Decorin in the spatial control of collagen mineralization.

Understanding the molecular mechanism by which the periodontal ligament (PDL) is maintained uncalcified between two mineralized tissues (cementum and bone) may facilitate the functional repair and regeneration of the periodontium complex, disrupted in the context of periodontal diseases. However, research that explores the control of type I collagen (COL I) mineralization fails to clarify the detailed mechanism of regulating spatial collagen mineralization, especially in the periodontium complex. In the present study, decorin (DCN), which is characterized as abundant in the PDL region and rare in mineralized tissues, was hypothesized to be a key regulator in the spatial control of collagen mineralization. The circular dichroism results confirmed that DCN regulated the secondary structure of COL I, and the surface plasmon resonance results indicated that COL I possessed a higher affinity for DCN than for other mineralization promoters, such as DMP-1, OPN, BSP and DSPP. These features of DCN may contribute to blocking intrafibrillar mineralization in COL I fibrils during the polymer-induced liquid-precursor mineralization process when the fibrils are cross-linked with DCN. This effect was more remarkable when the fibrils were phosphorylated by sodium trimetaphosphate, as shown by the observation of a tube-like morphology via TEM and mineral sheath via SEM. This study enhances the understanding of the role of DCN in mineralization regulation among periodontal tissues. This provides insights for the development of biomaterials for the regeneration of interfaces between soft and hard tissues.

The effect of prolonged holding time on the mechanical property and microstructural property of lithium disilicate glass-ceramic.

Repeat firing produces uncertainty about stabilizing lithium disilicate glass-ceramic (LDGC) material properties, even though prolonged holding time can enhance the mechanical property of LDGC during a single firing cycle. However, the effect of prolonged holding time and repeat firing on the mechanical property and microstructure of LDGC is not fully understood. In the present study, three groups of LDGC material were created: (i) extension of holding time (7 vs. 14 vs. 28 min) at 780-800 °C; (ii) holding time extension (7 vs. 14 min) and dual sintering at 800-820 °C, respectively; (iii) dual sintering with prolonged holding time (7 vs. 14 min) at 820-840 °C. The nano-indenter test revealed that prolonged holding time (14 and 28 min) promoted the enhancement of LDGC hardness and Young's modulus. X-ray photoelectron spectroscopy, X-ray diffraction and Fourier transform infrared spectroscopy confirmed that prolonged holding time increased and stabilized LD phase in LDGC, as well as induced residual compressive stress. Scanning electron microscopy showed that prolonged holding time increased LD crystal grains homogeneously and facilitated LDGC to form dense interlocking structure without enlarging crystal size grains significantly. In contrast, LDGC that dual sintered alone at 820-840 °C possessed inferior mechanical properties, coupled with heterogeneous crystal phases, residual tensile stress, and melted crystals grains in the porous microstructure. Interestingly, these deteriorated properties of LDGC caused by dual sintering alone could be counteracted by prolonging the holding time. Nevertheless, the LDGC materials displayed an excellent biocompatibility throughout the study. This study identified that prolonged holding time during repeated firing cycles stabilized LD phase and crystal grain size of LDGC, thus enhanced the mechanical properties, which provided a new insight to extend the repeat fired restoration longevity of LDGC. Graphical abstract.

Enhanced Healing and Antimicrobial Efficacy of Chitosan-g-Polyacrylamide in a Rat Model of Gingival Ulcers.

Patients in dental hospitals often experience oral ulcerative lesions, which lead to pain and affect the patient's quality of life. At present, the goal of treating oral ulcerative lesions with drugs is to reduce inflammation and promote ulcer healing. However, very few antibacterial and hemostatic drugs are designed to be suitable for the microenvironment of gingival ulcers. Based on this, we have designed a natural therapeutic agent for oral ulcerative lesions that meets the various requirements of oral ulcerative lesion medication. The chitosan-g-polyacrylamide (CP) copolymer is composed of chitosan as the main chain and polyacrylamide polymers as the side chains. Antibacterial experiments show that this polymer can effectively inhibit the proliferation of Gram-negative (Escherichia coli) and Gram-positive bacteria (Staphylococcus aureus). In vitro cell experiments also show that the CP copolymer is non-toxic, which is conducive to ulcer wound healing. Coagulation experiments prove that the CP copolymer can accelerate blood coagulation to stop bleeding. In experiments using a Wistar rat gingival ulcer model, the CP copolymer significantly promoted ulcer healing and shortened the healing time. These results indicate that the CP copolymer may serve as a potential therapeutic agent for oral ulcerative lesions.

microRNA-143-3p regulates odontogenic differentiation of human dental pulp stem cells through regulation of the osteoprotegerin-RANK ligand pathway by targeting RANK.

NEW FINDINGS: What is the central question of this study? What is the role of miR-143-3p during human dental pulp stem cell (hDPSC) differentiation. What is the main finding and its importance? miR-143-3p negatively regulates receptor activator of nuclear factor-κB (RANK). RANK ligand (RANKL) binds to RANK and stimulates the development of osteoclasts. Osteoprotegerin (OPG) inhibits the interaction between RANK and RANKL. The OPG-RANKL signalling pathway regulates odontogenic differentiation of hDPSCs. ABSTRACT: Human dental pulp stem cells (hDPSCs) are capable of differentiating into odontoblast-like cells, which secrete reparative dentin after injury, in which the role of microRNA-143-3p (miR-143-3p) has been identified. Therefore, we investigated the mechanism by which miR-143-3p influences odontoblastic differentiation of hDPSCs. The relationship between miR-143-3p and receptor activator of nuclear factor-κB (RANK) was initially identified by bioinformatics prediction and further verified by dual luciferase reporter gene assay. Gain- and loss-of-function analysis with miR-143-3p mimic and miR-143-3p inhibitor was subsequently conducted. Dentin sialophosphoprotein (DSPP), bone sialoprotein (BSP), alkaline phosphatase (ALP), osteocalcin (OCN) and osteopontin (OPN) mRNA levels were then evaluated by RT-qPCR. Osteoprotegerin (OPG), RANK ligand (RANKL), nuclear factor-κB (NF-κB) p65 protein levels and the extent of NF-κB p65 phosphorylation were examined by western blot analysis. Alizarin red staining was performed to assess the mineralization of hDPSCs. Cell apoptosis and cell cycle distribution were determined using flow cytometry. During odontoblastic differentiation of hDPSC, miR-143-3p had high expression, but RANK expression was low. miR-143-3p was found to target RANK, and its inhibition enhanced mineralization and hDPSC apoptosis, while blocking cell cycle entry. At the same time, miR-143-3p inhibition elevated the extent of NF-κB p65 phosphorylation, as well as the expression of RANK, RANKL, DSPP, BSP, ALP, OCN and OPN, while decreasing the OPG level. Silencing RANK had opposite effects on these markers. miR-143-3p regulates odontoblastic differentiation of hDPSCs via the OPG-RANKL pathway that targets RANK. The elucidation of the mechanisms of odontogenic differentiation of hDPSCs may contribute to the development of effective dental pulp repair therapies for the clinical setting.

The role of organic phosphate in the spatial control of periodontium complex bio-mineralization: an in vitro study.

The periodontal structure is a particularly exquisite model of hierarchical spatial control of mineralization. Extracellular matrix control in the selective mineralization of the periodontium complex remains elusive since the extracellular matrix is a set of mineralization promoters and inhibitors. The phosphorylated proteins, which are ubiquitous in the extracellular matrix of the periodontium complex, are well-documented as primary factors in the regulation of tissue mineralization. Whether organic phosphates are key regulators in defining the interfaces between dentin, cementum, periodontal ligament and alveolar bone is an issue worthy of research. Here, we investigated the in vitro remineralization process of demineralized and dephosphorylated periodontal tissue sections. When exposed to a metastable mineralization solution, a large number of calcospherulites deposited on the surface of the dephosphorylated sections and the tissue selective mineralization were disrupted. Interestingly, on adding a dentin matrix protein-1 analogue named polyacrylic acid, the surface mineralization rate in the dephosphorylated periodontal complex reduced dramatically. In contrast, hierarchical mineralization was displayed by the demineralized section at the tissue collagen fibrillar levels in both alveolar bone and dentin regions. These results demonstrated that the organic phosphate could prevent surface mineral deposition, and the minerals could penetrate the collagen fibrils to initiate a selective and hierarchal tissue mineralization with the assistance of the dentin matrix protein-1 analogue in the periodontal complex. This study enhances our understanding of the mineralization discrepancy in the periodontal tissues, which will provide some insight into the development of biomaterials for the regeneration of soft-hard tissue interfaces.

Aberrant activation of Wnt signaling pathway altered osteocyte mineralization.

Mineralization of bone is a dynamic process, involving a complex interplay between cells, secreted macromolecules, signaling pathways, and enzymatic reactions; the dysregulation of bone mineralization may lead to serious skeletal disorders, including hypophosphatemic rickets, osteoporosis, and rheumatoid arthritis. Very few studies have reported the role of osteocytes - the most abundant bone cells in the skeletal system and the major orchestrators of bone remodeling in bone mineralization, which is owed to their nature of being deeply embedded in the mineralized bone matrix. The Wnt/ß-catenin signaling pathway is actively involved in various life processes including osteogenesis; however, the role of Wnt/ß-catenin signaling in the terminal mineralization of bone, especially in the regulation of osteocytes, is largely unknown. This research demonstrates that during the terminal mineralization process, the Wnt/ß-catenin pathway is downregulated, and when Wnt/ß-catenin signaling is activated in osteocytes, dendrite development is suppressed and the expression of dentin matrix protein 1 (DMP1) is inhibited. Aberrant activation of Wnt/ß-catenin signaling in osteocytes leads to the spontaneous deposition of extra-large mineralized nodules on the surface of collagen fibrils. The altered mineral crystal structure and decreased bonding force between minerals and the organic matrix indicate the inferior integration of minerals and collagen. In conclusion, Wnt/ß-catenin signaling plays a critical role in the terminal differentiation of osteocytes and as such, targeting Wnt/ß-catenin signaling in osteocytes may serve as a potential therapeutic approach for the management of bone-related diseases.

Effect of ovariectomy on tissue-level changes in rat maxilla.

PURPOSE: It remains unclear whether estrogen deficiency affects the ultrastructure and tissue-level mechanical properties of the maxilla. The hypothesis of this study was that the ovariectomized rat could induce tissue-level changes of the maxilla. MATERIALS AND METHODS: Twelve 3-month-old female Sprague Dawley rats were acquired and randomly divided into two groups: ovariectomized and SHAM (control) (n = 6 for each group). Three months after the ovariectomy, implants were placed; the animals were sacrificed at day 28, and then samples were collected and prepared according to the previously established protocols. Advanced analytical equipment including scanning electron microscope with energy-dispersive spectrometry, transmission electron microscope, and nanoindentation were used to analyze bone quality. RESULTS: The results showed that the mature bone areas in the ovariectomized group were significantly affected in the mineral crystal and the microstructure. The micro-mechanical properties of the mature bone were also affected, showing significantly increased hardness (H) and reduced modulus (Er) in ovariectomized rats compared with the normal rats (P < .05). Differences in H and Er in new bone areas between the normal and ovariectomized rats were less significant. CONCLUSION: Ovariectomy affected maxilla bone tissue-level quality; however, the effects mainly existed in the mature bone areas, which were characterized by higher crystalline mineralization, hardness, and modulus.

Characterization of nano-structural and nano-mechanical properties of osteoarthritic subchondral bone.

BACKGROUND: Although articular cartilage is the primary tissues affected by osteoarthritis (OA), the underlying subchondral bone also undergoes noticeable changes. Despite the growing body of research into the biophysical and mechanical properties of OA bone there are few studies that have analysed the structure of the subchondral sclerosis at the nanoscale. In this study, the composition and nano-structural changes of human osteoarthritis (OA) subchondral bone were investigated to better understand the site-specific changes. METHODS: OA bone samples were collected from patients undergoing total knee replacement surgery and graded according to disease severity (grade I: mild OA; grade IV: severe OA). Transmission electron microscopy (TEM), Electron Diffraction, and Elemental Analysis techniques were used to explore the cross-banding pattern, nature of mineral phase and orientation of the crystal lattice. Subchondral bone nano-hydroxyapatite powders were prepared and characterised using high resolution transmission electron microscopy (HR-TEM) and fourier transform infrared spectroscopy (FTIR). Subchondal bone mechanical properties were investigated using a nano-indentation method. RESULTS: In grade I subchondral bone samples, a regular periodic fibril banding pattern was observed and the c-axis orientation of the apatite crystals was parallel to the long axis of the fibrils. By contrast, in grade IV OA bone samples, the bulk of fibrils formed a random and undulated arrangement accompanied by a circular oriented pattern of apatite crystals. Fibrils in grade IV bone showed non-hierarchical intra-fibrillar mineralization and higher calcium (Ca) to phosphorous (P) (Ca/P) ratios. Grade IV OA bone showed higher crystallinity of the mineral content, increased modulus and hardness compared with grade I OA bone. CONCLUSIONS: The findings from this study suggest that OA subchondral sclerotic bone has an altered mineralization process which results in nano-structural changes of apatite crystals that is likely to account for the compromised mechanical properties of OA subchondral bones.

Effectiveness of cysteine proteases on protein/pigment film removal.

OBJECTIVE: Theaflavin (TF) from the black tea can react to human salivary proline-rich proteins (PRPs) to form stains on exposed dental surfaces. Here, we employed a model of protein/pigment film using TF and dephosphorylated bovine ß-casein (Dß-CN), which has an extended conformation, similar to that of salivary PRPs, on a sensor surface to assess the efficacy of cysteine proteases (CPs) including papain, stem bromelain, and ficin, on removing TF bound to Dß-CN and the control TF readsorption on the residual substrate surfaces was also measured. METHODS: The protein/pigment complex film was built by using a quartz crystal microbalance with dissipation (QCM-D). The efficacies of CPs were assessed by Boltzman equation model. The surface details were detected by grazing angle infrared spectroscopy spectra, atomic force microscopy images, and contact angles. RESULTS: The efficacy order of CPs on hydrolyzing protein/pigment complex film is ficin>papain>bromelain. The results from grazing angle infrared spectroscopy spectra, atomic force microscopy images, and contact angles demonstrated that TF bound on the Dß-CN was effectively removed by the CPs, and the amount of TF readsorption on both the residual film of the Dß-CN/TF and the Dß-CN was markedly decreased after hydrolysis. CONCLUSION: This study indicates the potential application of the CPs for tooth stain removal and suggests that these enzymes are worthy of further investigation for use in oral healthcare.