Validation studies for germ-free Smad3-/- mice as a bio-assay to test the causative role of fecal microbiomes in IBD.

While the association between microbiomes and inflammatory bowel disease (IBD) is well known, establishing causal relationships between the two is difficult in humans. Germ-free (GF) mice genetically susceptible to IBD can address this question. Smad3-/- mice with defective transforming growth factor ß signaling are a model of IBD and colon cancer. They develop IBD upon colonization with Helicobacter under specific pathogen-free conditions, suggesting a role of the microbiome in IBD in this model. Thus, we rederived Smad3-/- mice GF to determine the potential of using these mice for testing the causative role of microbiomes in IBD. We found that fecal microbiomes from mice with IBD cause more severe gut inflammation in GF Smad3-/- and wild type mice compared to microbiomes from healthy mice and that Helicobacter induces gut inflammation within the context of other microbiomes but not by itself. Unexpectedly, GF Smad3+/+ and Smad3+/- mice given IBD microbiomes develop IBD despite their lack of disease in SPF conditions upon Helicobacter infection. This was not unique to the background strain of our Smad3 model (129); both wild type C57BL/6 and 129 strains developed IBD upon fecal transfer. However, wild type Swiss Webster stock was not susceptible, indicating that the genetic background of recipient mice influences the severity of IBD following fecal transfer. Our data suggest that the microbiome is an independent risk factor contributing to IBD development, and careful characterization of new GF models is needed to understand potential sources of confounding factors influencing microbiome studies in these mice.