Instructions for the use of L-PRF in different clinical indications.

Autologous platelet concentrates (APCs) have demonstrated clear benefits across various clinical applications, including alveolar ridge preservation, guided tissue regeneration, guided bone regeneration, sinus floor elevation (both lateral window approach and transcrestal technique), endodontic surgery, the treatment of medication-related osteonecrosis of the jaw bones, and periodontal plastic surgery. To ensure an optimal clinical outcome, clinicians must adhere strictly to the protocol to prepare the APCs and, especially follow evidence-based surgical guidelines, often simple but crucial, to minimize the likelihood of errors. The majority of clinical trials reported on second-generation APCs [the leukocyte- and platelet-rich fibrin (L-PRF) family, including its modifications (A-PRF, A-PRF+, CGF, T-PRF, H-PRF, etc.)]. These second-generation APCs offer additional benefits compared to the first-generation APCs, making them the preferred choice for the development of clinical recommendations. These recommendations have been formulated through a meticulous examination of the available clinical data and the clinical experience of the authors of this paper.

The effect of experimental diabetes and membrane occlusiveness on guided bone regeneration: A proof of principle study.

OBJECTIVES: To evaluate the effect of membrane occlusiveness and experimental diabetes on early and late healing following guided bone regeneration. MATERIAL AND METHODS: A total of 30 Wistar rats were randomly allocated to three groups: healthy (H), uncontrolled diabetic (UD) and controlled diabetic (CD). A critical size calvarial defect (CSD) was created at the mid-portion of one parietal bone, and it was treated with a double layer of e-PTFE membrane presenting 0.5 mm perforations. The animals were killed at 7 and 30 days of healing, and qualitative and quantitative histological evaluations were performed. Data were compared with the ones previously obtained from other 30 animals (10H, 10UD, 10 CD), where two CSDs were randomly treated with a double-layer e-PTFE occlusive membrane or left empty. RESULTS: Following application of cell occlusive or cell permeable membranes, significant regeneration can be observed. However, at 30 days in the H group occlusive compared to cell permeable membranes promoted enhanced bone regeneration (83.9 ± 7.3% vs. 52.5 ± 8.6%), while no significant differences were observed within the CD and UD groups. UD led to reduced regeneration compared to H when an occlusive barrier was applied, whereas comparable outcomes to H and CD were observed when placing perforated membranes. CONCLUSION: The application of cell permeable membranes may have masked the potentially adverse effect of experimental UD on bone regeneration. CLINICAL RELEVANCE: Membrane porosity might contribute to modulate the bone regenerative response in UD conditions. Future studies are needed to establish the degree of porosity associated with the best regenerative outcomes as well as the underlying molecular mechanisms.

Patterns of subgingival microbiota in different periodontal phenotypes.

OBJECTIVES: To compare the subgingival microbiota of patients with aggressive (AgP) or chronic periodontitis (CP) to healthy (H), non-periodontitis patients as well as to explore their relevant associations to different host genetic variants. METHODS: Following clinical examination, blood and subgingival plaque sampling of 471 study participants (125 AgP, 121 CP, 225 H), subgingival community analysis was performed by next generation sequencing of the 16S rRNA. Microbial data from 266 participants (75 AgP, 95 CP, 98 H) were available for analysis. SNPs in the IL6, IL6R and FTO gene were selected for genetic marker analyses. RESULTS: Combined periodontitis patients (AgP + CP), particularly those classified with AgP, exhibited lower alpha- and beta- diversity. Several genera (including Peptostreptococcaceae, Filifactor, Desulfobulbus, Tannerella and Lachnospiracee) and species were over-abundant in combined periodontitis vs. healthy individuals, while other genera such as Prevotella or Dialister were found to be more abundant in healthy cases. The only genus with difference in abundance between AgP and CP was Granulicatella. No associations between IL6, IL6RA and FTO genetic variants and microbial findings were detected. CONCLUSION: This study suggests that limited microbial differences existed between AgP and CP and challenges the current notion that periodontitis is associated with increased subgingival microbial diversity compared with periodontal health. CLINICAL SIGNIFICANCE: The findings of this study cast some doubts on the notion that the dysbiosis characteristic of periodontal disease is expressed as increased microbial diversity.

Periodontal status of children with primary immunodeficiencies: a systematic review.

OBJECTIVE: The aim of this systematic review was to appraise the existing literature on periodontal disease in children affected by different types of neutrophil-associated primary immunodeficiencies (PIDs). METHODS: A PRESS-validated search strategy was developed to search through databases MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, LILACS, Google Scholar and Open Grey. All included studies were assessed for methodological quality and risk of bias. RESULTS: One hundred eighteen articles reporting on 160 PID patients were included for qualitative analysis. The majority (70%) were individual case reports. Clinical and radiographic manifestations of the periodontal disease included poor oral hygiene, generalised alveolar bone loss, severe gingival inflammation, increased pocket depths, tooth mobility and gingival recession. For most studies, the primary intervention was periodontal treatment in the form of scaling and root planing or dental extractions. Stabilisation of the periodontal condition varied between different PIDs. In severe congenital neutropenia (SCN), 61% of cases reported stabilisation of the periodontal condition, while for all other PIDs, 'stability' was reported in less than 43% of cases. CONCLUSION: The published literature suggests that patients with PIDs can present with severe periodontitis and that conventional treatment approaches have limited benefits.

Expression of gingival crevicular fluid markers during early and late healing of intrabony defects after surgical treatment: a systematic review.

BACKGROUND: Surgical treatments such as guided tissue regeneration (GTR) and access flap surgery are widely employed for the treatment of intrabony defects. However, little is known regarding the postoperative expression of gingival crevicular fluid (GCF) markers. OBJECTIVE: The aim of this systematic review was to compare the expression of GCF markers following treatment of periodontal intrabony defects with guided tissue regeneration or access surgery. The association of the markers' expression with the clinical outcome was also assessed. METHODS: An electronic literature search was conducted in MEDLINE, EMBASE, OpenGrey, LILACS and Cochrane Library up to December 2018 complemented by a manual search. Human, prospective clinical studies were identified. The changes from baseline up to 30 days (early healing) and 3 months (late healing) were assessed. RESULTS: A total of 164 publications were identified and reviewed for eligibility. Of these, 10 publications fulfilled the inclusion criteria. The included studies evaluated 15 different GCF markers with a follow-up time between 21 and 360 days postoperatively. PDGF, VEGF and TIMP-1 changes were often investigated in the included studies; however, contrasting results were reported. Two studies agreed that both GTR and OFD lead to similar OPG level changes. TGF-ß1 is increased early postoperatively, irrespective of the surgical technique employed. CONCLUSION: There is limited evidence available on the expression of GCF markers after surgical interventions of intrabony periodontal defects. However, OPG and TGF-ß1 tend to increase early post-operatively, irrespective of the surgical technique employed, irrespective of the surgical technique employed. CLINICAL RELEVANCE: More well-designed, powered studies with sampling periods reflecting the regenerative process are needed, and future research should focus on employing standardised protocols for collecting, storing and analysing GCF markers.

Minimally invasive non-surgical vs. surgical approach for periodontal intrabony defects: a randomised controlled trial.

BACKGROUND: Periodontal intrabony defects are usually treated surgically with the aim of increasing attachment and bone levels and reducing risk of progression. However, recent studies have suggested that a minimally invasive non-surgical therapy (MINST) leads to considerable clinical and radiographic defect depth reductions in intrabony defects. The aim of this study is to compare the efficacy of a modified MINST approach with a surgical approach (modified minimally invasive surgical therapy, M-MIST) for the treatment of intrabony defects. METHODS: This is a parallel-group, single-centre, examiner-blind non-inferiority randomised controlled trial with a sample size of 66 patients. Inclusion criteria are age 25-70, diagnosis of periodontitis stage III or IV (grades A to C), presence of ≥ 1 'intrabony defect' with probing pocket depth (PPD) > 5 mm and intrabony defect depth ≥ 3 mm. Smokers and patients who received previous periodontal treatment to the study site within the last 12 months will be excluded. Patients will be randomly assigned to either the modified MINST or the M-MIST protocol and will be assessed up to 15 months following initial therapy. The primary outcome of the study is radiographic intrabony defect depth change at 15 months follow-up. Secondary outcomes are PPD and clinical attachment level change, inflammatory markers and growth factors in gingival crevicular fluid, bacterial detection, gingival inflammation and healing (as measured by geometric thermal camera imaging in a subset of 10 test and 10 control patients) and patient-reported outcomes. DISCUSSION: This study will produce evidence about the clinical efficacy and potential applicability of a modified MINST protocol for the treatment of periodontal intrabony defects, as a less invasive alternative to the use of surgical procedures. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03797807. Registered on 9 January 2019.

Osseointegration in osteoporotic-like condition: A systematic review of preclinical studies.

Osteoporosis is one of the most common skeletal disorders affecting a significant percentage of people worldwide. Research data suggested that systemic diseases such as osteoporosis could act as risk factors for osseointegration, jeopardizing the healing process and thus the predictability of dental implant success on compromised patients. It is well accepted that preclinical studies in animal models reproducing the osteoporotic condition are one of the most important stages in the research of new biomaterials and therapeutic modalities. The aim of this systematic review was to investigate whether osteoporosis compromises dental implant osseointegration in experimental osteoporotic-like conditions. A 3-stage systematic literature research was conducted in MEDLINE via OVID and EMBASE up to and including March 2017. Experimental studies reporting on dental implant osseointegration on different osteoporotic animal models were assessed. The studies had to report on the percentage of bone-to-implant contact (%BIC) as the primary outcome. ARRIVE guidelines for reporting on animal research were applied to evaluate the methodological quality and risk of bias of the studies. Fifty-seven studies met the inclusion criteria and were assessed qualitatively. The most adopted animal model was the rat. A variability of %BIC values was observed, ranging from 30% to 99% and from 26% to 94% for the healthy and osteoporotic group, respectively. The great majority (47) of the included studies concluded that estrogen deficiency significantly affects BIC values, 9 studies stated that it was not possible to observe statistical differences in BIC between ovariectomized and healthy groups and 1 study did not provide a comparison between the healthy and osteoporotic group. Owing to the great heterogeneity in implant surface, study design, observation time-points, site of implant placement and reported outcomes, a meta-analysis could not be performed. An overall high risk of bias was observed, owing to the limited information on animal housing and husbandry, baseline characteristics and health status, ethical statement and allocation to the experimental groups provided. Although the available studies seem to suggest a lower osseointegration in osteoporotic-like conditions, no robust conclusions can be drawn due to the great heterogeneity and overall low quality of the available studies. Future studies with emphasis on minimizing the possible sources of bias and evaluating osseointegration of dental implants placed into jawbones instead of long bones are warranted.

Pro-osteogenic properties of hydrophilic and hydrophobic titanium surfaces: Crosstalk between signalling pathways in in vivo models.

BACKGROUND: It is now generally accepted that the response to a particular signal, such as the surgical trauma following implant placement, is not the result of a single linear signalling pathway, but rather reflects pathway integration, which can occur at multiple levels. Although it is well documented that both SLA and SLActive surfaces are able to promote bone formation and osseointegration, it is still unclear which are the key signalling pathways involved and how surface hydrophilicity/hydrophobicity might affect pathway integration. OBJECTIVE: To combine gene and protein data from in vivo studies applying titanium hydrophobic (Sandblasting, Large-grit, Acid-etching, SLA) and hydrophilic (SLActive) surfaces to understand the molecular mechanisms responsible for the pro-osteogenic properties of these surfaces. METHODS: The Kyoto Encyclopedia of Genes and Genomes (KEGG® ) pathway database and the Ingenuity® Pathway Analysis (IPA® ) software were applied to the genomic and proteomic data of previous in vivo studies applying SLA and SLActive surfaces, with the specific aim to focus on bone formation-related signalling pathways. While gene data were derived from a human study on osseointegration, protein data originated from a preclinical study in rabbits. Data were available for the 4, 7 and 14 days of healing periods. RESULTS: Both genomic and proteomic data showed that the osteogenesis process takes place mainly at 7 and 14 days of healing on both SLA and SLActive surfaces. Surface hydrophilicity enhances bone formation at multiple levels, by directly promoting an earlier expression of pathways involved in cell proliferation and osteoblast precursor differentiation (eg, mitogen-activated protein kinase, phosphoinositide-3 kinase-AKT, Wnt, Notch, transforming growth factor-ß), but also by positively regulating angiogenesis, bone mineralization and bone remodelling. CONCLUSION: This study combined, for the first time, different 'omics' outputs to get new insights on the molecular mechanisms behind the influence of surface hydrophilicity on osseointegration/bone formation. Specific signalling pathways, such as Wnt, vascular endothelial growth factor and mitogen-activated protein kinase, were identified as differentially modulated by titanium surface hydrophilicity both at a genomic and proteomic level. These findings may be used in the future to monitor/predict the bone formation/osseointegration process, or as a screening tool towards the manufacture of new pro-osteogenic implant surfaces. In order to take into account the full complexity and interplay of cell signalling during bone formation, powerful bioinformatics tools integrating different 'omics' data and predicting signalling pathways trends should be applied by future studies.

Degradation pattern of a porcine collagen membrane in an in vivo model of guided bone regeneration.

BACKGROUND AND OBJECTIVE: Although collagen membranes have been clinically applied for guided tissue/bone regeneration for more than 30 years, their in vivo degradation pattern has never been fully clarified. A better understanding of the different stages of in vivo degradation of collagen membranes is extremely important, considering that the biology of bone regeneration requires the presence of a stable and cell/tissue-occlusive barrier during the healing stages in order to ensure a predictable result. Therefore, the aim of this study was to investigate the degradation pattern of a porcine non-cross-linked collagen membrane in an in vivo model of guided bone regeneration (GBR). MATERIAL AND METHODS: Decalcified and paraffin-embedded specimens from calvarial defects of 18, 10-month-old Wistar rats were used. The defects were treated with a double layer of collagen membrane and a deproteinized bovine bone mineral particulate graft. At 7, 14 and 30 days of healing, qualitative evaluation with scanning electron microscopy and atomic force microscopy, and histomorphometric measurements were performed. Markers of collagenase activity and bone formation were investigated using an immunofluorescence technique. RESULTS: A significant reduction of membrane thickness was observed from 7 to 30 days of healing, which was associated with progressive loss of collagen alignment, increased collagen remodeling and progressive invasion of woven bone inside the membranes. A limited inflammatory infiltrate was observed at all time points of healing. CONCLUSION: The collagen membrane investigated was biocompatible and able to promote bone regeneration. However, pronounced signs of degradation were observed starting from day 30. Since successful regeneration is obtained only when cell occlusion and space maintenance exist for the healing time needed by the bone progenitor cells to repopulate the defect, the suitability of collagen membranes in cases where long-lasting barriers are needed needs to be further reviewed.

The use of omics profiling to improve outcomes of bone regeneration and osseointegration. How far are we from personalized medicine in dentistry?

Increased life expectancy and broader restorative dental treatment alternatives for missing teeth have resulted in an increasing request of bone regeneration/augmentation procedures not only in healthy patients, but also in elderly and medically compromised ones. This is also combined with a growing demand for short implant loading protocols and for optimal aesthetic results. In order to meet these new dental needs, personalized treatment strategies tailored on each individual's characteristics and healing profile are warranted. Omics technologies are emerging as powerful tools to uncover molecules and signalling pathways involved in bone formation and osseointegration and to investigate differences in the molecular mechanisms between health and systemic diseases that could be targeted by future therapies. This review critically appraises the available knowledge on the application of omics technologies in the field of bone regeneration and osseointegration and explores their potential use for personalized medicine in the dento-maxillo-facial field. SIGNIFICANCE: The use of omics in personalising dental maxillo-facial treatments emerges as a desirable diagnostic and treatment strategy. Omics represent, in fact, powerful tools not only to shade light on the cascade of events taking place during bone formation/osseointegration, but also to identify specific signalling pathways and molecules that can be targeted by future therapies with the aim to enhance clinical outcomes in patients with compromised healing conditions.

Protein expression during early stages of bone regeneration under hydrophobic and hydrophilic titanium domes. A pilot study.

BACKGROUND AND OBJECTIVES: There is significant evidence that, during the early stages of osseointegration, moderately rough hydrophilic (SLActive) surfaces can accelerate osteogenesis and increase bone-to-implant contact in comparison to hydrophobic (SLA) surfaces. However, very little is known regarding the molecular mechanisms behind the influence that surface chemistry modifications to increase hydrophilicity determine on bone healing. The aim of this study was to describe for the first time the proteins and related signalling pathways expressed during early osseous healing stages under SLA and SLActive titanium domes for guided bone regeneration. MATERIAL AND METHODS: One SLA and 1 SLActive dome with an internal diameter of 5.0 mm and a height of 3.0 mm were secured to the parietal bones of nine 6-month-old male New Zealand rabbits. Three animals were randomly euthanized at 4, 7 and 14 days and the newly formed tissues retrieved under the domes were analysed with liquid chromatography-mass spectrometry/mass spectrometry. STRING and KEGG databases were applied for Gene Ontology and pathway analyses. RESULTS: A different modulation of several pathways was detected between the 2 groups at all healing times. The main differences in the osseous healing response associated to the 2 surfaces were related to pathways involved in regulating the inflammatory response, differentiation of osteoblast precursors and skeletogenesis. At day 7, the highest number of proteins and the highest cellular activity were observed in both groups, although a more complex and articulated proteome in terms of cellular metabolism and signal transduction was observed in SLActive samples. CONCLUSION: This is the first study describing the proteome expressed during early healing stages of guided bone regeneration and osseointegration. A combination of enhanced early osteogenic response and reduced inflammatory response were suggested for the hydrophilic group. Future studies are needed to corroborate these findings and explore the molecular effects of different titanium surfaces on the cascade of events taking place during bone formation.

The effect of experimental diabetes and glycaemic control on guided bone regeneration: histology and gene expression analyses.

OBJECTIVES: To investigate the effect of experimental diabetes and metabolic control on intramembranous bone healing following guided bone regeneration (GBR). MATERIAL AND METHODS: Ninety-three Wistar rats were allocated to three experimental groups, healthy (H), uncontrolled diabetes (D) and controlled diabetes (CD). Twenty one days following diabetes induction, a standardised 5-mm defect was created at the mid-portion of each parietal bone. In 75 animals (25H, 25D, 25CD), one defect was treated with an intracranial and extracranial membrane according to the GBR principle, and one defect was left empty (control); five animals per group were then randomly sacrificed at 3, 7, 15, 30 and 60 days and processed for decalcified histology. In 18 animals (6H, 6D, 6CD), both defects were treated according to the GBR principle; three animals from each group were then randomly sacrificed at 7 and 15 days of healing and employed for gene expression analysis. RESULTS: Application of the GBR therapeutic principle led to significant bone regeneration even in the D group. However, at 15 and 30 days, the osteogenesis process was impaired by uncontrolled diabetes, as shown by the significant reduction in terms of defect closure (38-42%) and newly formed bone (54-61%) compared to the healthy group. The comparison of the D vs. H group at 15 days of healing yielded the largest number of genes with significantly differential expression, among which various genes associated with the ossification process (bmp4, ltbp4, thra and cd276) were identified. CONCLUSIONS: Uncontrolled diabetes seems to affect early phases of the bone regeneration following GBR. A misregulation of genes and pathways related to cell division, energy production, inflammation and osteogenesis may account for the impaired regeneration process in D rats. Further studies are warranted to optimise the GBR process in this medically compromised patient population.

Effect of Wnt3a delivery on early healing events during guided bone regeneration.

OBJECTIVE: To evaluate the effect of recombinant Wnt3a delivery on the bone regeneration potential following application of the guided bone regeneration (GBR) principle. MATERIALS AND METHODS: A critical-size calvarial defect was created on each parietal bone of 14 Wistar strain rats. One defect was used as the test side and was treated with a collagen sponge carrying 2.0 µg of recombinant Wnt3a protein, whereas the contralateral side served as sham-operated control. Both defects were covered at both the extracranial and intracranial aspects with ePTFE non-resorbable membranes, according to the GBR principle. Following healing periods of 4 and 7 days, qualitative histological and histomorphometric evaluation of undecalcified sections was performed in subgroups of seven animals. The primary outcome parameter was the mean percentage of defect closure in the test and control defects. RESULTS: At 4 days of healing, a network of coagulum and fibrin was observed and initial signs of granulation tissue formation were evident with no apparent differences between the test and control groups. At 7 days of healing, the test group presented newly formed woven bone, originating from the borders of the defect, as opposed to the control group, whereby woven bone formation was not observed in any of the specimens. CONCLUSIONS: The delivery of mouse recombinant Wnt-3a protein in combination with GBR may promote woven bone formation in critical-size calvarial defects at 7 days of healing.

Prevalence and treatment of necrotizing ulcerative gingivitis (NUG) in the British Armed Forces: a case-control study.

OBJECTIVES: Necrotizing ulcerative gingivitis (NUG) has been seen in military populations throughout history. This study aims to determine the prevalence, treatment modality and risk factors associated with NUG in the British Armed Forces. MATERIALS AND METHODS: A whole population dataset of the British Armed Forces was searched to determine cases of NUG during the period 1 January to 31 December 2012. Individual case records were identified, and a case-control study undertaken with data gathered and analysed against a randomised control group, matched for age, sex and service. RESULTS: A prevalence rate for NUG of 0.11 % was determined against the whole military population. The majority of cases received (alone or in combination) the following: oral hygiene instruction (66.5 %), antibiotics (64.4 %) and a mouthwash (58.1 %). Of the cases, 48.7 % received debridement. Analgesics were only prescribed in 8.4 % of the cases, and smoking cessation advice was only given in 10.7 % of the cases. Analysis of risk factors against the control group showed an increase in odds ratios for diagnosis of NUG of 3.4 (95 % CI 2.0-5.7) for current smokers and 7.3 (95 % CI 1.9-28.0) for individuals with an overall Basic Periodontal Examination (BPE) score of 3. CONCLUSIONS: Whilst NUG is a rare disease, it is evident from this study that it still occurs within the British Armed Forces. A strong association was shown between NUG and current smokers and those cases with an overall BPE score of 3. CLINICAL RELEVANCE: This study provides prevalence data for NUG in the British Armed Forces and description of its treatment and associated risk factors. Oral hygiene and smoking must be addressed in patients with NUG and prescribing protocols should be carefully followed.

Microarray gene expression during early healing of GBR-treated calvarial critical size defects.

OBJECTIVES: To investigate the gene expression and molecular pathways implicated in the regulation of the osseous healing process following guided bone regeneration (GBR). MATERIAL AND METHODS: Six 6-month-old Wistar male rats were used. Standardized 5-mm critical size defects were created in the parietal bones of each animal and treated with an extracranial and intracranial ePTFE membrane, according to the GBR principle. Three animals were randomly sacrificed after 7 and 15 days of healing. Total RNA was extracted from each sample and prepared for gene expression analysis. RNA quality and quantity were assessed, followed by hybridization of the cRNA to Affymetrix GeneChip Rat Genome 230 2.0 Arrays. The Affymetrix data were processed, and first-order analysis, quality control and statistical analysis were performed. Biological interpretation was performed via pathway and Gene Ontology (GO) analysis. RESULTS: Between the 7- and 15-day samples, 538 genes were differently regulated. At day 7, inflammatory and immune responses were clearly upregulated. In addition, GO terms related to angiogenesis and cell cycle regulation were overexpressed. At day 15, a more complex cellular activity and cell metabolism were evident. The bone formation processes were significantly overexpressed, with several genes encoding growth factors, enzyme activity, and extracellular matrix formation found as upregulated. Remarkably, a negative regulation of Wnt signalling pathway was observed at 15 days. DISCUSSION: The gene expression profile of the cells participating in osseous formation varied depending on the healing stage. A number of candidate genes that seem differentially expressed during early stages of intramembranous bone regeneration was suggested.

The effect of experimental osteoporosis on bone regeneration: part 2, proteomics results.

OBJECTIVES: To identify and describe protein expression in a Wistar rat calvarial critical size defect (CSD) model following treatment with guided bone regeneration in healthy and osteoporotic conditions. MATERIAL AND METHODS: Thirty-six 10-month-old female Wistar rats were used. Half of them were ovariectomized (OVX) and fed with a low-calcium diet to induce an osteoporotic-like status. In each animal of both groups, two 5-mm calvarial CSDs were treated with deproteinized bovine bone mineral graft particles and a bilayer collagen membrane. Six OVX and six control rats were randomly euthanized at 7, 14, and 30 days. One defect/animal was randomly chosen for proteomic analysis. Differently expressed proteins between the two groups were identified with matrix-assisted laser desorption time-of-flight mass spectrometry and liquid chromatography-mass spectrometry/mass spectrometry. RESULTS: At 7 days, 29 and 27 proteins were, respectively, identified in the healthy and OVX animals. At 14 days, 103 proteins were detected in the healthy controls and 20 proteins in the OVX rats, while at 30 days, 31 and 75 proteins were identified, respectively. Only limited proteins known to play a role in the later stages of bone formation and maturation were identified within the animals 'proteomes. DISCUSSION: The osseous formation process was quite immature even at 30 days of healing. An overexpression of inflammatory and stress response pathways was detected in the OVX animals, as well as a tendency toward a delayed maturation of the osseous wound and a reduced/delayed differentiation of osteoblast cell precursors.

Leukocyte receptor expression in chronic periodontitis.

BACKGROUND AND OBJECTIVE: Microbial recognition in the periodontium through specific leukocyte receptors gives rise to the response which in susceptible individuals can lead to periodontal diseases. The aim of this study was to explore the expression of leukocyte receptors in the gingival tissues of chronic periodontitis patients and to analyse differences between diseased and control sites (sites with probing pocket depth <4 mm). MATERIAL AND METHODS: Thirty-seven chronic periodontitis patients were included in the study. Gingival biopsies were harvested from diseased and control sites and processed by flow cytometry for the determination of the expression of 16 leukocyte receptors (CD4, CD8, CD11b, CD14, CD16, CD19, CD25, CD28, CD49d, CD49e, CD62, CD71, CD80, CCR7, Ly6G and HLA-DR). RESULTS: Expression of all studied receptors was higher in test compared with control sites (p < 0.005). Sampled sites with less bleeding on probing exhibited higher expression of CD16 and CD14 receptors (p = 0.020 and 0.011, respectively). CONCLUSIONS: This study points towards considerable differences in the expression of leukocyte receptors between diseased and control sites in the same periodontal patients.

Nanoadhesion of Staphylococcus aureus onto Titanium Implant Surfaces.

Adhesion of bacteria to dental implant surfaces is the critical initial step in the process of biofilm colonization; however, the specific nanoadhesive interactions occurring during the first contact between bacterial cells and biomaterial substrates remain poorly understood. In this report, we utilize single-cell force spectroscopy to characterize the dynamics of the initial interaction between living Staphylococcus aureus cells and machined titanium surfaces at the nanoscale. Values for maximum adhesion force were found to increase from 0-s (-0.27 ± 0.30 nN) to 60-s (-9.15 ± 0.78 nN) surface delays, with similar results observed for total adhesion work (7.39 ± 2.38 and 988.06 ± 117.08 aJ, respectively). Single unbinding events observed at higher surface delays were modeled according to the wormlike chain model, obtaining molecular contour-length predictions of 314.06 ± 9.27 nm. Average single-bond rupture forces of -0.95 ± 0.04 nN were observed at increased contact times. Short- and long-range force components of bacterial adhesion were obtained by Poisson analysis of single unbinding event peaks, yielding values of -0.75 ± 0.04 and -0.58 ± 0.15 nN, respectively. Addition of 2-mg/mL chlorhexidine to the buffer solution resulted in the inhibition of specific adhesive events but an increased overall adhesion force and work. These results suggest that initial attachment of S. aureus to smooth titanium is mostly mediated by short-range attractive forces observed at higher surface delays.

In situ neutralisation of the antibacterial effect of 0.2% Chlorhexidine on salivary microbiota: Quantification of substantivity.

OBJECTIVE: To quantify the substantivity of a single 0.2% Chlorhexidine mouthwash in saliva after its neutralisation with tooth-brushing and 1% acetic acid, in order to identify the effect of Chlorhexidine substantivity in regard to the re-growing period of the salivary bacteria. METHODS: Unstimulated saliva samples were collected from a group of 15 healthy individuals at baseline (BS), and then 30s and 1, 3, 5 and 7h after the following protocols were performed: a single sterile water mouthwash (M-WATER) (negative control), a single 0.2% Chlorhexidine mouthwash (M-0.2% CHX) (positive control) and a single 0.2% Chlorhexidine mouthwash followed by a complete and detailed tooth-brushing, and a single 1% acetic acid mouthwash (M-0.2% CHX+NP). The samples were analysed using an epifluorescence microscope in combination with LIVE/DEAD(®) BacLight™ fluorescence solution. RESULTS: After the M-0.2% CHX treatment, the bacterial vitality was significantly lower than BS until 7h (87.6 ± 6.5% vs. 73.6 ± 8.8%; p<0.001). However, after M-0.2% CHX+NP, the bacterial vitality remained significantly lower until 3h with regard to BS (81.4 ± 3.8% vs. 68.1 ± 10.6%; p=0.001), increasing at 5 and 7h (no differences from BS). CONCLUSION: The immediate antibacterial effect of a single 0.2% Chlorhexidine mouthwash is so potent that the bacterial population needs more than 3h to return to baseline bacterial vitality levels. The substantivity of a 0.2% Chlorhexidine mouthwash is a property that significantly increases its antibacterial activity from the first hour and contributes to extend the duration of its effect by at least double.

Lipid subclasses profiles and oxidative stress in aggressive periodontitis before and after treatment.

BACKGROUND AND OBJECTIVE: Associations between dyslipidaemia, oxidative stress and periodontitis have emerged in recent years. However, there is a lack of studies investigating these associations in aggressive periodontitis (AgP) cases. The aim of this study was to investigate the lipid and oxidative stress profiles in patients with AgP, and to relate them to clinical variables and interleukin (IL)-6 genetic variants. MATERIAL AND METHODS: Twelve non-smoking Caucasian patients with AgP selected based on their IL6 haplotypes underwent periodontal non-surgical and surgical treatment. Peripheral blood samples taken at baseline and at six different time-points after treatment were processed to determine IL-6 circulating levels, lipid profiles (cholesterol, triglycerides, high-density lipoprotein [HDL] and low-density lipoprotein [LDL] subclasses) and oxidative stress markers (glutathione and total lipid hydroperoxide levels). RESULTS: HDLs were the most prevalent lipoproteins, followed by intermediate-density lipoprotein, very-low-density lipoprotein and LDL. The LDL subclasses consisted mainly of the less atherogenic large LDL. The lipid profile did not consistently change after treatment up to 3 mo after surgery. Periodontal disease severity was associated with LDL levels and size. The IL6 haplotypes were associated with total cholesterol, triglycerides, HDL and LDL subclasses after adjusting for confounders. IL-6 circulating levels were associated with both very-low-density lipoprotein and lipid hydroperoxide levels. CONCLUSION: Based on these data, we conclude that both periodontal disease severity and IL6 haplotypes may influence lipid profiles in AgP.