain Chemtob of Centre Hospitalier
Universitaire Sainte-Justine in Montreal, who has worked in the field for 35
years. The best predictor of preterm labor is whether a woman has experienced
it before. Other risk factors include carrying multiples, having a short cervix
and medical conditions such as diabetes or high blood pressure.

But
about half of preterm deliveries involve no known risk factors at all. “There’s
plenty of room to improve,” says Brice Gaudilliere, a physician-scientist at
Stanford University.

Gaudilliere
and others are looking to the human immune system for clues. “The immune system
is exquisitely sensitive to all sorts of environmental changes,” he says,
including the mother’s nutrition and stress. The immune system could be the
biological common denominator for the many known and suspected factors that
contribute to preterm labor.

Joy Degl as a ballerina
Today, Joy Degl is an active 7-year-old despite lingering breathing difficulties.Courtesy of J. Degl

Immune-related
genes and proteins involved in inflammation have been linked to preterm birth
for decades, but such links have not resulted in predictive tests or
treatments. “It’s one thing to say that the relationship between inflammation
and preterm birth is known,” says Nima Aghaeepour, a machine learning scientist
at Stanford. “It’s another thing to ask what are we going to do with this
information.”

To
bridge this gap, Gaudilliere and Aghaeepour are collaborating to examine the
immune system as a whole — dozens of cell types, hundreds of molecules and
thousands of genes. These researchers and others are using this systems
immunology approach to find ways to predict a woman’s risk of premature labor
based on a small sample of her blood, and then reduce that risk.

An inflammatory process

As soon as a woman becomes
pregnant, her immune system changes. Her body releases chemicals that keep
immune cells from attacking the embryo’s cells as foreign invaders. Once the
early ball of cells implants into the wall of the uterus, a thick layer of
tissue called the decidua starts to form between mother and embryo. For the
rest of the pregnancy, molecules released by the placenta and uterus, as well
as anti-inflammatory immune cells such as regulatory T cells, keep the immune
system at bay.

When
the pregnancy reaches full term, at 37 to 40 weeks, the uterus somehow switches
out of this immune suppression, says Sam Mesiano, a reproductive biologist at
Case Western Reserve University in Cleveland. Immune cells flood the area and
set off a chain reaction that ultimately triggers the uterus to contract.
Inflammation also causes cells to release enzymes that dissolve the membranes
surrounding the fetus, which break and release amniotic fluid. “All these
things get switched on by this inflammatory process,” Mesiano says. “That’s
what we want to happen.” But not before 37 weeks.

Some of the signs of inflammation linked to preterm birth differ from those found during full-term birth, says Nardhy Gomez-Lopez, a reproductive immunologist at Wayne State University in Detroit. For example, in 2017, she and colleagues reported in the American Journal of Reproductive Immunology that some proteins involved in inflammation, called cytokines, were present at higher than normal levels in amniotic fluid from a subset of women who delivered preterm. The earlier the women delivered their babies, the higher the cytokine levels. Infections, which are present in at least a quarter of preterm births, could be the cause, but inflammation and cytokine levels were also elevated when no infection was found.

Obstetricians
sometimes measure cytokine levels in amniotic fluid, but only when preterm
labor has already begun and an infection is suspected. Gomez-Lopez says
researchers have to back up and look for reliable immune markers that are
detectable in blood and tie them to the changes seen in the amniotic fluid. “We
think that by studying the systemic [immune] response in the mom, we can
predict these changes way earlier,” she says.

Cast a wide net

Part of the problem with
developing a predictive test is that preterm labor isn’t just one condition.
Thirty years ago, preterm labor was viewed simply as regular labor that
happened early, says perinatologist Roberto Romero at Wayne State, who directs
the perinatology research branch of the Eunice Kennedy Shriver National
Institute of Child Health and Human Development, or NICHD. Although scientists
now recognize that the biology of preterm labor is distinct, they still have to
grapple with the reality that it varies depending on the underlying cause. The
cells and molecules active during preterm labor brought on by infection, for
example, are different from those active in labor brought on by a drop in the
hormone progesterone.

Because
not all causes of preterm labor are known, it is hard to find biological
markers for each case, Romero says. A system-wide analysis can help because
researchers don’t have to know in advance which gene or protein to focus on, he
says. By examining, for instance, all the active genes in a woman’s white blood
cells, or all the proteins present in a blood sample, researchers can flag
differences in the immune systems of women who deliver prematurely versus those
who deliver full-term.

Wayne State and NICHD recently released gene activity data from the whole blood of 150 Detroit women, 71 of whom delivered preterm, and encouraged researchers to use the data to find predictors of preterm labor, as part of a crowdsourcing collaboration called the DREAM challenge. The challenge is expected to be completed in January 2020.

Aghaeepour
and Gaudilliere are taking the systems immunology approach a step further,
beyond measuring gene activity. Their teams are also collecting data on the
cells that contain those active genes, tracking fluctuations in the numbers of
those cells, studying which molecules are produced, how active each cell type
is and how those changes affect other immune factors. Casting a wide net is
important, Gaudilliere says, because if one type of immune cell is responding
to something, other types are probably also involved. “It makes little sense to
focus on one or the other cell type,” he says.

When
Gaudilliere joined the March of Dimes Prematurity Research Center at Stanford
in 2015, he quickly realized how little was known about immune cells throughout
pregnancy and labor. “From a basic physiological understanding of even normal
pregnancy, we’re just scratching the surface, especially in the domain of immunology,”
Gaudilliere says. One of the first things he and Aghaeepour, who joined in
2017, did was set up a study to establish what the immune system looks like
throughout full-term pregnancy. It was part of their homework, Gaudilliere
says.

The two recruited 21 women to donate three blood samples over the course of pregnancy and analyzed close to 1,000 features of the women’s immune systems at each time point. Features included measurements of 24 types of immune cells, the levels of immune-related molecules present in each cell type and the cells’ ability to respond to stimuli in laboratory experiments.

Putting
it all together, the team developed an “immune clock” of pregnancy — a
mathematical model that links the many immune parameters with how far along a
pregnancy is. The model, reported in 2017 in Science Immunology,
accurately estimated the gestational ages of a new set of 10 pregnancies. Now
the team is studying whether women who go on to deliver prematurely diverge
from the immune clock. With help from collaborators at the University of
Chicago, the group is refining the algorithm by incorporating immune changes
found in placentas collected after delivery.

Making connections

The Stanford team has been
trying to boost the clock’s accuracy by adding layers of data from outside the
immune system. “We established that the immune system does change throughout
pregnancy and that it’s very systematic,” Aghaeepour says. “But we know that
the immune system doesn’t act in isolation.”

The group recently integrated its immune measurements with several other data sources from 17 pregnant women: their gut, vaginal and oral microbiomes, blood levels of proteins and metabolism-related molecules, plus fetal genetic material released into the women’s blood. A machine learning algorithm found that the data as a whole were far more accurate at predicting gestational age than any one type alone. The study, published in January in Bioinformatics, included thousands of measurements.

Agheeapour
says the more samples they can collect to teach the algorithm, the more
accurate the program will be, and the better it can point researchers to the
important drivers of preterm birth risk. The plan is to pare down the test to
make it usable in resource-limited settings.

The
Stanford team is not alone in its attempt to combine different kinds of data
for one test. Instead of looking to blood, microbiologist Jennifer Fettweis of
Virginia Commonwealth University in Richmond and her colleagues are putting
together huge datasets from pregnant women’s vaginal microbiomes. The
researchers recently tracked microbiome composition and microbe gene activity
from vaginal swabs collected throughout the pregnancies of 597 women. The
researchers combined that data with periodic measurements of the women’s
vaginal cytokine levels. 

In a sample of mostly African-American women, 90 had delivered full-term and 45 had delivered preterm. The women who delivered preterm tended to have a more diverse mix of microbes than those who delivered at term, the group reported in June in Nature Medicine. The women who delivered early had much lower levels of Lactobacillus crispatus and they carried more of other types of bacteria, such as Lachnospiraceae BVAB1, that are linked to higher levels of cytokines that instigate inflammation and to vitamin D deficiency — two factors previously tied to preterm birth. The researchers suggest that microbiome changes could be a useful predictor of preterm labor risk. But because people’s microbiomes vary with geography and diet, among other things, no one microbiome profile will be predictive for everyone.

In an accompanying study, Fettweis and colleagues reported finding differences in microbiome diversity among African-American, Hispanic and white women who delivered full-term babies. The researchers don’t yet know why these differences exist, but they hope that the microbiome will hold clues about why African-American women are 1.5 times as likely to give birth prematurely and twice as likely to have very preterm infants (born before 34 weeks) as white women in the United States.

Many researchers think the microbiome differences are related to environmental factors, such as stress and nutrition. A group at Emory University in Atlanta is collecting environmental, microbiome and immune data from more than 500 pregnant African-American women to get some answers.

Drive for more data

As part of the Multi-Omic
Microbiome Study-Pregnancy Initiative, Fettweis and others are analyzing
microbiome samples from volunteers’ mouths, skin, vaginas and rectums, as well
as blood and urine samples and health data. Fettweis would also like to explore
how microbiome changes associated with preterm birth fit with other types of
data. It may be that some links between the activity of certain genes or cells
and preterm birth depend on the state of a mother’s microbiome, she suggests.

“We
need to start thinking about these things together,” she says. “We need
harmonization in the field.”

For data scientist Marina Sirota of the University of California, San Francisco, harmonizing data is a full-time occupation. She mines health data in search of relationships between risk factors and biological markers connected to preterm birth. In a meta-analysis of three studies, Sirota and collaborators found 210 genes with activity in white blood cells that differed in women who delivered preterm. Most of the affected genes were involved in immune responses, the researchers reported in 2018 in Frontiers in Immunology.

In a separate study, published in 2018 in Environment International, Sirota and colleagues matched California birth records with state data on environmental pollutants. Premature labor was more common among women living in areas with high levels of two drinking water contaminants, arsenic and nitrate. Sirota says many of the genes known to be affected by arsenic exposure are the same genes that she and colleagues found to be affected in their meta-analysis.

Work
like Sirota’s might eventually point to treatment options to reduce the risk of
preterm birth related to environmental exposures. After all, Gaudilliere says,
the end goal is to do more than find new risk factors; the aim is to come up
with therapies that target the different causes of preterm labor.

In Montreal, Chemtob has been working on a way to block the cytokine interleukin-1, which has been linked to preterm labor. On February 13 in Frontiers in Chemistry, he and colleagues described an IL-1 inhibitor that prevents inflammation-induced preterm birth in mice without hindering the cytokine’s normal anti-infection activities. The next step, Chemtob says, is lab studies testing women’s white blood cells to find the population of women who will most likely benefit.

He’s
working with Gaudilliere’s group to develop a lab test that can identify women
whose white blood cells change their activity when exposed to the inhibitor.
The researchers plan to pair the results of the blood test with the other
measurements to ultimately design a test that predicts preterm birth risk and
potential drug response at the same time. “That makes for ideal personalized
medicine,” Chemtob says.

“We
are at the beginning of an exciting period,” says Romero at Wayne State. The
field is now equipped to start studying preterm birth as a collection of
several different syndromes and seek out treatments to address each one, he
says.

This
shift could not come soon enough. For the youngest infants, even one extra day
in the womb can make a huge difference for their health.

After spending the last seven years speaking to and advocating for parents of preemies, and dealing with her daughter’s lingering health problems, Degl can attest that new ways to prevent prematurity are needed. If there was anything she could have done to extend her own pregnancy, she says, she would have done it without hesitation. “I think every mom would say that.”

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