Periodontitis is an inflammatory response to the commensal oral bacterial flora and represents one of the most prevalent infections in humans. Approximately 90% of the population exhibits some form of early disease (gingivitis), with 48% of the US adult population having periodontitis and the severe form of the disease affecting 15-20%. Periodontal disease is a polygenic condition that is associated with an exaggerated inflammatory response to the commensal biofilm that drives chronic biofilm dysbiosis at the biofilm-gingival interface. An elevated level of gingival crevicular fluid (GCF) interleukin 1beta (IL-1) has previously been established as a robust biomarker for a hyper- inflammatory phenotype and for mediating severe inflammation, bone loss and periodontal disease progression. We have recently completed a genome-wide association study (GWAS) of 4910 Caucasians with known levels of GCF-IL-1 to identify loci that are associated with high levels of GCF-IL-1. We have identified two novel quantitative trait loci with missense polymorphisms within the coding region of the anti-inflammatory gene IL37 that are both statistically significanty associated with high GCF-IL-1 levels (defined as upper quartile or as a continuous variable), p=6.8 X10-21. These missense polymorphisms at two IL-37 loci, are present in 30% and 8.5% of the population. Not only are both loci strongly associated with high local IL-1 levels, but the are also associated with more severe periodontal disease. Under normal conditions IL-37 activates Smad3 and attenuates the innate immune response to TLR agonists to include suppression of IL-1, IL-1?, TNF?, IL-6, MIP-2 (CXCL2) and GM-CSF. Thus, our central hypothesis is that these IL37 SNP variants cause a functional defect in IL-37 (either via altered mRNA splicing, protein synthesis, activation and/or bioactivity) that results in an excessive pro-inflammatory innate immune response to the commensal organisms of the oral cavity, thereby inducing greater clinical inflammation, bone loss and more severe clinical periodontal disease. Since mice do not express a murine homologue to hIL-37 we propose two aims in murine models to understand whether the two IL37 variants demonstrate impaired IL-37 function I.e. SMAD3 activation and suppression of the innate immune response. First we will transfect murine monocytic RAW cells with the human wild-type [IL37wt] or the two IL37 variants, IL37v1 or IL37v2, to assess the effects of these mutations on IL-37 function, stimulating cells with a range of TLR agonists. These experiments transfecting with IL37v1 or IL37v2 will be confirmed in the human monocytic line THP-1. We expect that the variants will have a hyper-inflammatory trait compared to hIL-37wt, measuring the levels of the mediators listed above, as the read-out. We will also create IL37 variant transgenic mice to measure the response to challenge with P gingivalis using a subcutaneous chamber model and an oral bone loss model. Finally, we will establish a new IL-37 genotyped cohort of subjects to study and confirm the role of these two IL37 variants on IL-37 function and response to TLR challenge using isolated human PBMC and hTERT immortalized cells. It is expected that understanding the role of this novel variant as a potential cause of the underlying hyper-inflammatory trait associated with severe periodontal disease, will ultimately improve diagnosis, prevention and treatment.

Public Health Relevance

High levels of IL-1 mediated gingival inflammation has been referred to as a hyper-inflammatory trait that is associated with severe periodontal disease, in a manner which is similar to high cholesterol serving as a risk indicator for cardiovascular disease. Using a Gene-Wide Association (GWA) approach with 4910 subjects with known levels of gingival crevicular fluid IL-1, we identified a novel gene locus with missense coding regions in a recently characterized cytokine, IL37. This cytokine is a fundamental suppressor of innate immunity and under normal conditions dampens the inflammatory response. Thus, we propose to test in animal models and in humans whether these highly significant genetic variants underlie the hyper-inflammatory trait seen among periodontitis patients.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE023836-04
Application #
9282271
Study Section
Oral, Dental and Craniofacial Sciences Study Section (ODCS)
Program Officer
Chander, Preethi
Project Start
2014-08-01
Project End
2019-05-31
Budget Start
2017-06-01
Budget End
2018-05-31
Support Year
4
Fiscal Year
2017
Total Cost
$377,249
Indirect Cost
$127,249
Name
University of North Carolina Chapel Hill
Department
Dentistry
Type
Schools of Dentistry
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Beck, James D; Moss, Kevin L; Morelli, Thiago et al. (2018) Periodontal profile class is associated with prevalent diabetes, coronary heart disease, stroke, and systemic markers of C-reactive protein and interleukin-6. J Periodontol 89:157-165
Beck, James D; Moss, Kevin L; Morelli, Thiago et al. (2018) In search of appropriate measures of periodontal status: The Periodontal Profile Phenotype (P3 ) system. J Periodontol 89:166-175
Sanders, A E; Sofer, T; Wong, Q et al. (2017) Chronic Periodontitis Genome-wide Association Study in the Hispanic Community Health Study / Study of Latinos. J Dent Res 96:64-72
Marchesan, Julie T; Jiao, Yizu; Moss, Kevin et al. (2017) Common Polymorphisms in IFI16 and AIM2 Genes Are Associated With Periodontal Disease. J Periodontol 88:663-672
Morelli, Thiago; Moss, Kevin L; Beck, James et al. (2017) Derivation and Validation of the Periodontal and Tooth Profile Classification System for Patient Stratification. J Periodontol 88:153-165
Zhang, Shaoping; Divaris, Kimon; Moss, Kevin et al. (2016) The Novel ASIC2 Locus is Associated with Severe Gingival Inflammation. JDR Clin Trans Res 1:163-170
Shungin, Dmitry; Cornelis, Marilyn C; Divaris, Kimon et al. (2015) Using genetics to test the causal relationship of total adiposity and periodontitis: Mendelian randomization analyses in the Gene-Lifestyle Interactions and Dental Endpoints (GLIDE) Consortium. Int J Epidemiol 44:638-50
Wu, X; Offenbacher, S; L?pez, N J et al. (2015) Association of interleukin-1 gene variations with moderate to severe chronic periodontitis in multiple ethnicities. J Periodontal Res 50:52-61
Abe, Toshiharu; AlSarhan, Mohammed; Benakanakere, Manjunatha R et al. (2015) The B Cell-Stimulatory Cytokines BLyS and APRIL Are Elevated in Human Periodontitis and Are Required for B Cell-Dependent Bone Loss in Experimental Murine Periodontitis. J Immunol 195:1427-35
Yu, N; Barros, S P; Zhang, S et al. (2015) Insulin Response Genes in Different Stages of Periodontal Disease. J Dent Res 94:194S-200S