BACKGROUND: Periodontitis is characterized by alveolar bone resorption, aggravated by osteoblast dysfunction, and associated with intracellular oxidative stress linked to the nuclear factor erythroid 2-related factor 2 (NRF2) level. We evaluated the molecular mechanism of periodontitis onset and development and the role of NRF2 in osteogenic differentiation. METHODS: Primary murine mandibular osteoblasts were extracted and exposed to Porphyromonas gingivalis lipopolysaccharide (Pg-LPS) or other stimuli. Reactive oxygen species (ROS) and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining were used to detect intracellular oxidative stress. Alkaline phosphatase staining and alizarin red S staining were used to detect the osteogenic differentiation of osteoblasts. Immunofluorescence and western blotting were used to determine the changes in the mitogen-activated protein kinase (MAPK) pathway and related molecule activities. Immunofluorescence colocalization and co-immunoprecipitation were performed to examine the nuclear translocation of NRF2 and its interaction with dual-specific phosphatase 1 (DUSP1) in cells. RESULTS: Ligated tissue samples showed higher alveolar bone resorption rate and lower NRF2 level than healthy periodontal tissue samples. Pg-LPS increased intracellular oxidative stress levels and inhibited osteogenic differentiation, whereas changes in NRF2 expression were correlated with changes in the oxidative stress and osteogenesis rate. NRF2 promoted the dephosphorylation of the MAPK pathway by nuclear translocation and the upregulation of DUSP1 expression, thus enhancing the osteogenic differentiation capacity of mandibular osteoblasts. The interaction between NRF2 and DUSP1 was observed. CONCLUSIONS: NRF2 and its nuclear translocation can regulate the osteogenic differentiation of mandibular osteoblasts under Pg-LPS conditions by interacting with DUSP1 in a process linked to the MAPK pathway. These findings form the basis of periodontitis treatment.
Animals ; NF-E2-Related Factor 2/physiology* ; Lipopolysaccharides/pharmacology* ; Osteoblasts/drug effects* ; Mice ; Porphyromonas gingivalis/chemistry* ; Cell Differentiation ; Osteogenesis ; Dual Specificity Phosphatase 1/metabolism* ; Mandible/cytology* ; Reactive Oxygen Species/metabolism* ; Oxidative Stress ; Periodontitis/metabolism* ; Cells, Cultured ; Male ; Cell Nucleus/metabolism*

BACKGROUNDThe thickness of the alveolar mucosa influences the probability of the occurrence of denture-induced irritations. Thick denture-supporting tissues offer relief from mucosal tenderness and ulcers; however, the uniformity of the thickness across the entire mandibular alveolar mucosa cannot be accurately determined in edentulous patients. This study aimed to assess the mucosal thickness of the denture-bearing area in the edentulous mandible.

METHODSTwenty-seven edentulous patients underwent cone-beam computed tomography scanning, wherein the patients wore a record base to retract soft tissues away from the alveolar mucosa. The measured regions were the central incisor (IC), lateral incisor (IL), canine (Ca), first premolar (P1), second premolar (P2), first molar (M1), and second molar (M2) regions. The thickness was measured in the alveolar ridge crest (T), buccal (B1-B4), and lingual (L1-L4) alveolar ridge mucosa. The average thickness of the mucosa at buccal sides (B) and lingual sides (L) were also assessed.

RESULTSThe differences in the mucosal thickness between the left and right sides were not significant. In the Ca-M2 regions, T was the thickest, and L3 was the thinnest of all the measured points in the same regions. L was significantly less than B in posterior regions (P < 0.01). On the other hand, M2 at L4 was thinnest of all the measured regions from Ca to M2 (P < 0.01), and was thicker than IC, IL, P1, and P2 at B2.

CONCLUSIONSSince the mucosal thickness of denture-bearing area in the edentulous mandible is not uniform; the tissue surface of the denture base or custom tray should be selectively relieved, which may reduce the risk of denture-induced irritations.

Aged ; Aged, 80 and over ; Alveolar Process ; anatomy & histology ; Dentures ; Female ; Humans ; Jaw, Edentulous ; Male ; Mandible ; anatomy & histology ; cytology ; Middle Aged ; Mucous Membrane ; cytology ; Retrospective Studies

OBJECTIVETo explore the effect of high glucose on proliferation of bone marrow stromal stem cells through Wnt/Β-catenin pathway.

METHODSBone marrow stormal cells were obtained from the mandible of Wistar rats and stimulated with different concentrations of glucose (5.5 and 16.5 mmol/L). Cell proliferation was evaluated with methyl thiazolyl tetrazolium assay (1, 3, 5, and 7 d)and cell cycle analysis by flow cytometry (5 d). Β-catenin and cyclin D1 protein levels were determined by Western blot. The mRNA expression of lymphoid enhancer binding factor-1 (LEF-1) and cyclin D1 were tested by real-time polymerase chain reaction.

RESULTSThe results of methyl thiazolyl tetrazolium assay indicated that the optical density values of two different concentrations of the glucose had no statistical difference on day 1 (P=0.700). On days 3, 5, and 7, the optical density values of the 16.5 mmol/L group were significantly lower than those in the 5.5 mmol/L group (P=0.006, P=0.002, and P=0.003). Cell cycle analysis indicated that high glucose concentration could reduced the progression from phase G1 to S, and the proliferation index values of the 16.5 mmol/L group were significantly lower than those of the 5.5 mmol/L group (P=0.014). The Β-catenin and cyclin D1 levels were lower in the 16.5 mmol/L group when compared with the 5.5 mmol/L group. High glucose condition also reduced the mRNA expressions of LEF-1 and cyclin D1.

CONCLUSIONHigh glucose can inhibit the proliferation of bone marrow stormal cells by suppressing the expressions of Β-catenin, LEF-1, and cyclin D1 in the Wnt/Β-catenin pathway.

Animals ; Bone Marrow Cells ; cytology ; Cell Proliferation ; drug effects ; Cyclin D1 ; metabolism ; Glucose ; pharmacology ; Lymphoid Enhancer-Binding Factor 1 ; metabolism ; Male ; Mandible ; cytology ; Mesenchymal Stromal Cells ; cytology ; Rats ; Rats, Wistar ; Wnt Signaling Pathway ; beta Catenin ; metabolism

OBJECTIVETo repair segmental mandibular defects with autologous bone marrow stromal cells (BMSCs) engineered bone.

METHODSIsolated BMSCs were expanded in vitro and osteogenic induced. In 12 canines, a 3 cm segmental mandibular defect at right mandible was created. 6 canine's defects were repaired with cell-scaffold constructs made from induced BMSCs and coral; others were repaired with coral as control. The engineered bone was evaluated by X-ray, CT, Dual Energy X-ray Absorptiometry (DXA), gross and histological examination, and biomechanical test post-operatively.

RESULTSInduced BMSCs grew well on coral scaffold. At 12 weeks, X-ray showed more callus formed in experimental group, while evident scaffold duration in control group. At 32 weeks, gross observation, X-ray and CT demonstrated well bony-union in experimental group, while bony-nonunion in control group. Also DXA revealed significantly higher bone mineral density of experimental group than control group. Histologically, mature bone were commonly observed and there were bony healing in experimental group, while fibrous healing occurred in control group. Biomechanical test revealed no significant difference between experimental group and normal group.

CONCLUSIONSCanine segmental mandibular defects can be repaired with the tissue-engineered bone generated by coral scaffold with autologous osteogenic BMSCs.

Animals ; Anthozoa ; Bone Marrow Cells ; cytology ; Bone Substitutes ; Cell Culture Techniques ; Dogs ; Mandible ; pathology ; surgery ; Mesenchymal Stromal Cells ; cytology ; Pilot Projects ; Tissue Engineering ; Tissue Scaffolds ; Transplantation, Autologous

OBJECTIVETo evaluate the possible signal transduction mechanism of the mechanical stress induced by the distraction procedure in osteocytes.

METHODSAn animal model of mandibular distraction osteogenesis in rabbits was established. The expressions of c-fos, OPG and OPGL were detected by ultrasensitive S-P immunohistochemical method.

RESULTAt 4 and 8 days after distraction, distraction zone showed strong positive staining of c-fos, which were apparently higher than that in distraction zone of 2, 4 and 6 weeks after consolidation. At 4 and 8 days after distraction and 2 weeks after consolidation, the expression of OPG was strong, and then wore off gradually at 4 and 6 weeks after consolidation. Weak signals of OPGL could be detected at 6 weeks after consolidation only.

CONCLUSIONc-fos, OPG and OPGL are important regulators in distraction osteogenesis. c-fos is interrelated with the mechanical stress induced by the distraction procedure closely, OPG promotes new bone formation, while OPGL plays a more active role in bone remodeling.

Animals ; Mandible ; cytology ; metabolism ; Osteocytes ; metabolism ; Osteogenesis, Distraction ; Osteoprotegerin ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; RANK Ligand ; biosynthesis ; genetics ; Rabbits ; Random Allocation

OBJECTIVETo repair segmental mandibular defects with autogenous bone marrow stromal cells (BMSCs) and coralline hydroxyapatite.

METHODSIsolated BMSCs were in vitro expanded and osteogenically induced. In 11 canines, a 3 cm segmental mandibular defect in right mandible was created. Five canine's defects were repaired with cell-scaffold constructs made from induced BMSCs and coralline hydroxyapatite (CHA); Others were repaired with CHA as control. The engineered bone was evaluated by X-ray, CT, gross and histological examination, biomechanical test 12, 26, 32 weeks post-operation respectively.

RESULTSBMSCs grew well on the CHA. X-ray and CT images showed better callus formation at connection sites in experimental group over time while worse formation at connection sites eventually in control group. At 32 weeks post-operation in experimental group, the defects were well repaired grossly. Histologically, there were bony healing and lamellar bone formation, in experimental group fibrous healing and woven bone formation in control group. Biomechanical test revealed no significant difference between experimental group and normal control group.

CONCLUSIONSCanine segmental mandibular defects can be ultimately repaired with the tissue-engineered bone generated by autogenous osteogenic BMSCs and CHA scaffold.

Animals ; Bone Marrow Cells ; cytology ; Bone Substitutes ; Ceramics ; therapeutic use ; Dogs ; Hydroxyapatites ; therapeutic use ; Mandible ; physiology ; Mandibular Injuries ; pathology ; surgery ; Mesenchymal Stromal Cells ; cytology ; Tissue Engineering ; Tissue Scaffolds

OBJECTIVETo observe the expression patterns of matrix metalloproteinase-3 and its inhibitor, tissue inhibitor metalloproteinases (TIMP)-1, in remodeling phase of mandibular distraction osteogenesis.

METHODSBilateral mandibular osteotomies were performed in 8 mature rabbits. The mandibles were lengthened 6 mm using a custom-made distractor with a rate of 1 mm/d. All animals were killed in 4 weeks after completion of distraction. The distracted calluses were harvested and processed for histology and immunohistochemistry of MMP-3 and TIMP-1.

RESULTSElevated expression of both MMP-3 and TIMP-1 was found in the distracted calluses resulting from mandibular lengthening. Positive staining for MMP-3 was seen in the osteoblasts and osteocytes, and TIMP-1 was mainly localized in osteoblasts.

CONCLUSIONMMP-3 and TIMP-1 appear to play important roles in the remodeling of new bone created by mandibular distraction osteogenesis.

Animals ; Bone Regeneration ; Bone Remodeling ; Female ; Male ; Mandible ; cytology ; metabolism ; surgery ; Matrix Metalloproteinase 3 ; biosynthesis ; Osteoblasts ; metabolism ; Osteogenesis, Distraction ; Rabbits ; Tissue Inhibitor of Metalloproteinase-1 ; biosynthesis

OBJECTIVEThe study was to investigate the circadian rhythm of the proliferation index (PI) of the mandibular osteoblast after distraction osteogenesis (DO) by a comparative study to explain the different rhythm characteristics.

METHODSForty-eight healthy goats were employed and divided randomly into DO group and control. Four weeks after the right side mandible of DO group was distracted, animals of the both groups were killed at 0:00, 4:00, 8:00, 12:00, 16:00, 20:00 time point subsequently. Samples of 2 cm x 2 cm x 0.5 cm bone tissue were excised from distracted zone of DO group and the right mental foramen zone of the control. Primary cell culture method was employed to isolate the osteoblast from bone tissue and a flow cytometric analysis was used to detect the PI of osteoblast. The data was analyzed with cosinor rhythmometry method and the different rhythm between the two groups were investigated.

RESULTSThe PI in the two groups demonstrated cosinor waves. The mesor and amplitude in DO group were significant higher compared with the control(P < 0.05), but there was no significant difference for the acrophase between the two groups (P > 0.05).

CONCLUSIONThe PI of osteoblast after DO showed significant circadian rhythm. It is suggested that DO is helpful to promote osteoblast proliferation and bone regeneration.

Animals ; Bone Regeneration ; Cell Division ; Cells, Cultured ; Circadian Rhythm ; Flow Cytometry ; Goats ; Mandible ; cytology ; Osteoblasts ; cytology ; Osteogenesis, Distraction ; Random Allocation

OBJECTIVEThe aim of this study was to investigate the circadian rhythm of the proliferative index of mandibular osteoblasts of goats in a normal situation.

METHODSTwenty four healthy goats were selected and randomly divided into six groups. They were killed, and a piece of bone tissue with 2 cm x 2 cm x 0.5 cm was obtained from right mandibular mental foramen region at 0:00, 4:00, 8:00, 12:00, 16:00 and 20:00 time points, after they had been fed for one week. The primary cell culture method was used to cultivate osteoblasts from the bone. The method of anti-5-bromodeoxyuridine monoclonal antibody and the flow cytometric analysis were used to detect the proliferative index of the mandibular osteoblasts at different time. The data were analyzed using the method of single cosine rhythmometry.

RESULTSThe proliferative indexes of the mandibular osteoblasts of goats were higher at the nighttime and lower in the daytime. The undulation of the proliferative indexes displayed a significant circadian rhythm (P < 0.01). The peak value was obtained at about 19:44, and the lowest value at about 7:44.

CONCLUSIONThe proliferative indexes of the mandibular osteoblasts of goats are not stable within 24 hours in a normal situation, but with significant circadian rhythm. There are different proliferative indexes at different time points.

Animals ; Cell Division ; Cells, Cultured ; Circadian Rhythm ; Flow Cytometry ; Goats ; Mandible ; cytology ; Osteoblasts ; cytology ; Random Allocation