Newly published research led by a BBRF grantee sheds important new light on how, during pregnancy, activation of the mother’s immune system—for example, in response to a viral infection—can set in motion a series of mechanisms that culminates, in some instances, in developmental impairment of the fetus that manifests as autism spectrum disorder (ASD) after birth.

The research, conducted in mice, is noteworthy not only for identifying an immune conflict between mother and fetus that in some cases can follow activation of the pregnant mother’s immune system, but also for revealing a resulting pathological process that occurs in some male fetuses—but not in females.

Diagnosed cases of ASD are about four times as commonplace in male offspring compared with female offspring, for reasons that are not yet well understood. ASD, of course, does occur in females, but the significantly lower reported incidence has led over the years to theories postulating a variety of protective factors unique to females. In contrast, the new research identifies biological processes that are unique to male fetuses and their interaction with the prebirth maternal environment, and may help account, at least in part, for the preponderance of male ASD cases.

Lucas Cheadle, Ph.D., a 2021 BBRF Young Investigator at Cold Spring Harbor Laboratory, led the research, which appeared in Science Advances. Irene Sanchez-Martin, Ph.D., was first author of the team’s paper.

ASD “arises from genetic and environmental risk factors,” Dr. Cheadle and colleagues note. “One environmental factor, maternal immune activation (MIA), in which infection during pregnancy increases ASD risk in offspring, disproportionately affects males.” Their research sought to discover a basis for this male-specific vulnerability and the mechanisms by which inflammatory signals across the maternal-fetal interface affect the male embryo.

It is already accepted that a majority of fetuses exposed to inflammation in the womb do not undergo atypical neurodevelopment. A majority of fetuses are resilient, and only a subset of all fetuses exposed to a mother’s inflammatory immune-related signals appears to be susceptible. But which fetuses, and why?

The team used a mouse model of maternal immune activation in which an immune-stimulating compound is injected into pregnant females 12.5 days into the embryonic development of the fetus, a point corresponding with the late second trimester in human pregnancy. This is around the time when maternal infections have been linked retrospectively to ASD in children.

The first finding of the research was that about 30% of mouse embryos in each litter displayed one or more developmental abnormalities following MIA. The abnormalities, measured beginning 24 hours after the immune challenge to the mother, ranged from decreased fetal weight to altered sensory organ development. “These abnormalities occur exclusively in a subset of males, never in females,” the team notes. No abnormalities emerged in fetuses in a control group in which mothers were injected with saline solution rather than the immune-activating compound.

In this part of the study, the team was able to conclude that in cases where maternal inflammation is involved, a male bias in abnormalities culminating in ASD arises during gestation—rather than, or in addition to, during the time following birth. While ASD-related pathologies traceable to maternal infection that affect the male-female disparity may begin postnatally in some portion of cases, the study specifically indicates how some begin prior to birth.

Perhaps even more significant were findings concerning the mechanisms through which maternal infection can, in vulnerable males, affect their neurodevelopment. The team identified the placenta as the primary target of maternal immune activation. The researchers propose that activation of the mother’s immune system in susceptible individuals causes a rapid shift in the placental environment, and specifically in the interface between mother and fetus, transforming it from one that suppresses immune activity to one that is proinflammatory. In the vulnerable subset of male embryos, this initiates processes that culminate in developmental deficits including those involved in the ultimate emergence of ASD.

In cases where a male fetus was impacted by MIA, the team found in the placental cells of the mother a robust activation of genes supporting proinflammatory processes. This occurred in a broad range of placental cell types, but most prominently in a type of cell called spongiotrophoblasts. These are specialized cells originating in the rodent fetus (male fetus, in this case) that help to form the cellular border between mother and fetus. (There are analogous cells in humans.)

The increased expression of proinflammatory genes in these cells had effects, the team observed, that coincided with a breakdown in the structural integrity of the placenta and an accumulation of immune cells and cytokines in the amniotic fluid, the fluid normally protecting the fetus within the amniotic sac. Cytokines are signaling proteins that call immune cells to the scene of an infection or other immune challenge. They can control or exacerbate inflammation, but in this case, cytokines including IL-6 were found to be directly associated with the emergence of abnormalities in the fetus. IL-6 was in fact found to be required for this process to proceed.

Connecting the various pieces of experimental and other evidence, Dr. Cheadle and colleagues propose that two factors are necessary to give rise to developmental abnormalities following MIA. The first is an unusual abundance of proteins originating from the male fetus—antigens—which can be read by the maternal immune system as “non-self.” The abundance of such proteins occurs in a placenta that has switched over from being an immunosuppressive to a proinflammatory microenvironment, due to the virus or other environmental factor that has triggered the mother’s immune system. The activation of immune cells injures the placenta, the theory suggests, and compromises fetal health.

In rodent mothers with activated immune systems carrying female embryos, there is likely a loss of some immunosuppressive properties as they are similarly exposed to the same immune-system elements as male embryos in the same situation. The difference, the team suggests, is that female fetuses carry a lower burden of antigens that are derived from the father, compared with male fetuses. The placenta remains either undamaged or is less damaged in such cases of MIA, and the female fetus continues to develop normally.

Immune incompatibility between mother and fetus is thought to contribute to human embryonic problems such as pre-eclampsia and death of the fetus in the womb. It also may be a factor in accounting for why ASD is more common in first-born children (the pregnancy is the first introduction of the maternal immune system to the fetus’s antigens) and to children born to an older father (increased mutations in the father’s sperm may create new fetal antigens with which a mother’s immune system must now cope). In most cases, these issues don’t arise. But they do in some significant number of cases, and this may help account for some of the total risk for ASD and other neurodevelopmental disorders.

The team cautions that all of their experiments were conducted in rodents, and are only suggestive of processes that may be in play in humans. But the long history of research in rodents on human illness has opened many doors, not only to the biology underlying human pathologies, particularly those occurring at the dawn of life, but also to logical paths along which to pursue therapeutics and preventive measures. In this case, close scrutiny of the maternal-fetal interface under conditions of maternal immune activation is leading to new insights into the potential origins of some forms of ASD pathology, the sex skew in its incidence, and perhaps also, potential paths of prevention, which may someday extend to addressing problems that occur prior to birth.