Differentiation Of MG-63 Cells Research Articles

Cytocompatibility, stability and osteogenic activity of powder metallurgy Ta-xZr alloys as dental implant materials.

Tantalum (Ta) and zirconium (Zr) alloys were found to had low elastic modulus and similar biomechanical characteristics as the human bone. However, the biocompatibility and osteogenic potential of Ta-xZr alloyswith different proportions (20, 30, 40 and 50% Zr by atom) remains to be investigated. In this study, the biocompatibility of Ta-xZr alloys and commercially pure titanium (cpTi) was evaluated in vitro by cell counting kit-8 assay. The adhesion of MG63 osteoblasts to the surface of the alloys was observed by fluorescence microscopy, and their morphology was analyzed by scanning electron microscopy (SEM). The expressions of alkaline phosphatase (ALP), Ki67, osteocalcin (OC), collagen-I (Col-I) and Integrin β1 mRNA in the cultured cells were determined by RT-PCR. As a result, Ta-xZr (x = 20, 30, 40 and 50 at%) alloys were non-toxic and supported proliferation of the MG63 cells. The osteoblasts adhered to the Ta-xZr alloys, and subsequently spread and proliferated rapidly. Furthermore, the cells grown on Ta-20Zr and Ta-30Zr expressed high levels of ALP, Col I and OC, indicating that the Ta-xZr alloys can induce osteogenesis. In conclusion, Ta-xZr alloys promoted the adhesion, proliferation and osteogenic differentiation of MG63 cells. The Ta-xZr composites with a higher proportion of Ta exhibited superior osteogenic activity, and Ta-30Zr is therefore a promising alternative for Ti implants.

Taurine Activates BMP-2/Wnt3a-Mediated Osteoblast Differentiation and Mineralization via Akt and MAPK Signaling

We aimed to elucidate the preventive effects of taurine against osteopenia in ovariectomized (OVX) rats and the mechanisms by which taurine regulates osteoblastogenesis in vitro and in vivo. The effects of the taurine on human osteoblast MG-63 cell differentiation and osteoblastogenesis effect in OVX rat were examined Konkuk University in 2018 by evaluating osteoblast differentiation, and expression of osteoblast-specific factors by western blotting analysis. Taurine supplementation significantly improved alkaline phosphatase (ALP) activity and mineralization in a concentration-dependent manner. Further, taurine induced the expression of osteogenic growth factors such as bone morphogenetic protein-2 (BMP-2), runt-related transcription factor 2 (RUNX2), small mothers against decapentaplegic 1/5/8 (SMAD1/5/8), wingless-type MMTV integration site family member 3A (Wnt3a), and collagen type 1 (COL-1) via mitogen-activated protein kinase (MAPK) and serine/threonine protein kinase (Akt). Moreover, the RUNX2 activity of the taurine-treated group was enhanced by protein-protein interactions such as Wnt3a-induced p-AKT/RUNX2 and BMP-mediated SMADs/MAPK/RUNX2 interactions. Our in vitro and in vivo results suggested that taurine can be considered as a potential therapeutic candidate agent for preventing bone loss in postmenopausal osteoporosis.

Enhanced bioactivity and osteogenic property of carbon fiber reinforced polyetheretherketone composites modified with amino groups

Polyetheretherketone (PEEK) is considered as a potential dental and orthopedic implant material owing to its favorable thermal and chemical stability, biocompatibility and mechanical properties. However, the inherent bio-inert and inferior osseointegration of PEEK have hampered its clinical application. In addition, carbon fiber is widely used as a filler to reinforce polymers for sturdy composites owing to its high strength, modulus, etc. In the study, carbon fiber reinforced PEEK (CPEEK) composites were fabricated and modified with amino groups by plasma-enhanced chemical vapor deposition surface modification technique. The surface characterization of composites was evaluated by FE-SEM, EDS, AFM, Water contact angle, XPS and FTIR, which revealed that amino groups were successfully incorporated on the modified CPEEK surface and significantly increased the hydrophilicity. In vitro study, cell adhesion, proliferation, ALP activity, ECM mineralization, real-time PCR analysis, and ELISA analysis showed the adhesion, proliferation and osteogenic differentiation of MG-63 cells on the amino group-modified CPEEK surface were higher than the CPEEK, equal to or better than pure titanium. Hence, the results indicated that the amino group-modified CPEEK possessed enhanced bioactivity and osteogenic property, which may be a potential candidate material for dental implants.

A Mg/Zn-co-doped composite coating on a titanium surface enhances osteogenic activity through the Wnt/β-catenin pathway

Although Ti alloys have good biocompatibility and mechanical properties, they lack bioactivity and cannot integrate with surrounding bone tissue when used as implants; thus, their clinical application is limited. In this study, a Mg/Zn-co-doped micro/nanocomposite coating (TZM) was prepared on a Ti alloy surface by plasma electrolytic oxidation (PEO), and the effect of the coating on the biological behaviour of osteoblasts on its surface was observed in experiments conducted in vitro and in vivo. Based on these findings, the role of the Wnt/β-catenin pathway in the effect of TZM on osteogenic differentiation was studied. The results show that TZM had a porous micro/nanostructure and was uniformly co-doped with Mg and Zn. TZM promoted the adhesion, spreading, proliferation and differentiation of MG-63 cells. In vivo experiments showed that Ti implants with the TZM coating stimulated new bone formation and promoted early implant osseointegration. More importantly, the addition of Wnt3a cells cultured on pure Ti (PT) significantly upregulated the level of β-catenin, and the addition of Dkk1 to cells cultured on the TZM surface significantly inhibited β-catenin activity. In conclusion, the preparation of a Mg/Zn-co-doped micro/nanocomposite coating through PEO is a feasible method for enhancing the osteogenic activity of Ti implants, and the Wnt/β-catenin signalling pathway may be involved in the mechanism by which the coating promotes osteoblast differentiation.

Effect of Silane-Coupling Modification on the Performance of chitosan-poly vinyl Alcohol-Hybrid Scaffolds in Bone Tissue Engineering

Biocompatibility and biodegradability characteristics of some polymers make them an excellent candidate to fabricate porous scaffolds for tissue engineering applications. However, the scaffold mechanical properties and biodegradation rate are vital for bone tissue replacement applications, which can be improved using proper fabrication techniques and cross-linker. In this investigation, Chitosan-polyvinyl alcohol scaffolds were prepared by freeze-drying technique utilizing various weight ratios of 3-Glycidoxypropyl trimethoxysilane (GPTMS) as a bioactive inorganic crosslinker. SEM micrographs indicated interconnected porous structures of cross-linked scaffolds while the average diameter of pores increased as a function of cross-linker enhancement. FTIR analysis was performed to confirm interactions among organic and inorganic components. The mechanical strength test represented that increasing GPTMS content improves the compressive strength of samples. The absorption capacity of the scaffolds in the PBS solution exhibited a decrease in water uptake and biodegradation by increasing silane coupling agent concentration. The formation of needle-like apatite particles proved suitable bioactivity of cross-linked samples. Moreover, MTT assay and ALP expression showed an acceptable adhesion, spreading, proliferation, and differentiation of MG-63 cells on the silane-contained scaffolds. Obtained results warrant further preclinical and clinical evaluations.

A Ca-deficientca-deficient hydroxyapatite (CDHA)/MgF2 bi-layer coating with unique nano-scale topography on biodegradable high-purity Mg

Enhanced corrosion resistance and accelerated new bone formation are desired to make Mg and its alloys to be ideal candidate for bone biomaterial. For this purpose, a CDHA/MgF2 bi-layer coating was prepared on high purity Mg by a combination of fluoride treatment and hydrothermal treatment. The coating exhibited a nanoscale surface topography. Enhanced adhesion strength and corrosion resistance was obtained for the CDHA/MgF2 bi-layer coating. In vitro cell experiment showed that the adhesion, proliferation and differentiation of MG63 cells were significantly improved on Mg with CDHA/MgF2 coating compared to that on Mg with HA coating and MgF2 coating. In conclusion, this study provides a promising surface modification method for Mg metal with enhanced corrosion resistance and superior osteogenic bioactivity.

Biosynthesis and characterization of zinc oxide nanoparticles from Artemisia annua and investigate their effect on proliferation, osteogenic differentiation and mineralization in human osteoblast-like MG-63 Cells

The Biocompatibility and stability of nanoparticles using plants have been widely investigated due to its applications in the biomedical industry. Currently, there is a growing interest in nanoparticles in bone remodelling. Artemisia annua is an herbal plant commonly used in the treatment of various ailments. This study investigated the zinc oxide nanoparticles (ZnO NPs) using the green synthesis technique from A. annua and the effects of A. annua ZnO-NPs on osteoblast differentiation and inhibition of osteoclast formation. The formulated ZnO-NPs from A. annua were characterized by using various spectroscopic and microscopic methods Fourier transform-infrared spectroscopy (FT-IR), transmission electron microscopy (TEM), X-ray diffraction (XRD), and UV–Visible spectroscopy. The disc diffusion method was adopted to test the antimicrobial efficacy of ZnO-NPs. The viability of MG-63 cells were assayed by MTT test and Osteogenic-related assays like Real-time PCR and Mineralization assay were adopted to determine the effects of A. annua ZnO-NPs on the multiplication and differentiation of human osteoblast-like MG-63 cells. The characterization of A. annua ZnO-NPs revealed the crystalline nature with high zinc content and the presence of bioactive compounds from A. annua extract. The synthesized A. annua ZnO-NPs indicate significant antimicrobial potential. Besides, A. annua ZnO-NPs enhanced the proliferation, differentiation, and mineralization without causing significant cytotoxic impact on MG-63 cells. These effects indicate that A. annua ZnO-NPs can both stimulate bone formation via the differentiation of MG-63 cells. Hence, it was concluded that A. annua ZnO-NPs can be a promising agent for the treatment of bone deformities and bone-related diseases, however further research also required to explore the clear mechanism of A. annua ZnO-NPs in the formation and differentiation of MG-63 cells.

Soluble expression and purification of high-bioactivity recombinant human bone morphogenetic protein-2 by codon optimisation in Escherichia coli.

We developed a simple method of preparing recombinant human bone morphogenetic protein-2 (rhBMP-2) with high biological activity. This rhBMP-2 was overproduced in Escherichia coli as a fusion protein with thioredoxin 6xHis-tag at its amino terminus. The cDNA fragment of human bone morphogenetic protein-2 (hBMP-2) fused to the secretion signal of alkaline phosphatase (PhoA) was expressed under T7 promoter in E. coli. After DNA sequence confirmation, the recombinant vector pETpho-bmp2 was transformed into E. coli BL21 (DE3). rhBMP-2 was produced by the recombinant strain pETpho-bmp2/BL21 (DE3) in a soluble form with an yield of 6.2mg/L culture. Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) results showed that the molecular weight of the product was approximately 28 kD. Moreover, rhBMP-2 was secreted as a dimer with a natural structure. rhBMP-2, purified by Ni Nitrilotriacetic acid Agarose (Ni-NTA) affinity chromatography, was used to examine osteosarcoma MG-63 cells and assay the alkaline phosphatase (ALP) activity. Results showed that rhBMP-2 induced MG-63 cell differentiation. When the final concentration was 500ng/mL, the effect was more remarkable and ALP activity reached 525% compared with that of the control group.

Study on the effect of curculigoside on osteoporosis

Objective: To investigate the regulation of curculigoside on osteogenic differentiation of MG63 and the protective effect on osteoporosis model mice. Methods: The effects of curculigoside on the survival rate of dexamethasone or H(2)O(2) treated MG63 were detected by methyl thiazolyl tetrazolium (MTT). The specimens were divided into six groups: blank control group, blank administration group, model group (dexamethasone or H(2)O(2) treatment group), low dose group (dexamethasone or H(2)O(2)+1.0 μmol/L curculigoside), medium dose group (dexamethasone or H(2)O(2)+2.5 μmol/L curculigoside) and high dose group (dexamethasone or H(2)O(2)+5.0 μmol/L curculigoside), the sample size of each group was 10. Western blotting was used to detect the expression of osteogenic differentiation-related proteins [type Ⅰ collagen, integrin β1, osteoblast-specific transcription factor (Osterix), osteocalcin and osteopontin] in MG63 cells after 1, 7 and 14 days incubated with 0, 1.0, 2.5 and 5.0 μmol/L of curculigoside. The sample size for each group at each time point was six. The experimental mice were divided into 4 groups: blank group, model group (dexamethasone treatment group), curculigoside low-dose group (dexamethasone+5 mg/kg curculigoside) and high-dose group (dexamethasone+45 mg/kg curculigoside), twenty each. After treatment, the tibia of the mice in each group were subjected to sacral HE staining. The number of osteoclasts was counted, and the levels of oxidative related factors in serum were determined by enzyme-linked immunosorbent assay (ELISA). Results: The MTT results showed that compared with the blank control group [(100±3.7)%], the cell survival rate decreased to (44.1±5.7)% after treatment with dexamethasone, and the survival rate increased to (79.7±3.8)% after treatment with 5.0 μmol/L of curculigoside. The cell survival rate decreased to (59.1±4.7)% after H(2)O(2) treatment, and the survival rate increased to (80.8±3.5)% after treatment with 2.5 μmol/L of curculigoside. The results of Western blotting showed that the expression of type Ⅰ collagen and integrin β1 in MG63 cells was significantly increased after 1.0, 2.5 and 5.0 μmol/L of curculigoside for 1, 7 and 14 days compared with 0 μmol/L of curculigo side for the same period. After increasing (P<0.05), the expression of Osterix and osteocalcin was significantly increased after 1 day of incubation (P<0.05). However, compared with 0 μmol/L curculigoside treatment, the expression of osteopontin in MG63 cells was not significantly different after incubation with 1.0, 2.5, 5.0 μmol/L of curculigoside for 7 and 14 days (P>0.05). Compared with the blank group, the number of tibia osteoclasts in the osteoporosis model group increased. In the low-dose and high-dose groups of curculigoside, the tibia cortex was more continuous and the number of osteoclasts decreased. Compared with the blank group, the activity of oxygen in the osteoporosis model group was significantly increased (P<0.05), and superoxide dimutase and catalase were significantly decreased (P<0.05). Conclusions: Curculigoside promotes the differentiation of MG63 cells by increasing the expression of osteoblast differentiation-related proteins, and has a certain therapeutic effect on dexamethasone-induced osteoporosis mice.

Antibodies in systemic lupus erythematosus: Prevalence and utility

Parathyroid hormone (PTH) acts as a regulator of calcium homeostasis and bone remodeling. Runx2, an essential transcription factor in bone, is required for osteoblast differentiation. Noncoding RNAs such as long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) play crucial roles in regulating gene expression in osteoblasts. In this study, we investigated the effects of PTH on osteoblast differentiation via Runx2, lncRNA, and miRNA expression in human bone marrow stromal cells (hBMSCs) and human osteoblastic cells (MG63). PTH-treatment of hBMSCs for 24 h, 7 days, and 14 days stimulated Runx2 mRNA expression. Using bioinformatics tools, we identified 17 lncRNAs originating from human Runx2 gene. Among these, lnc-SUPT3H-1:16 (RUNX2-AS1:32) expression was highly up-regulated by the 7 d PTH-treatment in hBMSCs. We also identified miR-6797-5p as the putative target of lnc-SUPT3H-1:16 and Runx2 using bioinformatics tools. PTH-treatment increased the expression of miR-6797-5p in hBMSCs, and overexpression of miR-6797-5p decreased osteoblast differentiation in MG63 cells, suggesting a role for lnc-SUPT3H-1:16 as sponge molecule. A luciferase gene reporter assay identified direct targeting of miR-6797-5p with lnc-SUPT3H-1:16 and 3′UTR Runx2 in MG63 cells. Thus, PTH stimulated the expression of lnc-SUPT3H-1:16, miR-6797-5p and Runx2, and due to the sponging mechanism of lnc- SUPT3H-1:16 towards miR-6797-5p, Runx2 was protected, resulting in the promotion of osteoblast differentiation.

Micro/nano-hierarchical structured TiO2 coating on titanium by micro-arc oxidation enhances osteoblast adhesion and differentiation.

Nano-structured and micro/nano-hierarchical structured TiO2 coatings were produced on polished titanium by the micro-arc oxidation (MAO) technique. This study was conducted to screen a suitable structured TiO2 coating for osteoblast adhesion and differentiation in dental implants. The formulation was characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and wettability testing. Adhesion, proliferation and osteogenic differentiation of MG63 cells were analysed by SEM, Cell Counting Kit-8 (CCK-8) and quantitative real-time PCR. The micro/nano-hierarchical structured TiO2 coating with both slots and pores showed the best morphology and wettability. XRD analysis revealed that rutile predominated along with a minor amount of anatase in both TiO2 coatings. Adhesion and extension of MG63 cells on the micro/nano-hierarchical structured TiO2 coating were the most favourable. MG63 cells showed higher growth rates on the micro/nano-hierarchical structured TiO2 coating at 1 and 3 days. Osteogenic-related gene expression was markedly increased in the micro/nano-hierarchical structured TiO2 coating group compared with the polished titanium group at 7, 14 and 21 days. These results revealed the micro/nano-hierarchical structured TiO2 coating as a promising surface modification and suitable biomaterial for use with dental implants.

Dual RGD-immobilized poly(L-lactic acid) by atmospheric pressure plasma jet for bone tissue engineering.

Surface treatment on PLA substrates by atmospheric pressure plasma jet (APPJ) for polymerization of dual RGD-peptides were investigated. Peptide-modified surfaces have been highlighted as the most promising approach to improve the integration of implants into surrounding bones. By varying the RF power, PLA substrates treated by APPJ process have a tendency to form a hydrophobic surface. The effects on the proliferation and differentiation of MG63 cells were evaluated and osteocalcin (OCN) expression was analyzed using RT-PCR. The water contact angle of the W/APPJ process PLA was approximately 54% of that of the W/O APPJ process PLA substrates. W/APPJ process significantly increased cell proliferation, improved the functionality of the material without using a complicated procedure. We believe that pretreatment using the APPJ processes and dual RGD grafting can be more appropriate than traditional surface modification methods, with more potential for application to bone materials.

Effects of ultraviolet treatment and alendronate immersion on osteoblast-like cells and human gingival fibroblasts cultured on titanium surfaces

In this study, we evaluated the effects of ultraviolet (UV) treatment and alendronate (ALN) immersion on the proliferation and differentiation of MG-63 osteoblast-like cells and human gingival fibroblasts (HGFs) cultured on titanium surfaces. MG-63 cells were used for sandblasted, large grit, and acid-etched (SLA) titanium surfaces, and HGFs were used for machined (MA) titanium surfaces. SLA and MA specimens were subdivided into four groups (n = 12) according to the combination of surface treatments (UV treatment and/or ALN immersion) applied. After culturing MG-63 cells and HGFs on titanium discs, cellular morphology, proliferation, and differentiation were evaluated. The results revealed that UV treatment of titanium surfaces did not alter the proliferation of MG-63 cells; however, HGF differentiation and adhesion were increased in response to UV treatment. In contrast, ALN immersion of titanium discs reduced MG-63 cell proliferation and changed HGFs into a more atrophic form. Simultaneous application of UV treatment and ALN immersion induced greater differentiation of MG-63 cells. Within the limitations of this cellular level study, simultaneous application of UV treatment and ALN immersion of titanium surfaces was shown to improve the osseointegration of titanium implants; in addition, UV treatment may be used to enhance mucosal sealing of titanium abutments.

The Influence of Mg2+ Ions on the In Vitro Efficacy of Chitosan-Titanium Dioxide Nanotubes (CTNTs) Scaffolds

Low mechanical strength and lack of osteoconductive cues are problems associated with chitosan-based scaffolds. This research aimed to fabricate reinforced chitosan-titanium dioxide (TiO2) nanotubes (CTNTs) scaffolds attributed to the enhanced biocompatibility and physical properties of TiO2 nanotubes (TNTs). The incorporation of hydrothermally synthesized TNTs at weight percent of 16 into chitosan was achieved via direct blending and lyophilization. CTNTs scaffolds were further subjected to 24-h adsorption in MgCl2 solutions of 0.5 mM, 1 mM, 2.5 mM, and 5 mM at physiological pH. The adsorption affinity of CTNTs towards Mg2+ ions was high and mainly attributed to the macroporosity of scaffolds and nanocavities of TNTs. The maximum monolayer adsorption capacity of CTNTs for Mg2+ ions was 8.8 mg/g scaffolds. Its adsorption isotherm fitted well with Langmuir isotherm by showing R2 of 0.9995. Fluorescence-based staining, cell viability, and alkaline phosphatase assays indicated that the adsorbed Mg2+ ions onto CTNTs scaffolds aided in promoting higher proliferation and early differentiation of MG63 cells than scaffolds without Mg2+ ions in a concentration-dependent manner. Based on current results, CTNTs scaffolds with Mg2+ ions may be a potential biomaterial for bone regeneration.

Parathyroid hormone-stimulation of Runx2 during osteoblast differentiation via the regulation of lnc-SUPT3H-1:16 (RUNX2-AS1:32) and miR-6797-5p

Parathyroid hormone (PTH) acts as a regulator of calcium homeostasis and bone remodeling. Runx2, an essential transcription factor in bone, is required for osteoblast differentiation. Noncoding RNAs such as long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) play crucial roles in regulating gene expression in osteoblasts. In this study, we investigated the effects of PTH on osteoblast differentiation via Runx2, lncRNA, and miRNA expression in human bone marrow stromal cells (hBMSCs) and human osteoblastic cells (MG63). PTH-treatment of hBMSCs for 24 h, 7 days, and 14 days stimulated Runx2 mRNA expression. Using bioinformatics tools, we identified 17 lncRNAs originating from human Runx2 gene. Among these, lnc-SUPT3H-1:16 (RUNX2-AS1:32) expression was highly up-regulated by the 7 d PTH-treatment in hBMSCs. We also identified miR-6797-5p as the putative target of lnc-SUPT3H-1:16 and Runx2 using bioinformatics tools. PTH-treatment increased the expression of miR-6797-5p in hBMSCs, and overexpression of miR-6797-5p decreased osteoblast differentiation in MG63 cells, suggesting a role for lnc-SUPT3H-1:16 as sponge molecule. A luciferase gene reporter assay identified direct targeting of miR-6797-5p with lnc-SUPT3H-1:16 and 3′UTR Runx2 in MG63 cells. Thus, PTH stimulated the expression of lnc-SUPT3H-1:16, miR-6797-5p and Runx2, and due to the sponging mechanism of lnc- SUPT3H-1:16 towards miR-6797-5p, Runx2 was protected, resulting in the promotion of osteoblast differentiation.

The effect of collector type on the physical, chemical, and biological properties of polycaprolactone/gelatin/nano-hydroxyapatite electrospun scaffold.

Electrospinning is considered a powerful method for the production of fibers in the nanoscale size. Small pore size results in poor cell infiltration, cell migration inhibition into scaffold pores and low oxygen diffusion. Electrospun polycaprolactone/gelatin/nano-hydroxyapatite (PCL/Gel/nHA) scaffolds were deposited into two types of fiber collectors (novel rotating disc and plate) to study fiber morphology, chemical, mechanical, hydrophilic, and biodegradation properties between each other. The proliferation and differentiation of MG-63 cells into the bone phenotype were determined using MTT method, alizarin red staining and alkaline phosphatase (ALP) activity. The rates for disc rotation were 50 and 100 rpm. The pore size measurement results indicated that the fibers produced by the disc rotation collector with speed rate 50 rpm have larger pores as compared to fibers produced by disc rotation at 100 rpm and flat plate collectors. A randomly structure with controlled pore size (38.65 ±0.33 μm) and lower fiber density, as compared to fibers collected by disc rotation with speed rate 100 rpm and flat plate collectors, was obtained. Fibers collected on the rotating disc with speed rate 50 rpm, were more hydrophilic due to larger pore size and therefore, faster infiltration of water into the scaffold and the rate of degradation was higher. These results demonstrate that PCL/Gel/nHA scaffolds made through a rotating disc collector at 50 rpm are more feasible to be used in bone tissue engineering applications due to appropriate pore size and increased adhesion and proliferation of cells, ALP activity and mineral deposits. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 933-950, 2019.

Nanostructured titanium surfaces fabricated by hydrothermal method: Influence of alkali conditions on the osteogenic performance of implants

Hydrothermal method is an easy-to-use approach for creating nanostructured surfaces on titanium (Ti). However, whether the alkali conditions of this method influence the osteogenic potential of the modified surfaces remains unknown. In this study, we fabricated nanostructured surfaces, termed the Ti-1, Ti-5, and Ti-10 groups, by using the hydrothermal method in 1 M, 5 M, and 10 M NaOH aqueous solutions, respectively. An untreated Ti surface served as a control. The osteogenic performance of modified surfaces was systemically investigated, including the proliferation and osteogenic differentiation of human osteoblast-like MG63 cells in vitro and the osteointegration of implants in a rabbit femoral condyle defect model. After hydrothermal treatment, the hydrophilicity of modified surfaces was greatly enhanced. The Ti-1 group showed a nanowire-like topography, while the Ti-5 and Ti-10 groups exhibited a nanopetal-like topography with different pore sizes. Compared with the untreated Ti surface, the modified surfaces showed good cytocompatibility and enhanced the osteogenic differentiation of MG-63 cells. Compared with the other modified surfaces, the Ti-5 group was the most favourable for the osteogenic differentiation of cells, showing higher levels of alkaline phosphatase activity, osteogenic gene expression, mineralization and osteoprotegerin secretion. Twelve weeks after implantation at the bone defects, the Ti-5 group showed superior peri-implant bone regeneration and higher peak push-out force than the other groups. Overall, this study revealed the crucial role of alkali conditions of hydrothermal method in modulating the material characteristics of modified surfaces and their osteogenic performance in vitro and in vivo, highlighting the need for optimizing the processing conditions of hydrothermal method for enhanced osteointegration.

Osteoblastic Differentiation of Functionalized Biphasic Hydroxyapatite and b-Tricalcium Phosphate with Recombinant Human Growth and Differentiation (rhGDF-5)

The purpose of this research was to examine the effect of biphasic hydroxyapatite and b-tricalcium phosphate (HAp-bTCP) immobilized GDF-5 to assess the hypothesis that osteoblast is enhanced and bone formation is promoted when compared to biphasic HAp-bTCP in vitro and in vivo. First, we characterized the release profiles from HAp-bTCP and tested their cell viability, differentiation. After that, we loaded GDF-5 to HAp-bTCP. In the animal study, 4 full thickness critical-size calvarial defects were prepared in a total of 16 male rabbits. The cranial defects were filled with HAP-bTCp, or heparized HAP-bTCP. At 5 and 10 weeks, dissected specimens were processed for histological and histochemical analysis. GDF-5 was considered to be released in a sustained mode from HAp-bTCP. In vitro results provided that the proliferation and differentiation of MG-63 cells to HAp-bTCP-GDF-5 was higher than that of the control group. In addition, in vivo animal studies showed that HAp-bTCP-GDF-5 has a significant improvement to bone formation. HAp-bTCP-GDF-5 is thought to be an excellent biomaterial that delivers osteogenic differentiation factors such as GDF-5, and GDF-5 can be possibly useful as an effective alternative to avail new bone formation.

Phenotype and Viability of MLO-Y4 Cells Is Maintained by TGFβ₃ in a Serum-Dependent Manner within a 3D-Co-Culture with MG-63 Cells.

The osteocyte network inside the bone matrix is of functional importance and osteocyte cell death is a characteristic feature of pathological bone diseases. Osteocytes have emerged as key regulators of bone tissue maintenance, yet maintaining their phenotype during in vitro culture remains challenging. A 3D co-culture system for osteocytes with osteoblasts was recently presented, enabling the determination of more physiological effects of growth factors on cells in vitro. MLO-Y4 cells were embedded within a type I collagen gel and cultured in the presence of surface MG-63 cells. Co-culture was performed in the presence or absence of TGFβ3. Gene expression by quantitative PCR, protein expression by fluorescent immunohistochemistry and cell viability tests were performed. The 3D co-culture induced cell differentiation of MG-63 cells seen by increased type I collagen and osteocalcin mRNA expression. TGFβ3 maintained osteocyte differentiation of MLO-Y4 cells during co-culture as determined by stable E11 and osteocalcin mRNA expression till day 4. Interestingly, most of the effects of TGFβ3 on co-cultured cells were serum-dependent. Also, TGFβ3 reduced cell death of 3D co-cultured MLO-Y4 cells in a serum-dependent manner. This study shows that 3D co-culture upregulates differentiation of MG-63 cells to a more mature osteoblast-like phenotype; while the addition of TGFβ3 maintained the characteristic MLO-Y4 osteocyte-like phenotype and viability in a serum-dependent manner.

Semaphorin3A enhances osteogenesis of MG63 cells through interaction with Schwann cells invitro.

Bone remodeling is under the control of various signals and systems in the body, including the nervous system. Semaphorin (Sema) 3A is a chemorepellent protein which regulates bone mass. Schwann cells, having a pivotal role following nerve injury, interact with Sema3A under numerous circumstances. The present study established a co‑culture system of MG63 and Schwann cells to investigate the role of the interaction between Sema3A and Schwann cells in osteogenesis. The results from the alkaline phosphatase assay, calcium nodule staining and the analysis of the osteogenic gene expression revealed that Sema3A inhibits osteogenic differentiation of MG63 cells in single‑cell culture and promotes osteogenic differentiation of MG63 cells in co‑culture with Schwann cells, in a concentration‑dependent manner. These findings suggest that the presence of Schwann cells induces Sema3A‑associated osteogenic differentiation in bone cells, and also reveals the pivotal role of Sema3A as a regulator in the skeletal and nervous systems, thus contributing to a better understanding of the interaction between these systems.