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Spotlight on Research

CHRONIC LUNG INFECTIONS INCREASE INFLAMMATION OVER TIME IN PEOPLE WITH CF

A team of researchers from across Canada and the US, including Cystic Fibrosis Canada-funded Dr. Dao Nguyen from McGill University, recently published their study on the impact of chronic lung infections on lung inflammation. Specifically, they looked at the role of Pseudomonas aeruginosa bacterial adaptations in promoting lung inflammation.

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Lung disease is the leading cause of symptoms and early death in people with cystic fibrosis (CF). Most people with CF have lungs that are chronically infected with the bacteria Pseudomonas aeruginosa for decades. The body’s immune system releases white blood cells in an attempt to kill these bacteria, but is unable to wipe them out. Instead, the white blood cells cause inflammation in the lungs, making things worse. 

Over time in the lungs of CF patients, Pseudomonas aeruginosa bacteria evolve and adapt to the CF lung environment. Perhaps unexpectedly, Pseudomonas bacteria become less toxic over time; however, these less toxic bacteria are associated with more advanced lung disease. The current research attempted to understand this paradox.

Scientists have observed that over time within each person with a chronic Pseudomonas infection, the bacteria can develop mutations in the lasR gene, which produces a protein that controls the production of many secreted bacterial enzymes. At least 1/3 of people with chronic infections of Pseudomonas carry bacteria with lasR mutations. Such mutant bacteria secrete fewer enzymes, yet they are associated with higher levels of inflammation and poorer lung function. 

The current study examined the effects of Pseudomonas LasR mutant bacteria on the lungs of people with CF. Researchers performed a variety of experiments to better understand the interactions between Pseudomonas bacteria and the human body, and the implications of these interactions for the health of people with CF.

How did they test the effects of Pseudomonas bacteria on the body?

In the lungs of people with CF, Pseudomonas bacteria grow as a slime layer within the mucus that covers the lung surface. In response to these bacteria, lung cells send inflammation signals that activate the immune system to send white blood cells to the area, in an attempt to get rid of the bacteria.

To replicate these conditions in the lab, scientists grew Pseudomonas bacteria in a synthetic gel that mimics CF lung secretions. Human lung cells were then exposed to different Pseudomonas bacteria recovered from CF patients at various stages of their lives, and scientists measured the inflammatory response of the lung cells. Specifically, they measured the amounts of the pro-inflammatory molecules interleukin-8 and interleukin-6, which are inflammatory signals that recruit white blood cells to the area to kill the bacteria. These white blood cells then cause damage in the body’s tissues. 

They also studied the response of human lung cells to Pseudomonas bacteria that had been genetically engineered to contain mutations commonly found in later stage chronic infections – lasR mutations, which interfere with the bacteria’s ability to produce certain secreted enzymes.

What did they find?

Scientists found that human lung cells produce higher levels of the pro-inflammatory molecules interleukin-8 and interleukin-6 in response to Pseudomonas bacteria recovered from late-stage chronic infections from CF patients. They found similar results when they tested Pseudomonas bacteria with lasR mutations. Later stage chronic lung infections with Pseudomonas bacteria in people with CF often contain LasR mutant bacteria, so this finding is consistent with the observation that later stage infections in CF patients are associated with higher levels of inflammation, and implicates lasR mutations as the possible mechanism behind this link.

How did they use living mice to support their work?

To test their hypothesis that Pseudomonas bacteria with lasR mutations are connected with greater levels of inflammation, scientists infected mice with Pseudomonas bacteria with and without lasR mutations. The mice infected with the LasR mutant bacteria showed higher levels of lung inflammation and damage compared to mice infected with Pseudomonas bacteria without the lasR mutations.

What else did this study find?

Researchers found that Pseudomonas bacteria with lasR mutations did not secrete enzymes that directly broke down interleukin-8 and interleukin-6. Furthermore, one particular bacterial enzyme, LasB, which is regulated by lasR, is required for the breakdown of interleukin-8 and interleukin-6.

In addition, researchers found that the Pseudomonas bacterial enzymes are better at breaking down interleukin-8 and interleukin-6 than enzymes secreted by the human body itself. When these pro-inflammatory signal molecules are not broken down, they will accumulate to induce greater inflammation in the lung. Therefore, when the LasB bacterial enzymes are not present due to lasR mutations, there will be higher levels of interleukin-8 and interleukin-6 present in the body, causing the body to recruit more white blood cells to the area, leading to more inflammation. 

Did they test their findings in people?

Researchers tested their findings in a group of 17 adults with CF who were chronically infected with Pseudomonas bacteria. They analyzed mucus samples from each person to determine the amount of LasR mutant bacteria, and measured the levels of interleukin-8 in their blood. As predicted, they found that people carrying LasR mutant bacteria had higher levels of interleukin-8, which is a marker of inflammation. Therefore, they were able to demonstrate an association between lung infections with LasR mutant Pseudomonas bacteria and inflammation in the lungs of people with cystic fibrosis. 

Overall, this study advances our understanding of the role of chronic Pseudomonas bacterial infections in increasing lung inflammation over time in people with CF. Identifying that Pseudomonas bacteria often develop lasR mutations over time, and that these mutations interfere with bacteria’s ability to break down inflammatory signals provides a reason for this link. When fewer bacterial enzymes are available to kill the body’s own internally generated inflammatory signals, interleukin-8 and interleukin-6, inflammation ensues, contributing to worsening lung disease.  

To access the article in Science Advances journal, please click here.

Nomenclature note: lasR refers to the gene, while LasR refers to the bacteria

Bio:

Dao Nguyen is an Assistant professor in the Department of Medicine at McGill University, a scientist at both the Research Institute of the McGill University Health Centre and Meakins Christie Laboratories, and a clinician-scientist in the Division of Respiratory Medicine at the McGill University Health Centre. Her research is focused on CF microbiology, particularly the molecular microbiology of Pseudomonas aeruginosa, antibiotic tolerance and biofilms, as well as host-pathogen interactions in CF pulmonary infections. She is the recipient of a CFC Scholarship, CIHR Clinician scientist award, and Burroughs Wellcome Fund Career Award for Medical Scientist. 

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