Maternal immunity protects fetuses from harm from Oropouche

Maternal immunity protects fetuses from harm from Oropouche

Experiments with animals show the importance of health care during pregnancy and monitoring for epidemics.

A mother's love is a fortress, and it seems her immune system is too. That's what a study from Unicamp, recently published in the journal, suggests. iScience, from the Cell group. The study, conducted by researchers from the Institute of Biology (IB), investigated the possible effects of congenital infection — from mother to child — by the Oropouche virus (Orov), finding that, when the pregnant woman has a good immune system, the infection reaches her reproductive system, but causes almost no damage to the fetus.

According to the study, this protection occurs because, upon contact with the pathogen, cells of the reproductive system produce different cytokines, including type I and III interferons, signaling proteins that constitute one of the main components of the innate immune response. When released, these cytokines bind to interferon receptors present on other cells and trigger a signaling pathway that culminates in the expression of genes with antiviral activity, helping to control viral replication.

Conversely, if maternal interferon signaling is impaired, the virus passes to the fetus in greater quantities, causing more damage to the child, even if its immune system is intact. The reasons for this effect were not evaluated by the study, but it is possible that, even if the fetus possesses a genetic framework to respond to the virus, the lack of maturity of this response prevents a more effective action by the fetus. “The mother's body is the habitat, the barrier that controls what passes through. So, when she controls the infection, she can protect the baby,” considers IB professor José Luiz Módena, supervisor of the thesis that originated the publication.

This article is the result of research conducted by biomedical scientist Stéfanie Primon Muraro. In her doctoral studies at the IB (Institute of Biosciences), the author investigated the mechanisms behind the transmission of Oropouche virus between mother and child, seeking to fill a gap in experimental research on this topic. Since the 1980s, reports and observational studies have pointed to an association between pregnant women infected with Oropouche and cases of spontaneous abortion and fetal malformation, but no analysis had been done to detect the virus in fetuses.

Common in forested areas, Oropouche virus has been known in Brazil since the 1960s and is widely transmitted among wild animals such as sloths, primates, rodents, and birds. In humans, it is usually transmitted through... Culicoides paraensis, a mosquito from the black fly family popularly known as maruim or "porvinha". From time to time, cases of transmission between humans by the vector insect occur in urban areas, but, because it is a disease associated with economically vulnerable regions, especially in the North of the country, its study has historically been neglected.

The main symptoms of Oropouche are fever, joint pain, and skin rashes that disappear after a week. However, some patients develop serious complications, such as neurological sequelae and hemorrhage, which can even result in death. However, the "clinical similarity between Oropouche infection and that caused by other arboviruses, such as dengue, Zika, and chikungunya, creates diagnostic difficulties and leads to underreporting of Oropouche cases," states Muraro in his thesis.

The experiment

To arrive at the results, Muraro, with the support of the team from the Laboratory for the Study of Emerging Viruses (LEVE), led by Professor Módena, developed assays in cells and in pregnant mice. In the first case, the author evaluated whether the pathogen was capable of multiplying in placental cells of humans and mice—which was confirmed—and what types of immune response these cells would eventually trigger—in this case, the release of interferons. Furthermore, using non-pregnant mice, she observed that the virus can reach female reproductive organs, such as the uterus and ovaries.

In the second trial, the researcher worked with a crossbreeding and infection model in pregnant mice. At this stage, the aim of the experiment was to evaluate whether the virus can infect pregnant females and, if so, whether it is capable of crossing the placenta and causing harm to the fetus. "This is not guaranteed, because when pregnant, the female undergoes several changes in her physiology and hormonal levels that could affect the virus's ability to infect and transmit," says the professor.

To this end, experiments were conducted with three mouse models: the wild-type, which has two copies, maternal and paternal, of the gene responsible for expressing the interferon receptor — homozygous positive — and therefore responds to this cytokine in its entirety; the homozygous negative, which has no copies of this gene and, therefore, does not respond to interferon signaling; and a heterozygote, which has only one copy of the gene, but also expresses the interferon receptor.

The experiment demonstrated that, for the wild-type mouse—which mated with a wild-type mouse and produced only wild-type fetuses—the virus reaches the placenta and fetus at low levels, but without causing significant changes in size or neurodevelopment. On the other hand, the negative mice developed a very severe Oropouche infection and died within three days of infection. This demonstrated the importance of interferons for defense against the virus, but made it impossible to monitor the pregnancy and the effects of the disease on the fetuses due to their premature death.

Given this result, the experiment turned its attention to heterozygous females. Their single copy of the gene allows them to survive infection by the virus, but when mating with a negative male, he will not transmit a copy of the gene to the offspring. Therefore, the female mouse can produce offspring that are responsive to the receptor (heterozygous) if the fetus receives the positive gene from her, and non-responsive offspring (homozygous negative) if the offspring do not receive it. This allowed them to monitor the infection in the double-negative fetuses.

With this step, the study confirmed the importance of the maternal defense system, because, although the fetuses presented lower weight and size, this difference was not statistically significant when compared to those in the control group—which were not infected. It is worth noting that the analysis of the placentas from this crossbreeding revealed multiple areas of calcification, which the authors believe indicate locations with active viral replication. This is a phenomenon observed in other types of viral and parasitic infections, such as rubella and toxoplasmosis, and affects functions such as the exchange of nutrients between the mother and the fetus—which would impact the offspring's growth—but this relationship was not confirmed by the study.

As a final step to prove the dependence of maternal interferon on the immune pathway, the researchers treated wild-type mice with different doses of an antibody that "switches off" the interferon receptor and prevents the antiviral response mediated by this cytokine. This allowed them to reduce the mother's immune response and observe the virus reaching the double-positive fetuses and multiplying more easily. "The viral load in the placenta increases significantly, even with very low doses of the antibody, and we see an increase in fetal viral load and a tendency for the fetus to be absorbed by the mother, showing that in this scenario, abortion or growth restriction may eventually occur," says Módena.