Luciana M. Shaddox

Microbiological Characterization in Children with Aggressive Periodontitis

The objective of this study was to characterize the subgingival microbiota of African-American children with Localized Aggressive Periodontitis (LAP). Fifty-one children were included. Subgingival plaque samples were taken from diseased (DD) and healthy sites (DH) in LAP and from healthy sites in HS and HC and analyzed by 16S rRNA-based microarrays. Aggregatibacter actinomycetemcomitans (Aa) was the only species found to be both more prevalent (OR = 8.3, p = 0.0025) and abundant (p < 0.01) in DD. Filifactor alocis (Fa) was also found to be more prevalent in DD (OR 2.31, CI 1.06-5.01, p = 0.03). Most prevalent species in healthy sites were Selenomonas spp, Veillonella spp, Streptococcus spp, Bergeyella sp, and Kingella oralis. Overall, Streptococcus spp, Campylobacter gracilis, Capnocytophaga granulosa, Haemophilus parainfluenzae, and Lautropia mirabilis were most abundant in healthy children, while Aa, Fa, Tannerella sp, Solobacterium moorei, Parvimonas micra, and Capnocytophaga sp were most abundant in LAP. Based on a comprehensive analysis with 16S rRNA-based microarrays, Aa was strongly associated and site-specific in LAP. In contrast, other species were found to be associated with healthy sites and individuals (ClinicalTrials.gov number CT01330719). Abbreviations: healthy site in healthy sibling (HS); healthy site in healthy control child (HC).

Hyper-responsive Phenotype in Localized Aggressive Periodontitis

The ‘hyper-responsive’ trait is an increased inflammatory response upon stimulation of innate immune receptors. Our objective was to determine if a hyper-reactive trait is present in a cohort diagnosed with aggressive periodontitis (LAgP). Peripheral blood was collected from 30 LAgP, 10 healthy unrelated, and 10 healthy sibling participants and stimulated with lipopolysaccharide (LPS) from E. coli and P. gingivalis. Cyto/chemokine response profiles were evaluated and analyzed by ANOVA. Elevated levels of pro-inflammatory cyto/chemokines were detected in E. coli and P. gingivalis LPS-stimulated LAgP cultures when compared with those of healthy unrelated control individuals. Periodontally healthy siblings presented with attenuated hyper-inflammatory cyto/chemokine profiles. Regression analysis demonstrated the hyper-reactive trait to be concomitant expression of pro-inflammatory cyto/chemokines and an absence of anti-inflammatory mediator expression. Our findings demonstrate hyper-responsive trait in a LAgP cohort, along with an attenuated hyper-responsiveness in healthy siblings, which can be induced in response to multiple TLR ligations.

Local Inflammatory Markers and Systemic Endotoxin in Aggressive Periodontitis

While much research has focused on local and systemic factors contributing to periodontal disease, little is known regarding mechanisms linking these factors. We have previously reported a systemic hyper-inflammatory response to bacterial endotoxin in localized aggressive periodontitis (LAP). The objectives of this study were to delineate cyto/chemokines in gingival crevicular fluid (GCF) and evaluate systemic levels of endotoxin associated with LAP. Clinical parameters, GCF, and peripheral blood were collected from: 34 LAP, 10 healthy siblings, and nine healthy unrelated control individuals. Cyto/chemokines were quantified in GCF, systemic endotoxin levels were quantified in plasma, and correlation analysis was performed among all parameters. Nine mediators were elevated in LAP diseased sites as compared with healthy sites (TNFα, INFγ, IL1β, IL2, IL6, IL10, Il12p40, GMCSF, and MIP1α, p < 0.001), while MCP1, IL4, and IL8 were elevated in healthy sites (p < 0.01). Four- to five-fold-higher endotoxin levels were detected in LAP plasma compared with that from healthy participants (p < 0.0001), which correlated with all clinical parameters and most cyto/chemokines analyzed. In conclusion, higher systemic levels of endotoxin were found in LAP, which correlates with an exacerbated local inflammatory response and clinical signs of disease. (Clinicaltrials.gov number, NCT01330719).

Local levels of inflammatory mediators in uncontrolled type 2 diabetic subjects with chronic periodontitis

AIM The aim of this study was to evaluate the levels of a wide panel of cyto/chemokines in the gingival crevicular fluid (GCF) of uncontrolled type 2 diabetic subjects as compared with non-diabetic subjects with periodontitis. METHODS Twenty-six uncontrolled type 2 diabetic subjects (glycated haemoglobin levels >7.5%) and 20 non-diabetic subjects with chronic periodontitis were enrolled in this study. The levels of 14 cyto/chemokines were measured in the GCF of healthy and diseased sites of the diabetic and non-diabetic subjects using multiplex bead immunoassays. RESULTS The concentrations of eotaxin, macrophage inflammatory protein-1α, granulocyte-macrophage colony-stimulating factor, interleukin (IL)-6, tumour necrosis factor-α and IL-12 were higher in healthy and diseased sites of diabetic than non-diabetic subjects, after adjustment for multiple comparisons (p < 0.0035). CONCLUSION Uncontrolled type 2 diabetes mellitus modulated the local levels of several cyto/chemokines at both healthy and diseased periodontal sites in favour of a proinflammatory profile, which may partially explain the greater susceptibility of diabetic subjects to periodontal breakdown.

Autoreactivity of Serum Immunoglobulin to Periodontal Tissue Components: A Pilot Study

BACKGROUND Periodontal diseases are inflammatory diseases resulting in the destruction of tissues of the periodontium. Although bacteria must be present for periodontal disease to occur, a susceptible host is also required, which is determined by genetic, environmental, and acquired factors. One such factor, autoimmunity, may play a role in the tissue destruction. Data indicate that some antibodies that occur in the gingival lesion are directed to host tissue components, such as type I collagen, although investigations of other periodontal autoimmune targets are limited. METHODS Histologic sections and extracts from periodontally healthy teeth and the associated soft tissues were probed with serum from localized aggressive periodontitis (LAgP), chronic periodontitis (CP), and periodontally healthy subjects to determine autoreactivity to components of the periodontium. Any autoreactivity observed was characterized further by mass spectrometry and enzyme-linked immunosorbent assay. RESULTS Autoreactivity to components of the periodontium was observed in CP and LAgP. Known autoimmune targets, such as collagen and heat shock protein, were identified along with multiple potential autoimmune targets, including members of the extracellular matrix, such as vimentin, spectrin, filamin, actin, lamin, keratin, and tubulin. Finally, it was determined that the autoreactivity observed in LAgP was more severe and diverse than that observed in CP. CONCLUSION These data demonstrated that autoimmune reactivity can play a role in the tissue destruction of periodontal disease but that the nature of the autoreactivity may differ based on the type and/or stage of periodontal disease.

LPS-induced Inflammatory Response after Therapy of Aggressive Periodontitis

We have reported a lipopolysaccharide (LPS)-induced hyper-inflammatory response in localized aggressive periodontitis (LAP). It is unknown whether treatment is able to modulate this LPS responsiveness. Fifty-nine individuals with LAP were treated by mechanical debridement and systemic antibiotics. Clinical parameters and cyto/chemokine responsiveness of whole blood stimulated with Porphyromonas gingivalis or Escherichia coli LPS were monitored at baseline and 3, 6, and 12 months post-treatment. Overall, clinical parameters were improved following treatment. Additionally, P. gingivalis LPS induction of eotaxin, IFNγ, IL10, IL12p40, IL1β, IL6, IP10, MCP1, MIP1α, GM-CSF, and TNFα was significantly decreased (p < .05). Similarly, induction of eotaxin, INFγ, IL10, IL12p40, GM-CSF, and TNFα by E. coli LPS was also reduced post-treatment. These reductions correlated with decreases in clinical parameters. Importantly, these reductions in LPS responsiveness were most robust at 3 months, and some lost significance at 6 to 12 months post-treatment. In conclusion, LPS-induced hyper-inflammatory response in LAP can be partially modulated by periodontal therapy. Conversely, rebound in the hyper-responsiveness of some mediators, in the presence of improved clinical parameters, suggests that this phenotype could be partially influenced by a genetic trait and play a role in future disease recurrence (ClinicalTrials.gov, NCT01330719).

Treating chronic periodontitis: current status, challenges, and future directions.

Periodontitis affects roughly one-third of the US population. A timely diagnosis of chronic periodontitis at its earliest stage is essential to avoid more challenging severe stages of the disease. Most cases of slight and moderate chronic periodontitis can be successfully managed by mechanical removal and/or reduction of subgingival bacterial biofilms and calculus. However, any factor that affects either the local environment or the host response may contribute to progression of the disease and a poor treatment response. Thus, it is essential that clinicians are aware of etiologic and risk factors associated with disease development and progression in order to plan and execute a successful treatment. This paper reviews a variety of risk factors, both local and systemic, that can impact the successful treatment of chronic periodontitis.

Systemic inflammatory responses in patients with type 2 diabetes with chronic periodontitis

Objective The objective of this case–control study was to quantify the immune responsiveness in individuals with type 2 diabetes (T2D) as compared with patients without diabetes (NT2D) diagnosed with periodontitis. Research Design and Methods Peripheral blood was collected from 20 patients with moderate-to-severe chronic periodontitis (10 T2D, 10 NT2D). Blood samples were stimulated with ultrapure Porphyromonas gingivalis and Escherichia coli lipopolysaccharide (LPS) for 24 hours. 14 cytokines/chemokines were quantified in culture supernatants using multiplex technology. Results T2D individuals demonstrated higher unstimulated levels of interleukin 6 (IL-6), IL-1β, tumor necrosis factor α, interferon γ, IL-10, IL-8, macrophage inflammatory protein 1α (MIP1α), and 1β (MIP1β), and higher stimulated levels of IL-6, IL-8, IL-10, MIP1α and MIP1β, along with lower unstimulated and stimulated levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) when compared with NT2D (p<0.05). Importantly, the LPS-induced levels of IL-6, IL-8, IL-10 and MIP1α strongly correlated with severity of disease, measured by pocket depths (PD), within the T2D group (r2≥0.7, p<0.05), but not within NT2D. Conclusions Among patients with chronic periodontitis, patients with T2D seem to have an enhanced LPS-induced immune responsiveness than individuals without diabetes, which correlates with periodontal disease severity, concomitant with a less robust GM-CSF response. This data may in part explain the higher predisposition to periodontitis in this population.

Benefits of early systemic antibiotics in localized aggressive periodontitis: a retrospective study

BACKGROUND Treatment of localized aggressive periodontitis (LAP) may include systemic antibiotics, yet it is unclear at what stage of treatment planning antibiotics are most effective. AIM This retrospective analysis compared immediate versus delayed antibiotic therapy on clinical parameters and gingival crevicular fluid (GCF) inflammatory mediators. MATERIAL AND METHODS At baseline, 3 months and 6 months after treatment, clinical parameters [probing depth (PD), clinical attachment level (CAL), bleeding on probing (BoP) and plaque] and GCF were collected from LAP participants, who received a 7-day antibiotic regimen immediately (ImA) or 3 months following (DelA) mechanical therapy. RESULTS Although both groups presented significant CAL reductions at 6 months, only ImA resulted in a reduction in mean PD at both 3 and 6 months, along with reductions in CAL and BoP at 3 months following therapy. In addition, GCF mediators were higher in DelA group at 3 months post mechanical treatment, but were significantly reduced 6 months following antibiotic therapy. CONCLUSIONS ImA and DelA regimens were both effective in improving CAL by 6 months post therapy. However, ImA allowed for better improvement in overall clinical parameters early in the course of treatment, concomitant with lower levels of inflammatory mediators within the GCF.

Correlation of Aggregatibacter actinomycetemcomitans Detection with Clinical/Immunoinflammatory Profile of Localized Aggressive Periodontitis Using a 16S rRNA Microarray Method: A Cross-Sectional Study

Objective The objective of this study was to determine whether the detection of Aggregatibacter actinomycetemcomitans (Aa) correlates with the clinical and immunoinflammatory profile of Localized Aggressive Periodontitis (LAP), as determined by by 16S rRNA gene-based microarray. Subjects and Methods Subgingival plaque samples from the deepest diseased site of 30 LAP patients [PD ≥ 5 mm, BoP and bone loss] were analyzed by 16S rRNA gene-based microarrays. Gingival crevicular fluid (GCF) samples were analyzed for 14 cyto/chemokines. Peripheral blood was obtained and stimulated in vitro with P.gingivalis and E.coli to evaluate inflammatory response profiles. Plasma lipopolysaccharide (LPS) levels were also measured. Results Aa was detected in 56% of LAP patients and was shown to be an indicator for different bacterial community structures (p<0.01). Elevated levels of pro-inflammatory cyto/chemokines were detected in LPS-stimulated blood samples in both Aa-detected and Aa-non-detected groups (p>0.05). Clinical parameters and serum LPS levels were similar between groups. However, Aa-non-detected GCF contained higher concentration of IL-8 than Aa-detected sites (p<0.05). TNFα and IL1β were elevated upon E.coli LPS stimulation of peripheral blood cells derived from patients with Aa-detected sites. Conclusions Our findings demonstrate that the detection of Aa in LAP affected sites, did not correlate with clinical severity of the disease at the time of sampling in this cross-sectional study, although it did associate with lower local levels of IL-8, a different subgingival bacterial profile and elevated LPS-induced levels of TNFα and IL1β.