Márcio Zaffalon Casati

Impact of smoking on inflammation: overview of molecular mechanisms

BackgroundInflammation is a critical component of normal tissue repair, as well as being fundamental to the body’s defense against infection. Environmental factors, such as smoking, have been reported to modify the host response and hence modify inflammation progression, severity and outcome. Therefore, a comprehensive understanding of the molecular mechanisms by which smoking affects inflammation is vital for preventive and therapeutic strategies on a clinical level.AimThe purpose of the present article is to review the potential biological mechanisms by which smoking affects inflammation, emphasizing recent developments.ResultsSmoking is reported to effect a number of biological mediators of inflammation through its effect on immune-inflammatory cells, leading to an immunosuppressant state. Recent evidence strongly suggests that the molecular mechanisms behind the modulation of inflammation by smoking mainly involve the nuclear factor-kappa B (NF-kB) family, through the activation of both an inhibitor of IkB kinase (IKK)-dependent and -independent pathway. In addition to NF-kB activation, a number of transcriptional factors including GATA, PAX5 and Smad 3/4, have also been implicated.ConclusionMultiple mechanisms may be responsible for the association of smoking and inflammation, and the identification of potential therapeutic targets should guide future research.

Mesenchymal Stem Cell Properties of Periodontal Ligament Cells From Deciduous and Permanent Teeth

BACKGROUND Human postnatal stem cells have been identified in periodontal ligaments (PDLs). In this study, the in vitro biologic properties of CD105(+) enriched cell subsets from PDLs harvested from deciduous (DePDL) and permanent (PePDL) teeth are comparatively assessed. METHODS PDL tissue was obtained from 12 teeth (six primary and six permanent) from which CD105(+) CD34(-) CD45(-) cells were isolated by magnetic cell sorting. To identify and quantitatively compare the stem cell markers, DePDL and PePDL cells were assessed for CD166 surface antigen expression by flow cytometry, real-time polymerase chain reaction, and immunostaining for Stro-1 and Oct-4, osteogenic and adipogenic differentiation, and proliferation rate by trypan blue method. RESULTS Magnetic cell sorting isolated cell populations containing 23.87% (+/- 11.98%) and 11.68% (+/- 6.27%) of CD105(+) expressing cells from PePDL and DePDL, respectively. Flow cytometric analysis demonstrated a higher proportion of CD105(+) cells coexpressing CD166 surface antigen in PePDL, whereas immunostaining and real-time polymerase chain reaction analysis demonstrated that both cell subsets expressed Stro-1 and Oct-4. DePDL-CD105(+) subsets were more proliferative compared to PePDL subsets, and both cell populations showed multipotential capabilities to differentiate in vitro to osteoblast/cementoblast- and adipocyte-like cells. However, a higher expression of adipogenic-related genes was observed in DePDL cells, whereas PePDL-CD105(+) cell subset presented a more homogeneous osteoblast/cementoblast response. CONCLUSION These findings demonstrate that highly purified mesenchymal progenitor cell subsets can be obtained from the PDLs of both deciduous and permanent teeth, and further indicate phenotype dissimilarities that may have an impact on their clinical applications.

Subgingival biodiversity in subjects with uncontrolled type‐2 diabetes and chronic periodontitis

BACKGROUND AND OBJECTIVE There is a bidirectional relationship between periodontal disease and type-2 diabetes mellitus (DM). Inflammatory mediators may negatively affect glycemic control, and increased glucose levels and resultant glycation end-products may alter the host response against bacterial infection. However, no agreement has been reached regarding the effect of DM on periodontal subgingival microbiota. Therefore, the purpose of the present study was to compare the subgingival biodiversity in deep periodontal pockets of subjects with chronic periodontitis and either uncontrolled type-2 diabetes or no diabetes using 16S rRNA gene cloning and sequencing. MATERIAL AND METHODS Twelve subjects with uncontrolled type-2 diabetes (glycated hemoglobin > 8%) and eleven nondiabetic subjects presenting severe and generalized chronic periodontitis were selected. Subgingival biofilm from periodontal pockets > 5 mm were assessed using the 16S rRNA gene cloning and sequencing technique. RESULTS Significant differences were observed in subgingival microbiota between diabetic and nondiabetic subjects. Diabetic subjects presented higher percentages of total clones of TM7, Aggregatibacter, Neisseria, Gemella, Eikenella, Selenomonas, Actinomyces, Capnocytophaga, Fusobacterium, Veillonella and Streptococcus genera, and lower percentages of Porphyromonas, Filifactor, Eubacterium, Synergistetes, Tannerella and Treponema genera than nondiabetic individuals (p < 0.05). Moreover, some phylotypes, such as Fusobacterium nucleatum, Veillonella parvula, V. dispar and Eikenella corrodens were detected significantly more often in diabetic subjects than in nondiabetic subjects (p < 0.05). CONCLUSION Subjects with uncontrolled type-2 diabetes and chronic periodontitis presented significant dissimilarities in subgingival biodiversity compared with nondiabetic subjects.

Periodontal debridement as a therapeutic approach for severe chronic periodontitis: a clinical, microbiological and immunological study

AIM To clinically, microbiologically and immunologically characterize periodontal debridement as a therapeutic approach for severe chronic periodontitis. MATERIAL AND METHODS Twenty-five patients presenting at least eight teeth with a probing pocket depth (PPD) of >or=5 mm and bleeding on probing (BOP) were selected and randomly assigned to quadrant-wise scaling and root planing or one session of full-mouth periodontal debridement. The following clinical outcomes were assessed: plaque index, BOP, position of gingival margin, relative attachment level (RAL) and PPD. Real-time PCR was used for quantitative analysis of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Tannerella forsythia. The enzyme-linked immunosorbent assay permitted the detection of IL-1beta, prostaglandin E(2), INF-gamma and IL-10 in gingival crevicular fluid (GCF). All the parameters were evaluated at baseline, and at 3 and 6 months after treatment. RESULTS Both the groups had similar means of PPD reduction and attachment gain over time. Besides a significant reduction in the bacterial level after treatment in both groups, microbiological analysis failed to demonstrate significant differences between them. Finally, no difference was observed between groups with respect to the levels of inflammatory mediators in GCF. CONCLUSION Periodontal debridement resulted in a similar clinical, microbiological and immunological outcome when compared with standard scaling and root planing and therefore may be a viable approach to deal with severe chronic periodontitis.

Current perspective of the impact of smoking on the progression and treatment of periodontitis

This literature review provides an overview of the current scenario regarding the impact of smoking on the progression and treatment of periodontitis; clinical, microbiological and immunological data from studies from our and other groups are presented. In general, preclinical and clinical data are unanimous in demonstrating that smokers present increased susceptibility, greater severity and faster progression of periodontal disease compared with nonsmokers. The evidence further demonstrates that smokers lose more teeth and have a less favorable response to therapy than do nonsmokers. Although it is well established that smoking significantly impacts on the onset, progression and outcome of periodontal disease, the mechanisms involved remain unclear. More importantly, some of the reported deleterious effects of smoking on periodontal tissues have been reported to be reversible upon participation in smoking-cessation programs. Therefore, clinicians should strongly advise smokers to enroll in cessation strategies, even temporarily, in order to improve the overall outcome.

Antimicrobial Photodynamic Therapy as an Adjunct to Non-Surgical Treatment of Aggressive Periodontitis: A Split-Mouth Randomized Controlled Trial

BACKGROUND The management of aggressive periodontitis (AgP) represents a challenge for clinicians because there are no standardized protocols for an efficient control of the disease. This randomized controlled clinical trial evaluated the effects of repeated applications of antimicrobial photodynamic therapy (aPDT) adjunctive to scaling and root planing (SRP) in patients with AgP. METHODS Using a split-mouth design, 20 patients with generalized AgP were treated with aPDT + SRP (test group) or SRP only (control group). aPDT was applied at four periods. All patients were monitored for 90 days. Clinical, microbiologic, and immunologic parameters were statistically analyzed. RESULTS In deep periodontal pocket analysis (probing depth [PD] ≥ 7 mm at baseline), the test group presented a decrease in PD and a clinical attachment gain significantly higher than the control group at 90 days (P < 0.05). The test group also demonstrated significantly less periodontal pathogens of red and orange complexes and a lower interleukin-1β/interleukin-10 ratio than the control group (P < 0.05). CONCLUSION The application of four sessions of aPDT, adjunctive to SRP, promotes additional clinical, microbiologic, and immunologic benefits in the treatment of deep periodontal pockets in single-rooted teeth in patients with AgP.

Full-Mouth Ultrasonic Debridement Associated With Amoxicillin and Metronidazole in the Treatment of Severe Chronic Periodontitis

BACKGROUND The purpose of this study was to evaluate the adjunctive clinical, microbiologic, and immunologic effects of the systemic administration of amoxicillin and metronidazole in the full-mouth ultrasonic debridement of patients with severe chronic periodontitis. METHODS Twenty-five patients presenting at least eight teeth with probing depth (PD) > or =5 mm and bleeding on probing (BOP) were selected and randomly assigned to full-mouth ultrasonic debridement + placebo (control group) or full-mouth ultrasonic debridement + amoxicillin and metronidazole (test group). The clinical outcomes evaluated were visible plaque index, BOP, position of the gingival margin, relative attachment level (RAL), and PD. Real-time polymerase chain reaction (PCR) was used for quantitative analysis of Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans), Porphyromonas gingivalis, and Tannerella forsythia (previously T. forsythensis). The enzyme-linked immunosorbent assay (ELISA) technique permitted the detection of prostaglandin E(2,) interleukin-1beta, and interferon-gamma levels in gingival crevicular fluid. All parameters were evaluated at baseline and at 3 and 6 months post-treatment. RESULTS At 6 months, the test treatment resulted in lower BOP and an additional reduction (0.83 mm) in PD (P <0.05). Data also showed RAL gain > or =2 mm at 43.52% of sites in control patients compared to 58.03% of sites in test patients (P <0.05). However, both groups had similar mean RAL gain (1.68 and 1.88 mm for the control and test groups, respectively). Real-time PCR and ELISA failed to identify significant differences between the groups. CONCLUSIONS Both treatments resulted in significant clinical improvements; however, there was a slight, but significantly greater, improvement in BOP and the percentage of sites with PD > or =5 mm exhibiting RAL gain > or =2 mm in the test group. Nevertheless, no improvement in the microbiologic or immunologic outcome was observed with the adjunctive use of systemic amoxicillin and metronidazole.

Tooth-derived stem cells: Update and perspectives

Tissue engineering is an emerging field of science that focuses on creating suitable conditions for the regeneration of tissues. The basic components for tissue engineering involve an interactive triad of scaffolds, signaling molecules, and cells. In this context, stem cells (SCs) present the characteristics of self-renewal and differentiation capacity, which make them promising candidates for tissue engineering. Although they present some common markers, such as cluster of differentiation (CD)105, CD146 and STRO-1, SCs derived from various tissues have different patterns in relation to proliferation, clonogenicity, and differentiation abilities in vitro and in vivo. Tooth-derived tissues have been proposed as an accessible source to obtain SCs with limited morbidity, and various tooth-derived SCs (TDSCs) have been isolated and characterized, such as dental pulp SCs, SCs from human exfoliated deciduous teeth, periodontal ligament SCs, dental follicle progenitor cells, SCs from apical papilla, and periodontal ligament of deciduous teeth SCs. However, heterogeneity among these populations has been observed, and the best method to select the most appropriate TDSCs for regeneration approaches has not yet been established. The objective of this review is to outline the current knowledge concerning the various types of TDSCs, and discuss the perspectives for their use in regenerative approaches.

Photodynamic Therapy During Supportive Periodontal Care: Clinical, Microbiologic, Immunoinflammatory, and Patient-Centered Performance in a Split-Mouth Randomized Clinical Trial

BACKGROUND This study investigates the effect of photodynamic therapy (PDT) as monotherapy during supportive periodontal therapy. METHODS A split-mouth, randomized controlled trial was conducted in patients with chronic periodontitis (N = 22) presenting at least three residual pockets (probing depth [PD] ≥5 mm with bleeding on probing [BOP]). The selected sites randomly received the following: 1) PDT; 2) photosensitizer (PS); or 3) scaling and root planing (SRP). At baseline and 3 and 6 months, clinical, microbiologic (real-time polymerase chain reaction analyses), cytokine pattern (multiplexed bead immunoassay), and patient-centered (regarding morbidity) evaluations were performed. RESULTS All therapies promoted similar improvements in clinical parameters throughout the study (P <0.05), except that BOP was not reduced in the PS protocol (P >0.05). Lower levels of Aggregatibacter actinomycetemcomitans were observed in the PDT and SRP protocols at 3 months when compared with the PS protocol (P <0.05). An inferior frequency detection of Porphyromonas gingivalis was observed in the PDT protocol at 3 and 6 months and in the SRP protocol at 6 months from baseline (P <0.05). In addition, PDT protocol presented inferior frequency of P. gingivalis at 3 months when compared with the other therapies (P <0.05). Only patients in the PDT protocol exhibited augmented levels of anti-inflammatory interleukin (IL)-4 and reduced proinflammatory IL-1β and IL-6 throughout the study (P <0.05). Intergroup analyses showed reduced IL-10 and increased interferon-γ and IL-1β levels in the PS protocol when compared with the other therapies during follow-ups (P <0.05). No differences in morbidity were observed between the therapies (P >0.05), although the need for anesthesia was higher in SRP-treated sites (P <0.05). CONCLUSION PDT as an exclusive therapy may be considered a non-invasive alternative for treating residual pockets, offering advantages in the modulation of cytokines.

DNA Methylation Status of the IL8 Gene Promoter in Aggressive Periodontitis

BACKGROUND Studies evaluating the methylation status of cytokine genes may have relevance for inflammatory diseases in which the expression of some cytokines is altered, such as periodontitis. This study observes the DNA methylation status in the interleukin-8 (IL8) gene promoter in cells of the oral epithelium of subjects affected by generalized aggressive periodontitis (AgP) and compares it to those of control subjects. METHODS Genomic DNA from epithelial oral cells of 37 generalized AgP patients and 37 controls were purified and modified by sodium bisulphite. Modified DNA was submitted by methylation-specific polymerase chain reaction, electrophoresed on 10% polyacrylamide gels, and stained. RESULTS Subjects who presented generalized AgP have a higher frequency of hypomethylation of the IL8 gene promoter in oral epithelium cells than that of controls (86.5% in the generalized AgP group versus 62% in the control group; P = 0.016; chi(2) test). CONCLUSIONS A marked hypomethylated status is found in the oral epithelial cells of subjects presenting with generalized AgP, compared to controls, in the promoter region of the IL8 gene. This hypomethylated status may reflect a generalized condition of oral epithelial cells, including gingival epithelium, because gingival epithelial cells were also collected during mouthwash use.