“Our line of research aims to identify the biochemical pathways involved in this neuroimmune regulation,” Gabanyi told Agência FAPESP. “We believe the results will help us understand and treat chronic intestinal diseases such as irritable bowel syndrome, for example.”
As the article in Cell explains, the intestine is constantly exposed to a variety of antigens, from those present in food to the thousands of microorganisms that make up the gut flora. Mechanisms are therefore required in the intestine to control the immune response to these antigens because excessive inflammation can be harmful to tissue.
“We discovered that macrophages in the intestine specialize. When these defense cells are close to the gut lumen, they are able to perceive the presence of pathogens and have a more pro-inflammatory profile, whereas the macrophages in the intestinal wall have no contact with potential invaders and display a more anti-inflammatory profile,” Gabanyi said.
The researchers also found that in the presence of potentially pathogenic bacteria, a specific group of neurons is activated and releases the neurotransmitter noradrenaline, which induces the expression of anti-inflammatory genes when it comes into contact with intestinal wall macrophages.
Assembling the jigsaw puzzle
To reach these conclusions, the group led by Daniel Mucida, a Brazilian researcher affiliated with Rockefeller University’s Laboratory of Mucosal Immunology, performed several experiments in mouse models. Thanks to a research internship abroad scholarship from FAPESP, their work was also assisted by Frederico Azevedo da Costa Pinto, a professor at the University of São Paulo’s School of Veterinary Medicine & Animal Science (FMVZ-USP) in Brazil.
A huge number of attenuated Salmonella bacteria were administered orally to mice. While not capable of causing infection, these microorganisms were recognized, and they triggered an immune response. Two hours later, the researchers analyzed the various layers of the murine intestine.
“After this period, we could already see a clear difference between the two groups of macrophages in terms of gene expression,” Gabanyi said. “Expression of anti-inflammatory genes was augmented in those cells farthest from the lumen and bacteria, which we call muscularis macrophages, whereas the pattern of gene expression changed little in those cells closest to the lumen, which are termed lamina propria macrophages and naturally perform pro-inflammatory regulation.”
According to Gabanyi, imaging had already shown the muscularis macrophages to be very close to intestinal tissue neurons, and the researchers suspected they might be communicating with each other.
“The change in gene expression took place so quickly that we suspected it might be happening via a neural pathway,” Gabanyi said.
Analysis of the muscularis macrophages showed that the proteins most expressed on the cells’ surfaces included beta-2 adrenergic receptors, which are precisely the receptors that respond to noradrenaline.
“We found that the muscularis macrophages expressed a much larger quantity of beta-2 adrenergic receptors than the other intestinal macrophages,” Gabanyi said.
When they repeated the experiment using mice that had been genetically modified so as not to express beta-2 adrenergic receptors, the researchers observed that there was no increase in the expression of anti-inflammatory genes, confirming that the pathway was not activated without the actions of noradrenaline on the macrophages.
The next step was to identify which neurons were releasing the neurotransmitters that modulated the immune response. According to Gabanyi, intrinsic neurons, i.e., neurons whose cell bodies are located inside the wall of the small intestine, are known not to release noradrenaline.
“Therefore, we thought it might be the sympathetic ganglion neurons, which are located near the spine,” she said. “They’re a large group of neurons that belong to the peripheral nervous system, and their axons enter the intestine.”
The suspicion was again confirmed by experiments with genetically modified mice. In this case, they expressed fluorescent proteins that enabled the activation of these neurons to be observed under a microscope.
The researchers are now trying to determine how sympathetic ganglion neurons perceive the presence of bacteria and other pathogens. To do this, they must detect the first stimulus to which the neurons respond by releasing noradrenaline.
“We’re also investigating the role of the muscularis macrophages during an inflammatory response in this tissue,” Gabanyi said. “We suspect one of these defense cells’ functions is precisely to protect neurons.”
She went on to note that there is probably a significant parallel between neuroimmune regulation mechanisms in mice and in humans.
“No one has studied this in humans, but there’s evidence of a parallel analogy of this kind. For example, some people develop irritable bowel syndrome after an episode of intestinal infection,” she said.
Gabanyi believes it will be possible in the future to identify ways of activating this macrophage-neuron communication pathway, which would be useful in treating patients with chronic intestinal diseases.