The Interconnection between Mouth and Gut: Understanding the Microbial and Host Factors Influencing Susceptibility to Periodontitis and Colitis

Oral Health is fundamentally tied to general health. However, oral health and general health are perceived as separate entities. Pathologies that arise in the mouth can have a profound impact on systemic health.

Periodontitis is a microbiome mediated oral inflammatory disease that affects nearly 10% of the global population and 50% of Americans aged ?30-year.

Periodontitis and associated bacteria have been reported to contribute to the pathogenesis of several systemic diseases, like inflammatory bowel disease (IBD), diabetes, endocarditis, rheumatoid arthritis, Alzheimer?s Disease, preterm birth, and osteoporosis.

IBD is a chronic inflammatory disease of the gastrointestinal tract, and primarily includes Crohn's disease and ulcerative colitis. IBD has a high prevalence in Western societies and >3 million people are affected by it in the US. Both periodontitis and IBD impose enormous social, psychological, and economic burden on people and health services.

To date, the etiology of IBD remains largely unknown, and the potential causal role of periodontitis and associated bacteria in IBD remains poorly investigated.

The long-term goal of the proposed research is it to elucidate the molecular mechanisms governing periodontitis process and the potential causal role of periodontitis and associated bacteria in IBD.

This project leverages complementary in vitro and in vivo mechanistic approaches and next generation sequencing to pursue the following aims- a) determine the transcriptional and epigenetic landscapes that shape oral microbiota and periodontal disease process; and b) identify oral and gut bacteria involved in eliciting IBD and the associated molecular mechanisms.

Our project defines complex interrelationship between periodontitis and IBD.

Results from our study will not only guide in effectively improving oral health but will also help in minimizing the risk of IBD in susceptible individuals. Thus, our study will have a far-reaching impact. Specific Aims: Our grant (1R01DK131277-01) was reviewed favorably by the ODCS study section.

While the reviewers found the application very promising from an Early-Stage Investigator, they indicated few minor/ moderate weaknesses.

Hence, we seek interim research support to gather more preliminary data to revise and improve the current application, which includes performing shotgun metagenomics sequencing, flow cytometry, and single-cell RNA-seq and ATAC-seq.

Additionally, as recommended by the reviewers, we plan to generate specific-pathogen-free Irf8 cKO mice and germ-free Irf8 gKO mice and conduct preliminary studies. Pathologies that arise in the mouth can have a profound impact on systemic health.

Periodontitis, a common oral inflammatory disease, is associated with the pathogenesis of Inflammatory Bowel Disease (IBD). Increased prevalence of periodontitis and periodontal pathogens has been noted in IBD patients.

Certain similarities exist between the two diseases in terms of mediators of immune responses and bacterial colonization, however, the interrelationship between the two diseases is ill defined.

To date, the etiology of IBD remains largely unknown, and the potential causal role of periodontitis and associated bacteria in IBD remains poorly studied.

Genome- wide association studies have identified SNPs in the Interferon Regulatory Factor 8 (IRF8) gene as significant risk factors for IBD. We have shown that IRF8 is critical for periodontal homeostasis and osteoclast regulation.

However, to date, no studies have investigated the direct role of IRF8 in periodontitis mediated IBD, which represents a significant knowledge gap that is addressed in this proposal.

The long-term goal of the proposed research is it to elucidate the molecular mechanisms governing periodontitis processes and the potential causal role of periodontitis and associated bacteria in IBD.

Based on our preliminary data, we hypothesize that Irf8 plays a key role in governing periodontal and intestinal homeostasis, and Irf8 deficiency along with dysbiotic oral and gut microbiota promote colitis in Irf8 KO mice.

To achieve our goal, we will utilize Irf8 deficient mice (Irf8 global KO and conditional KO), which serve as a unique pre-clinical model to study periodontitis and IBD pathogenesis.

We will leverage complementary in vitro and in vivo mechanistic approaches and single cell sequencing to define a) how Irf8 deficiency shapes the oral microbiota and transcriptional and epigenetic landscapes that regulate periodontal disease processes; and b) how Irf8 deficiency along with dysbiotic gut microbiota creates an opportunistic environment for oral bacteria to colonize the gut and elicit intestinal inflammation.

This will enable us to identify how Irf8 shapes a) genes and signaling mechanisms that regulate immunoinflammatory responses in periodontitis and IBD; and b) the potential cellular and humoral responses by which oral bacteria elicit IBD.

Aim 1: To determine the molecular mechanisms shaping oral microbiota & periodontitis in Irf8 KO mice Rationale: Our data shows that Irf8 gKO mice exhibit reduced periodontal inflammation but increased oral dysbiosis, which we hypothesize is due to Irf8 impact on immune cells and transcription & epigenetic signatures.

We will determine the transcriptional and epigenetic landscapes that shape oral microbiota and periodontal disease process in Irf8 gKO mice by performing single-cell RNA-seq and ATAC-seq.

Rationale: Our data shows that periodontitis induced Irf8 gKO mice exhibit colitis and have higher abundance of oral bacteria in their gut.

We hypothesize that the dysbiotic oral bacteria along with dysbiotic gut bacteria in Irf8 gKO mice exhibit increased colitogenic potential.

We will identify specific oral and gut bacteria involved in eliciting IBD in Irf8 KO mice by performing shotgun metagenomics sequencing and inoculate the identified bacteria in Irf8 cKO mice and Irf8 gKO germ-free mice, followed by flow cytometry analysis.

Our study elucidates the distinct inflammatory responses occurring in periodontal and intestinal tissues, and how co-existence of Irf8 deficiency along with dysbiotic oral and gut microbiota creates a susceptible environment for developing IBD.

Our data indicates that transmigration of oral bacteria to the gut via enteral route (dissemination via swallowing) and subsequent alteration of gut microbiome plays a major role in IBD pathogenesis.

The new knowledge derived from this project has the potential to significantly advance the fields of periodontology & IBD.

It uncovers immunoinflammatory responses regulated by Irf8 & pathways up/down-stream of Irf8 that could be potentially targeted to develop intervention and therapeutic approaches for periodontitis and IBD.

Elucidation of the causal role of oral bacteria in IBD in this pre-clinical model will lay the platform for future clinical/translational research.

IBD patients at UMB and IRF8 mutation patients at the NIH, as well as Irf8 point mutation mice will be examined in the future for in-depth understanding of IRF8 role in periodontitis and IBD. Collectively, our study will contribute to improving oral health while concurrently minimizing the risk of IBD.

This represents a significant step towards providing integrated oral and systemic health care, which is in alignment with NIH?s vision.