DMP1 and IFITM5 Regulate Osteogenic Differentiation of MC3T3-E1 on PEO-Treated Ti-6Al-4V-Ca2+/Pi surface.

Despite numerous studies on various surface modifications on titanium and its alloys, it remains unclear what kind of titanium-based surface modifications are capable of controlling cell activity. This study aimed to understand the mechanism at the cellular and molecular levels and investigate the in vitro response of osteoblastic MC3T3-E1 cultured on the Ti-6Al-4V surface modified by plasma electrolytic oxidation (PEO) treatment. A Ti-6Al-4V surface was prepared by PEO at 180, 280, and 380 V for 3 or 10 min in an electrolyte containing Ca2+/Pi ions. Our results showed that PEO-treated Ti-6Al-4V-Ca2+/Pi surfaces enhanced the cell attachment and differentiation of MC3T3-E1 compared to the untreated Ti-6Al-4V control but did not affect cytotoxicity as shown by cell proliferation and cell death. Interestingly, on the Ti-6Al-4V-Ca2+/Pi surface treated by PEO at 280 V for 3 or 10 min, MC3T3-E1 showed a higher initial adhesion and mineralization. In addition, the alkaline phosphatase (ALP) activity significantly increased in MC3T3-E1 on the PEO-treated Ti-6Al-4V-Ca2+/Pi (280 V for 3 or 10 min). In RNA-seq analysis, the expression of dentin matrix protein 1 (DMP1), sortilin 1 (Sort1), signal-induced proliferation-associated 1 like 2 (SIPA1L2), and interferon-induced transmembrane protein 5 (IFITM5) was induced during the osteogenic differentiation of MC3T3-E1 on the PEO-treated Ti-6Al-4V-Ca2+/Pi. DMP1 and IFITM5 silencing decreased the expression of bone differentiation-related mRNAs and proteins and ALP activity in MC3T3-E1. These results suggest that the PEO-treated Ti-6Al-4V-Ca2+/Pi surface induces osteoblast differentiation by regulating the expression of DMP1 and IFITM5. Therefore, surface microstructure modification through PEO coatings with Ca2+/Pi ions could be used as a valuable method to improve biocompatibility properties of titanium alloys.

A new link between apoptosis induced by the metformin derivative HL156A and autophagy in oral squamous cell carcinoma.

The metformin derivative HL156A exerts antitumoral effects in various cancers. Despite evidence in the literature, the underlying molecular mechanisms have not been clearly elucidated. Here, we examined the antiproliferative role and mechanism of HL156A in oral squamous cell carcinoma (OSCC). Using MTT and colony formation assays, we found that HL156A exerts an antiproliferative effect in oral cancer cells in a concentration-dependent manner. Flow cytometry was used to analyze the cell cycle distribution and apoptosis. Exposure to HL156A induced cell cycle arrest at the G2/M transition and increased apoptosis rates, associated with the increased caspase-3/PARP activity. On the other hand, HL156A induced autophagy, as demonstrated by autophagic vacuole staining and quantification of autolysosome-associated LC3BI/II proteins. Interestingly, inhibition of autophagy with chloroquine (CQ) increased the extent of apoptosis and promoted the antiproliferative effect of HL156A in OSCC cell lines, suggesting that autophagy mitigates HL156A-induced apoptosis. The relevance of these observations was confirmed in an in vivo system, as cotreatment with HL156A and CQ inhibited tumor growth in a xenograft mouse model of oral cancer. These results showed that HL156A has an antiproliferative effect associated with cell cycle arrest and apoptosis and induces autophagy to protect cells against apoptosis.

Nano/Micro-Sized Morphologies of Hydroxyapatite Coatings Containing Mn and Si on an Oxidized Ti-6Al-4V Alloy Surface for Dental Implants.

To improve the surface characteristics of Ti-6Al-4V dental implants and the binding between the bone and implant surface, biocompatible oxide films were formed by plasma electrolytic oxidation (PEO). The PEO treatment was performed using electrolyte solutions containing Ca (calcium acetate monohydrate), P(calcium glycerophosphate), Mn (manganese(II) acetate tetrahydrate), and Si (sodium metasilicate nonahydrate), which are the major constituents of bone, for 3 min at 280 V. The morphology and crystalline phase of the PEO-treated surfaces were characterized using field-emission scanning electron microscopy, energy-dispersive X-ray spectrometry, X-ray diffraction, and Fourier transform infrared spectroscopy. All the obtained PEO-treated samples exhibited a morphology comprising porous structures. Oval and irregular pore structures were observed as the Mn content increased. As the Si content increased, the areas occupied by the pores increased. When both, Si and Mn were used for the PEO treatment, the number of nano- to micro-sized pores gradually decreased with the increasing ratios of the constituents.

Plasma Electrolytic Oxidation on Ti-xNb-2Ag-2Pt Alloys for Nano- and Micro-Pore Formation in Electrolyte with Ca and P Ions for Dental Implant Use.

In this study, plasma electrolytic oxidation (PEO) on Ti-xNb-2Ag-2Pt alloys for nano- and micro-pore formation in electrolyte with Ca and P ions for dental implant use was studied using various experimental equipment. The Ti-xNb-2Ag-2Pt alloys were fabricated using a vacuum arc melting furnace, and micro-pores were created through PEO-treatment in an electrolyte containing Ca and P ions. The PEO-treated surface was observed by X-ray diffractometer (XRD), field-emission scanning electron microscopy, and energy dispersive X-ray spectroscopy (EDS). The microstructure of Ti- xNb-2Ag-2Pt alloys showed the transformation of needle-like structure to equiaxed structure, as Nb content increased. Adding small amounts of Ag and Pt to Ti-xNb alloys, microstructure was not observed the significantly difference compared to Ti-xNb. The nano- and micro-pore sizes increased as the Nb content increased after PEO-treatment of Ti-xNb. In the case of Ti-50Nb-2Ag-2Pt, groove structure was observed, also the Ca/P ratio increased as the Nb content increased. The oxide layer thickness of Ti-xNb-2Ag-2Pt alloys was increased, as the Nb content increased.

Electrochemical Analysis of Nano- and Micro-Sized Pore Formed Ti-6Al-4V Alloys in Solution Containing Ca, P, Mn, and Si Ions via Plasma Eletrolytic Oxidation for Bio-Implant Materials.

The purpose of this study was to investigate electrochemical analysis of nano- and micro-sized pore formed Ti-6Al-4V alloys in solution containing Ca, P, Mn and Si ions via plasma eletrolytic oxidation for bio-implant materials. The coatings were produced on Ti-6Al-4V alloy for dental implant using the plasma electrolytic oxidation (PEO) method in electrolytes with the various concentration of 0, 5, and 20% Mn and Si, respectively. Electrochemical potentiodynamic polarization and AC impedance behaviors were carried out in 0.9% NaCl solution at 36.5 ± 1 °C using potentiostat (Potentiostat, EG&G, 362) and electrochemical impedance spectroscope (EIS, EG&G, 1025). The potentiodynamic polarization test with a scan rate of 1.667 mV s-1 was carried out from -1500 mV to 2000 mV. The frequency range used for EIS was 10²-105 Hz. The amplitude of AC signal was 10 mV and 5 points per decade was used. From the potentiodynamic polarization test, PEO treated alloy in electrolyte containing Ca, P, Mn, and Si show a lower corrosion potential than that on the bulk surface. In the case of Mn and Si doped surface, the corrosion resistance increase compared to non-doped surface with Mn and Si elements, and the current density was lower than that of the bulk surface. From the AC impedance test, in the case of Mn and Si doped surface, polarization resistance values were higher than other specimens, and nano- and micro-sized pores were covered with corrosion product consisted Mn and Si elements.

Nanotube Morphology Changes of Ti-xTa-Ag-Pt Alloys with Ta Content for Biomaterials.

In this study, nanotube morphology changes of Ti-xTa-Ag-Pt alloys with Ta content for biomaterials were researched using various experimental instruments. Ti-xTa-Ag-Pt alloys were manufactured in an Ar atmosphere using a vacuum arc-melting furnace with Ta contents of 10 and 50, and then heat-treated at 1100 °C for 1 hr. Nanotube formation of Ti-xTa-Ag-Pt (x = 10, 50 wt%) alloys were performed using a DC power of 30 V in 1.0 M H3PO4 + 0.8 wt% NaF electrolyte solution. Surface characteristics were investigated using an optical microscope, X-ray diffractometer, field-emission scanning electron microscope, energy-dispersive X-ray spectroscopy, and Image analyzer (Image J). Ti-10Ta-Ag-Pt alloy had a needle-like structures, and Ti-Ti-50Ta-Ag-Pt showed the mixed structure (equiaxed and needle-like structures). As the Ta content increased, the α-phase decreased and the ß-phase increased. The highly ordered nanotubes were formed on the ß-phase, whereas disordered nanotubes were formed on needle-like structure of α-phase in Ti-10Ta-Ag-Pt alloy. As the Ta content increases, large and small nanotube diameters became smaller in size. Anatase and rutile phases were formed on the alloy surface. Ta, Ag, and Pt elements were uniformly distributed over the entire surface and at the edge or inside of the nanotube.

Corrosion Behaviors of Zn, Si, and Mn-Doped Hydroxyapatite Coatings Formed on the Ti-6Al-4V Alloy by Plasma Electrolytic Oxidation.

Ti alloy for implant materials is not good biocompatibility between metal implants and natural bone without surface modification. In order to improve the bone attachment ability, implant surface modification is required, and corrosion resistance is also important when a surface modified implant is inserted into the body. In this study, corrosion behaviors of anodic oxide film formed on Ti-6Al-4V alloy with concentration of Mn and Zn ions were studied using various experimental techniques. Ti-6Al-4V ELI disk were used as the substrate for PEO treatment. The sample was used a anodic electrode and PEO treatment was carried out by varying the content of each of the electrolytic solutions containing Zn, Mn, Si, Ca and P, using a Pt bar as a cathodic electrode. Pulsed DC power was used at 280 V for 3 min. For the corrosion behaviors of the specimens, the potentiodynamic polarization and AC impedance test were carried out in 0.9% NaCl solution. From the result of experiments, as Zn ion content increases, the number of pores and the pore size increase. As the Mn ion increases, the number of pores decreases, while the pore size increases. As the ions of Mn increase, the corrosion potential decreases. As Zn ion content increased, the polarization resistance increased.

Nanotube Morphology Changes on the Ti-xNb-Ag-Pt Alloy with Nb Contents.

The purpose of this study was to investigate the nanotube morphology changes of Ti-xNb-Ag-Pt alloys with Nb content. Ti-xNb-Ag-Pt was fabricated using arc melting vacuum furnace. The Ti- xNb-Ag-Pt ingot was further homogenized in an Ar atmosphere at 1100 °C for 1 h in a vacuum and then quenched at 0 °C. Nanotube formation on the samples was performed using anodization method with a DC power supply at 30 V for 2 h in 1 M H3PO4 +0.8 wt.% NaF at 25 °C. The surface morphology was observed using OM, FE-SEM, EDS, and XRD. In the microstructure of Ti-xNb-Ag-Pt alloy, needle-like structures on α and α″ gradually disappeared with increasing Nb, ß-phase equilibrium structure appeared, and particle size decreased. The nanotube morphology of the Ti-xNb-Ag-Pt alloy changed according to the content of Nb. As the Nb content increases, the highly ordered nanotubes have changed to irregular nanotubes. The difference in dissolution area at the bottom of the nanotube was depending on the Nb content.

Surface Observation of Plasma Electrolytic Oxidation-Treated Ti-6Al-4V Alloy After 2-Step Nano-Mesh Formation.

The purpose of this study was to investigate the surface observation of PEO-treated Ti-6Al-4V alloy after 2-step nano-mesh formation was investigated by FE-SEM, EDS, and XRD. Anodic oxidation treatment was performed on the electrolyte containing 0.8 wt.% Na/F and 1M H3PO4 to form a nanotube structure on the Ti-6Al-4V alloy. After removing the nanotube layer, PEO-treatment was performed on the electrolyte containing Mg and Zn ions. After forming the nanotubes, the nanomesh surface was obtained by removing the layer, and the surface roughness increased with cycle number of nanotube formation. Also, as the number of nanotubes increased, the anatase peak increased.

Highly Ordered Nanotube Formation on Beta Typed Ti-xTa Alloy Surface.

In this study, the highly ordered nanotube formation on beta typed Ti-xTa alloy surface was investigated. The Ti-xTa binary alloys were manufactured using a vacuum arc-melting furnace with varying Ta contents (10, 30, and 50 wt%), and then homogenized by heat treatment at 1050 °C for 1 h. The nanotube formation of Ti-xTa (x = 10-50 wt%) alloys were performed using a DC power source of 30 V in 1.0 M H3PO4 + 0.8 wt% NaF electrolyte solution for 2 hrs. The surface characterization was performed using field-emission scanning electron microscopy and energy-dispersive X-ray spectroscopy. The microstructure of Ti-xTa alloy showed martensite structure α', α″-phase, and a '-phase structure. As the Ta content increased, the needle-like structures of α' and α'-phase gradually disappeared and only the equiaxed structure of ß-phase appeared. Nanotube morphology of Ti-xTa alloy changed according to Ta content. As the Ta content increased, the size of the nanotubes decreased and the number of the smaller nanotubes increased. In the cross-sectioned nanotube layer, the gap size between the nanotubes decreased as the Ta content increased.

Functional Elements Coatings on the Plasma Electrolytic Oxidation-Treated Ti-6Al-4V Alloy by Electrochemical Precipitation Method.

In this study, functional elements coatings on the plasma electrolytic oxidation (PEO)-treated Ti-6Al-4V alloy by electrochemical precipitation method were investigated. Ti-6Al-4V ELI disks were used as specimens for PEO and HA coating. The applied voltage and time were selected to be 280 V and 3 minutes for PEO treatment, respectively. The electrochemical precipitation on the Ti-6Al-4V alloy was carried out using cyclic voltammetry with cycle of 3, 5, 10, and 20 from -1.5 V to 0 V (vs. SCE electrode) in electrolyte containing Ca, P, Mg, Mn, Sr, Zn, and Si ions by cyclic voltammetry after PEO treatment. The morphology changes of the coatings on the PEO treated Ti-6Al-4V alloy surface were observed using FE-SEM and EDS. PEO surface has a uniformly distributed circular shape and a porous surface and the deposition of low cycles in electrolyte containing Mg, Mn, Sr, Zn, and Si-HA coated surfaces show uniform circular and granular structures. The precipitates of Mg, Mn, Sr, Zn, and Si-HA on the PEO treated surface showed a large number of circular particles as the number of deposition cycles increases with a mixture of rod-shaped particles and petal-shaped particles around pores. The precipitate nucleates around the pore and grows rapidly as the cycles increase.

Functional Elements Coatings on Ti-6Al-4V Alloy by Plasma Electrolytic Oxidation for Biomaterials.

In this study, functional elements coatings on Ti-6Al-4V alloy by plasma electrolytic oxidation for biomaterials were studied using various experimental techniques. For this study, Ti-6Al-4V ELI disk (grade 5, Timet Co. Ltd., Japan diameter; 10 mm) were used as the substrate for PEO treatment in the electrolyte containing Ca, P, Si, Zn, and Mn ions. The PEO treatment was performed using a pulsed DC power supply at 280 V for 3 min. The PEO-treated surface was observed with field-emission scanning electron microscope (FE-SEM), and energy dispersive X-ray spectrometry (EDS; Oxford ISIS 310, England), Image Analyzer software, and X-ray diffractometer (TF-XRD, X'pert Philips, Netherlands). The number of pores increases, as the Zn ion content increases. And the number of pores decrease, as the Mn ion content increases. The addition of Zn and Mn ions affects the number and size of pores and the area occupied by the pores. Mn and Zn are distributed around pores and in the pores. Anatase and rutile peaks appear and the HA peak shifted to the left in the case of Mn and Zn ion additions.

Hydroxyapatite Coatings Containing Mn and Si on the Oxidized Ti-6Al-4V Alloy for Dental Applications.

The purpose of this study was to investigate hydroxyapatite coatings containing Mn and Si on the oxidized Ti-6Al-4V alloy for dental applications. Dental implant fixture and Ti-6Al-4V ELI disk were used as substrates for plasma electrolytic oxidation (PEO) treatment. PEO treatment was performed at 280 V for 3 min in various solutions. The surface morphologies of the specimens after PEO treatment were observed with a field-emission scanning electron microscope, energy-dispersive X-ray spectroscopy, X-ray diffractometer, and Fourier transform infrared spectroscopy. The breakdown potential for pore formation depended on the added ions in electrolytes. Rough surface with micro-pores was formed after plasma discharge in the electrolytes containing Si and Mn ions. The surface morphologies of implant fixtures were covered with manganese-silicon compounds, as Mn concentration increased. From the XRD analysis, anatase peaks decreased, as Mn and Si contents increased. From the results of FT-IR analysis, Si-HA and Mn-HA was formed on the implant surface.

Pore Shape Changes and Apatite Formation on Zn and Si Ion-Doped HA Films of Ti-6Al-4V After Plasma Electrolytic Oxidation Treatment.

In this study, pore shape changes and apatite formation on zinc (Zn) and silicon (Si) ion-doped hydroxyapatite (HA) films of Ti-6Al-4V by plasma electrolytic oxidation (PEO) treatment has been investigated by several techniques. The PEO films and the Ti-6Al-4V surface after immersion in SBF were observed by X-ray spectroscopy, field-emission scanning electron microscopy, and energy dispersive X-ray spectroscopy. The number of pores decreased as Zn ion concentration increased from 5Zn to 10Zn. The maximum size of pores were increased from 5Zn to 20Zn concentration, whereas, the minimum size of pores decreased. The amount of bone-like apatite formation for the 5Zn/5Si sample was higher than those of other samples immersed in SBF.

Morphology Changes of Plasma Electrolytic Oxidized Ti-6Al-4V Alloy in the Electrolytes Containing Sr and Si Ions.

In this study, morphology changes of plasma electrolytic oxidized (PEO) Ti-6Al-4V alloy in the electrolytes containing Sr and Si ions were researched using field-emission scanning electron microscope, Image J program thin film X-ray diffraction, and energy dispersive X-ray spectroscopy. Pulsed DC power supplied Ti-6Al-4V alloy was used at a 280 V for 3 minutes in the electrolyte containing Sr and Si ions. To determine cell growth, human embryonic kidney cells 293 were grown at a density of 2 × 105 cells per 1.0 ml in well plates. The pore size decreased, as the content of Sr ion increased, whereas, the number of pores per area increased. The Ca/P ratio of PEO treated in electrolyte containing high Sr concentration showed the higher than those of in electrolyte containing low Sr concentration. The peak of HA is shifted to left side, as the concentration of Sr increased. Also, the number of cells increased, as the concentration of Sr increased.

Electrochemical Deposition of Hydroxyapatite Substituted with Magnesium and Strontium on Ti-6Al-4V Alloy.

In the present study, electrochemical deposition of hydroxyapatite substituted with magnesium and strontium on Ti-6Al-4V alloy have investigated. Mg and Sr-doped HAp was coated using subsequently pulsed electrochemical deposition process at 85 °C in the solution contained Ca, Mg, Sr, and P ion. The morphology of Mg/Sr-HAp formed on implant was investigated using scanning electron microscopy and X-ray diffraction. HAp grain size and XRD intensity decreased with Mg2+ and Sr2+ ions. The initial current density was changed with addition of Mg and Sr ion concentration when the constant voltage was applied to specimens. The morphologies and phase of HAp coated layers were affected by the Mg and Sr ion concentration. Results suggest that Mg/Sr-HAp layer formed on Ti can be a potential candidate for dental materials application.

A Simulated Body Fluid Evaluation of TiO2 Barrier Oxide Layer Formed by Electrochemical Reaction.

The modified surface during implantation is considered to be an effective strategy to improve high adhesion of bone cell and osseointegration activity. In this paper, the TiO2 barrier oxide Layer has been fabricated by using the ion characteristic of an electrolytic solution, the surface characteristics have been investigated by EDS, XPS, and FE-SEM. From the analysis of the chemical states, phosphorus and calcium were observed in the TiO2 barrier layer, which were penetrated from the electrolyte into the oxide layer during deposit process. In addition, Ca 2p spectrum was identified into two peaks for Ca 2p3/2 and 2p1/2 at 347.4 and 351.3 eV, which are related to hydroxyapatite. Also, P spectrum was confirmed into two peaks for P1/2 and P3/2 levels with binding energy 134.2 and 133.4 eV, respectively. Thus, the incorporated phosphate species were found mostly in the forms of HPO-4, PO-3. From the result of biological evaluation in simulated body fluid (SBF), the apatite morphologies were effective for bioactive property on the modified surface.

Biocompatibility and Degradation of a Low Elastic Modulus Ti-35Nb-3Zr Alloy: Nanosurface Engineering for Enhanced Degradation Resistance.

In this study, the biocompatibility and degradation behavior of a low elastic modulus Ti-35Nb-3Zr alloy were investigated and compared with that of the conventional orthopedic and dental implant materials, i.e., commercially pure titanium (Cp-Ti) and Ti-6Al-4V alloy. The biocompatibility test results suggested that cells proliferate equally well on Ti-35Nb-3Zr and Cp-Ti. The degradation rates of Cp-Ti and Ti-6Al-4V were ∼68% (p < 0.05) and ∼84% (p < 0.05) lower as compared to Ti-35Nb-3Zr, respectively. Interestingly, the passive current density (ipass (1000mv)) of the Ti-35Nb-3Zr alloy was ∼29% lower than that of Cp-Ti, which suggests that the alloying elements in the Ti-35Nb-3Zr alloy have contributed to its passivation behavior. Nanosurface engineering of the Ti-35Nb-3Zr alloy, i.e., a two-step electrochemical process involving anodization (producing nanoporous layer) and calcium phosphate (CaP) deposition, decreased the degradation rate of the alloy by ∼83% (p < 0.05), and notably, it was similar to the conventional Ti-6Al-4V alloy. Hence, it can be suggested that the nanosurface-engineered low elastic modulus Ti-35Nb-3Zr alloy is a promising material for orthopedic and dental implant applications.

Phenomena of Nano- and Micro-Pore Formation on Ti-(10~50)Ta Alloys by Plasma Electrolytic Oxidation for Dental Implants.

In this study, the phenomena of nano and micro-pore formation on Ti-(10~50)Ta alloys by plasma electrolytic oxidation for dental implants was investigated using various experimental techniques. The Ti­xTa alloys having Ta contents of 10, 30, and 50 wt.% were prepared using arc-melting vacuum furnace. Micro-pore formation was performed using a potentiostat in 1 M H3PO4 electrolyte by using a potentiostat at various applied voltage (180 V, 210 V, and 240 V). The microstructure of Ti­xTa alloys changed from α' phase to ß + α″ phase with Ta content increased. The applied potential increased, the numbers of micro-pore per unit area decreased, whereas the area ratio of occupied by micro-pores increased. The Ta contents increased, the numbers of micro-pore per unit area decreased, whereas the area ratio of occupied by micro-pores increased at 210 V and 240 V. The thickness of oxide layer and micro-pore size can be controlled by applied potential.

Surface Characteristics of Nanotube Formed Ti­25Nb­xZr Alloys.

The purpose of this study was to investigate surface characteristics of nanotube formed Ti­25Nb­xZr alloys. Ti­25Nb­xZr ternary alloys with 0­15 wt% Zr were manufactured in a vacuum arcmelting furnace. Ingots of the alloys were homogenized in an Ar atmosphere at 1000 °C for 12 h, followed by quenching in ice water. Nanotube layers were formed by an electrochemical method in 1 M H3PO4 and 0.8 wt% NaF at 30 V for 1 h. As the Zr content of the Ti­25Nb­xZr Alloys increased, the crystals transformed from a needle-like to equiaxed structure. Two nanotube sizes were obtained on the Ti­25Nb­xZr alloys. As the Zr content was increased, the diameters of the larger tubes decreased, whereas those of the smaller tubes increased. However, increased Zr content in the Ti­25Nb­xZr alloys had little effect on wettability.