Biomarkers, Metagenomics, and Metabolomics in Grade C Molar Incisor Pattern Periodontitis: A Multidimensional Approach to Understand C/MIP Pathogenesis

Abstract

Objective: This project aimed to clarify grade C molar incisor pattern periodontitis (C/MIP) pathogenesis in young cases by comparing them to age-matched periodontitis-free controls in a multidimensional approach.

Material and Methods: (1) The literature on saliva, GCF, serum, and blood biomarkers in systemically healthy and non-smoking young C/MIP (≤25 years old) was systematically searched for promising biomarkers. Six databases were searched, including PubMed, Embase, Web of Science, Scopus, Virtual Health Library and ProQuest, for cross-sectional, case-control, and cohort studies with more than 10 subjects in the cases group. Meta-analysis was done on biomarkers that were assessed using the same detection methods and sample type in at least two papers. (2) After that, a case series on 31 C/MIP young cases (≤25 years old) was compiled to evaluate grade C periodontitis through clinical data and radiographs. Following that, a series of four analyses were conducted on the same group of patients, the 31 C/MIP young cases in which (3) human total immunoglobulin G (IgG) levels were investigated in saliva, gingival crevicular fluid (GCF), and serum using an Enzyme-Linked Immunosorbent Assay (ELISA); (4) the saliva and serum metabolites profiles of C/MIP and controls were analysed using a combination of nuclear magnetic resonance (NMR) and mass spectrometry (MS) respectively; (5) DNA was extracted from plaque and saliva of the same group using Qigen PowerSoil kit. (6) metabolomics and metagenomics data were then integrated using R software and Cytoscape to identify significant microbe–metabolite interactions.

Results: (1) Eighty-seven biomarkers were assessed, with the majority being higher in cases than in controls. Only the meta-analysis of total serum IgG with low heterogeneity value revealed a statistically significant increase in its levels in C/MIPs compared to controls (standardised mean difference: 1.08; 95% CI: 0.76, 1.40). (2) A considerable portion of MIPs were female, and almost half of the MIPs were Afro/Caribbean, with more than half reported having a family history of periodontitis with tooth loss in either siblings, parents, and/or grandparents at a young age. Most affected incisors and molars had grade 1 mobility and class I furcation involvement, with most cases having vertical bone loss in at least one molar. (3) After adjusting for covariates, cases had higher Immunoglobulin G levels in saliva (p=0.005) and gingival crevicular fluid (p<0.001) than controls; however, serum IgG levels did not reach the significance threshold (p=0.137) for differences between test and controls. Among other factors contributing to IgG levels, males had higher serum IgG than females (p=0.018), and serum IgG levels increased with age (p= 0.033). (4) Of the 42 salivary metabolites detected by NMR, dimethylamine, proline, and glycine were significantly lower in C/MIP than controls after adjusting for covariates. Conversely, the MS method detected 349 metabolites in serum, among which Bis(hydroxymethyl) propionic acid, 1-[(9Z)-octadecenoyl]-2-tetradecanoyl-sn-glycero-3-phosphocholine, decanoylcarnitine, 4-Picoline, tranexamic acid, 2-Hydroxy-5-vinylphenyl hydrogen sulfate, and L-Serine were lower in C/MIP compared to controls, whereas methyl indole-3-acetate, N-(3-acetamidopropyl) pyrrolidin-2-one, 4-Ethyl-2-hydroxy-6-methoxyphenyl hydrogen sulfate, and sulfosalicylic acid were higher in C/MIP than controls (5) C/MIP had a more diverse and distinct bacterial profile than controls in plaque and saliva. A novel unknown group of bacteria, labelled GGB, showed significant associations with C/MIP. The following bacteria were consistently identified in all analyses: C/MIP-associated taxa in plaque: Desulfobulbus oralis, Anaerolineaceae bacterium oral taxon 439, GGB4333 SGB5935, Treponema SGB3604, Campylobacter rectus, Treponema denticola, Eubacterium nodatum, Parvimonas micra, Eubacterium brachy, Slackia exigua, and Fretibacterium fastidiosum. Oral-health-associated taxa: Rothia mucilaginosa in saliva, and GGB10485 SGB49305 and Leptotrichia hongkongensis in plaque. Desulfobulbus oralis and GGB10485 SGB49305 showed considerable power as biomarkers in plaque. (6) Notable clusters of key periodontal pathogens were noticed including Porphyromonas gingivalis, Tannerella forsythia, Filifactor alocis and Abiotrophia defectiva and among metabolites, taurine, glutamine, N-Phenylacetylglutamine, and tetraacetylethylenediamine. Conclusion: This study identified for the first time the critical microbial and metabolic signatures associated with the pathogenesis of C/MIP in young cases using a combination of omics techniques. Integration of metagenomics and metabolomics revealed new insights into host-microbe interaction and metabolic pathways driving the disease, thus opening up new avenues for better diagnostic and therapeutic approaches.