Assessment of the anti-inflammatory and biological properties of Bioroot Flow: A novel bioceramic sealer.

INTRODUCTION: BioRoot Flow (BRF) is a novel premixed bioceramic sealer indicated for endodontic treatments, but the biological and immunomodulatory effects of this endodontic sealer on human periodontal ligament stem cells (hPDLSCs) have not been elucidated. METHODS: To ascertain the biological impact of BRF, TotalFill BC Sealer (TFbc), and AH Plus (AHP) on human Periodontal Ligament Stem Cells (hPDLSCs), assessments were conducted to evaluate the cytocompatibility, cellular proliferation, migratory capacity, osteo/cementogenic differentiation potential, the ability to form mineralized nodules, and the immunomodulatory characteristics of hPDLSCs following treatment with these endodontic sealers. RESULTS: Biological assays showed adequate cell metabolic activity and cell migration in BRF, while SEM assay evidenced that TFbc and BRF groups demonstrated a superior cell adhesion process, including substrate adhesion, cytoskeleton development, and spreading on the niche-like structures of the cement as compared to the AHP group. TFbc and BRF-treated groups exhibited a significantly lower IL6 and IL8 production than AHP (* p <.05). The bioceramic sealers stimulated heightened expression of BSP, CEMP-1, and CAP genes within a 7-14 day period. Notably, BRF and TFbc demonstrated a significant enhancement in the mineralization of hPDLSCs when compared to the negative control. Among these, cells treated with BRF showed a more substantial accumulation of calcium (*** p < .001). CONCLUSIONS: Taken together, these findings indicate that BRF can potentially enhance cell differentiation by promoting the expression of essential genes related to bone and cement formation. In addition, BRF and TFbc displayed anti-inflammatory effects.

Radiological assessment of periodontal ligament space visibility on third molars for forensic age assessment - a comparison study of three different staging scales.

Various staging scales have been proposed for the assessment of the visibility of the periodontal ligament space of mandibular third molars on dental panoramic radiographs (PANs) for forensic age assessment in living individuals. However, up to now, there has been no systematic comparison between these staging scales available. We directly compared the 2010 staging scale proposed by Olze et al. with the 2017 staging by Lucas et al. and the 2020 staging by Guo et al. in a German study population. We evaluated 233 PANs from 115 females and 118 males aged 20.0 to 40.9 years using three independent examiners, with one examiner conducting two assessments. We examined the correlation between age and stage, as well as the inter- and intra-rater reliabilities. While the point estimates for the correlation coefficient and the reliability measures were lowest for the Guo scale and highest for the Olze scale, confidence intervals showed a large overlap, particularly for the scales of Olze et al. and Lucas et al. The correlation coefficients between stage and age were consistently lower in females than in males across all methods. In summary, we showed that the staging scales of Olze et al. and Lucas et al. were very similar. The Olze method showed higher point estimates across all analyses, and because there are more reference data available for this method, we argue that it should be preferred as the method of choice for further studies in the field. However, Guo method could be considered for instances, in which the inter-radicular periodontal ligament is not evaluable.

Use of ultrasound imaging for assessment of the periodontium: A systematic review.

The objective of this study was to systematically review the literature regarding diagnostic applications of ultrasound imaging for evaluation of the periodontium in humans. The search was conducted on Medline, EMBASE, Web of Science, Scopus, Cochrane, and PubMed up to April 3, 2023. The studies included were exclusively human studies that assessed the periodontium with ultrasound (US) imaging (b-mode). Outcomes measured included alveolar bone level, alveolar bone thickness, gingival thickness, and blood flow quantification. References were imported to Covidence. Two reviewers conducted phases 1 and 2. The JBI risk assessment tool for cross-sectional studies was used. Extracted data included the transducer and measurements used and the study's outcomes. The search yielded 4892 studies after removing duplicates. From these, 25 studies were included and selected for extraction. Included studies retrieved outcomes from US examinations of the periodontal tissues. From the selected studies, 15 used US on natural teeth, 4 used US on implants, 2 used US on edentulous ridges, and 4 used color flow/power in US to evaluate the blood flow. The results of the present systematic review suggest that US might be a feasible and valuable diagnostic tool for the periodontium, with the potential to complement shortfalls of current radiographic technologies.

Cortical and Trabecular Bone Stress Assessment during Periodontal Breakdown-A Comparative Finite Element Analysis of Multiple Failure Criteria.

Background and Objectives: This numerical analysis investigated the biomechanical behavior of the mandibular bone as a structure subjected to 0.5 N of orthodontic force during periodontal breakdown. Additionally, the suitability of the five most used failure criteria (Von Mises (VM), Tresca (T), maximum principal (S1), minimum principal (S3), and hydrostatic pressure (HP)) for the study of bone was assessed, and a single criterion was identified for the study of teeth and the surrounding periodontium (by performing correlations with other FEA studies). Materials and Methods: The finite element analysis (FEA) employed 405 simulations over eighty-one mandibular models with variable levels of bone loss (0-8 mm) and five orthodontic movements (intrusion, extrusion, tipping, rotation, and translation). For the numerical analysis of bone, the ductile failure criteria are suitable (T and VM are adequate for the study of bone), with Tresca being more suited. S1, S3, and HP criteria, due to their distinctive design dedicated to brittle materials and liquids/gas, only occasionally correctly described the bone stress distribution. Results: Only T and VM displayed a coherent and correlated gradual stress increase pattern for all five movements and levels of the periodontal breakdown. The quantitative values provided by T and VM were the highest (for each movement and level of bone loss) among all five criteria. The MHP (maximum physiological hydrostatic pressure) was exceeded in all simulations since the mandibular bone is anatomically less vascularized, and the ischemic risks are reduced. Only T and VM displayed a correlated (both qualitative and quantitative) stress increase for all five movements. Both T and VM displayed rotation and translation, closely followed by tipping, as stressful movements, while intrusion and extrusion were less stressful for the mandibular bone. Conclusions: Based on correlations with earlier numerical studies on the same models and boundary conditions, T seems better suited as a single unitary failure criterion for the study of teeth and the surrounding periodontium.

Assessment of the Maximum Amount of Orthodontic Force for PDL in Intact and Reduced Periodontium (Part I).

This study examines 0.6 N and 1.2 N as the maximum orthodontic force for periodontal ligament (PDL) at multiple levels of periodontal breakdown, and the relationships with the ischemic, necrotic, and resorptive risks. Additionally, this study evaluates if Tresca failure criteria is more adequate for the PDL study. Eighty-one 3D models (from nine patients; nine models/patients) with the 2nd lower premolar and different degrees of bone loss (0-8 mm) where subjected to intrusion, extrusion, rotation, translation, and tipping movements. Tresca shear stress was assessed individually for each movement and bone loss level. Rotation and translation produced the highest PDL stresses, while intrusion and extrusion determined the lowest. Apical and middle third PDL stresses were lower than the cervical stress. In intact periodontium, the amount of shear stress produced by the two investigated forces was lower than the 16 KPa of the maximum physiological hydrostatic pressure (MHP). In reduced periodontium (1-8 mm tissue loss), the apical amount of PDL shear stress was lower than MHP for both applied forces, while cervically for rotation, translation and tipping movements exceeded 16 KPa. Additionally, 1.2 N could be used in intact periodontium (i.e., without risks) and for the reduced periodontium only in the apical and middle third of PDL up to 8 mm of bone loss. However, for avoiding any resorptive risks, in the cervical third of PDL, the rotation, translation, and tipping movements require less than 0.2-0.4 N of force after 4 mm of loss. Tresca seems to be more adequate for the study of PDL than other criteria.

Assessment of the Best FEA Failure Criteria (Part I): Investigation of the Biomechanical Behavior of PDL in Intact and Reduced Periodontium.

The accuracy of five failure criterions employed in the study of periodontal ligaments (PDL) during periodontal breakdown under orthodontic movements was assessed. Based on cone-beam computed tomography (CBCT) examinations, nine 3D models of the second lower premolar with intact periodontium were created and individually subjected to various levels of horizontal bone loss. 0.5 N of intrusion, extrusion, rotation, tipping, and translation was applied. A finite Elements Analysis (FEA) was performed, and stresses were quantitatively and qualitatively analyzed. In intact periodontium, Tresca and Von Mises (VM) stresses were lower than maximum physiological hydrostatic pressure (MHP), while maximum principal stress S1, minimum principal stress S3, and pressure were higher. In reduced periodontium, Tresca and VM stresses were lower than MHP for intrusion, extrusion, and the apical third of the periodontal ligament for the other movements. 0.5 N of rotation, translation and tipping induced cervical third stress exceeding MHP. Only Tresca (quantitatively more accurate) and VM are adequate for the study of PDL (resemblance to ductile), being qualitatively similar. A 0.5 N force seems safe in the intact periodontium, and for intrusion and extrusion up to 8 mm bone loss. The amount of force should be reduced to 0.1-0.2 N for rotation, 0.15-0.3 N for translation and 0.2-0.4 N for tipping in 4-8 mm periodontal breakdown. S1, S3, and pressure criteria provided only qualitative results.

Assessment of cell phone effect on dental socket healing in rat.

BACKGROUND: Electromagnetic waves can cause biological effects on repair process. Due to the proximity of the jawbone and the soft tissue around it in a part of the face with which it has the closest contact during the cell phone use, this study aims to investigate the effect of mobile waves on socket healing after tooth extraction in rats. METHODS: This experimental study was conducted on 32 rats. The case group was exposed to a 900 MHz frequency electromagnetic field for 30 min/d. Sacrificed eight rats from the case group and 7 rats from the control group on day 14, and 8 rats from the case group and 7 rats from the control group were sacrificed on day 28 at the end of the radiation period and CBCT and microscopic examinations on the maxillary bones and soft tissue were performed. RESULTS: According to the findings, the healing process was significantly different in two groups in terms of the percentage of new bone formation on day 14 after the end of radiation (P=0.014). The other measured parameters including the degree of inflammation, thickness of the formed bone, number of osteoblasts and Gray Scale had no significant difference between the two groups in any of the 14-day and 28-day intervals. CONCLUSIONS: The results of this study showed that intermittent exposure to high frequency electromagnetic fields over a period of 20 hours has no significant effect on the healing process of alveolar socket after tooth extraction in rats.

Assessment of proliferation, clonogenic assay, and osteogenic differentiation of human periodontal ligament stem cells following application of orthodontic forces.

Context: The proliferation and differentiation of human periodontal ligament stem cells (hPDLSC) into other cell types are also mediated by mechanical stresses; they might offer therapeutic benefits in tissue regeneration and angiogenesis. Objectives: The study was planned to assess the proliferation, clonogenic potential, and osteogenic differentiation of human periodontal ligament stem cells (PDLSC) following the application of light and heavy orthodontic forces. Materials and Methods: A couple forces of 50 gm (light force) were applied on the 1st premolar on the one side and 250 gm (heavy force) on the contralateral side in the upper arch of patients requiring orthodontic treatment with extraction of all 1st premolars. After 30 days, periodontal tissues were scrapped from extracted teeth for the establishment of PDLSC in vitro. PDLC from the lower premolar teeth where no orthodontic force was applied acted as the control group. Morphology, viability, proliferating rate and population doubling time, clonogenicity, and alkaline phosphatase activity were analysed. Result: The osteogenic potential was confirmed by Alizarin red staining and the expression of the osteogenic markers by qRT-PCR. The morphology, growth kinetics, potency, and osteogenic lineage characteristics inferred the application of high force reduced the proliferative ability and osteogenesis of PDLSC, though the difference was not significant. Conclusion: The established PDLSCs demonstrated their MSC-like properties based on morphology, growth kinetics, colony forming ability, and AP activity. The culture-expanded PDLSCs showed their differentiation potential into osteocytes. The application of high force reduced the proliferative ability and osteogenesis of PDLSCs, variations were not significant.

Dental age assessment based on the radiographic visibility of the periodontal ligament in lower third molars in a Thai sample.

The objective of the present study was to analyze the radiographic visibility of the periodontal ligament in completed root formation lower third molars in a sample of lower northern Thai population. Digital panoramic images from 800 patients with ages ranging from 16 to 26 years were used in this study. The visibility status of the periodontal ligament of lower third molars with completed root formation including apical closure was assessed. For each stage, the minimum age, maximum age, median, mean, and standard deviation were calculated. The minimum age found in stage 0 was 16.17 years in males and 17.00 years in females. Stage 1 was first achieved at the age of 16.17 years in males and 17.08 years in females. The earliest onset of stage 2 was 17.00 years in males and 18.17 years in females. The incidence of stage 3 was first observed at 19.17 years in males and 18.83 years in females. It may be concluded that the radiographic visibility of the periodontal ligament in lower third molars may be a useful approach in the dental age assessment in a Thai population. In case the periodontal ligament visibility is found to be in stage 2, it may be confirmed that the individual is at least 18 years of age.

Comparative assessment of the rate of orthodontic tooth movement in adolescent patients undergoing treatment by first bicuspid extraction and en mass retraction, associated with low-frequency mechanical vibrations in passive self-ligating and conventional brackets: A randomized controlled trial.

BACKGROUND: Low-frequency vibrations are one of the many non-surgical modalities aimed at increasing the rate of orthodontic tooth movement. OBJECTIVE: The present trial was conducted to assess the efficacy of low-frequency vibrations in increasing the rate of orthodontic tooth movement in adolescent patients undergoing fixed mechanotherapy with passive self-ligating brackets and conventional brackets. MATERIALS AND METHODS: Setting and sample population: department of orthodontics and dentofacial orthopaedics in a nationally accredited dental college. Participants, study design and methods: 65 patients were randomly allocated to three groups. Two experimental groups consisted of passive self-ligating and conventionally ligated appliances received low-frequency vibrations. The control group did not receive any vibrations. Allocation ratio was 1:1:1.32. Eligibility criteria: adolescent patients with sound and healthy dentition, incisor irregularity<5mm. PRIMARY OUTCOME: rate of orthodontic tooth movement in mm/month. Randomization and blinding: computer-generated random allocation sequencing was done and data assessor was blinded. STATISTICS: the Q-Q plot and Shapiro-Wilks test judged the normality of the data. The parametric test included ANCOVA and post-hoc analysis. RESULTS: No statistically significant enhancement of tooth movement was seen in the experimental groups, when comparison was done with the control group P>0.05. Comparison between the two experimental groups did not reveal any significant difference either. CONCLUSION: No statistically significant increase of orthodontic tooth movement was seen with low-frequency vibrations and the mode of ligation did not have any effect in increasing the rate of tooth movement either.

Electromechanical impedance measurements for bone health monitoring through teeth used as probes of a Piezo-device.

Bone is a dynamic biological tissue that acts as the primary rigid support of the body. Several systemic factors are responsible for pathologies that negatively affect its structural attributes. Although the bone is in continuous renewal by osteogenesis, metabolic diseases are the most common affectations that alter its natural equilibrium. Different techniques based on ionizing radiation are used for the bone diagnosis restrictively. However, if these are not used adequately, the application could present risks for human health. In this paper, it is proposed and explored a new technique to apply an early-stage diagnosis of bone variations. The technique evaluates bone structural conditions from the teeth (used as probes) by applying a structural health monitoring (SHM) methodology. An experimental procedure is described to identify the stiffness variations produced by mechanical drillings done in prepared bone samples. The identification is carried out applying the electromechanical impedance technique (EMI) through a piezo-actuated device in the frequency spectrum 5-20kHz. Three bone samples with incorporated teeth (three teeth, two teeth, and one tooth) were prepared to emulate a mandibular portion of alveolar bone-PDL (periodontal ligament)-tooth system. Piezo-device was attached to the crown of the tooth with an orthodontic bracket allowing the teeth to act as probes. The electrical resistance measurements were computed with an electrical decoupling approach that improved the detection of the drillings; it was due to the increment of the sensitivity of the signals. The results showed that the bone mass reduction is correlated with statistical indices obtained in specific frequency intervals of the electrical resistance. This work suggests the possibility of a future application addressed to a bone diagnosis in a non-invasive way.

Assessment of platelet-rich fibrin in the maintenance and recovery of cell viability of the periodontal ligament.

This study analyzed the efficacy of autologous platelet-rich fibrin (PRF) in maintaining and recovering cell viability of the periodontal ligament (PDL). The PDL cells were isolated from 45 extracted teeth randomly distributed among 6 groups: 5 min, 1 h, 2 h, PRF 30 min, PRF 1 h and PRF 2 h. In the groups 5 min, 1 h and 2 h (n = 5), the teeth were kept dry in extra-alveolar times of 5 min, 1 h and 2 h respectively. The teeth of the groups PRF 30 min, PRF 1 h and PRF 2 h (n = 10) were kept dry at extra-alveolar times of 30 min, 1 and 2 h followed by immersion in PRF for 45 min. PDL cells were isolated by enzymatic digestion with type II collagenase and dispase, counted and analyzed for viability with Trypan blue vital dye in Neubauer chamber. The variables total number of cells and cell viability demonstrated that in the 5 min, 1 h and 2 h groups there was a decrease after the extra-alveolar dry times of 1 and 2 h. In comparison with the total number of cells, group 1 h, considered immediate reimplantation, did not present statistical difference when compared to the groups PRF 30 min, PRF 1 h and 2 h, a result that demonstrates that PRF assists in cell maintenance and recovery. PRF provided increased cell viability in relation to the different dry extra-alveolar times analyzed (p < 0.001). Autologous PRF presented effectiveness in maintaining and recovering PDL cells from extracted teeth and kept dry for up to 2 h.

Examination of regressive features of third molars for the purpose of age assessment in the living by means of rescaled regression analyses.

The main criterion of dental age assessment in living adolescents and young adults is the evaluation of third molars' mineralization. Concerning forensic age assessment after the completion of third molars' mineralization, apposition of secondary dentine and narrowing of the periodontal membrane as seen as decreasing radiolucent areas in the radiographs for mandibular third molars have already been described as regressive features. The present study examines the combination of both these features for the purpose of age assessment in regression analyses after rescaling the data to make it on the interval scale. To this end, a total of 1245 orthopantomograms was evaluated, taken from 606 females and 639 males in the age group of 15-40 years. The apposition of secondary dentine and narrowing of the periodontal membrane as seen as decreasing radiolucent areas in the radiographs were determined for the lower third molars. The correlation of the features with the chronological age was assessed by means of rescaled regression analyses. Furthermore, regression formulas for age assessment were established. The values of the standard error of estimate ranged between 3.55 and 4.52 years. In general, the rescaled regression of the examined features appears to be suited for forensic age assessment. A limitation of the present study is the comparatively low number of evaluable teeth in the examined age group. Due to an incomplete development or a lack of the mandibular third molars, only a mere half of the respective teeth could be included in the statistical analysis.

Morpho-histological assessment of the periodontal support structures under the action of excessive occlusal forces and under the influence of nicotine.

Smoking and occlusal trauma are two factors that can interfere with bone homeostasis. The aim of this study was to evaluate the histocellular changes occurring in the periodontal ligament and alveolar bone during the action of excessive occlusal forces, and to assess the influence of nicotine on the alveolar bone loss in teeth subjected to occlusal trauma. MATERIALS AND METHODS: Fifty-six Wistar rats were randomized into seven groups (n=8). Animals were exposed to nicotine and occlusal trauma for 7, 14 and 30 days. Three groups were exposed to occlusal trauma alone, another three groups were exposed to occlusal trauma and nicotine, and one group was not exposed to any treatment. RESULTS: Periodontal lesions induced in the first stage (7-14 days) manifested by a moderate increase of the periodontal space, a multiplication, thickening and elongation of periodontal fibers, as well as their condensation in the middle area of the periradicular space. Regarding bone changes induced by occlusal trauma, groups 5 and 7 (occlusal trauma and nicotine administration) had higher bone losses compared to groups 1, 2, 3, 4 and 6. This study demonstrated that nicotine significantly affected the alveolar bone. CONCLUSIONS: The induced occlusal trauma caused obvious tissue damage. At the same time, it was found that nicotine enhanced alveolar bone resorption, increased tooth mobility and induced an exacerbation of inflammatory processes.

Assessment of static and perfusion methods for decellularization of PCL membrane-supported periodontal ligament cell sheet constructs.

OBJECTIVES: Decellularization aims to harness the regenerative properties of native extracellular matrix. The objective of this study was to evaluate different methods of decellularization of periodontal ligament cell sheets whilst maintaining their structural and biological integrity. DESIGN: Human periodontal ligament cell sheets were placed onto melt electrospun polycaprolactone (PCL) membranes that reinforced the cell sheets during the various decellularization protocols. These cell sheet constructs (CSCs) were decellularized under static/perfusion conditions using a) 20 mM ammonium hydroxide (NH4OH)/Triton X-100, 0.5% v/v; and b) sodium dodecyl sulfate (SDS, 0.2% v/v), both +/- DNase besides Freeze-thaw (F/T) cycling method. CSCs were assessed using a collagen quantification assay, immunostaining and scanning electron microscopy. Residual fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) were assessed with Bio-plex assays. RESULTS: DNA removal without DNase was higher under static conditions. However, after DNase treatment, there were no differences between the different decellularization methods with virtually 100% DNA removal. DNA elimination in F/T was less efficient even after DNase treatment. Collagen content was preserved with all techniques, except with SDS treatment. Structural integrity was preserved after NH4OH/Triton X-100 and F/T treatment, while SDS altered the extracellular matrix structure. Growth factor amounts were reduced after decellularization with all methods, with the greatest reduction (to virtually undetectable amounts) following SDS treatment, while NH4OH/Triton X-100 and DNase treatment resulted in approximately 10% retention. CONCLUSIONS: This study showed that treatment with NH4OH/Triton X-100 and DNase solution was the most efficient method for DNA removal and the preservation of extracellular matrix integrity and growth factors retention.

[Construction and assessment of a three-dimensional finite element model of mandibualr second molar mesialization using customized lingual appliance and mini-implant].

Objective: To construct a three-dimensional (3D) finite element model and analyze the biomechanical characteristics during mandibular second molar mesialization using customized lingual appliances and mini-implant. Methods: One adult student volunteer from The First Affiliated Hospital of Zhengzhou University with lower left first molar extraction was selected. After CT scanning, Mimics, Geomagic, ANSYS were employed to develop a 3D finite element model including customized lingual brackets, stainless steel lingual arch wire, buccal buckles, lower dentition, periodontal ligaments and alveolar bone. Four different loading methods (1. the force of 1.470 N loaded at the lingual side only; 2, 3, 4. the forces of 0.490, 0.735 and 0.980 N loaded at both buccal and lingual sides, respectively.) were included. The initial displacements of the lower second molar and stress distribution in the periodontal ligaments were analyzed. Results: More uniform stress distributions in the periodontal membrane of mandibular left second molar were found when the mesial force were loaded at both buccal and lingual sides than the force loaded at lingual side only. More inclination of the second molar was observed when force loaded at both sides than at lingual side only. With the values of the force increased, the mesial inclination potential of the second molar, the initial movement of the second molar and the stress distribution in the periodontal membrane increased. Under the force of 0.735 N at both buccal and lingual sides, mesially inclined initial displacements of the mandibuar second molar were observed and the stress values of von Mises was in the suitable area. Conclusions: Less second molar rotation was found when the force loaded at both buccal and lingual sides than loaded at lingual side only. However, force loaded at both sides would increase the measial inclination potential of the second molar.

Assessment of periodontal mechano-nociceptive function in healthy Chinese individuals.

OBJECTIVE: Few clinical techniques are available for the description of mechano-nociceptive function in human periodontal tissues. The objective of this study was to test a new technique for assessment of periodontal mechano-nociceptive function by direct application of controlled forces to the teeth and quantify site, side and gender differences in healthy individuals. DESIGN: Twenty healthy young adults (ten males and ten females) participated. A handheld pressure algometer was used to assess pressure pain threshold (PPT) on the maxillary and mandibular central incisors, canines and first premolars on the left and the right side from two directions: lateral and vertical. Statistical analysis was performed using analyses of variance (ANOVA) with repeated measures to compare data. RESULTS: There were significant gender differences of PPT for both lateral (Plateral=0.007) and vertical (Pvertical=0.001) directions with lower thresholds in females (higher sensitivity) than in males. A significant site effect (Plateral=0.002, Pvertical<0.001) was observed with less sensitivity at the first premolar compared to the anterior teeth. No significant right-to-left side differences (P=0.082) were found for the PPTs. There were significant PPT differences between maxillary and mandibular teeth (Plateral=0.020, Pvertical=0.041,) and significant differences between lateral and vertical direction (P<0.001). CONCLUSION: The novel application of PPTs directly to the teeth indicated an adequate and practical method with potential value for clinical assessment of painful conditions affecting the periodontal ligament.

Assessment of cellular materials generated by co-cultured 'inflamed' and healthy periodontal ligament stem cells from patient-matched groups.

Recently, stem cells derived from the'inflamed' periodontal ligament (PDL) tissue of periodontally diseased teeth (I-PDLSCs) have been increasingly suggested as a more readily accessible source of cells for regenerative therapies than those derived from healthy PDL tissue (H-PDLSCs). However, substantial evidence indicates that I-PDLSCs exhibit impaired functionalities compared with H-PDLSCs. In this study, patient-matched I-PDLSCs and H-PDLSCs were co-cultured at various ratios. Cellular materials derived from these cultures were investigated regarding their osteogenic potential in vitro and capacity to form new bone following in vivo transplantation. While patient-matched I-PDLSCs and H-PDLSCs could co-exist in co-culture systems, the proportion of I-PDLSCs tended to increase during in vitro incubation. Compared with H-PDLSC monoculture, the presence of I-PDLSCs in the co-cultures appeared to enhance the overall cell proliferation. Although not completely rescued, the osteogenic and regenerative potentials of the cellular materials generated by co-cultured I-PDLSCs and H-PDLSCs were significantly improved compared with those derived from I-PDLSC monocultures. Notably, cells in co-cultures containing either 50% I-PDLSCs plus 50% H-PDLSCs or 25% I-PDLSCs plus 75% H-PDLSCs expressed osteogenesis-related proteins and genes at levels similar to those expressed in H-PDLSC monocultures (P>0.05). Irrespective of the percentage of I-PDLSCs, robust cellular materials were obtained from co-cultures with 50% or more H-PDLSCs, which exhibited equivalent potential to form new bone in vivo compared with sheets generated by H-PDLSC monocultures. These data suggest that the co-culture of I-PDLSCs with patient-matched H-PDLSCs is a practical and effective method for increasing the overall osteogenic and regenerative potentials of resultant cellular materials.

Avaliação dos eixos ECA / ANG II / AT1 e ECA-2 / ANG (1-7) / MAS em fibroblastos de gengiva e ligamento periodontal humanos estimulados por IL-1ß e sua contribuição na produção e regulação de citocinas e quimiocinas / Evaluation ACE /ANG II /AT1 and ACE-2 /ANG (1-7) /MAS axis in gingival and periodontal ligament human fibroblasts stimulated with IL-1ß and its contribution in the production of cytokines and chemokines

O Sistema renina-angiotensina (SRA) tem sido relatado como um importante modulador de processos inflamatórios e imunológicos, incluindo a doença periodontal (DP). Estudos sugerem neste sistema um eixo alternativo (ECA-2 /ANG(1-7) /MAS) que atuaria como um contra-regulador de efeitos mediados pelo clássico eixo (ECA /ANGII /AT1). Sabe-se que bactérias periodontopatogênicas, como a Porphyromonas gingivalis (Pg), possuem componentes bioativos de membrana (ex. lipopolissacarídeos-LPS) capazes de induzir uma forte resposta imune no hospedeiro devido à liberação de citocinas nas células, entre elas Interleucina (IL)- 1ß. Neste contexto, fibroblastos são as células mais abundantes nos tecidos periodontais e possuem em sua superfície celular receptores necessários para o reconhecimento da invasão bacteriana, ativando cascatas intracelulares, que levam à produção de citocinas. O objetivo deste estudo foi verificar se os eixos ECA/ ANGII/ AT1 e ECA-2/ ANG(1-7)/ MAS contribuem para a produção e/ ou regulação de citocinas inflamatórias (CI) por fibroblastos de gengiva humana (HGF) e ligamento periodontal humano (HPLF) estimulados por IL-1ß. Após o pré-tratamento com Losartan e Ang (1-7) ou silenciamento mediado por RNA de interferência (RNAi) de AT1, HGF e HPLF foram estimulados por IL-1ß por 3 horas (RNAm) ou 24 horas (proteína). Expressão de RNAm para AT1, MAS, ECA, ECA-2, IL-1ß, TNF-α, IL-6, IL-8, IL-10, TGF-ß, CXCL12, RANK-L e OPG foram avaliados por RT-qPCR e das proteínas IL-6, IL-8, ECA e ECA-2 por ELISA. Foi realizado também Western Blot para detecção de AT1 e ECA nos extratos celulares e dosagem de nitrito no sobrenadante das culturas. Ambos os subtipos de fibroblastos mostraram aumento da expressão de RNAm para AT1, IL-1ß, IL-6, IL-8, TNF-α e OPG, quando estimulados por IL-1ß. No entanto, apenas em HPLF foi observado aumento para MAS, ECA e TGF-ß. Losartan e Ang (1-7) não modularam o transcrito, a secreção de CI e nem a produção de nitrito no sobrenadante das culturas, tanto em HGF como em HPLF. O silenciamento do receptor AT1 reduziu a secreção de IL-6 e IL-8 induzida por IL-1ß em cultura de HGF e HPLF e aumentou a expressão gênica de OPG somente em HGF. Estes resultados sugerem que o silenciamento de AT1, mas não o bloqueio farmacológico deste receptor pelo antagonista Losartan, em HGF e HPLF, pode controlar a produção de IL-6 e IL-8, que por sua vez contribuem para a patogênese periodontal.(AU)

The renin-angiotensin system (RAS) has been reported as an important modulator of inflammatory and immune responses, including periodontal disease (PD). Studies suggest an alternative axis as part of this system (ACE-2 / ANG (1-7) / MAS) that would act as counter-regulatory to the classical axis (ECA / ANGII / AT1). It is known that periodontal bacteria such as Porphyromonas gingivalis (Pg) have bioactive components in their membrane (such as lipopolysaccharide-LPS) capable of inducing a strong immune response in the host due to the release of cytokines in cells, including interleukin (IL) - 1ß. In this regard, fibroblasts are the most abundant cells in periodontal tissues and receptors needed for the recognition of bacterial invasion by activating intracellular cascades that lead to cytokine production. The aim of this study was to determine whether the axes ACE / ANGII / AT1 and ACE-2 / ANG (1-7) / MAS contribute to the production and / or regulation of inflammatory cytokines (IC) by fibroblasts of human gingiva (HGF) and human periodontal ligament (HPLF) stimulated IL-1ß. After pre-treatment with Losartan, Ang (1-7) or silencing mediated by RNA interference (RNAi) of AT1, HGF and HPLF were stimulated by IL-1ß for 3 hours (RNAm) or 24 hours (protein). Expression mRNA for AT1, MAS, ACE, ACE-2, IL-1ß, TNF-α, IL-6, IL-8, IL-10, TGF-ß, CXCL12, RANK-L and OPG was assessed by RT- qPCR and proteins IL-6, IL-8, ACE and ACE-2 by ELISA. Western Blot for the detection of AT1 and ECA and dosage of nitrite was also performed. Experiments stimulated by IL-1ß showed a positive control for gene expression AT1, IL-1ß, IL-6, IL-8, TNF-α and OPG in HGF and HPLF and MAS, ACE and TGF-ß only HPLF. Losartan and Ang (1-7) did not modulate the transcription and secretion of IC and no nitrite production in the culture supernatant of HGF and HPLF. The silencing AT1 reduced IL-6 secretion and IL-8 induced by IL- ß in cultured HGF and HPLF and increased OPG gene expression only HGF. These results suggest that silencing AT1, but not pharmacological blockade of this receptor by Losartan in HPLF and HGF, can control the production of IL-6 and IL-8, which in turn contribute to the pathogenesis of periodontal disease.(AU)

Dynamic assessment of Capparis spinosa buds on survival of periodontal ligament cells using a real-time cell analysis method.

BACKGROUND: Tooth avulsion is the most severe type of traumatic dental injuries and it results in the complete displacement of the tooth out of its socket in alveolar bone. Reimplantation of the tooth is considered to be a best treatment modality due to its biological and psychological advantages. Its prognosis depends on the extra alveolar time, the storage medium, and the patient's general health. OBJECTIVE: The aim of this study was to evaluate the effect of Capparis spinosa (C. spinosa) in maintaining the viability of human periodontal ligament (PDL) cells using a real-time cell analysis method. MATERIALS AND METHODS: Periodontal ligament cells were obtained from healthy human third molars extracted for orthodontic purposes. The storage media tested were: Dulbecco's Modified Eagle Medium (DMEM), C. spinosa, Hank's Balanced Salt Solution (HBSS), and light milk. A real-time cell analyzer system was used to evaluate cell viability. After seeding cell suspensions into the wells of the E-plate 96, PDL cells were treated with each of tested media and monitored for every 5 min for 26 h. Statistical analysis of the data was accomplished using one-way analysis of variance complemented by the Tukey test. The level of significance was set at P < 0.05. RESULTS: Dulbecco's Modified Eagle Medium (control) and C. spinosa groups had significantly higher cell index values compared with the HBSS and light milk (P < 0.05). Although, C. spinosa showed better results than DMEM (control), but this difference was not found statistically significant. CONCLUSION: Capparis spinosa can be a suitable, alternative storage medium for avulsed teeth.