Researchers describe the role played by phosphatidylinositol 3-kinase gamma (PI3Ky) in regulating the immune response against the protozoan parasite Trypanosoma cruzi, which causes Chagas disease.
The team, affiliated with the University of São Paulo (USP), Brazil, described how molecules able to control the signalling pathway mediated by this enzyme could be tested as a treatment for Chagas disease.
Lead author and researcher at the University of São Paulo’s Ribeirão Preto Medical School Maria Claudia da Silva mentioned, “Our results show that the expression of PI3Kγ increases during infection by T. cruzi in both mice and humans. This appears to be essential to avoid excessive inflammation that might injure the organism and also to control heart parasitemia”.
According to the WHO, Chagas disease affects 8 million people worldwide, with the majority of cases in Latin America, where it remains one of the biggest health problems, causing more than 10,000 deaths per year.
The parasite, transmitted via insect bites from triatomine, blood transfusions with people already infected, or by ingesting contaminated food, stays with infected individuals for life. The initial phase of the disease may be asymptomatic, or may cause nausea, headache, fever, rash, swollen eyelids, or enlarged liver and spleen.
Should the disease not be treated, complications such as heart ventricle enlargement or oesophagus/colon enlargement may occur. In turn, these would lead to heart failure, loss of peristaltic movement and sphincter dysfunction.
Supervising the research, Professor João Santana da Silva said, “In our experimental model, we used a strain of the parasite that prefers heart tissue. The initial tests showed that PI3Kγ-deficient mice developed severe cardiomyopathy in the acute phase and died after a short time, but we had no idea why this happened.”
The team also identified that in genetically modified mice, who are not able to express PI3Ky, the levels of parasite were the same as in wild mice. However, as the wild mice could express the enzyme, they survived the condition.
“When we looked at their [the mice] hearts, we found that PI3Kγ-deficient mice had far higher parasitemia levels and much more severe inflammation [myocarditis]. The immune system was producing proinflammatory molecules in an uncontrolled manner, injuring the heart tissue, and even so, it was unable to kill the parasites efficiently,” said Prof da Silva, after describing how the team expected the mice that died to have larger numbers of parasites circulating in the bloodstream than those organisms that survived.
The immune response to the parasite was also investigated by the team, along with how it changes in the absence of PI3Ky. Professor Thiago Mattar Cunha described how other studies show the enzyme in a signalling pathway vital to the migration of defence cells to sites of inflammation within the organism.
Under ordinary conditions, chemokines are produced by the parasite when it infects host cells. Macrophages and dendritic cells are activated by the chemokines, which migrate to the site to attack the invading cells.
Though not totally efficient, this defence mechanism manages to keep levels of the parasite low, with most individuals having no symptoms in the acute phase.
“Our results show that when the signalling pathway mediated by PI3Kγ is not active in macrophages, these defence cells lose their capacity to kill the parasites and control the inflammation,” Prof Cunha said. “To prove that the problem resided specifically in macrophages, we used an animal model called conditional knockout, in which PI3Kγ is missing only in macrophages.”
The team found that without the enzyme, the macrophages produce less nitric oxide, which is necessary to kill the parasites, and acts together with a pro-inflammatory cytokine, interferon gamma (IFNγ).
Should the macrophages not express PI3Ky, they cannot kill parasites in the presence of IFNγ, explained Prof da Silva.
In partnership with Edecio Cunha Neto, a researcher at USP’s Medical School in São Paulo (FM-USP), the team studied tissue from patients who developed cardiopathy in the chronic phase of Chagas disease and who underwent biopsy or heart transplant. They also analysed a database containing information on all molecules expressed in the heart tissue.
Individuals with higher levels of PI3Kγ had lower levels of heart parasitemia than those who expressed less PI3Kγ, although both groups presented with myocarditis. In addition, all had higher levels of PI3Kγ and of all molecules in the pathway mediated by this enzyme than non-Chagas patients with chronic congestive heart failure.
Prof da Silva explained, “These findings suggest that this enzyme is also involved in [the] control of the parasite in humans. In our in vitro experiments, we observed that human macrophages failed to kill the intracellular pathogen when infected with T. cruzi after treatment with the PI3Kγ inhibitor. How this happens is something we have yet to understand.”
Preliminary results indicate that patients who develop cardiopathy in the chronic phase of infection by T. cruzi, have a higher incidence of a polymorphism that may be associated with lower activity of PI3Kγ than in patients who develop chronic disease in other organs, such as the spleen or intestine.
“We’re currently writing another paper in which we raise the hypothesis that people with a certain polymorphism in the gene that encodes PI3Kγ run a greater risk of developing cardiopathy in the chronic phase,” Prof Cunha said.
He went of to say that another possible line of investigation would be to determine whether rare cases of patients who die suddenly of myocarditis are associated with lower levels of PI3Ky, as the team observed in the mouse models.
Prof da Silva describes interest in investigating the signalling pathways that modulate PI3Kγ production in the human organism.
“Drugs are available to inhibit PI3Kγ production but not to stimulate it,” he said. “We now need to investigate the regulatory pathways mediated by PI3Kγ in search of molecules that induce the release of these substances. We’ve already performed lab tests on some molecules that display this type of action. It’s basic science, but there are potential applications for the control of parasitemia in both the acute and chronic phases.”
According to Prof Cunha, activating the enzyme “is no easy task” and could have implications for other pathological conditions. “Inhibitors of the PI3Kγ pathway are being tested for the treatment of cancer and inflammatory diseases,” he said.
The study was published in Nature Communications.