Nurcan Buduneli

Metatranscriptomics of the Human Oral Microbiome during Health and Disease

ABSTRACT The human microbiome plays important roles in health, but when disrupted, these same indigenous microbes can cause disease. The composition of the microbiome changes during the transition from health to disease; however, these changes are often not conserved among patients. Since microbiome-associated diseases like periodontitis cause similar patient symptoms despite interpatient variability in microbial community composition, we hypothesized that human-associated microbial communities undergo conserved changes in metabolism during disease. Here, we used patient-matched healthy and diseased samples to compare gene expression of 160,000 genes in healthy and diseased periodontal communities. We show that health- and disease-associated communities exhibit defined differences in metabolism that are conserved between patients. In contrast, the metabolic gene expression of individual species was highly variable between patients. These results demonstrate that despite high interpatient variability in microbial composition, disease-associated communities display conserved metabolic profiles that are generally accomplished by a patient-specific cohort of microbes. IMPORTANCE The human microbiome project has shown that shifts in our microbiota are associated with many diseases, including obesity, Crohn’s disease, diabetes, and periodontitis. While changes in microbial populations are apparent during these diseases, the species associated with each disease can vary from patient to patient. Taking into account this interpatient variability, we hypothesized that specific microbiota-associated diseases would be marked by conserved microbial community behaviors. Here, we use gene expression analyses of patient-matched healthy and diseased human periodontal plaque to show that microbial communities have highly conserved metabolic gene expression profiles, whereas individual species within the community do not. Furthermore, disease-associated communities exhibit conserved changes in metabolic and virulence gene expression. The human microbiome project has shown that shifts in our microbiota are associated with many diseases, including obesity, Crohn’s disease, diabetes, and periodontitis. While changes in microbial populations are apparent during these diseases, the species associated with each disease can vary from patient to patient. Taking into account this interpatient variability, we hypothesized that specific microbiota-associated diseases would be marked by conserved microbial community behaviors. Here, we use gene expression analyses of patient-matched healthy and diseased human periodontal plaque to show that microbial communities have highly conserved metabolic gene expression profiles, whereas individual species within the community do not. Furthermore, disease-associated communities exhibit conserved changes in metabolic and virulence gene expression.

Adipokines and Inflammatory Mediators After Initial Periodontal Treatment in Patients With Type 2 Diabetes and Chronic Periodontitis

BACKGROUND This study was performed to evaluate the effects of initial periodontal treatment on clinical periodontal measurements, glycemic control, and systemic inflammatory mediator levels in patients with type 2 diabetes and chronic periodontitis. METHODS Thirteen well-controlled (glycated hemoglobin [HbA1c] <7%) and 12 poorly controlled (HbA1c > or =7%) patients with type 2 diabetes and chronic periodontitis and 15 systemically healthy patients with chronic periodontitis were enrolled. Blood samples were collected at baseline from all patients and 1 and 3 months after the initial periodontal treatment from patients with diabetes. Serum levels of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, C-reactive protein (CRP), soluble intercellular adhesion molecule-1, adiponectin, and leptin were analyzed by enzyme-linked immunosorbent assay. RESULTS The study groups showed similar improvements in clinical periodontal variables at all evaluation times (P <0.05). HbA1c levels in the poorly controlled group with diabetes decreased significantly at 3 months after completion of the initial periodontal treatment (P <0.05), whereas no significant changes were evident in the well-controlled group. There were insignificant decreases in TNF-alpha and CRP levels (P >0.05). IL-6 levels decreased in well-controlled patients with diabetes and in the systemically healthy group (P <0.05). Adiponectin levels increased in the systemically healthy group (P <0.05). Leptin levels increased at 1 month in well-controlled patients with diabetes (P <0.05). CONCLUSIONS Within the limits of this study, patients with type 2 diabetes and chronic periodontitis exhibited similar clinical periodontal improvements as their systemically healthy counterparts. Initial periodontal treatment appeared to improve glycemic control in poorly controlled patients with diabetes. Decreases in levels of IL-6, TNF-alpha, CRP, and leptin and an increase in adiponectin levels after periodontal therapy may be a function of glycemic control in patients with type 2 diabetes.

Saliva concentrations of RANKL and osteoprotegerin in smoker versus non-smoker chronic periodontitis patients

OBJECTIVES To compare the salivary receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) concentrations in smokers versus non-smokers with chronic periodontitis. MATERIAL AND METHODS Whole saliva samples were obtained from 67 untreated chronic periodontitis patients, of whom 34 were smokers, and from 44 maintenance patients, of whom 22 were smokers. Full-mouth clinical periodontal measurements were recorded. Saliva cotinine, sRANKL and OPG concentrations were determined by ELISA. Statistical analysis was performed using the Mann-Whitney U test, Bonferronis correction for multiple comparisons and Spearmans correlations. RESULTS Untreated smokers exhibited significantly higher values of clinical periodontal recordings than untreated non-smokers (all p<0.05). Salivary cotinine level correlated with clinical attachment level (p=0.023). Smoker versus non-smoker maintenance groups showed no significant differences in clinical parameters. There were significant differences in sRANKL and OPG concentrations between untreated and maintenance groups (all p<0.01). Salivary OPG concentration was significantly lower (all p<0.01) and the sRANKL/OPG ratio was higher (all p<0.01) in smokers than in non-smokers. OPG concentration correlated positively with probing depth, clinical attachment level and bleeding on probing (all p<0.005) and negatively with pack-year, and cotinine level (p<0.05). CONCLUSION Salivary RANKL and OPG concentrations are suggested to be affected by smoking as not only the untreated but also the treated smokers exhibited higher RANKL and lower OPG concentrations than non-smokers.

Gingival Crevicular Fluid MMP-8 and -13 and TIMP-1 Levels in Patients With Rheumatoid Arthritis and Inflammatory Periodontal Disease

BACKGROUND The purpose of this study was to compare gingival crevicular fluid (GCF) levels of matrix metalloproteinase (MMP)-8 and -13 and tissue inhibitor of MMP (TIMP)-1 in patients with rheumatoid arthritis (RA) and systemically healthy counterparts with inflammatory periodontal disease. METHODS Subjects (N = 74) were divided into five groups: 12 patients with RA and gingivitis; 13 patients with RA and periodontitis; 12 systemically healthy patients with gingivitis; 13 systemically healthy patients with periodontitis; and 24 periodontally and systemically healthy volunteers. Full-mouth clinical periodontal measurements were performed at six sites/tooth. GCF samples obtained from two sites in single-rooted teeth were analyzed by immunofluorometric assay and enzyme-linked immunosorbent assay. Data were assessed statistically by parametric tests. RESULTS The total amounts of MMP-8 were lower in the healthy control group than in RA-gingivitis, RA-periodontitis, and healthy-periodontitis groups (P <0.05). MMP-13 levels were similar in all five study groups (P >0.05). Patients with RA and gingivitis or periodontitis exhibited levels of MMP-8 and -13 and TIMP-1 that were similar to systemically healthy counterparts (P >0.05). CONCLUSIONS The coexistence of RA and periodontitis did not significantly affect the investigated parameters. GCF MMP-8 levels increased with periodontal inflammation. Despite the long-term usage of corticosteroids and non-steroidal anti-inflammatory drugs, similar GCF MMP-8 and -13 levels in patients with RA and systemically healthy counterparts suggest that RA may create a tendency to overproduce these enzymes.

Chlorhexidine decreases the risk of ventilator-associated pneumonia in intensive care unit patients: a randomized clinical trial.

BACKGROUND AND OBJECTIVE The aim was to evaluate whether oral swabbing with 0.2% chlorhexidine gluconate (CHX) decreases the risk of ventilator-associated pneumonia (VAP) in intensive care unit (ICU) patients. MATERIAL AND METHODS Sixty-one dentate patients scheduled for invasive mechanical ventilation for at least 48 h were included in this randomized, double-blind, controlled study. As these patients were variably incapacitated, oral care was provided by swabbing the oral mucosa four times/d with CHX in the CHX group (29 patients) and with saline in the control group (32 patients). Clinical periodontal measurements were recorded, and lower-respiratory-tract specimens were obtained for microbiological analysis on admission and when VAP was suspected. Pathogens were identified by quantifying colonies using standard culture techniques. RESULTS Ventilator-associated pneumonia developed in 34/61 patients (55.7%) within 6.8 d. The VAP development rate was significantly higher in the control group than in the CHX group (68.8% vs. 41.4%, respectively; p = 0.03) with an odds ratio of 3.12 (95% confidence interval = 1.09-8.91). Acinetobacter baumannii was the most common pathogen (64.7%) of all species identified. There were no significant differences between the two groups in clinical periodontal measurements, VAP development time, pathogens detected or mortality rate. CONCLUSION The finding of the present study, that oral care with CHX swabbing reduces the risk of VAP development in mechanically ventilated patients, strongly supports its use in ICUs and indeed the importance of adequate oral hygiene in preventing medical complications.

Smoking and matrix metalloproteinases, neutrophil elastase and myeloperoxidase in chronic periodontitis.

OBJECTIVES To investigate possible relationship between smoking and serum concentrations of matrix metalloproteinase-8,-9 (MMP-8, MMP-9), tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), neutrophil elastase (NE), myeloperoxidase (MPO) in chronic periodontitis (CP) patients relative to periodontally healthy subjects. METHODS Serum samples were obtained from 111 subjects before initiation of any periodontal intervention. Fifty-five CP patients (39 non-smokers, 16 smokers) and 56 periodontally healthy subjects (39 non-smokers, 17 smokers) were recruited. Serum concentrations of MMP-8 were determined by IFMA and MPO, MMP-9, TIMP-1, NE concentrations by ELISA. ANCOVA and Pearson correlation analysis was utilized for statistical analysis. RESULTS Serum MPO, NE concentrations were higher in smoker CP than non-smoker CP patients (P=0.002 and P<0.001, respectively), whereas these were similar in smoker, non-smoker periodontally healthy groups (P > 0.05). TIMP-1 concentration was higher in non-smoker CP than smoker CP group (P<0.05). MMP-9/TIMP-1 ratios were higher in smoker CP than non-smoker CP group (P=0.01). MMP-8 concentrations, MMP-8/TIMP-1 and MMP-9/TIMP-1 ratios in CP group were not significantly different from those in periodontally healthy group (P>0.05). CONCLUSIONS Our findings of significantly elevated serum MMP-9, MPO, NE together with decreased TIMP-1 in smoker CP patients than non-smokers support that smoking together with periodontal destruction may expose/predispose to cardiovascular diseases.

Salivary Antioxidants in Patients With Type 1 or 2 Diabetes and Inflammatory Periodontal Disease: A Case-Control Study

BACKGROUND The purpose of this study was to evaluate and compare salivary concentrations of reduced, oxidized glutathione, uric acid, ascorbic acid, and total antioxidant capacity in subjects with diabetes and systemically healthy subjects with inflammatory periodontal disease. METHODS Sixteen patients with type 1 diabetes mellitus (DM), 25 patients with type 2 DM, and 24 systemically healthy patients, all with inflammatory periodontal disease, were recruited. Whole-saliva samples were obtained, and full-mouth clinical periodontal measurements, including plaque index, probing depth, gingival recession, clinical attachment level, and bleeding on probing, were recorded at six sites per tooth. Saliva flow rate and salivary levels of reduced and oxidized glutathione, vitamin C, uric acid, and total antioxidant capacity were determined. Data were analyzed statistically by non-parametric tests. RESULTS The subjects with type 2 DM had fewer teeth and more sites with probing depths >4 mm than the patients with type 1 DM (both P <0.01). The mean salivary reduced-glutathione concentration was lower in patients with type 1 DM than in the other two groups (both P <0.05). No significant differences in the salivary concentrations of the other antioxidants measured were found among the groups (P >0.05). Oxidized glutathione levels in the patients with type 1 DM were significantly lower than in the systemically healthy group (P = 0.007). In both groups with diabetes, salivary reduced-glutathione levels correlated positively with probing depth, and total antioxidant capacity correlated with salivary flow rate (P <0.01). CONCLUSION The decrease in salivary reduced-glutathione levels in patients with type 1 DM may have a role in periodontal tissue destruction by predisposing tissues to oxidative stress.

Interleukin‐17 and interleukin‐18 levels in saliva and plasma of patients with chronic periodontitis

BACKGROUND AND OBJECTIVE This study was planned to investigate whether patients with chronic periodontitis exhibit different salivary and/or plasma concentrations of interleukin (IL)-17 and IL-18 compared with clinically healthy subjects. MATERIAL AND METHODS Whole saliva and blood samples, together with full-mouth clinical periodontal recordings, were obtained from 22 otherwise healthy untreated nonsmokers with chronic periodontitis and from 21 systemically and periodontally healthy control subjects. The concentrations of IL-17 and IL-18 in saliva and plasma were determined using ELISAs. RESULTS The healthy control group exhibited significantly lower values in all clinical periodontal measurements (p < 0.001). The salivary concentration of IL-17 was significantly lower, and that of IL-18 significantly higher, in patients from the chronic periodontitis group compared with healthy control subjects (p = 0.025 and p = 0.009, respectively). Plasma IL-17 and IL-18 concentrations were similar in the two study groups (p > 0.05). CONCLUSION Within the limits of the present study, it may be suggested that an elevated salivary IL-18 level in untreated nonsmoker chronic periodontitis patients has the potential to be a biomarker for periodontal tissue destruction.

Interleukin-17, RANKL, and Osteoprotegerin Levels in Gingival Crevicular Fluid From Smoking and Non-Smoking Patients With Chronic Periodontitis During Initial Periodontal Treatment

BACKGROUND This study was performed to evaluate the effects of initial periodontal treatment on the gingival crevicular fluid (GCF) levels of interleukin (IL)-17, soluble receptor activator of nuclear factor-kappa B ligand (sRANKL), and osteoprotegerin (OPG) in smoking and non-smoking patients with chronic periodontitis. METHODS At baseline, GCF samples were obtained from 10 smoking and 10 non-smoking systemically healthy patients with chronic periodontitis. Initial periodontal treatment, consisting of motivation and instruction for daily plaque control and scaling and root planing (SRP), was performed. GCF sampling and clinical periodontal measurements were repeated 4 weeks after completion of SRP. The data were tested statistically by the Student t and Wilcoxon matched-pairs test and Spearman correlation analysis. RESULTS All clinical periodontal measurements had decreased significantly 4 weeks after SRP (P <0.001). GCF volume and the total amount and concentration of OPG decreased in smokers and non-smokers after SRP, whereas the IL-17 concentration increased (P <0.05). sRANKL levels did not differ between groups or with SRP (P >0.05). Significant correlations were found between baseline IL-17 and receptor activator of nuclear factor-kappa B ligand (RANKL) levels and between baseline papilla bleeding index and OPG levels (P <0.001 and P <0.05, respectively). CONCLUSIONS Neither smoking nor periodontal inflammation seemed to influence GCF RANKL levels in systemically healthy patients with chronic periodontitis. Smoking and non-smoking patients with chronic periodontitis were not affected differently by the initial periodontal treatment with regard to GCF IL-17 and OPG concentrations.

Diabetes-related molecular signatures in infrared spectra of human saliva

BackgroundThere is an ongoing need for improvements in non-invasive, point-of-care tools for the diagnosis and prognosis of diabetes mellitus. Ideally, such technologies would allow for community screening.MethodsIn this study, we employed infrared spectroscopy as a novel diagnostic tool in the prediction of diabetic status by analyzing the molecular and sub-molecular spectral signatures of saliva collected from subjects with diabetes (n = 39) and healthy controls (n = 22).ResultsSpectral analysis revealed differences in several major metabolic components - lipid, proteins, glucose, thiocyanate and carboxylate - that clearly demarcate healthy and diseased saliva. The overall accuracy for the diagnosis of diabetes based on infrared spectroscopy was 100% on the training set and 88.2% on the validation set. Therefore, we have established that infrared spectroscopy can be used to generate complex biochemical profiles in saliva and identify several potential diabetes-associated spectral features.ConclusionsInfrared spectroscopy may represent an appropriate tool with which to identify novel diseases mechanisms, risk factors for diabetic complications and markers of therapeutic efficacy. Further study into the potential utility of infrared spectroscopy as diagnostic and prognostic tool for diabetes is warranted.