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Controlled response

Anti-inflammatory drugs may fight the immune system's exaggerated reaction to malaria


When the microscopic malaria-causing parasite invades a person’s blood, it attacks and destroys the red cells, causing anemia. The damaged cells then adhere to the walls of the blood vessels and, in more severe cases, can even obstruct the blood flow, leading to brain damage. In response, the immune system is activated and completes the list of symptoms, bringing on fever, muscle pains, severe shaking and even convulsions. This disease, which is one of the major public health challenges, infects some 250 million people worldwide every year and is still  scarcely understood in humans. A measure of this lack of knowledge is the discovery of the group of immunologist Ricardo Gazzinelli, from the René Rachou Research Center, in the state of Minas Gerais: the immune system has an exaggerated response to this parasite, the plasmodium; this is exactly the opposite of what was thought. The results also indicate that drugs that can control this excessive inflammatory reaction –  which are currently not part of the arsenal used against malaria – may become valued allies against the disease.

Expectations regarding how the immune system might react to the infection resulted from what we know about bacterial sepsis, which consists of generalized infection and inflammation and which is in some ways similar to malaria. In extreme cases, the immune system enters a state that experts have  named “immune paralysis,” in which the defense cells stop reacting, much like a muscle that contracts to the point of making movement impossible. It is suspected that this process is driven by proteins that are the protagonists of innate immune responses, the receptors known as TLRs (toll-like receptors). On the membrane of defense cells, TLRs can recognize invading microorganisms and send signals to the other cells that are involved with the inflammation response that helps to fight the infection.

In an article published in April in the journal Proceedings of the National Academy of Sciences (PNAS), Gazzinelli’s group showed what happened when they cultivated cells from patients with acute Plasmodium falciparum infection, this being the most lethal form of malaria, in the presence of compounds that activate the TLRs, or agonists. “We expected to see cell tolerance to the agonists of these receptors,” the researcher tells us. This expectation was in line with the immune paralysis hypothesis. However, what they actually observed was the opposite: “In malaria’s acute phase, the response of the TLRs to the aggressor was highly increased, and we detected very high levels of inflammatory mediators in circulation, such as several cytokines.”

As part of a collaboration effort with parasitologist Luiz Hildebrando Pereira da Silva, the cells had come from 57 patients seen at the malaria clinic at the Rondonia State Tropical Medicine Research Center in the city of Porto Velho, which is now part of the Oswaldo Cruz Foundation, as is the case with the René Rachou institute. Though surprised, the team had no doubts regarding their findings. “The result was highly reproducible among patients,” explains Gazzinelli, “which shows that our initial understanding was wrong.” Upon measuring the content of inflammatory substances (the cytokines) in the blood, the group also found that when patients were treated for or cured of the parasite, this immune and inflammatory response returned to its normally low level.

The next step was to understand how the defense strategy against malaria is orchestrated by a patient’s genes. To this end, Gazzinelli’s group resorted to micro-arrangements, i.e., chips in which they could analyze at one time the level of activity of 20 thousand genes per patient, before and after treatment. Among those with malaria they discovered a larger expression of the genes that control the expression of toll-like receptors. Moreover, they found that this genetic activity is induced by a cytokine called interferon-gamma (γ).

During the research, which led to Bernardo Franklin getting a PhD under the guidance of Gazzinelli, the group also infected mice with Plasmodium chabaudi, the parasite species that causes the rodent version of malaria. When they analyzed the spleen of the mice seven days after the infection, the researchers found that interferon-gamma was being produced at a rate 20 times greater than that of mice free of malaria – a result that was very similar to what was detected in the blood of human patients. Finally, using genetically modified mice, the study characterized the sequence of events that leads the immune system to produce an exaggerated response. When the parasite enters the bloodstream, it activates one of the types of toll-like receptors, the TLR-9 kind, which, in turn, induces T lymphocytes – one of the types of defense cells – to produce interferon-gamma. This substance transmits a signal to the immune cells for them to express the genes of other varieties of TLRT, causing the defense system to respond strongly to the plasmodium.

Next steps
But this is not where things end. In his pursuit of a means to control the disease, the immunologist from the state of Minas Gerais selected more active genes among patients with malaria fever and, with this as a starting point, expects to develop biological markers to enable him to forecast each person’s resistance or susceptibility to malaria. Another line of work in this project, i.e., the study of malaria in rodents, may uncover strategies for blocking the over-activation of the immune system when faced with the disease. For the time being, the results indicate that anti-inflammatory drugs that interfere with the signaling path of the toll-like receptors may become major allies in the fight against malaria. Anti-inflammatory drugs of this kind are not yet being marketed, but some compounds are in the  pre-clinical and clinical trials stage.

The possibility of resorting to anti-inflammatory drugs is yet one more difference between the disease caused by the plasmodium and sepsis, which is also typified by an exaggerated inflammatory reaction. However, in the case of sepsis, without this inflammation, the bacterial invasion that gave rise to the problem can come out the winner (see Pesquisa FAPESP issue 146); regarding malaria, however, Gazzinelli and Franklin showed, in an article published in 2007 in Microbes and Infection, that the reaction to toll-like receptors in mice is not important in terms of controlling plasmodium infection. Therefore, though they will not cure malaria, these drugs may help avoid the disease’s symptoms.

Scientific articles
FRANKLIN, B.S. et al. Malaria primes the innate immune response due to interferon-γ induced enhancement of toll-like receptor expression and function. PNAS. v. 106, n. 14, p. 5.789-5.794. abr. 2009.
FRANKLIN, B.S. et al. MyD88-dependent activation of dendritic cells and CD4+ T lymphocytes mediate symptoms, but is not required for the immunological control of parasites during rodent malaria. Microbes and Infection. v. 9, p. 881-890. June 2007.