Biological Behavior Of Osteoblasts Research Articles

Peroxiredoxin 1 promotes proliferation and inhibits differentiation of MC3T3-E1 cells via AKT1 / NF-κB signaling pathway

ObjectivesOsteoporosis is the most common metabolic bone disease worldwide. The decrease in bone mass is primarily accompanied by a decrease in the number and activity of osteoblasts. Peroxiredoxins (PRDXs) are proteins that detect extremely low peroxide levels and act as sensors that regulate oxidation signals, thereby regulating various cellular functions. This study aimed to evaluate the effects of PRDX1 and estrogen on the biological behavior of osteoblasts, including their proliferation and differentiation. MethodsOvariectomized (OVX) mice were used to establish a model of osteoporosis and perform morphological and immunohistochemical analyses. Prdx1 gene knockout and overexpression were performed in mouse MC3T3-E1 pre-osteoblasts to assess proliferation and osteogenic differentiation using the cell counting kit-8, quantitative reverse transcription polymerase chain reaction, western blotting (WB), Alizarin Red S staining, etc. ResultsThe OVX mice exhibited osteoporosis and PRDX1 expression increased. In vitro experiments showed that during the osteogenic differentiation of osteoblasts, PRDX1 expression decreased, while the expression of COL1 and RUNX2 increased. After Prdx1 knockout, the proliferation of osteoblasts decreased; expression of Runx2, ALP, and COL1 increased; and mineralization increased. However, after Prdx1 overexpression, osteoblast proliferation was enhanced, whereas osteogenic differentiation and mineralization were inhibited. Estrogen inhibits the H2O2-induced decrease in osteoblastic differentiation and increase in PRDX1 expression. WB revealed that when LY294002 inhibited the AKT signaling pathway, the levels of p-AKT1, p-P65, and PRDX1 protein in MC3T3-E1 cells decreased. However, when pyrrolidine dithiocarbamate (PDTC) inhibited the NF-κB signaling pathway, the expression of p-AKT1 and PRDX1 did not change except for a significant reduction of p-P65 expression. Furthermore, PDTC reversed the decreased expression of RUNX2, ALP, and COL1 caused by PRDX1 overexpression. ConclusionsPRDX1 promotes the proliferation of osteoblasts and inhibits osteogenic differentiation. Estrogen regulated osteoblastic differentiation by affecting the expression of PRDX1 in osteoblasts, and the effect is related to the AKT1/NF-κB signaling pathway.

Gastrodin improves osteoblast function and adhesion to titanium surface in a high glucose environment

ObjectiveTo investigate the effects of gastrodin on the biological behavior of osteoblasts and osseointegration on the surface of the titanium plate in a high glucose environment, and to explore the possible regulatory mechanisms involved. MethodsA high glucose-induced oxidative damage model of MC3T3-E1 cells was established in vitro to observe the effects of gastrodin on cellular oxidative stress, cell viability, osteogenic differentiation, mineralization, migration, and adhesion ability on the titanium surface. ResultsHigh glucose environment can cause oxidative stress damage to MC3T3-E1 cells, leading to a decrease in cell viability, osteogenesis, migration, adhesion and other functions. Gastrodin can upregulate the expression of antioxidant enzymes (Nrf2 and HO-1) and osteogenic differentiation related proteins (RUNX2 and BMP2) in MC3T3-E1 cells in high glucose environment, thereby inhibiting the excessive production of intracellular reactive oxygen species (ROS), reversing the decrease in cell viability, and improving the osteogenic differentiation and mineralization ability of osteoblasts. And gastrodin alleviated the decline in cell migration ability, improved the morphology of the cytoskeleton and increased the adhesion ability of osteoblasts on the surface of titanium plates in high glucose environment. However, gastrodin itself did not affect the cell viability, osteogenic differentiation and mineralization ability of osteoblasts in normal environment. ConclusionsGastrodin may protect MC3T3-E1 cells osteogenesis and osseointegration on the surface of the titanium plate in vitro by upregulating antioxidant enzymes expression, and attenuating high glucose-induced oxidative stress. Therefore, gastrodin may be a potential drug to address the problem of poor implant osseointegration in patients with diabetes.

The Combined Effects of Material Surface Chemistry and Vitamin D3 on MC3T3-E1 Cells

Separate stimulation by vitamin D3 and material surface chemistry has long witnessed and proved remarkable effects on biological behavior of osteoblasts, but combined stimulation effects of these two have not been reported yet. In this project, the surface of hydroxyl, amino and methyl groups were constructed by silicon-alkyl self-assembly technology. Then, chemical surfaces were to be integrated with vitamin D3 at different concentrations (10−7 mol/L, 10−8 mol/L, 10−9 mol/L) to stimulate MC3T3-E1 osteoblasts. It turned out to be quite successful in establishing the research platform for combined stimulation of material surface chemistry and vitamin D3. Meanwhile, indicators such as osteoblast adhesion spots, cytoskeleton formation, proliferation, and differentiation have been fully examined in the process for us to explore general rules and possible mechanism of combined stimulation effects of these two on early responses of osteoblasts. The results suggest that the combined effects of vitamin D3 and material surface chemistry can induce early responses of osteoblasts on different material surfaces, with differing response degrees. Combined stimulation of material surface chemistry-vitamin D3 causes adhesion plaque and cytoskeleton differences, leading to different early cell behaviors. These findings will provide a strong theoretical basis for the treatment of osteoporosis and the construction of tissue-engineered bone models in vitro.

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.

Long non-coding RNA TUG1 knockdown repressed the viability, migration and differentiation of osteoblasts by sponging miR-214.

The maxillofacial region in the human body is susceptible to fracture and corresponding soft tissue injury. In the current study, the effect of long non-coding RNA (lncRNA) taurine upregulated gene 1 (TUG1) on maxillofacial fracture development was investigated. In total, 50 patients diagnosed with maxillary fracture and 50 healthy volunteers were enrolled in this study. Participants' TUG1 expression level in serum was measured using reverse transcription-quantitative (RT-q)PCR. After transfection with small interfering (si)-TUG1, microRNA (miR)-214 mimic, miR-214 inhibitor, bone morphogenetic protein 2 (BMP2) mimic or a combination, the biological behavior of osteoblasts was evaluated using MTT, Transwell assays, RT-qPCR, flow cytometry and western blot analysis. Recovery experiments were used to explore the potential mechanism. Results demonstrated that TUG1 expression was decreased in the serum of patients with maxillary fractures. Knockdown of TUG1 repressed viability, migration and differentiation and induced apoptosis of osteoblasts. StarBase v2.0 revealed that TUG1 served as a sponge for miR-214 and BMP2 is a direct target of miR-214. Altogether, it was revealed that TUG1 expression was decreased in patients with maxillary fractures and TUG1 knockdown repressed the biological process of osteoblasts by sponging miR-214.

Research progress on the osseointegration of titanium implants promoted by cold atmospheric plasma

The application of cold atmospheric plasma to titanium surface modification has recently become a research focus in the area of material modification. Previous studies found that cold atmospheric plasma can affect the colonization of bacteria and biological behaviors of osteoblasts by changing the surface characteristics of titanium in vitro. In vivo studies reveal that cold atmospheric plasma can promote the process of osseointegration of titanium implants. This review focuses on research on the effects of the surface modification of titanium implants with cold atmospheric plasma on osseointegration.

Effect of low-intensity pulsed ultrasound on the biological behavior of osteoblasts on porous titanium alloy scaffolds: An in vitro and in vivo study

Low-intensity pulsed ultrasound (LIPUS) has been used in patients with fresh fractures, delayed union and non-union to enhance bone healing and improve functional outcome. However, there were few studies concerning the effects of LIPUS on the biological behavior of osteoblasts on porous scaffolds. This study aimed to evaluate the effects of LIPUS on the biological behavior of osteoblasts on porous titanium-6aluminum-4vanadium (Ti6Al4V) alloy scaffolds in vitro and in vivo. Scaffolds were randomly divided into an ultrasound group and a control group. Mouse pre-osteoblast cells were cultured with porous Ti6Al4V scaffolds in vitro. The effects of LIPUS on the biological behavior of osteoblasts were evaluated by observing the adhesion, proliferation, differentiation and ingrowth depth on porous Ti6Al4V scaffolds. In addition, scaffolds were implanted into rabbit mandibular defects in vivo. The effects of LIPUS on bone regeneration were evaluated via micro-CT, fluorescent staining and toluidine blue staining. The results revealed that osteoblast adhered well to the scaffolds, and there was no significant difference in the methyl thiazolyl tetrazolium value between the ultrasound group and the control group (p>0.05). Compared with the control group, ultrasound promoted the alkaline phosphatase activity, osteocalcin levels and ingrowth depth of the cells on the scaffolds (p<0.05). In addition, micro-CT and histomorphological analysis showed that the volume and amount of new bone formation were increased and that bone maturity was improved in the ultrasound group compared to the control group. These results indicate that LIPUS promotes osteoblast differentiation as well as enhances bone ingrowth in porous Ti6Al4V scaffolds, and promotes bone formation and maturity in porous Ti6Al4V scaffolds.

Superiority of calcium-containing nanowires modified titanium surface compared with SLA titanium surface in biological behavior of osteoblasts: A pilot study

ObjectiveThe aim of this study was to present a procedure of preparing calcium-containing nanowires on pretreated micro-structured titanium substrate and explore the superiority of this modified titanium surface compared with SLA titanium surface in biological behavior of osteoblasts. Materials and methodsIn this study, micro-structured titanium substrate was prepared via acid etching. Through hydrothermal treatment with a NaOH solution, nanowires were synthesized on the pretreated micro-structured titanium substrate to prepare the nanowires modified titanium surface (Ti-NW). Then, the Na+ existing in the nanowires was replaced by Ca2+ through secondary hydrothermal treatment with a CaCl2 solution to prepare the calcium-containing nanowires modified titanium surface (Ti-NW-Ca). The sandblasting and acid etching (SLA) titanium surface, which was most commonly used for clinical oral implants, was prepared as the control group. Physical properties of different titanium surfaces including topography, elemental composition, surface roughness and wettability were measured. Assays of spreading, proliferation and differentiation abilities of MC3T3-E1 cells on different titanium surfaces were investigated as well. ResultsThe modified titanium surface was successfully prepared and appeared as calcium-containing nanowire networks, which revealed excellent hydrophilicity and could sustainedly release Ca ions into phosphate buffer saline. The results of in vitro study showed that the Ti-NW-Ca surface had satisfactory osteocompatibility and upregulated the cell spreading, proliferation and differentiation in comparison with the SLA titanium surface. ConclusionsCompared with the SLA titanium surface, the calcium-containing nanowires modified titanium surface demonstrated the ability to enhance osteoblast behavior and suggested the potential to accelerate the dental implant osseointegration.

Effect and mechanism of leptin on bone metabolism in postmenopausal women

Objective To investigate the effects of Leptin on the base of anthropometry, densitometry, and biochemistry and reveal the role of Leptin in bone metabolism in postmenopausal women with osteoporosis (OP). Methods From January 2008 to January 2014, data of 82 postmenopausal females with or without OP were analyzed. They were randomized into two groups: OP (n=40) and conditional control group (CON) (n=42). The expression of Leptin receptors in osteoblasts was observed by immunohistochemistry. 1×10 ng/ml, 1×102 ng/ml, 1×103 ng/ml, 1×104 ng/ml of Leptin were added respectively. Then the proliferation, differentiation and mineralization of osteoblasts were measure at 24 h, 48 h and 96 h. The gene expressions of bone marrow mesenchymal stem cells (BMSCs) derived osteoblasts and adipocytes treated with Leptin were examined by quantitative analysis of Real time-PCR. Results The expression of Leptin receptors in osteoblasts was observed on plasma membranes and cytoplasm in the normal group and osteoporosis group. In osteoporosis group, different concentrations of Leptin enhanced cell proliferation, cell differentiation and cell mineralization. The growth rate of MTT and the concentration of ALP in the serum of were significantly increased with the effects of 1×10 ng/ml, 1×102 ng/ml and 1×103 ng/ml of Leptin, and the action of 1×102 ng/ml Leptin was the most powerful with time dependence. Within 96 hours, the growth rate of osteoblasts increased gradually, and the concentration of ALP increased gradually in 3 weeks. The expression of RANKL/OPG in bone marrow BMSCs derived osteoblasts treated with Leptin were significantly increased. Conclusion Leptin receptor is present in osteoblasts, and Leptin affects biological behaviors of osteoblasts through receptors. The direct effect of Leptin may relate the expression of RANKL/OPG. Then, the balance between bone resorption and bone formation was broken and finally osteoporosis occurred. Lepin may promote osteogenesis and inhibit bone resorption in the differentiation of BMSCs. Key words: Osteoporosis, postmenopausal; Leptin; Bone diseases, metabolic

In vitro study on new bioactive bone screws affecting biological behavior of osteoblasts

Objective To detect the effect of a new bioactive bone screws composed of nanohydroxyapatite/polyamid 66/glass fiber (n-HA/PA66/GF) biomaterial on biological behavior of osteoblasts with a view to a later clinical application of the screws.Methods Mice osteoblasts were co-cultured with n-HA/PA66/GF biomaterial or its extract.Cell growth was detected under a microscope,cell proliferation using MTT assay,apoptosis by flow cytometry,osteocalcin by ELISA method,cell migration using Transwell invasion assay,cell adhesion and growth by scanning electron microscope,and cytoskeleton and actin filament distribution using immunofluorescene.Results Direct contact test demonstrated n-HA/PA66/GF material had no obvious cytotoxicity to osteoblasts.Extract of n-HA/PA66/GF material stimulated osteoblast proliferative activity over time with absorbance value of 0.96 ± 0.14,1.54 ± 0.15,and 2.39 ±0.27 respectively after 2,4,and 6 hours of coculture (P ＜ 0.05).The n-HA/PA66/GF material induced more osteoblasts to entering S period without obvious influence on apoptosis rate and promoted osteocalcin secretion.Migrated cells in medium supplied with n-HA/PA66/GF material or FBS was 8.73 ± 3.26 and 9.47 ± 3.29 in each visual field,but there was no significant difference (P ＞ 0.05).Osteoblasts had a regular form on the surface of n-HA/PA66/GF material,closely adhered to the surface,and reproduced and aggregated with each other to form stratified cell layers.However,nHA/PA66/GF material exhibited no obvious influence on distribution of cytoskeleton and actin filament.Conclusion The new n-HA/PA66/GF screws has excellent cytocompatibility with positive regulatory effect on cell growth,proliferation,secretion,adhesion,cycle,and osteocalcin secretion. Key words: Cell biology; Osteoblasts; Nano-hydroxyapatite/polyamide 66/glass fiber

Surface Modification of NiTi Alloy via Cathodic Plasma Electrolytic Deposition and its Effect on Ni Ion Release and Osteoblast Behaviors

To reduce Ni ion release and improve biocompatibility of NiTi alloy, the cathodic plasma electrolytic deposition (CPED) technique was used to fabricate ceramic coating onto a NiTi alloy surface. The formation of a coating with a rough and micro-textured surface was confirmed by X-ray diffraction, scanning electron microscopy, and energy-dispersive X-ray spectroscopy, respectively. An inductively coupled plasma mass spectrometry test showed that the formed coating significantly reduced the release of Ni ions from the NiTi alloy in simulated body fluid. The influence of CPED treated NiTi substrates on the biological behaviors of osteoblasts, including cell adhesion, cell viability, and osteogenic differentiation function (alkaline phosphatase), was investigated in vitro. Immunofluorescence staining of nuclei revealed that the CPED treated NiTi alloy was favorable for cell growth. Osteoblasts on CPED modified NiTi alloy showed greater cell viability than those for the native NiTi substrate after 4 and 7 days cultures. More importantly, osteoblasts cultured onto a modified NiTi sample displayed significantly higher differentiation levels of alkaline phosphatase. The results suggested that surface functionalization of NiTi alloy with ceramic coating via the CPED technique was beneficial for cell proliferation and differentiation. The approach presented here is useful for NiTi implants to enhance bone osseointegration and reduce Ni ion release in vitro.