Osteonectin Expression Research Articles

Xanthium strumarium seed extract boosts osteogenesis in human dental pulp stem cell model

BackgroundIn traditional medicine, Xanthium strumarium is used as an anti-inflammatory and anti-arthritic plant-based medicine. Human Dental Pulp Stem Cells (hDPSCs) are an ideal in vitro model for drug and bioactive compound screening. This study assessed the potential of X. strumarium aqueous extract on hDPSCs differentiation towards the osteogenic lineage. Materials and methodsHDPSCs were isolated and cultured by explant method and characterized by surface marker expression, Colony Forming units fibroblasts (CFU-F), Population Doubling time (PDT), and tri-lineage differentiation. X. strumarium aqueous seed extract (XSE) was prepared and its cytotoxic effect on hDPSCs was examined by MTT assay. The effect of XSE on hDPSC differentiation into osteocytes was investigated by biochemical staining and gene expression. ResultsThe hDPSCs were positive for CD73, CD90, and CD105 and negative for CD34, CD45, and HLA-DR surface markers. The cells had a colony-forming ability with a PDT of 44.91 h. The hDPSCs differentiated into osteocytes, chondrocytes, and adipocytes. The XSE concentration of 15 μg/ml had a significant increase in hDPSC viability. Alizarin Red S staining revealed that XSE treatment enhanced calcium accumulation and matrix mineralization in hDPSCs. XSE treatment also increased osteonectin and IL-6 transcript expression in osteogenesis-induced hDPSCs. ConclusionX. strumarium aqueous extract is a suitable candidate for bone repair because it promotes osteogenic differentiation in hDPSCs. Therefore this could be explored further in the treatment of bone disorders.

Prenatal exposure to crude oil vapor reduces differentiation potential of rat fetal mesenchymal stem cells by regulating ERK1/2 and PI3K signaling pathways: Protective effect of quercetin

It has been indicated that crude oil vapor (COV) induces tissue damage by several molecular mechanisms. Quercetin (QT) as an important component of food with antioxidant properties has a protective role against cell toxicity caused by many pollutants. However, data related to the adverse effects of crude oil vapor (COV) on stem cell fate and differentiation and the role of quercetin (QT) in protecting stem cells against the toxicity caused by these pollutants is very limited. This study aimed to explore the protective effect of QT against the adverse effects of COV on fetal mesenchymal stem cells (fMSCs) differentiation. Twenty-four pregnant Wistar rats were categorized into 4 groups including the control, COV, COV+QT, and QT. Rats were exposed to COV from gestational day (GD) 0–15 and received QT by gavage. The fMSCs were isolated from fetuses, and cell proliferation, differentiation potential, expression of osteogenesis and adipogenesis-related genes, and phosphorylation of PI3K and ERK1/2 signaling proteins were evaluated. The results showed that COV reduced the proliferation and differentiation of fMSCs through the activation of PI3K and ERK1/2 signaling pathways. Also, COV significantly decreased the expression of osteonectin, ALP, BMP-6, Runx-2, PPARγ, and CREBBP genes in differentiated cells. QT treatment increased the proliferation and differentiation of fMSCs in COV-exposed rats. In conclusion, our findings suggest that prenatal exposure to COV impaired fMSCs differentiation and QT reduced the adverse effects of COV by regulating ERK1/2 and PI3K signaling pathways.

Evaluation of the effects of Momordica charantia on tibial defect injury in rats

We aimed to examine the histological and immunohistochemical effects of Momordica charantia on bone repair, which provides positive regulation such as blood sugar, blood pressure, antilipidemic, anticarcinogenic, antibacterial, antioxidant, anti-inflammatory, wound healing, and tissue regeneration in rats with tibial bone defect. Momordica charantia (MC) (commonly called bitter melon; Goya; bitter apple; bitter gourd; bitter squash; balsam-pear; with many more names listed below) is a tropical and subtropical vine of the family Cucurbitaceae, widely grown in Asia, Africa, and the Caribbean for its edible fruit. Its many varieties differ substantially in the shape and bitterness of the fruit. In this study, 32 male Sprague Dawley rats, 12 weeks old and weighing 250-300gr, were used. 8 rats in each group randomly, 1st group control, 2nd group defect (Sham), 3rd group defect + MC group (14 days), 4th group defect + MC group (28 days) separated into the group. In this experimental study, 600 mg/kg/day MC (bitter melon) extract was mixed into the drinking water of groups 3 and 4 and administered to rats by oral tube. One 6 mm diameter cylindrical defects were created in the body of the right tibia bone. No action was taken in Group 1. In Group 2, only tibial defect was made. For 14 days in group 3 and 28 days in group 4, 600 mg/kg/day MC extract was mixed with drinking water and given by oral gavage. Elevated Malondialdehyde (MDA) levels and Myeloperoxidase (MPO) decreased. In groups 3 and 4, osteoblastic activity and osteocyte development increased, while osteopontin and osteonectin expression were found to be positive, while osteoclastic activity decreased compared to the sham group. It is a good antioxidant in the groups where MC is applied. It has been observed that it can have a positive effect on bone repair.
 Keywords: Momordica charantia; Tibial defect; Fracture healing; Bone repair; Antioxidant

Evaluating the Effect of Jointly Administering Synthetic Graft and Ankaferd Blood Stopper for Bone Recovery in Created Bone Deformities in Diabetic Rats

Objective: In the present paper, it was purposed to examine the late impacts of bone tissue recoveries in combination with synthetic grafts and Ankaferd Blood Stopper (ABS), which are effective in bone wound healing in diabetic rats.
 Methods: A total of 64 Wistar albino male rats with diabetes were studied. A bone deformity was generated in the calvarium of diabetic rats. These diabetic rats are divided into 4 different groups. Only saline was applied to the bone defect in group 1, Beta-tricalcium phosphate (β-TCP) graft was administered to Group 2, ABS solution to Group 3, (β-TCP+ABS) were administered to group 4. Eight rats from each group were sacrificed on the 28th day and another 8 rats on the 56th day. Immunohistochemical, histopathological, and Dual Energy X-ray Absorptiometry (DEXA) analyzes of the obtained samples were made.
 Results: In histopathological measurements, osteoblastic activity and bone regeneration were considerably higher in the group treated with group 2 and group 4 when compared to the control group on the 28th and 56th day (p˂ .05). Western blotting findings showed that the osteopontin (OP) and osteonectin (ON) expression at 28th day increased dramatically in the treated with group 4. DEXA analyzes revealed that BMC values in groups treated with group 2, group 3 and group 4 on 28th day were considerably higher than the control group (p˂ .05).
 Conclusion: We can believe and conclude that ABS in combination with a β-TCP bone graft will produce more successful outcomes on wound
 healing and formation of new bone in diabetic rats.

An in vitro study into three different PRF preparations for osteogenesis potential.

To investigate the effect of Advanced Platelet-Rich Fibrin (A-PRF+), Leukocyte Platelet-Rich Fibrin (L-PRF), and injectable Platelet-Rich Fibrin (i-PRF) on osteogenesis of a human osteoblast-like cell line in vitro. Different PRF protocols are used in clinical dentistry in the last years. Recent literature documented the positive impact of PRF derivatives in vivo and in vitro, on different types of cells. However, hardly any literature comparing the new protocols for PRF (the A-PRF+ and i-PRF) with the original protocol of PRF (L-PRF) is present for osteoblast-like cells. A-PRF+, L-PRF, and i-PRF were prepared from six male donors and pre-cultured with 10mL culture medium for 6 days. 5 x103 cells/ml osteoblasts from the osteoblast cell line (U2OS) were seeded and cultured either with conditioned medium derived from the different PRF conditions or with regular culture medium. At five different time points (0, 7, 14, 21, 28 days), the osteogenic capacity of the cells was assessed with Alizarin Red S to visualize mineralization. Also in these cells, the calcium concentration and alkaline phosphatase activity were investigated. Using qPCR, the expression of alkaline phosphatase, osteocalcin, osteonectin, ICAM-1, RUNX-2, and collagen 1a was assessed. In osteoblast-like cells cultured with conditioned medium, the A-PRF+ conditioned medium induced more mineralization and calcium production after 28 days of culturing compared with the control (p< .05). No significant differences were found in the extent of cell proliferation between the different conditions. RUNX-2 and osteonectin mRNA expression in the cells were lower in all PRF-stimulated cultures compared with control at different time points. The i-PRF-conditioned medium induced more ALP activity (p< .05) compared with control and osteoblasts-like cells differentiated more compared with osteoblasts cultured with L-PRF. The three PRF preparations seem to have the capacity to increase the osteogenic potential of osteoblast-like cells. A-PRF+ seems to have the highest potential for mineralization, while i-PRF seems to have the potential to enhance early cell differentiation.

Effects of electric charge on fracture healing

Fracture nonunion is a common and challenging complication. Although direct current stimulation has been suggested to promote fracture healing, differences in cell density near the positive and negative electrodes have been reported during direct current stimulation. This study aimed to explore the effects of these differences on osteoblast proliferation and fracture healing. MC3T3-E1 cells were stimulated by positive and negative charges to observe cell proliferation, apoptosis, and osteogenic factor expression in vitro, while positive and negative charges were connected to the Kirschner wires of the fractures in an in vivo double-toe fracture model in New Zealand white rabbits and fracture healing was assessed in digital radiography (DR) examinations performed on days 1, 15, 30. Bone tissue samples of all rabbits were analysed histologically after the last examination. The results showed that in comparison with the control group, after DC stimulation, the number of cells near the positive electrode decreased significantly (P < 0.05), apoptosis increased (P < 0.05), the expression of osteocalcin, osteoblast-specific genes, and osteonectin decreased significantly near the positive electrode (P < 0.05) and increased significantly at the negative electrode (P < 0.05). The fracture at the positive electrode junction of New Zealand white rabbits did not heal. Histomorphological analysis showed more bone trabeculae and calcified bone in the bone tissue sections of the control group and the negative electrode group than in the positive electrode group. The bone trabeculae were thick and showed good connections. However, positive charge inhibited osteoblast proliferation and a positive charge at fracture sites did not favour fracture healing. Thus, a positive charge near the fracture site may be a reason for fracture nonunion.

Electrospun PCL/fibrin scaffold as a bone implant improved the differentiation of human adipose-derived mesenchymal stem cells into osteo-like cells

Bone remodeling is not able to repair the major injuries caused by trauma, cancer, etc. Bone tissue engineering is aimed to provide implants for bone repair or replacement. To approach such a purpose, polymeric scaffolds along with stem cells are used to fabricate a supportive matrix mimicking in vivo microenvironment. Here, we developed an in vivo-like matrix based on polycaprolactone (PCL) and fibrin to enhance the differentiation of human adipose-derived mesenchymal stem cells into osteo-like cells (AD-MSCs). PCL scaffold was fabricated by electrospinning and coated with fibrin. AD-MSCs were cultured and differentiate on PCL and PCL/fibrin. The viability of cultured cells in comparison with tissue culture plate (TCPS) was evaluated by MTT assay on days 1, 2, and 3 of differentiation. The differentiation was compared to TCPS on days 7, 14, and 21 of differentiation. To evaluate the differentiation efficiency, calcium content, alkaline phosphatase (ALP), and expression of RUNX-2, Collagene-I, Osteocalcin, and Osteonectin were assessed in differentiated cells. A slight increase in cell viability was observed in the TCPS group on day 3 of differentiation. Higher calcium content and ALP activity were observed in PCL and PCL/fibrin groups, respectively. The expression of osteogenic genes was higher in the PCL/fibrin group on days 7, 14, and 21 of differentiation. It seems PCL acts synergistically with fibrin to make an in vivo-like microenvironment and enhanced the differentiation of AD-MSCs into osteo-like cells.

Histological and Immunohistochemical Analysis of Peri-Implant Soft and Hard Tissues in Patients with Peri-Implantitis.

Currently, researchers are focused on the study of cytokines as predictive biomarkers of peri-implantitis (PI) in order to obtain an early diagnosis and prognosis, and for treatment of the disease. The aim of the study was to characterize the peri-implant soft and hard tissues in patients with a peri-implantitis diagnosis. A descriptive observational study was conducted. Fifteen soft tissue (ST) samples and six peri-implant bone tissue (BT) samples were obtained from 13 patients who were diagnosed with peri-implantitis. All the samples were processed and embedded in paraffin for histological and immunohistochemical analyses. A descriptive and quantitative analysis of mast cells and osteocytes, A proliferation-inducing ligand (APRIL), B-cell activating factor (BAFF), osteonectin (ON), and ∝-smooth muscle actin (∝-SMA) was performed. We observed the presence of mast cells in peri-implant soft tissue in all samples (mean 9.21 number of mast cells) and osteocytes in peri-implant hard tissue in all samples (mean 37.17 number of osteocytes). The expression of APRIL-ST was 32.17% ± 6.39%, and that of APRIL-BT was 7.09% ± 5.94%. The BAFF-ST expression was 17.26 ± 12.90%, and the BAFF-BT was 12.16% ± 6.30%. The mean percentage of ON was 7.93% ± 3.79%, and ∝-SMA was 1.78% ± 3.79%. It was concluded that the expression of APRIL and BAFF suggests their involvement in the bone resorption observed in peri-implantitis. The lower expression of osteonectin in the peri-implant bone tissue can also be associated with a deficiency in the regulation of bone remodeling and the consequent peri-implant bone loss.

Diabetes Medication Metformin Inhibits Osteoclast Formation and Activity in In Vitro Models for Periodontitis.

Diabetes and periodontitis are comorbidities and may share common pathways. Several reports indicate that diabetes medication metformin may be beneficial for the periodontal status of periodontitis patients. Further research using appropriate cell systems of the periodontium, the tissue that surrounds teeth may reveal the possible mechanism. Periodontal ligament fibroblasts anchor teeth in bone and play a role in the onset of both alveolar bone formation and degradation, the latter by inducing osteoclast formation from adherent precursor cells. Therefore, a cell model including this type of cells is ideal to study the influence of metformin on both processes. We hypothesize that metformin will enhance bone formation, as described for osteoblasts, whereas the effects of metformin on osteoclast formation is yet undetermined. Periodontal ligament fibroblasts were cultured in the presence of osteogenic medium and 0.2 or 1 mM metformin. The influence of metformin on osteoclast formation was first studied in PDLF cultures supplemented with peripheral blood leukocytes, containing osteoclast precursors. Finally, the effect of metformin on osteoclast precursors was studied in cultures of CD14+ monocytes that were stimulated with M-CSF and receptor activator of Nf-κB ligand (RANKL). No effects of metformin were observed on osteogenesis: not on alkaline phosphatase activity, Alizarin red deposition, nor on the expression of osteogenic markers RUNX-2, Collagen I and Osteonectin. Metformin inhibited osteoclast formation and accordingly downregulated the genes involved in osteoclastogenesis: RANKL, macrophage colony stimulating factor (M-CSF) and osteoclast fusion gene DC-STAMP. Osteoclast formation on both plastic and bone as well as bone resorption was inhibited by metformin in M-CSF and RANKL stimulated monocyte cultures, probably by reduction of RANK expression. The present study unraveling the positive effect of metformin in periodontitis patients at the cellular level, indicates that metformin inhibits osteoclast formation and activity, both when orchestrated by periodontal ligament fibroblasts and in cytokine driven osteoclast formation assays. The results indicate that metformin could have a systemic beneficiary effect on bone by inhibiting osteoclast formation and activity.

Enhancement of Osteoblast Differentiation Using No-Ozone Cold Plasma on Human Periodontal Ligament Cells.

Periodontitis is an inflammatory disease that leads to periodontal tissue destruction and bone resorption. Proliferation and differentiation of cells capable of differentiating into osteoblasts is important for reconstructing periodontal tissues destroyed by periodontitis. In this study, the effects of the nozone (no-ozone) cold plasma (NCP) treatment on osteoblastic differentiation in periodontal ligament (PDL) cells were investigated. To test the toxicity of NCP on PDL cells, various NCP treatment methods and durations were tested, and time-dependent cell proliferation was analyzed using a water-soluble tetrazolium salts-1 assay. To determine the effect of NCP on PDL cell differentiation, the cells were provided with osteogenic media immediately after an NCP treatment to induce differentiation; the cells were then analyzed using alkaline phosphatase (ALP) staining, an ALP activity assay, real time PCR, and Alizarin Red S staining. The NCP treatment without toxicity on PDL cells was the condition of 1-min NCP treatment immediately followed by the replacement with fresh media. NCP increased ALP, osteocalcin, osteonectin, and osteopontin expression, as well as mineralization nodule formation. NCP treatment promotes osteoblastic differentiation of PDL cells; therefore, it may be beneficial for treating periodontitis.

3D-Printed Regenerative Magnesium Phosphate Implant Ensures Stability and Restoration of Hip Dysplasia.

Osteoarthritis of the hip is a painful and debilitating condition commonly occurring in humans and dogs. One of the main causes that leads to hip osteoarthritis is hip dysplasia. Although the current surgical methods to correct dysplasia work satisfactorily in many circumstances, these are associated with serious complications, tissue resorption, and degeneration. In this study, a one-step fabrication of a regenerative hip implant with a patient-specific design and load-bearing properties is reported. The regenerative hip implant is fabricated based on patient imaging files and by an extrusion assisted 3D printing process using a flexible, bone-inducing biomaterial. The novel implant can be fixed with metallic screws to host bone and can be loaded up to physiological loads without signs of critical permanent deformation or failure. Moreover, after exposing the hip implant to accelerated in vitro degradation, it is confirmed that it is still able to support physiological loads even after losing ≈40% of its initial mass. In addition, the osteopromotive properties of the novel hip implant is demonstrated as shown by an increased expression of osteonectin and osteocalcin by cultured human mesenchymal stem cells after 21 days. Overall, the proposed hip implant provides an innovative regenerative and mechanically stable solution for hip dysplasia treatment.

Immobilization of cobalt‐loaded laponite/carboxymethyl chitosan on polycaprolactone nanofiber for improving osteogenesis and angiogenesis activities

AbstractDesigning scaffolds with therapeutic potential to stimulate angiogenesis and osteogenesis is considered a promising strategy in the regeneration of large skeletal defects. In this study, the cobalt ions (Co2+) were incorporated in laponite (LAP) nanodiscs (0.65, 1.3, and 2.6 g/L) and, in combination with carboxymethyl chitosan (CMC), were immobilized on the electrospun polycaprolactone (PCL) nanofiber to fabricate scaffolds for simultaneous enhancing osteogenesis and angiogenesis inductions to human bone marrow mesenchymal stem cells. The Fourier transform infrared, energy dispersive X‐ray, and inductively coupled plasma atomic emission spectrometries confirmed the Co2+‐loading on the LAP nanodiscs and the chemical immobilization on the surface of PCL nanofibers. The result of MTT assay, scanning electron microscopy, and fluorescence staining displayed that Co2+‐loaded scaffolds were biocompatible. Furthermore, the incorporation of Co2+ ions into LAP nanodiscs had significant effects on the calcium deposition and expression of osteogenic markers, alkaline phosphatase and osteonectin, as well as, the expression of angiogenic markers, hypoxia‐inducible factor‐1α and vascular endothelial growth factor of the cells seeded on the scaffolds (p ≤ 0.05). These results suggested that the combination of Co2+ ions with CMC/LAP could result in a synergetic effect on the osteogenesis and angiogenesis stimulations.

Sol-Gel Synthesis, in vitro Behavior, and Human Bone Marrow-Derived Mesenchymal Stem Cell Differentiation and Proliferation of Bioactive Glass 58S

Background:Bioactive glasses 58S, are silicate-based materials containing calcium and phosphate, which dissolved in body fluid and bond to the bone tissue. This type of bioactive glass is highly biocompatible and has a wide range of clinical applications. Methods:The 58S glass powders were synthesized via sol-gel methods, using tetraethyl orthosilicate, triethyl phosphate, and calcium nitrate, as precursors. Upon the analyses of phase and chemical structures of bioactive glass in different gelation times (12, 48, and 100 h), the appropriate heat treatment (at 525, 575, and 625 °C) was performed to eliminate nitrate compounds and stabilize the glass powder samples. The in vitro assay in SBF solution revealed the bioactivity of the synthesized 58S glass through the morphological (SEM), chemical structure (FTIR), release of calcium, phosphorous and silicon elements, pH variations, and weight loss measurements. The behavior of MSCs in the presence of bioactive glass powders was studied by MTT cytotoxicity, cell staining, ALP activity and biomineralization tests, as well as by the evaluation of ALP, osteocalcin, osteonectin, collagen I, and RUNX2 gene expression. Results: The results confirmed a gelation time of 100 h and a calcination temperature of 575 °C at optimal conditions for the synthesis of nitrate-free bioactive glass powders. Conclusion:The glass spherical nanoparticles in the range of 20-30 nm possess the improved bioactivity and osteogenic properties as demanded for bone tissue engineering.

Tetracalcium Phosphate/Monetite/Calcium Sulfate Hemihdrate Biocement Powder Mixtures Prepared by the One-Step Synthesis for Preparation of Nanocrystalline Hydroxyapatite Biocement-Properties and In Vitro Evaluation.

A modified one-step process was used to prepare tetracalcium phosphate/monetite/calcium sulfate hemihydrate powder cement mixtures (CAS). The procedure allowed the formation of monetite and calcium sulfate hemihydrate (CSH) in the form of nanoparticles. It was hypothesized that the presence of nanoCSH in small amounts enhances the in vitro bioactivity of CAS cement in relation to osteogenic gene markers in mesenchymal stem cells (MSCs). The CAS powder mixtures with 15 and 5 wt.% CSH were prepared by milling powder tetracalcium phosphate in an ethanolic solution of both orthophosphoric and sulfuric acids. The CAS cements had short setting times (around 5 min). The fast setting of the cement samples after the addition of the liquid component (water solution of NaH2PO4) was due to the partial formation of calcium sulfate dihydrate and hydroxyapatite before soaking in SBF with a small change in the original phase composition in cement powder samples after milling. Nanocrystalline hydroxyapatite biocement was produced by soaking of cement samples after setting in simulated body fluid (SBF). The fast release of calcium ions from CAS5 cement, as well as a small rise in the pH of SBF during soaking, were demonstrated. After soaking in SBF for 7 days, the final product of the cement transformation was nanocrystalline hydroxyapatite. The compressive strength of the cement samples (up to 30 MPa) after soaking in simulated body fluid (SBF) was comparable to that of bone. Real time polymerase chain reaction (RT-PCR) analysis revealed statistically significant higher gene expressions of alkaline phosphatase (ALP), osteonectin (ON) and osteopontin (OP) in cells cultured for 14 days in CAS5 extract compared to CSH-free cement. The addition of a small amount of nanoCSH (5 wt.%) to the tetracalcium phosphate (TTCP)/monetite cement mixture significantly promoted the over expression of osteogenic markers in MSCs. The prepared CAS powder mixture with its enhanced bioactivity can be used for bone defect treatment and has good potential for bone healing.

Use of Therapeutic Pathogen Recognition Receptor Ligands for Osteo-Immunomodulation.

Therapeutic pathogen recognition receptor (PRR) ligands are reaching clinical practice following their ability to skew the immune response in a specific direction. We investigated the effects of various therapeutic PRR ligands on bone cell differentiation and inflammation. Following stimulation, alkaline phosphatase (ALP) activity (Day 10), osteocalcin, osteonectin expression (Day 14), and calcium deposition (Day 21) were quantified in bone marrow-derived human mesenchymal stem cells (hMSCs). The osteoclastogenic response was determined by measuring tartrate-resistant acid phosphate (TRAP) activity in human monocytes. TNF-α, IL-6, IL-8, and IL-10 expressions were measured by enzyme-linked immunosorbent assay as an indicator of the ligands’ inflammatory properties. We found that nucleic acid-based ligands Poly(I:C) and CpG ODN C increased early ALP activity in hMSCs by 4-fold without affecting osteoclast formation. These ligands did not enhance expression of the other, late osteogenic markers. MPLA, Curdlan, and Pam3CSK4 did not affect osteogenic differentiation, but inhibited TRAP activity in monocytes, which was associated with increased expression of all measured cytokines. Nucleic acid-based ligands are identified as the most promising osteo-immunomodulators, as they favor early osteogenic differentiation without inducing an exaggerated immune-cell mediated response or interfering in osteoclastogenesis and thus can be potentially harnessed for multifunctional coatings for bone biomaterials.

Activation of Human Osteoblasts via Different Bovine Bone Substitute Materials With and Without Injectable Platelet Rich Fibrin in vitro.

IntroductionThe aim of the in vitro study was to compare the effect of four bovine bone substitute materials (XBSM) with and without injectable platelet-reach fibrin for viability and metabolic activity of human osteoblasts (HOB) as well as expression of alkaline phosphatase (ALP), bone morphogenetic protein 2 (BMP-2), and osteonectin (OCN).Materials and MethodsCerabone® (CB), Bio-Oss® (BO), Creos Xenogain® (CX) and MinerOss® X (MO) ± i-PRF were incubated with HOB. At day 3, 7, and 10, cell viability and metabolic activity as well as expression of ALP, OCN, and BMP-2, was examined.ResultsFor non-i-PRF groups, the highest values concerning viability were seen for CB at all time points. Pre-treatment with i-PRF increased viability in all groups with the highest values for CB-i-PRF after 3 and 7 and for CX-i-PRF after 10 days. For metabolic activity, the highest rate among non-i-PRF groups was seen for MO at day 3 and for CB at day 7 and 10. Here, i-PRF groups showed higher values than non-i-PRF groups (highest values: CB + i-PRF) at all time points. There was no difference in ALP-expression between groups. For OCN expression in non-i-PRF groups, CB showed the highest values after day 3, CX after day 7 and 10. Among i-PRF-groups, the highest values were seen for CX + i-PRF. At day 3, the highest BMP-2 expression was observed for CX. Here, for i-PRF groups, the highest increase was seen for CX + i-PRF at day 3. At day 7 and 10, there was no significant difference among groups.ConclusionXBSM sintered under high temperature showed increased HOB viability and metabolic activity through the whole period when compared to XBSM manufactured at lower temperatures. Overall, the combination of XBSM with i-PRF improved all cellular parameters, ALP and BMP-2 expression at earlier stages as well as OCN expression at later stages.

The osteogenic differentiation of human dental pulp stem cells in alginate-gelatin/Nano-hydroxyapatite microcapsules

BackgroundMicrocapsule is considered as a promising 3D microenvironment for Bone Tissue Engineering (BTE) applications. Microencapsulation of cells in an appropriate scaffold not only protected the cells against excess stress but also promoted cell proliferation and differentiation. Through the current study, we aimed to microcapsulate the human Dental Pulp Stem Cells (hDPSCs) and evaluated the proliferation and osteogenic differentiation of those cells by using MTT assay, qRT-PCR, Alkaline phosphatase, and Alizarine Red S.ResultsThe SEM results revealed that Alg/Gel microcapsules containing nHA showed a rough and more compact surface morphology in comparison with the Alg/Gel microcapsules. Moreover, the microencapsulation by Alg/Gel/nHA could improve cell proliferation and induce osteogenic differentiation. The cells cultured in the Alg/Gel and Alg/Gel/nHA microcapsules showed 1.4-fold and 1.7-fold activity of BMP-2 gene expression more in comparison with the control group after 21 days. The mentioned amounts for the BMP-2 gene were 2.5-fold and 4-fold more expression for the Alg/Gel and Alg/Gel/nHA microcapsules after 28 days. The nHA, addition to hDPSCs-laden Alg/Gel microcapsule, could up-regulate the bone-related gene expressions of osteocalcin, osteonectin, and RUNX-2 during the 21 and 28 days through the culturing period, too. Calcium deposition and ALP activities of the cells were observed in accordance with the proliferation results as well as the gene expression analysis.ConclusionThe present study demonstrated that microencapsulation of the hDPSCs inside the Alg/Gel/nHA hydrogel could be a potential approach for regenerative dentistry in the near future.Graphical abstract

The Osteogenetic Potential of Chitosan Coated Implant: An In Vitro Study.

Objective: Chitosan is a promising polymer that has been used for coating dental implants. However, research concerning coatings with implant surfaces other than commercially pure titanium is limited. Therefore, this study aims to clarify the chitosan material's effect with two degrees of deacetylation (DDA) as coatings for laser surface microtopographic implants. Methods: Sixty-three Laser-Lok (LL) implant discs were divided into three groups (21 in each group), and two groups were coated with either 80 or 95 DDA chitosan. The groups were categorized as LL 95, LL 80, or LL control. Then, hMSC-TERT 20 cells were used to evaluate the cell morphology, viability, and osteogenic capacity of the chitosan material 7 and 14 days after culture. Two-way ANOVA followed by one-way analysis of variance (ANOVA) and Tukey's post hoc test were used. Results: All samples were biocompatible and allowed cell attachment. However, cell spreading and attachment were noticeably increased in the LL 95 group. There was a significant increase in the expression of osteogenic markers in chitosan-coated samples compared to the control group. The 95 DDA-coated group exhibited higher ALP, Runx2, osteocalcin, and osteonectin expression compared to the 80 DDA and control groups on days 7 and 14. Conclusion: A high DDA of chitosan promotes biomineralization and osteoblast formation. Therefore, this combination of laser surface and chitosan can enhance future dental implant healing processes and osseointegration.

Effects of alloplastic graft material combined with a topical ozone application on calvarial bone defects in rats.

This study presents the evaluation of the damage in the bone tissue resulting from a calvarial defect in rats and the efficiency of exposure to an ozone application with an alloplastic bone graft on the calvarial bone damage. Wistar male rats (n = 56) were divided into four groups: a control group (n = 14), defect and ozone group (n = 14), defect and graft group (n = 14), and defect, graft, and ozone group (n = 14). Under anaesthesia, a circular full-thickness bone defect was created in all groups, and the experimental groups were further divided into two sub-groups, with 7 rats in each group sacrificed at the end of the 4th and 8th weeks. Bone samples were dissected, fixed in 10% formalin solution, and decalcified with 5% ethylene-diamine-tetraacetic acid (EDTA). After the routine follow-up on tissues, immunostaining of osteopontin and osteonectin antibodies was applied to sections and observed under a light microscope. The control group exhibited osteopontin and osteonectin expression in fibroblasts and inflammatory cells at the end of the 4th week with an acceleration at the 8th week. Ozone administration elucidated new trabecular bone formation by increasing osteoblastic activity. Lastly, our observations underscore that a combination of allograft and ozone application increased the osteoblast, osteocyte, and bone matrix development at the 4th and 8th weeks. Exposure to an ozone application with an alloplastic bone graft on calvarial bone damage may induce osteoblastic activity, matrix development, mature bone cell formation, and new bone formation in rats.

Supplementation of Calcium and Fluoride-Free Water Mitigates Skeletal Fluorosis in Fluoride-Intoxicated Rats.

Fluorosis is a public health concern in 25 countries around the globe. The present study is about the mitigation of fluoride (F) toxicity by giving F-free water (FFW) and calcium (Ca). A study was conducted by taking 76 Wistar rats in two phases, phase I (6 months), where rats were randomly divided into four groups: normal-Ca diet (NCD) 0.5%; low-Ca diet (LCD) 0.25%; NCD + 100 ppm F and LCD + 100 ppm F in groups 1, 2, 3 and 4, respectively. F and Ca were given through water and diet respectively. Phase II is the reversal of fluorosis for 3 months, where LCD group 2 was treated with NCD. Groups 3 and 4 were divided into two subgroups each: 3X and 3Y, and 4X and 4Y, respectively. Groups 3X and 4X received FFW with NCD. Group 3Y continued as phase I and 4Y NCD and F. The biochemical expression, gene expression, biomechanical properties and DXA were studied by standard methods. The results revealed that in phase I, bone turnover was significantly increased whereas bone mineral content and biomechanical properties of group 4 were significantly decreased (p ≤ 0.05) as compared with that of all other groups. Trabecular separation and total porosity increased in groups 2 and 4. Expression of osteocalcin, osteonectin and osteopontin genes was significantly downregulated in group 4. Bone turnover in group 4X was normalised. Expressions of osteocalcin, osteonectin and osteopontin were upregulated after providing NCD and FFW. In conclusion, low calcium aggravates skeletal fluorosis which could be mitigated on supplementation of Ca and FFW.