Interaction Between Immune Cells and Bacteria Associated With Periodontitis

This study evaluates the interaction between host immune cells and bacteria associated with periodontitis. It comprises biological material from donors with and without periodontal disease. Specifically, we collect a spit and blood sample to conduct in vitro stimulations and measurements of selected parameters related to periodontitis to clarify obscure areas in the immunologic pathogenesis of this disease.

Study Overview

• Status: Completed
• Conditions: Inflammation; Rheumatoid Arthritis; Periodontitis; Periodontal Diseases; Dysbiosis; Aggressive Periodontitis; Chronic Periodontitis; Periodontal Pocket; Generalized Chronic Periodontitis; Microbial Disease; Inflammation Gum; Immunologic Disease; Generalized Aggressive Periodontitis
• Intervention / Treatment: Other: In vitro stimulation of blood with periodontitis-associated- and control bacteria; Diagnostic test: Anti-CCP- and anti-P.g.-antibodies titers; Genetic: Analysis of selected single nucleotide polymorphisms (SNPs); Diagnostic test: periodontitis-associated bacteria presence

Detailed Description

Periodontitis is a prevalent, multifactorial inflammatory disease characterized by the interaction between microorganisms organized in biofilms on tooth surfaces and host immune cells, leading to an inflammatory destruction of the tooth-supporting tissues and - if left untreated - eventually tooth loss. Periodontitis affects up to 50% of the population in the United States of America, and is classified in an aggressive and a chronic form depending on genetic factors, age of onset, speed and severity of attachment loss.

The onset of periodontitis is caused by an immunologic imbalance between host immune cells and residing microorganisms in subgingival pockets. The host immune cells are capable of enhancing both a protective and a destructive inflammatory response towards the microorganisms through the release of inflammatory mediators e.i. proinflammatory and antiinflammatory cytokines.

The role of antibodies in periodontitis is also unclear. Some studies show an excessive antibody level against bacteria associated with periodontitis e.g. Porphyromonas gingivalis (P.g.).

In general, this study contributes to a profound understanding of the host immune cells role in the onset and pathogenesis of periodontitis by comparing healthy versus diseased donors immunologic responses toward pathogene and apathogene microorganisms and their genetic background.

• Study Type: Observational
• Enrollment (Actual): 90

Contacts and Locations

Study Locations

• Denmark
  • Copenhagen, Denmark, 2200
    • Institute for Inflammation Research, Center for Rheumatology and Spine Diseases, Rigshospitalet, Copenhagen University Hospital.
  • Copenhagen, Denmark, 2200
    • Section for Periodontology, Microbiology and Community Dentistry, Department of Odontology, Faculty of Health and Medical Sciences, University of Copenhagen

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 19 years to 60 years (Adult)
• Accepts Healthy Volunteers: Yes
• Genders Eligible for Study: All

• Sampling Method: Non-Probability Sample

Study Population

This study includes 170 participants, which are legally competent e.i. above 18 years of age and general decision competent. The participants are divided into three groups: 1) Participants with chronic periodontitis (n=35), 2) participants with aggressive periodontitis (n=35) and 3) healthy participants without periodontal disease (n=100). The participants are recruited from the Department of Odontology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark and assigned to a group in accordance with the periodontal disease classification system in 1999 American Academy of Periodontology.

Description

Inclusion criteria for chronic periodontitis donors:

• 50-60 years of age.
• Interproximal attachment loss at minimum 3 teeth besides molars and incisors.
• Clinical attachment loss at minimum 10 sites identified by bleeding and pus upon probing.
• Visible radiographic bone loss.
• Medically healthy donors.

Inclusion criteria for aggressive periodontitis donors:

• 19-40 years of age.
• Interproximal attachment loss at minimum 3 teeth besides molars and incisors.
• Clinical attachment loss at minimum 10 sites identified by bleeding and pus upon probing.
• Visible radiographic bone loss.
• Medically healthy donors.

Inclusion criteria for healthy donors: (age: 19-40 years; 50-60 years)

• No sign of inflammatory conditions or other general systemic diseases.
• Medically healthy donors.

Exclusion criteria for all groups:

• Pregnant and breastfeeding.
• Antibiotic treatment within 6 months.
• Suffer from periodontal manifestations caused by systemic diseases e.i. genetic diseases, haematologic anomalies or syndromes.

Study Plan

How is the study designed?

Number of groups / cohorts

3

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Chronic periodontitis donors; Donors are medically healthy.; Slow to moderate attachment loss and bone destruction.; Good correlation between etiological factors and serverity of attachment loss.	Other: In vitro stimulation of blood with periodontitis-associated- and control bacteria; Peripheral mononuclear blood cells are stimulated with periodontitis-associated- and control bacteria to measure the amount of positive cytokine-producing cells.; Diagnostic test: Anti-CCP- and anti-P.g.-antibodies titers; Anitbody titers will be measured in saliva and serum samples.; Genetic: Analysis of selected single nucleotide polymorphisms (SNPs); DNA obtained from saliva samples will be used to determine the genotype of the participants for selected SNPs.; Diagnostic test: periodontitis-associated bacteria presence; Determination of the presence of periodontitis-associated bacteria e.i. Porphyromonas gingivalis in saliva and blood samples.
Aggressive periodontitis donors; Donors are medically healthy.; Rapid attachment loss and bone destruction.; Familial aggregation.; No correlation between etiological factors and serverity of attachment loss.	Other: In vitro stimulation of blood with periodontitis-associated- and control bacteria; Peripheral mononuclear blood cells are stimulated with periodontitis-associated- and control bacteria to measure the amount of positive cytokine-producing cells.; Diagnostic test: Anti-CCP- and anti-P.g.-antibodies titers; Anitbody titers will be measured in saliva and serum samples.; Genetic: Analysis of selected single nucleotide polymorphisms (SNPs); DNA obtained from saliva samples will be used to determine the genotype of the participants for selected SNPs.; Diagnostic test: periodontitis-associated bacteria presence; Determination of the presence of periodontitis-associated bacteria e.i. Porphyromonas gingivalis in saliva and blood samples.
Control donors; Donors are medically healthy.; No sign of inflammatory conditions.	Other: In vitro stimulation of blood with periodontitis-associated- and control bacteria; Peripheral mononuclear blood cells are stimulated with periodontitis-associated- and control bacteria to measure the amount of positive cytokine-producing cells.; Diagnostic test: Anti-CCP- and anti-P.g.-antibodies titers; Anitbody titers will be measured in saliva and serum samples.; Genetic: Analysis of selected single nucleotide polymorphisms (SNPs); DNA obtained from saliva samples will be used to determine the genotype of the participants for selected SNPs.; Diagnostic test: periodontitis-associated bacteria presence; Determination of the presence of periodontitis-associated bacteria e.i. Porphyromonas gingivalis in saliva and blood samples.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
periodontitis-associated- and control bacterial stimulation of host immune cells.	Identification and determination of the amount of cytokine-producing immune cells when stimulated with bacteria associated with periodontitis including pro- and antiinflammatory cytokines. Genuses: Porphyromonas, Prevotella, Eikenella, Aggregatibacter, Actinomyces, Lactobacillus, Bifidobacterium, Rothia.	Aug. 2020
Anti-cyclic citrullinated peptides (anti-CCP) antibodies titers.	Determination of the prevalence of anti-CCP-positive periodontitis patients through measure of anti-CCP antibody titers in serum samples and correlation with the level of antibodies to P. gingivalis, and to the abundancy of the bacterium in saliva.	Aug. 2020
P. gingivalis presence and related antibodies.	Determination of P. gingivalis presence in saliva and serum samples through RT-qPCR and determination of the level of antibodies towards the bacterium using in-house Luminex-based technology.	Aug. 2020
Single nucleotide polymorphism (SNP) analysis.	Investigation of the potential association between periodontitis and selected polymorphisms in the PADI genes using multiplex bead-based SNP assays with the Luminex technology.	Aug. 2020

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Cytokine profile in saliva.	Determination of the cytokine profile in saliva samples from subject with periodontitis and healthy controls with Luminex based technology.	Aug. 2020
Presence of other periodontal bacteria.	Determination of the presence of other periodontal bacteria through RT-qPCR assays.	Aug. 2020

Collaborators and Investigators

• Sponsor: University of Copenhagen
• Collaborators: Copenhagen University Hospital, Denmark
• Investigators: Study Director: Palle Holmstrup, DDS, PhD, Dr Odont, University of Copenhagen; Study Director: Claus Henrik Nielsen, PhD, MSc, MD, Institute for Inflammation Research, Center for Rheumatology and Spine Diseases, Rigshospitalet, Copenhagen University Hospital, Copenhagen, Denmark.

Publications and helpful links

General Publications

• Kinane DF, Preshaw PM, Loos BG; Working Group 2 of Seventh European Workshop on Periodontology. Host-response: understanding the cellular and molecular mechanisms of host-microbial interactions--consensus of the Seventh European Workshop on Periodontology. J Clin Periodontol. 2011 Mar;38 Suppl 11:44-8. doi: 10.1111/j.1600-051X.2010.01682.x.
• Eke PI, Dye BA, Wei L, Slade GD, Thornton-Evans GO, Borgnakke WS, Taylor GW, Page RC, Beck JD, Genco RJ. Update on Prevalence of Periodontitis in Adults in the United States: NHANES 2009 to 2012. J Periodontol. 2015 May;86(5):611-22. doi: 10.1902/jop.2015.140520. Epub 2015 Feb 17.
• Bartold PM, Van Dyke TE. Periodontitis: a host-mediated disruption of microbial homeostasis. Unlearning learned concepts. Periodontol 2000. 2013 Jun;62(1):203-17. doi: 10.1111/j.1600-0757.2012.00450.x.
• Neely AL, Holford TR, Loe H, Anerud A, Boysen H. The natural history of periodontal disease in man. Risk factors for progression of attachment loss in individuals receiving no oral health care. J Periodontol. 2001 Aug;72(8):1006-15. doi: 10.1902/jop.2001.72.8.1006.
• Armitage GC. Development of a classification system for periodontal diseases and conditions. Northwest Dent. 2000 Nov-Dec;79(6):31-5.
• Haffajee AD, Socransky SS, Patel MR, Song X. Microbial complexes in supragingival plaque. Oral Microbiol Immunol. 2008 Jun;23(3):196-205. doi: 10.1111/j.1399-302X.2007.00411.x.
• Damgaard C, Holmstrup P, Van Dyke TE, Nielsen CH. The complement system and its role in the pathogenesis of periodontitis: current concepts. J Periodontal Res. 2015 Jun;50(3):283-93. doi: 10.1111/jre.12209. Epub 2014 Jul 5.
• Liu YC, Lerner UH, Teng YT. Cytokine responses against periodontal infection: protective and destructive roles. Periodontol 2000. 2010 Feb;52(1):163-206. doi: 10.1111/j.1600-0757.2009.00321.x. No abstract available.
• Pussinen PJ, Jousilahti P, Alfthan G, Palosuo T, Asikainen S, Salomaa V. Antibodies to periodontal pathogens are associated with coronary heart disease. Arterioscler Thromb Vasc Biol. 2003 Jul 1;23(7):1250-4. doi: 10.1161/01.ATV.0000072969.71452.87. Epub 2003 Apr 24.
• Pussinen PJ, Nyyssonen K, Alfthan G, Salonen R, Laukkanen JA, Salonen JT. Serum antibody levels to Actinobacillus actinomycetemcomitans predict the risk for coronary heart disease. Arterioscler Thromb Vasc Biol. 2005 Apr;25(4):833-8. doi: 10.1161/01.ATV.0000157982.69663.59. Epub 2005 Feb 3.
• Berglundh T, Donati M. Aspects of adaptive host response in periodontitis. J Clin Periodontol. 2005;32 Suppl 6:87-107. doi: 10.1111/j.1600-051X.2005.00820.x.
• Fillatreau S. Cytokine-producing B cells as regulators of pathogenic and protective immune responses. Ann Rheum Dis. 2013 Apr;72 Suppl 2:ii80-4. doi: 10.1136/annrheumdis-2012-202253. Epub 2012 Dec 19.
• Belstrom D, Paster BJ, Fiehn NE, Bardow A, Holmstrup P. Salivary bacterial fingerprints of established oral disease revealed by the Human Oral Microbe Identification using Next Generation Sequencing (HOMINGS) technique. J Oral Microbiol. 2016 Jan 14;8:30170. doi: 10.3402/jom.v8.30170. eCollection 2016.
• Belstrom D, Holmstrup P, Bardow A, Kokaras A, Fiehn NE, Paster BJ. Comparative analysis of bacterial profiles in unstimulated and stimulated saliva samples. J Oral Microbiol. 2016 Mar 16;8:30112. doi: 10.3402/jom.v8.30112. eCollection 2016.

Study record dates

Study Major Dates

• Study Start (Actual): February 1, 2017
• Primary Completion (Actual): October 12, 2018
• Study Completion (Actual): March 23, 2020

Study Registration Dates

• First Submitted: July 19, 2017
• First Submitted That Met QC Criteria: July 19, 2017
• First Posted (Actual): July 21, 2017

Study Record Updates

• Last Update Posted (Actual): March 24, 2020
• Last Update Submitted That Met QC Criteria: March 23, 2020
• Last Verified: October 1, 2018

More Information

Terms related to this study

• Keywords: Inflammation; Periodontitis; Cytokines; Bacteria; Saliva; Aggressive periodontitis; Periodontal disease; Porphyromonas gingivalis; Dysbiosis; Anti-CCP; Regulation; Single nucleotide polymorphisms; Peptidyl arginine deiminases
• Additional Relevant MeSH Terms: Behavioral Symptoms; Pathologic Processes; Stomatognathic Diseases; Mouth Diseases; Aggression; Periodontitis; Inflammation; Periodontal Diseases; Aggressive Periodontitis; Chronic Periodontitis; Immune System Diseases; Dysbiosis; Periodontal Pocket; Physiological Effects of Drugs; Immunologic Factors; Antibodies
• Other Study ID Numbers: H-16024734

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: NO
• IPD Plan Description: IPD will not be available to other researchers in accordance with our approved Human Subjects Protection application in Denmark.

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No