The researchers used 4 groups of IL-10-KO mice in the study. In the first round, they had younger mice enrolled at 4 weeks of age who ate their standard mouse chow the whole time, as well as mice who ate the mouse chow with raw broccoli sprouts mixed in.
In the second round, they had the same 2 diet groups, but mice were enrolled at 7 weeks of age. The researchers were particularly interested in understanding the development of IBDs in early life, which is why they studied the Crohn’s mouse models at the juvenile stage (4-6 weeks old) and at the adolescence stage (7-9 weeks old) with hopes to better understand how host-diet-microbial community interactions and disease severity differ by age.
The mice were fed for 7 days to acclimate to their respective diets before the researchers triggered symptoms, and the mice stayed on their diets for the following 2 weeks while the disease progressed. To trigger symptoms, new healthy mice that host more microbes were added to the cage. Since the IL-10-KO mice in the study can’t produce IL-10, their immune systems have trouble tolerating gut microbiota, and the new microbes in the cage triggered colitis and Crohn’s symptoms. For the next 15-16 days after infection, the researchers regularly weighed the mice and collected fecal samples to assess for signs of colitis development.
Analyzing Microbial Communities and Biogeography
At the end of the study, the researchers examined the gut tissues of the euthanized mice and microbial communities present throughout their intestines, as well as the presence of certain markers of inflammation and broccoli metabolites in the blood. The researchers wanted to know what types of microbes were living in particular parts of the gut. In other words, they wanted to understand how the broccoli sprout diet affected microbial biogeography in the Crohn’s models, since they cannot study this in humans.
DNA was extracted from the intestinal tissue samples collected from the mice and sent for sequencing to identify the bacteria present. Once the sequencing data was returned, the researchers used bioinformatics software and human ingenuity to study the gut microbial ecology of our mouse models.
“We found many exciting results from this study. First, we show that the mice that ate the broccoli sprout diet had a greater concentration of an anti-inflammatory metabolite called sulforaphane in their blood. Even though our mice were immunocompromised and had colitis, this increase in sulforaphane protected them from severe disease symptoms like weight loss, fecal blood and diarrhea,” said Lola Holcomb, lead author and a Ph.D. Candidate in the Graduate School of Biomedical Sciences and Engineering at the University of Maine. Lola is a member of a lab led by Suzanne Ishaq, Ph.D., a corresponding study author and assistant professor of animal and veterinary sciences at the University of Maine, School of Food and Agriculture, Orono, Maine.
Interestingly, the researchers found that the younger group of mice, the juveniles, responded better to the broccoli sprout diet than their adolescent counterparts did. The younger mice had milder disease symptoms and richer gut microbial communities. Furthermore, the younger mice showed stronger bacterial community similarity to each other (aka, stronger beta-diversity), and stronger adherence to location-specific community composition throughout different parts of the gut.