Maria Lúcia Rubo de Rezende

Differential production of macrophage inflammatory protein-1alpha, stromal-derived factor-1, and IL-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide from P. gingivalis.

BACKGROUND Fibroblasts are considered important cells in periodontitis. When challenged by different agents, they respond through the release of cytokines that participate in the inflammatory process. The aim of this study is to evaluate and compare the expression and production of macrophage inflammatory protein (MIP)-1alpha, stromal-derived factor (SDF)-1, and interleukin (IL)-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide (LPS) from Porphyromonas gingivalis. METHODS Fibroblasts were cultured from biopsies of gingival tissue and periodontal ligament of the same donors and used on the fourth passage. After confluence in 24-well plates, the culture medium alone (control) or with 0.1 to 10 microg/ml of LPS from P. gingivalis was added to the wells, and after 1, 6, and 24 hours, the supernatant and the cells were collected and analyzed by enzyme-linked immunosorbent assay and real-time polymerase chain reaction, respectively. RESULTS MIP-1alpha, SDF-1, and IL-6 protein production was significantly greater in gingival fibroblasts compared to periodontal ligament fibroblasts. IL-6 was upregulated in a time-dependent manner, mainly in gingival fibroblasts (P <0.05), which secreted more MIP-1alpha in the lowest concentration of LPS used (0.1 microg/ml). In contrast, a basal production of SDF-1 that was inhibited with the increase of LPS concentration was detected, especially after 24 hours (P <0.05). CONCLUSION The distinct ability of the gingival and periodontal ligament fibroblasts to secrete MIP-1alpha, SDF-1, and IL-6 emphasizes that these cells may differently contribute to the balance of cytokines in the LPS-challenged periodontium.

Effects of TGF-β1, PDGF-BB, and IGF-1 on the rate of proliferation and adhesion of a periodontal ligament cell lineage in vitro

BACKGROUND Considering the role of growth factors in periodontal regeneration, the aim of this study was to evaluate the influence of platelet-derived growth factor (PDGF)-BB, insulin-like growth factor (IGF)-1, and transforming growth factor-beta 1(TGF-β1), alone or in combination, on the rate of proliferation and adhesion of periodontal ligament (PDL) cells in vitro. METHODS After establishment and characterization of a primary culture of PDL cells, 72 culture dishes were plated with 103 cells distributed among four test groups and a control group. Test groups had PDGF-BB, TGF-β1, IGF-1, or a combination of all three added to the culture medium, whereas the control group received no growth factor. The samples were counted in triplicate 1, 3, 5, and 7 days after seeding. For the adhesion assay, 14 patients provided 30 root fragments distributed among 10 groups: scaling and root planing (SRP), SRP + growth factors, SRP + citric acid plus tetracycline (CA+T), and SRP + (CA+T) + growth factors. The data were evaluated statistically by analysis of variance complemented by Tukey, Dunnett, and Student-Newman-Keuels methods. RESULTS Maximum rates of proliferation were observed at day 3 for all groups. TGF-β1 induced a 344.17% ± 58.80% increased proliferation rate over control (P <0.05), followed by the combination (277.5% ± 29.38%), PDGF-BB (238.79% ± 5.79%), and IGF-1 (233.16% ± 19.19%). Groups treated by (CA+T) showed increased numbers of cells attached to root fragments, especially SRP + (CA+T) + combination (13.25 ± 1.79), with significant differences (P <0.05) from groups treated only by SRP. CONCLUSION This combination of growth factors stimulated a mitogenic response and favored the adhesion of PDL cells in vitro, suggesting its possible role in periodontal regeneration.

Long-term evaluation of periodontal parameters and implant outcomes in periodontally compromised patients: a systematic review.

BACKGROUND The aim of this systematic review is to evaluate the long-term outcomes of patients with periodontitis submitted to periodontal therapy/maintenance and implant placement. METHODS Studies reporting clinical and/or long-term implant outcomes from partially edentulous patients with periodontitis who were treated and followed periodontal maintenance for ≥5 years were considered eligible for the review. Screening of the articles, data extraction, and quality assessment were conducted independently and in duplicate. RESULTS Search of MEDLINE, EMBASE, and CENTRAL databases resulted in 959 papers, and of them 931 were excluded after title/abstract assessment. The full texts of 28 potentially eligible publications were screened, but only 10 studies met inclusion criteria. Most of the included studies (77.8%) presented a medium/high methodologic quality. The results demonstrated that patients with a diagnosis of periodontitis had satisfactory implant outcomes. Implant survival was high (92.1%) within studies reporting 10 years of follow-up. Parameters related to probing depth, clinical attachment level, and bone loss around teeth increased the occurrence of peri-implantitis and implant loss. Non-attendance to periodontal maintenance and smoking habits were also associated with less favorable implant outcomes. CONCLUSIONS This systematic review confirmed that implant therapy can be successfully used in patients with a diagnosis of periodontitis who underwent proper therapy and regular periodontal maintenance. Residual pockets, non-attendance to the periodontal maintenance program, and smoking were considered to be negative factors for the long-term implant outcomes.

Comparison among four commonly used demineralizing agents for root conditioning: a scanning electron microscopy

Dental roots that have been exposed to the oral cavity and periodontal pocket environment present superficial changes, which can prevent connective tissue reattachment. Demineralizing agents have been used as an adjunct to the periodontal treatment aiming at restoring the biocompatibility of roots. Objective This study compared four commonly used demineralizing agents for their capacity of removing smear layer and opening dentin tubules. Methods Fifty fragments of human dental roots previously exposed to periodontal disease were scaled and randomly divided into the following groups of treatment: 1) CA: demineralization with citric acid for 3 min; 2) TC-HCl: demineralization with tetracycline-HCl for 3 min; 3) EDTA: demineralization with EDTA for 3 min; 4) PA: demineralization with 37% phosphoric acid for 3 min; 5)Control: rubbing of saline solution for 3 min. Scanning electron microscopy was used to check for the presence of residual smear layer and for measuring the number and area of exposed dentin tubules. Results Smear layer was present in 100% of the specimens from the groups PA and control; in 80% from EDTA group; in 33.3% from TC-HCl group and 0% from CA group. The mean numbers of exposed dentin tubules in a standardized area were: TC-HCl=43.8±25.2; CA=39.3±37; PA=12.1±16.3; EDTA=4.4±7.5 and Control=2.3±5.7. The comparison showed significant differences between the following pairs of groups: TC-HCl and Control; TC-HCl and EDTA; CA and Control; and CA and EDTA. The mean percentages of area occupied by exposed dentin tubules were: CA=0.12±0.17%; TC-HCl=0.08±0.06%; PA=0.03±0.05%; EDTA=0.01±0.01% and Control=0±0%. The CA group differed significantly from the others except for the TC-HCl group. Conclusion There was a decreasing ability for smear layer removal and dentin tubule widening as follows: AC>TC-HCl>PA>EDTA. This information can be of value as an extra parameter for choosing one of them for root conditioning.

Influence of combined oral contraceptives on the periodontal condition

Most studies investigating the impact of oral contraceptives have been performed some years ago, when the level of sexual hormones was greater than the actual formulations. Objective The aim of this study was to evaluate the effects of current combined oral contraceptives (COC) on periodontal tissues, correlating the clinical parameters examined with the total duration of continuous oral contraceptive intake. Material and methods Twenty-five women (19-35 years old) taking combined oral contraceptives for at least 1 year were included in the test group. The control group was composed by 25 patients at the same age range reporting no use of hormone-based contraceptive methods. Clinical parameters investigated included pocket probing depth (PD), clinical attachment level (CAL), sulcular bleeding index (SBI) and plaque index (Pl.I). Data were statistically evaluated by unpaired t test, Pearsons correlation test and Spearmans correlation test. Results The test group showed increased PD (2.228±0.011 x 2.154±0.012; p<0.0001) and SBI (0.229±0.006 x 0.148±0.005, p<0.0001) than controls. No significant differences between groups were found in CAL (0.435±0.01 x 0.412±0.01; p=0.11). The control group showed greater Pl.I than the test group (0.206±0.007 x 0.303±0.008; p<0.0001). No correlation between the duration of oral contraceptive intake, age and periodontal parameters was observed. Conclusions These findings suggest that the use of currently available combined oral contraceptives can influence the periodontal conditions of the patients, independently of the level of plaque accumulation or total duration of medication intake, resulting in increased gingival inflammation.

Demineralization of the Contacting Surfaces in Autologous Onlay Bone Grafts Improves Bone Formation and Bone Consolidation

BACKGROUND Autologous bone grafts are usually well consolidated after 4 to 5 months but can be incompletely interlocked with the native bone. This study investigated the effect of acid demineralization of the graft-bed interface on graft consolidation. METHODS Onlay bone grafts were performed on the calvaria of 36 guinea pigs. Half of the animals had the graft-bed contacting surfaces demineralized with 50% citric acid (pH 1.0) for 3 minutes (test group). The other half received no demineralization (control group). The bone grafts were immobilized by a resorbable membrane glued to the recipient bed with cyanoacrylate. After 7, 30, and 90 days, specimens (n = 6) were obtained for light microscopy. Data from qualitative analysis and computerized histomorphometry were statistically processed at a significance level of 5%. RESULTS Osteogenesis was not seen at the interface after 7 days. After 30 days, the test group showed 34.39% ± 13.4% of the interface area filled with mineralized tissue, compared to 17.14% ± 8.6% in the control group (P = 0.026). After 90 days, the mean percentages of mineralized tissue at the interface in the test and control specimens were 54.00% ± 11.23% and 38.65% ± 7.76% (P = 0.041), respectively. Within groups, a higher percentage of the area filled with mineralized tissue was seen at 90 days compared to 30 days (P = 0.004 for control and 0.041 for test). CONCLUSIONS Demineralization of the contacting surfaces between autologous bone graft and bone bed improved new bone formation and bone consolidation. These data need to be confirmed in humans.

Evaluation of oral and nasal odor in patients with and without cleft lip and palate: preliminary report.

Objective To evaluate oral and nasal halitosis parameters in patients with and without clefts. Design Randomized and prospective study. Patients with and without clefts were evaluated as to oral and nasal halitosis. Setting University of São Paulo, Bauru Dental School and Hospital for Rehabilitation of Craniofacial Anomalies, Bauru, Brazil. Patients Twelve patients with clefts and 12 without clefts were evaluated, and no exclusion criteria were followed. Interventions Nasal and oral halitosis were measured with the use of a portable sulfide monitor during a single visit. Results One hundred percent of the patients with clefts had altered values for volatile sulfur compounds. Only 33.3% of the patients without clefts had at least one strong value for nasal halimeter measurements, and 58.3% of these patients showed alteration in the nasal values. Statistical evaluation, made using the Mann-Whitney U test, classifying the nasal halimeter values as normal, weak, and strong, showing a statistical significant group difference (p = .003). There was no significant difference in the oral halimeter values between the two groups. Conclusions Patients with cleft lip and palate had a tendency to present higher values for the nasal halimeter measurements, when compared with patients without clefts.

Inhibitory Signals Mediated by Programmed Death-1 Are Involved With T-Cell Function in Chronic Periodontitis

BACKGROUND Inhibitory signals mediated via molecules such as programmed death-1 (PD-1) play a critical role in downmodulating immune responses and maintaining peripheral tolerance. We investigated the involvement of cytokines and PD-1 engagement in mediating the T-cell unresponsiveness to bacterial and ubiquitous antigens in periodontal diseases. METHODS Gingival and peripheral blood samples from healthy individuals and patients with chronic periodontitis were collected and used for the subsequent assays. Leukocytes in the lesion site and blood were evaluated using flow cytometry. The production of interferon-gamma, interleukin-10, and transforming growth factor-beta proteins was evaluated by enzyme-linked immunosorbent assay (ELISA), and the presence of PD-1+ cells in the inflamed gingiva was confirmed by immunofluorescence confocal microscopy for CD4 and PD-1 colocalization. RESULTS T cells from patients with chronic periodontitis proliferated poorly in response to Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans) antigen. T-cell unresponsiveness was not associated with imbalanced cytokine production. However, T cells from patients with chronic periodontitis expressed significantly higher levels of PD-1 either upon isolation or after culture with antigens. Moreover, PD-1 blocking did not result in significant T-cell proliferation in cells cultured with phytohemagglutinin or bacterial antigens. The blockade of PD-1 resulted in the increased production of IFN-gamma. In addition, CD4+ and CD8+ T cells expressing PD-1 accumulated in lesions with chronic periodontitis. CONCLUSION These data show that PD-1 engagement could be involved in the modulation of IFN-gamma production by T cells in patients with chronic periodontitis.

A modified approach for vestibuloplasty in severely resorbed mandible using an implant-retained postoperative stent: a case report

BACKGROUND Severely resorbed mandibles often present a short band of keratinized tissue associated with a shallow vestibule. As a result, prominent muscle insertions are present, especially in the mental region of the mandible. This case report describes the deepening of the vestibular sulcus in an atrophic mandible by combining free gingival grafts harvested from the palate and a postoperative acrylic resin stent screwed on osseointegrated implants placed at the anterior region of the mandible. STUDY DESIGN During the second-stage surgery, a split-thickness labial flap was reflected and apically sutured onto the periosteum. Two free gingival grafts were obtained and then sutured at this recipient site. A previously custom-made acrylic stent was then screwed onto the most distally positioned implants. To document the procedures stability over time, a metal ball was placed in the most apical part of the vestibule and standardized cephalometric radiographs were taken before and 6 months after the procedure. Linear measurements of vestibular depths over the observation time were realized using specific software for radiographic analysis. RESULTS The proposed technique augmented the band of attached masticatory mucosa, deepened the vestibule and prevented the muscle reinsertion. The difference between the 2 measurements of vestibular depths was 9.39 mm (initial 20.88 mm, final 11.49 mm) after a 6-month postoperative period. CONCLUSION The technique, in combination with palatal mucosal graft and use of a postoperative stent, decreased the pull of mentalis muscle and provided a peri-implantally stable soft tissue around implants.

Bone demineralization with citric acid enhances adhesion and spreading of preosteoblasts.

BACKGROUND Previous studies have demonstrated that bone demineralization can improve consolidation in bone grafts. The biologic mechanisms underlying this phenomenon remain unclear. METHODS Twelve adult male guinea pigs were used in this experiment. Forty-five bone samples removed from the calvaria of nine animals were divided in groups (n = 9) according to the time of demineralization with citric acid (50%, pH 1): 15, 30, 90, and 180 seconds and non-demineralized samples (control). Preosteoblasts (MC3T3-E1) were cultured on the bone samples for 24, 48, and 72 hours (n = 3). Fifteen samples removed from the remaining three animals were analyzed by scanning electron microscopy/energy dispersive spectrometry (SEM/EDS) after demineralization (n = 3). RESULTS The number of preosteoblasts increased significantly with time in all groups. The bone surface area covered by these cells increased with time, except in the control group. Intragroup differences occurred between 24 and 72 hours (P < 0.05). Samples demineralized for 30 seconds showed greater area covered by preosteoblast cells than for the other times of demineralization in all periods of cell culture (P < 0.05) without a statistically significant difference compared with 15 seconds. SEM/EDS showed diminished content of calcium (Ca) after 15 seconds of demineralization, but the Ca content increased after 180 seconds of demineralization (P < 0.05). The phosphorus (P) amount increased significantly only after 30 seconds of demineralization (P < 0.5). The sulfur (S) content was increased in demineralized samples in relation to non-demineralized ones, reaching the highest level after 90 seconds, when the difference became significant in relation to all the other times of demineralization (P < 0.05). Magnesium (Mg) content did not differ significantly between demineralized and non-demineralized samples. CONCLUSIONS Bone surfaces demineralized for 30 seconds increased the spreading of preosteoblasts as well as the surface area covered by these cells. Bone demineralization deserves to be studied in periodontal and maxillofacial regenerative procedures.