Activity Of MG63 Cells Research Articles

Comparison of properties of HAp and CaTiO3-HAp ceramic composite fabricated by different sintering conditions

A hydroxyapatite (HAp) ceramic and a 20% wt CaTiO3-HAp ceramic composite for potential application as biomaterials were produced using single-step sintering (SS) and two-step sintering (TSS) in air. The sample pellets were SS sintered at 1250 ?C with a holding time of 1 h. For TSS, the pellets were heated at 10?C/min to 1250?C (T1) and held at this temperature for 20 min (t1). The temperature was then rapidly lowered at 20?C/min to 1150?C (T2), and held at this temperature for 10 min (t2). The 20% wt CaTiO3-HAp ceramic composite obtained from TSS had a fine grain of 0.56 ? 0.04 mm due to the restraint of grain growth caused by the fast cooling from T1 to T2 and the hard CaTiO3 particles dispersed in the HAp matrix. The microstructure refinement of the 20% wt CaTiO3-HAp ceramic composite fabricated by TSS produced suitable sintered density due to increased locking at triple junctions. This ceramic composite exhibited fracture toughness of 2.10 MPa.m1/2 with a brittleness of 1.97 mm-1/2, indicating that crack propagation was suppressed. MG-63 osteoblast cells were able to adhere to the surface of both HAp and the 20% wt CaTiO3-HAp ceramic composite produced by TSS and were no different between 3 and 7 days of culture. The alkaline phosphatase activity of MG-63 cells was higher on both ceramics cultured in osteogenic differentiation medium than in complete growth medium, which indicated more efficient osteoinductivity.

CeO2 Nanoparticle Bioactive Materials Promote MG-63 Osteogenic Differentiation and Antioxidant Activity Through NRF2 Signaling.

The incidence of bone-related diseases is higher in the elderly population, which greatly affects the patients' quality of life. Throughout this research, we synthesized a biocomposite nanomaterial of CeO2. The unique structural characteristics of CeO2 nanoparticles (CeO2 NPs) were studied by means of XRD, TEM, and SEM. Nanoparticles of an osteosarcoma cell line (MG-63) were assayed for ALP enzyme levels, key proteins in osteoblasts, and stained with Alizarin Red S to assess the physical properties, bioactivity, and calcium deposition of the osteosarcoma cell line. Moreover, we used H2O2 to construct an oxidative stress model to evaluate the antioxidant activity of CeO2 NPs. Experimental data showed that the CeO2 NPs increased the antioxidant capacity of MG-63 cells and significantly increased alkaline phosphatase activity, calcium deposition, and bone growth as manifested by increased expression of bone differentiation proteins BMP2, OCN, OPN, and type I collagen. Interestingly, RNA interference and functional recovery experiments confirmed that CeO2 NPs enhanced the antioxidant activity of MG-63 cells related to NRF2 signaling. In conclusion, the material is expected to be a potential treatment for bone-related diseases.

Use of Various UVC Photofunctionalization Times to Modify Surface Characteristics and Enhance the Biologic Activity of SLA Titanium.

To evaluate the effect of ultraviolet-C light (UVC) photofunctionalization treatment time on the biologic activity of airborne particle-abraded and acid-etched (SLA) titanium surfaces and to analyze its physical and chemical mechanisms. SLA titanium was treated with UVC light for different lengths of time (10 minutes, 20 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, and 24 hours), and then changes to its surface characteristics were evaluated via electron microscope scanning, X-ray photoelectron spectroscopy (XPS), water contact angle measurement, and zeta potential measurement. The effect of UVC photofunctionalization on the biologic processes of SLA titanium surfaces was assessed by analyzing the bovine serum albumin adsorption, adhesion, proliferation, morphology, and alkaline phosphatase activity of MG-63 cells. UVC irradiation did not change the topography of SLA titanium surfaces. As treatment times increased, the water contact angle decreased from 120 degrees to 0 degrees, and the hydrocarbon content decreased. Zeta potential testing showed increased surface potential of photofunctionalized titanium. In vitro testing showed that cell adhesion, proliferation, morphology, and alkaline phosphate (ALP) activity on titanium surfaces were significantly improved by UVC photofunctionalization. UVC photofunctionalization can improve the biologic activity of SLA titanium surfaces by removing hydrocarbons and increasing the surface potential of titanium.

Investigation of physical, mechanical and biological properties of polyhydroxybutyrate-chitosan/graphene oxide nanocomposite scaffolds for bone tissue engineering applications

Mechanical properties appropriate to native tissues, as an essential component in bone tissue engineering scaffolds, plays a significant role in tissue formation. In the current study, Poly-3 hydroxybutyrate-chitosan (PC) scaffolds reinforced with graphene oxide (GO) were made by the electrospinning method. The addition of GO led to a decrease in fibers diameter, an increase in thermal capacity and an improvement in the surface hydrophilicity of nanocomposite scaffolds. A significant increase in the mechanical properties of PC/GO (PCG) nanocomposite scaffolds was achieved due to the inherent strength of GO as well as its uniform dispersion throughout the polymeric matrix owing to hydrogen bonding and polar interactions. Also, lower biological degradation of the scaffolds (~30% in 100 days) due to the presence of GO provides essential mechanical support for bone regeneration. In addition, the bioactivity results showed that GO reinforcement significantly increases the biomineralization on the surface of the scaffolds. Evaluating cell adhesion and proliferation, as well as ALP activity of MG-63 cells on PC and PCG scaffolds indicated the positive effect of GO on scaffolds' biocompatibility. Overall, the improvement of physicochemical, mechanical, and biological properties of GO-reinforced scaffolds shows the potential of PCG nanocomposite scaffolds for bone tissue engineering.

Effect of Ishophloroglucin A Isolated from Ishige okamurae on In Vitro Osteoclastogenesis and Osteoblastogenesis.

The balance between bone-resorbing osteoclasts and bone-forming osteoblasts is essential for the bone remodeling process. This study aimed to investigate the effect of Ishophloroglucin A (IPA) isolated from Ishige okamurae on the function of osteoclasts and osteoblasts in vitro. First, we demonstrated the effect of IPA on osteoclastogenesis in receptor activator of nuclear factor κB ligand (RANKL)-induced RAW 264.7 cells. IPA inhibited the tartrate-resistant acid phosphatase (TRAP) activity and osteoclast differentiation in RANKL-induced RAW 264.7 cells. Moreover, it inhibited the RANKL-induced osteoclast-related factors, such as TRAP, matrix metalloproteinase-9 (MMP-9), and calcitonin receptor (CTR), and transcription factors, such as nuclear factor of activated T cells 1 (NFATc1) and c-Fos. IPA significantly suppressed RANKL-activated extracellular signal-regulated kinase (ERK), and NF-κB in RAW 264.7 cells. Our data indicated that the ERK and NF-κB pathways were associated with the osteoclastogenesis inhibitory activity of IPA. Next, we demonstrated the effect of IPA on osteoblastogenesis in MG-63 cells. IPA significantly promoted alkaline phosphatase (ALP) activity in MG-63 cells, along with the osteoblast differentiation-related markers bone morphogenetic protein 2 (BMP2), type 1 collage (COL1), p-Smad1/5/8, and Runx2, by activating the MAPK signaling pathways. Taken together, the study indicated that IPA could be effective in treating bone diseases, such as osteoporosis.

Investigation on characterization of novel anti-bacterial chitosan/gelatin composite membranes loaded with quercetin via electrophoretic deposition

Peri-implantitis is characterized by inflammation resulting from bacterial infections in peri-implant connective tissue. The purpose of this study was to prepare and characterize chitosan/gelatin (CSG)-based membranes with antibacterial agents to functionalize the surface of titanium (Ti) implants. CSG membranes were prepared on Ti substrates via electrophoretic deposition (EPD). Quercetin, an active flavonoid responsible for fulfilling various plant functions, was introduced as an antibacterial agent to be loaded into the membrane during preparation. The fabrication of quercetin-loaded CSG membranes via EPD was also investigated. Fluorescent microscope, Attenuated Total Reflection Fourier transform infrared spectroscopy, and X-ray diffraction results verified the entrapment of quercetin. The membranes swelled by 150% of mass after rehydration. The antibacterial effects of quercetin on Gram-positive bacteria, such as Staphylococcus aureus and methicillin-resistant Staphylococcus aureus, were verified by spread-plate, scanning electron microscopy, and live/dead staining. Cytological experiments showed that the biocompatibility of rat bone marrow mesenchymal stromal cells was promoted by quercetin-loaded membranes, exclusively in the group with the highest content of quercetin. The quercetin-loaded groups also enhanced the antineoplastic activity of MG-63 cells. These results suggested that quercetin-loaded CSG membranes were successfully fabricated via EPD. Thus, biocompatible and antibacterial membranes could be a potential strategy to functionalize Ti implants.

LBODP016 Lithocholic Acid Induced 25-hydroxyvitamin D-1alpha Hydroxylase (cyp27b1) Expression And Activities In Vitro And In Vivo

Abstract Our previous study reported that oleanolic acid, a natural agonist for bile acid membrane receptor Takeda G protein coupled receptor 5 (TGR5), could exert bone protective effects, increase circulating 1, 25(OH)2D3 level and induce renal CYP27B1 expression and activities. However, it is unclear if bile acids, as the endogenous agonist of bile acid receptors, could regulate CYP27B1 expression and activities in kidney and bone tissues. The present study aimed to characterize the effects of lithocholic acid (LCA), a secondary bile acid, on CYP27B1 expression and activities in vitro using human proximal tubule (HKC-8) and osteosarcoma (MG-63) and in vivo using mature (6-month-old) male C57BL/6 mice. HKC-8 and MG-63 cells were treated with either LCA (10-10 to 10-5 M), PTH (10-7M), or its vehicle for 4 hours. For MG-63 cells, LCA significantly induced CYP27B1 protein expression at 10-10 and 10-9M (P<0. 001) and CYP27B1 mRNA expression at 10-9M (P<0. 001). For HKC-8 cells, LCA significantly induced CYP27B1 protein expression at 10-10 to 10-6M (P<0. 01) and CYP27B1 mRNA expression at 10-10M (P<0. 001). In addition, LCA significantly induced CYP27B1 activity in MG-63 cells (10-10M) (P<0. 05) and in HKC-8 cells (10-6M) (P<0. 01). For the in vivo study, male C57BL/6 mice were fed with 25mg/kg LCA or its vehicle for three consecutive days and sacrificed 5 hours after the last gavage. Serum, kidney, and iliac crests were collected for subsequent analysis. LCA significantly induced circulating 1,25(OH)2D3 levels (P<0. 01) and CYP27B1 protein expression in both kidney and iliac crests in mice (P <0. 05). In addition, the time-dependent change (2, 5, 8 hours) in serum 1,25(OH)2D3 levels were determined upon feeding the male mice with a single dose of 25mg/kg LCA. LCA significantly increased serum 1,25(OH)2D3 levels in male mice at 2 and 5 hours (P <0. 01 and P <0. 05, respectively), and serum 1,25(OH)2D3 levels returned to basal level by 8 hours. LCA significantly increased CYP27B1 protein expression in kidney in mice at 8 hours after treatment (vs. control group, P<0. 05). In summary, our results showed that LCA significantly induced circulating 1,25(OH)2D3 levels, possibly by its actions to induce CYP27B1 expressions and activities in renal and extra-renal (bone) tissues. The study indicated that bile acids and other TGR5 ligands might play an important role in the regulation of vitamin D metabolism. Presentation: No date and time listed

Development of macroporous eggshell derived apatite bone cement for non-load bearing defect repair in orthopedics

A novel and easy approach was attempted to developed a ready to use injectable macroporous apatite bone cement derived from eggshell under physiological conditions where, the solid phase contains hydroxyapatite and eggshell derived β-tricalcium phosphate and the liquid phase is the biopolymeric solution (gelatin and chitosan for improving injectability) with disodium hydrogen phosphate (as binding accelerator) in diluted acetic acid. Also, polysorbate as liquid porogen is incorporated in liquid phase (to enhance cement porosity) and it was compared with the cements containing mannitol as solid porogen. All are mixed in an optimized composition to get desired bone cement. The so-formed cements set within clinically acceptable setting time (≤20 min) and are good injectable (>75%), along with stability at physiological pH (∼7.3–7.4). The apatite phased bone cement formed when the after-set cement immersed in phosphate buffer solution (PBS) and incubated for 7 days at physiological conditions, confirmed by X-ray diffraction and Fourier transform Infrared spectroscopy analysis. The cements hold acceptable compressive strength (2.5–4 MPa), within the range of trabecular bone and are also degradable (19–25%) in PBS and simulated body fluid within 70 days. The average pore size of the eggshell derived apatite bone cements (ESDAPCs) falls in between 50 and 250 μm with interconnectivity, confirmed by scanning electron microscopy and micro-CT analysis verified its macroporous nature. The viability and alkaline phosphatase activity of MG63 cells incubated with the ESDAPCs was found to be significantly higher after 3rd and 14th day when compared to their respective controls. Also, the MG63 cells were fully grown over the surface of the ESDAPCs with increased proliferation and extended filopodia. In conclusion, the developed ESDAPC has the ability to become a potential material for repairing low or non-load bearing defects in orthopaedic applications.

Application value of a selenium-hydroxyapatite nanodelivery system as osteosarcoma treatment

To investigate the anticancer and osteogenic properties and mechanism of action of a selenium (Se4+) hydroxyapatite (HA) nanodelivery system for osteosarcoma (OS) therapy. We loaded different concentrations of Na2SeO3 (3% and 6%) into HA nanoparticles (HANP) for OS therapy. Simultaneously, we tested the stability and pH responsiveness of Se-HANP in vitro experiments, and OS MG-63 cells and mice BMSCs were cultured to further evaluate its permeability, anticancer ability and mineralization ability [alkaline phosphatase (ALP), osteocalcin (OCN)]. In addition, We conducted in vivo experiments by constructing OS mouse models to further investigate the anticancer mechanism of Se-HANP. Se-HANP showed good stability in blood; an acidic environment (pH 5) was more conducive to Se4+ ion release, and could effectively penetrate into OS MG-63 cells. In cell experiments, compared with HANP, Se-HANP could more effectively improve BMSCs viability and enhanced their mineralization performance, such as improving ALP and OCN levels. In addition, Se-HANP could effectively inhibit the activity and invasiveness of MG-63 cells; 6% Se-HANP showed stronger anticancer effects. In OS in vivo mouse experiments, Se-HANP could effectively inhibit tumor tissue growth (volume and weight) in a concentration dependent manner. In addition, Se4+ activated endogenous and exogenous apoptotic pathways by increasing the level of reactive oxygen species (8-OHdG) in tumor cells, leading to tumor cell apoptosis. Se-HANP could effectively penetrate OS cells, and its good stability and pH responsiveness could improve the anticancer efficiency of Se4+. Moreover, its excellent mineralization ability could effectively repair the bone defects caused by OS. Accordingly, Se-HANP can have great potential in OS treatment.

Therapeutic Potential of Naringenin Nanosuspension: In Vitro and In Vivo Anti-Osteoporotic Studies.

Naringenin (NRG) is a flavonoid and has been reported as an anti-osteoporotic agent. However, poor bioavailability may limit the anti-osteoporotic potential of the drug. The purpose of the study was to compare the anti-osteoporotic activity of naringenin nanosuspension (NRG-NS) with the NRG and standard therapeutic drug, raloxifene hydrochloride (RLX). Here, NRG-NS showed anti-osteoporotic activity in MG-63 cells by upregulating the osteocalcin levels. The in vivo anti-osteoporotic activity of NRG-NS was further investigated in an osteoporotic rat model to mimic the post-menopausal condition. The animals were randomized and separated into six groups. The animals were treated with RLX (p.o., 5.4 mg/kg), NRG (p.o., 20 mg/kg), NRG-NS (p.o., 20 mg/kg), and blank-NS for 60 days after completion of a 30-day post-surgery period and compared with control and ovariectomized (OVX) groups. After the treatment, body and uterine weights, biochemical estimation in serum (calcium, phosphorus, acid phosphatase, alkaline phosphatase, osteocalcin), bone parameters (length, diameter, dry weight, density, ash weight, bone mineral content) and bone microarchitecture by histopathology were determined. The results showed the protective effects of NRG-NS on osteoblast-like MG-63 cells. The biochemical estimations confirmed the normalization of parameters viz., alkaline phosphatase, calcium concentrations, and bone density with a decrease in levels of acid phosphatase and inorganic phosphorus with NRG-NS as compared to plain NRG. The results indicated that the oral administration of NRG-NS could be a potential therapeutic formulation for the treatment of osteoporosis.

The Effect of Heat Treatment of β-Tricalcium Phosphate-Containing Silica-Based Bioactive Aerogels on the Cellular Metabolism and Proliferation of MG63 Cells.

β-Tricalcium phosphate was combined with silica aerogel in composites prepared using the sol–gel technique and supercritical drying. The materials were used in this study to check their biological activity and bone regeneration potential with MG63 cell experiments. The composites were sintered in 100 °C steps in the range of 500–1000 °C. Their mechanical properties, porosities, and solubility were determined as a function of sintering temperature. Dissolution studies revealed that the released Ca-/P molar ratios appeared to be in the optimal range to support bone tissue induction. Cell viability, ALP activity, and type I collagen gene expression results all suggested that the sintering of the compound at approximately 700–800 °C as a scaffold could be more powerful in vivo to facilitate bone formation within a bone defect, compared to that documented previously by our research team. We did not observe any detrimental effect on cell viability. Both the alkaline phosphatase enzyme activity and the type I collagen gene expression were significantly higher compared with the control and the other aerogels heat-treated at different temperatures. The mesoporous silica-based aerogel composites containing β-tricalcium phosphate particles treated at temperatures lower than 1000 °C produced a positive effect on the osteoblastic activity of MG63 cells. An in vivo 6 month-long follow-up study of the mechanically strongest 1000 °C sample in rat calvaria experiments provided proof of a complete remodeling of the bone.

Effects of lysophosphatidic acid (LPA) signaling via LPA receptors on cellular functions associated with ATP reduction in osteosarcoma cells treated with ethidium bromide.

Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) exhibits a variety of malignant properties in cancer cells. Intracellular ATP depletion leads to the development of necrosis and apoptosis. The present study aimed to evaluate the effects of LPA receptor-mediated signaling on the regulation of cancer cell functions associated with ATP reduction. Long-term ethidium bromide (EtBr) treated (MG63-EtBr) cells were established from osteosarcoma MG-63 cells. The intracellular ATP levels of MG63-EtBr cells were significantly lower than that of MG-63 cells. LPAR2, LPAR3, LPAR4 and LPAR6 gene expressions were elevated in MG63-EtBr cells. The cell motile and invasive activities of MG63-EtBr cells were markedly higher than those of MG-63 cells. The cell motile activity of MG-63 cells was increased by LPA4 and LPA6 knockdowns. In cell survival assay, cells were treated with cisplatin (CDDP) every 24h for 3 days. The cell survival to CDDP of MG63-EtBr cells was lower than that of MG-63 cells. LPA2 knockdown decreased the cell survival to CDDP of MG-63 cells. The cell survival to CDDP of MG-63 cells was inhibited by (2S)-OMPT (LPA3 agonist). Moreover, the cell survival to CDDP of MG-63 cells was enhanced by LPA4 and LPA6 knockdowns. These results indicate that LPA signaling via LPA receptors is involved in the regulation of cellular functions associated with ATP reduction in MG-63 cells treated with EtBr.

Surface Free Energy of Titanium Disks Enhances Osteoblast Activity by Affecting the Conformation of Adsorbed Fibronectin

This study evaluated the influence of surface free energy (SFE) of titanium disks on the adsorption and conformation of fibronectin (FN) and the biological behavior of osteoblasts cultured on the FN-treated modified surfaces. High [H]-SFE titanium disks were irradiated by a 30 W UV light, while low (L)-SFE titanium disks received no treatment. The surface characteristics of the titanium disks were examined using scanning electron microscope, optical surface profilometer, X-ray photoelectron spectroscopy, and contact angle measurements. Adsorbed FN on different groups was investigated using attenuated total reflection-Fourier transform infrared spectroscopy. MG-63 cells were cultured on FN-treated titanium disks to evaluate the in vitro bioactivity. The experiment showed H-SFE titanium disks adsorbed more FN and acquired more ß-turn content than L-SFE group. MG-63 cells cultured on FN-treated H-SFE titanium disks showed better osteogenic responses, including adhesion, proliferation, alkaline phosphatase activity and mineralization than that on FN-treated L-SFE titanium disks. Compared to L-SFE titanium disks, integrin-β1, integrin-α5 and Rac-1 mRNA levels were significantly higher in MG-63 cells on FN-treated H-SFE after 3 h of culture. These findings suggest that the higher SFE of H-SFE compared to L-SFE titanium disks induced changes in the conformation of adsorbed FN that enhanced the osteogenic activity of MG-63 cells.

Antibacterial ability and osteogenic activity of polyphenol-tailored calcium silicate bone cement.

Calcium silicate-based cement (CSC) has attracted much interest because of its favourable osteogenic effect that supports its clinical use. Although CSC has antibacterial activity, this activity still needs to be improved when used in an infected bone defect. Natural polyphenols have been considered antimicrobial reagents. To this end, three different types of polyphenols (gallic acid (GA), pyrogallol (PG) and tannic acid (TA)) with different concentrations as a liquid phase were mixed with bioactive calcium silicate to enhance the antibacterial activity of CSC. The setting time, antibacterial activity, and osteogenic activity of CSC were studied. Evaluation of antibacterial ability and reactive oxygen species (ROS) was performed using Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus) bacteria, while a human osteoblast-like cell line (MG63) was used to examine osteogenic activity. The experimental results showed that the addition of polyphenols did not remarkably affect the phase composition and morphology of CSC, but changed the setting time and diametral tensile strength. At the same concentration of 1 wt%, the setting time of TA (21 min) was significantly shorter than that of PG (26 min) and GA (68 min), and was indistinguishable from the control cement (20 min). GA had a significantly higher antioxidant activity than PG and TA. As expected, higher concentrations of polyphenols had a more positive impact on ROS generation. More importantly, the incorporation of polyphenols greatly enhanced the antibacterial activity of CSC against E. coli and S. aureus, but had little effect on the in vitro osteogenic activity of MG63 cells and the cytotoxicity of L929 cells. It was concluded that among the three phenolic compounds, the optimal concentration of the liquid phase in the hybrid cement was 5 wt% TA in terms of setting time, strength, antibacterial activity and in vitro osteogenic activity.

Decarboxylated osteocalcin, a possible drug for type 2 diabetes, triggers glucose uptake in MG63 cells.

BACKGROUNDUncarboxylated osteocalcin (GluOC) has been reported to improve glucose metabolism, prevent type 2 diabetes, and decrease the severity of obesity in mice with type 2 diabetes. GluOC can increase glucose uptake in a variety of cells. Glucose metabolism is the main source of energy for osteoblast proliferation and differentiation. We hypothesized that decarboxylated osteocalcin (dcOC), a kind of GluOC, can increase glucose uptake in MG63 cells (osteoblast-like osteosarcoma cells) and influence their proliferation and differentiation.AIMTo investigate the effects of dcOC on glucose uptake in human osteoblast-like osteosarcoma cells and the possible signaling pathways involved.METHODSMG63 cells (human osteoblast-like osteosarcoma cells) were treated with dcOC (0, 0.3, 3, 10, or 30 ng/mL) for 1 and 72 h, and glucose uptake was measured by flow cytometry. The effect of dcOC on cell proliferation was measured with a CCK-8 assay, and alkaline phosphatase (ALP) enzyme activity was measured. PI3K was inhibited with LY294002, and hypoxia-inducible factor 1 alpha (HIF-1α) was silenced with siRNA. Then, GPRC6A (G protein-coupled receptor family C group 6 subtype A), total Akt, phosphorylated Akt, HIF-1α, and glucose transporter 1 (GLUT1) levels were measured by Western blot to elucidate the possible pathways by which dcOC modulates glucose uptake.RESULTSThe glucose uptake of MG63 cells was significantly increased compared with that of the paired control cells after short-term (1 h) treatment with dcOC at different concentrations (0.3, 3, and 10 ng/mL groups, P < 0.01; 30 ng/mL group, P < 0.05). Glucose uptake of MG63 cells was significantly increased compared with that of the paired control cells after long-term (72 h) treatment with dcOC at different concentrations (0.3, 3, and 10 ng/mL groups, P < 0.01; 30 ng/mL group, P < 0.05). DcOC triggered Akt phosphorylation in a dose-dependent manner, and the most effective stimulatory concentration of dcOC for short-term (1 h) was 3 ng/mL (P < 0.01). LY294002 abolished the dcOC-mediated (1 h) promotion of Akt phosphorylation and glucose uptake without affecting GLUT1 protein expression. Long-term dcOC stimulation triggered Akt phosphorylation and increased the protein levels of HIF-1α, GLUT1, and Runx2 in a dose-dependent manner. Inhibition of HIF-1α with siRNA abolished the dcOC-mediated glucose uptake and substantially decreased GLUT1 protein expression. DcOC intervention promoted cell proliferation in a time- and dose-dependent manner as determined by the CCK-8 assay. Treatment with both 3 ng/mL and 10 ng/mL dcOC affected the ALP activity in MG63 cells after 72 h (P < 0.01).CONCLUSIONShort- and long-term dcOC treatment can increase glucose uptake and affect proliferation and ALP activity in MG63 cells. This effect may occur through the PI3K/Akt, HIF-1α, and GLUT1 signaling factors.

In vitro Studies of Polycaprolactone Nanofibrous Scaffolds Containing Novel Gehlenite Nanoparticles.

Background:Recently, many studies have been done on the physicochemical properties and biocompatibility of polycaprolactone (PCL) scaffolds containing ceramic reinforcers in the field of bone tissue engineering. In this study, the physical, mechanical and biological properties of electrospined-fabricated PCL scaffolds containing gehlenite (GLN) nanoparticles (NPs) as a novel bioceramic were investigated.Methods:To obtain the appropriate mechanical properties, the solution contains 3%, 5%, 7%, and 10% wt. of GLN NPs were prepared. Fiber morphology was investigated by scanning electron microscopy. In order to evaluate the NPs distribution, Energy Dispersive X-Ray Spectroscopy, X-ray diffraction, and Fourier Transform Infrared Spectroscopy spectroscopy were used. The scaffold hydrophilicity was measured by the water contact angle test. The tensile test was used to check the mechanical strength of the scaffold. The proliferation of MG-63 cells was evaluated by the MTT test. Alkaline phosphatase (ALP) activity of MG-63 cells was also examined.Results:Average fibers' diameters and porosity of PCL/GLN7% were obtained 150–500 nm and 80%, respectively. An increase in the scaffold hydrophilicity was observed by the addition of GLN NPs. The strength of PCL/GLN7% was higher than the blank PCL scaffold. Cell proliferation of scaffolds containing GLN was higher than the blank PCL scaffold. A significant increase in the secretion of ALP for GLN-loaded scaffolds was seen.Discussion:The results showed that PCL/GLN7% composite scaffold could be a good candidate for bone tissue engineering.Conclusion:The overall results indicate that the scaffold (PCL /GLN7%) has suitable mechanical properties, a great cell compatibility for bone tissue regeneration.

Fabrication and Characterization of silk fibroin scaffold containing ascorbic acid-loaded chitosan nanoparticles for bone regeneration applications

The aim of this study was to fabricate and characterize silk fibroin scaffold containing ascorbic acid-loaded chitosan nanoparticles. Therefore, ascorbic acid-loaded chitosan nanoparticles were first fabricated using the ionic gelation method. Scanning electron microscope (SEM) images and dynamic light scattering (DLS) results showed the spherical nanoparticles with an average particle size of 200 nm. Then, different amounts of nanoparticles were placed in the silk fibroin solution, and finally, the scaffolds were prepared by the freeze-drying method. The effect of nanoparticle concentrations on various properties such as morphology, structural changes, water absorption, drug release, toxicity, adhesion, and alkaline phosphatase activity of MG63 cells were studied. Morphological examinations of the cross-section of the scaffold showed that all scaffolds have a porous structure with interconnected pores. The results of infrared fourier transform spectroscopy(FTIR) confirmed the presence of nanoparticles in the scaffold. The mean size of the pores and porosity percentages reduced as the nanoparticle content rose. The release of ascorbic acid in all samples started with the burst release in the first 24 hours and then continued with a controlled release for up to 14 days. Higher amounts of ascorbic acid were released from the scaffold containing more nanoparticles. Cellular studies showed that the scaffold was non-toxic and that MG63 cells adhered well onto the surface of the scaffold and the pore wall. Also, the proliferation and alkaline phosphatase activity of MG63 cells increased with increasing ascorbic acid amounts.

Synergistic Photoantimicrobial Chemotherapy of Methylene Blue-Encapsulated Chitosan on Biofilm-Contaminated Titanium.

Intensive efforts have been made to eliminate or substantial reduce bacterial adhesion and biofilm formation on titanium implants. However, in the management of peri-implantitis, the methylene blue (MB) photosensitizer commonly used in photoantimicrobial chemotherapy (PACT) is limited to a low retention on the implant surface. The purpose of this study was to assess enhancive effect of water-soluble quaternary ammonium chitosan (QTS) on MB retention on biofilm-infected SLA (sandblasted, large grid, and acid-etched) Ti alloy surfaces in vitro. The effectiveness of QTS + MB with different concentrations in eliminating Gram-negative A. actinomycetemcomitans or Gram-positive S. mutans bacteria was compared before and after PACT. Bacterial counting and lipopolysaccharide (LPS) detection were examined, and then the growth of human osteoblast-like MG63 cells was evaluated. The results indicated that the synergistic QTS + MB with retention ability significantly decreased the biofilm accumulation on the Ti alloy surface, which was better than the same concentration of 1 wt% methyl cellulose (MC). More importantly, the osteogenic activity of MG63 cells on the disinfected sample treated by QTS + MB-PACT modality was comparable to that of sterile Ti control, significantly higher than that by MC + MB-PACT modality. It is concluded that, in terms of improved retention efficacy, effective bacteria eradication, and enhanced cell growth, synergistically, PACT using the 100 μg/mL MB-encapsulated 1% QTS was a promising modality for the treatment of peri-implantitis.

Preparation and Characterization of Vancomycin-Loaded Microsphere Composite Scaffolds

Objective: To fabricate and characterize sustained release antibacterial microsphere composite scaffolds for bone tissue engineering. Methods: First, poly(lactic acid glycolic acid) (PLGA) copolymer, chitosan (CS), and hyaluronic acid (HA) were used to prepare porous microspheres by the emulsion method. Vancomycin (VA) hydrochloride was loaded onto the microspheres (VA@PLGA-CS-HA) for sustained release of drug, after which hydroxyapatite (HAP) was added (VA@PLGA-CS-HA/HAP). The detailed in vitro characterization of the antibacterial sustained-release microspheres was performed using various techniques including the Cell Counting Kit-8 assay; hemolysis, osteogenic, and antibacterial experiments; scanning electron microscopy (SEM); and mechanical testing. Results: SEM showed that the sustained-release PLGA-CS-HA microspheres had a regular spherical shape, and the composite scaffolds of VA@PLGA-CS-HA/HAP microspheres were compact in structure. The relative colony-forming unit was less than 3% after 14 d of intervention. Cytotoxicity testing revealed that the relative growth rate of MC3T3-E1 and MG-63 osteoblast cells was more than 80% when the concentration of VA did not exceed 50 mg/mL. The mechanical strength of the scaffolds was comparable to that of human cancellous bone. The composite scaffolds enhanced the alkaline phosphatase activity of MG-63 cells and promoted the mRNA expression of osteogenic genes such as runt-related transcription factor 2, osteocalcin, osteopontin, and collagen. Conclusion: VA@PLGA-CS-HA microspheres can control the slow and sustained release of VA in vitro. The VA@PLGA-CS-HA/HAP microsphere composite scaffolds has a slower degradation rate and good antibacterial and mechanical properties in vitro.

Roles of endothelial cells in the regulation of cell motility via lysophosphatidic acid receptor-2 (LPA2) and LPA3 in osteosarcoma cells

Lysophosphatidic acid (LPA) signaling via LPA receptors (LPA1 to LPA6) exhibits a variety of biological responses. In tumor microenvironment, endothelial cells promote cancer cell functions. In this study, we investigated the roles of endothelial cells in the regulation of cell motile activity via LPA2 and LPA3 in human osteosarcoma MG-63 cells. In cell motility assay, the cell motile activity of MG-63 cells was markedly increased by the supernatants of endothelial F2 cells. MG-63 cell motility elevated by the supernatants was enhanced by GRI-977143 (LPA2 agonist) and reduced by (2S)-OMPT (LPA3 agonist). LPAR2 and LPAR3 expressions were increased in highly migratory MG63-CR7(F2) cells, which were generated from MG-63 cells by co-culture with F2 cell supernatants. MG63-CR7(F2) cell motility was stimulated by LPA treatment. In the presence of F2 cell supernatants, MG63-CR7(F2) cell motility was markedly enhanced by GRI-977143 and suppressed by (2S)-OMPT. Autotaxin (ATX) enzymatically converts lysophosphatidylcholine (LPC) to LPA. ATX expression was higher in MG63-CR(F2) cells than in MG-63 cells. MG63-CR7(F2) cell motility was markedly increased by LPC in comparison with MG-63 cells. In addition, MG63-CR(F2) cell motility was significantly stimulated by the supernatants of LPC treated F2 cells. The present results suggest that the activation of LPA signaling via LPA2 and LPA3 by endothelial cells is involved in the modulation of cell motile activity of MG-63 cells.