Application of a Promising Bone Graft Substitute in Bone Tissue Regeneration: Characterization, Biocompatibility, and In Vivo Animal Study.

The purpose of the present study was to investigate the effect of local hydroxyapatite (HA) combined with extracted sea cucumber (Stichopus hermanni) collagen as a promising bone graft substitute on bone remodeling. Fourier-transform infrared spectroscopy, X-ray diffractometry, transmission electron microscopy, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and Sprague-Dawley rat model were used to characterize the microstructure, in vitro cytotoxicity, and in vivo bone-healing properties of the investigated biocomposite material. Analytical results found that the hydrothermal reaction-synthesized local HA had a hexagonal close-packed structure. The addition of extracted S. hermanni collagen did not influence the microstructure and functional groups of the local HA. Moreover, the MTT assay indicated that the investigated biocomposite material possessed a good in vitro biocompatibility. The in vivo animal study also revealed that the investigated biocomposite material exhibited the highest number of osteoblasts after 14 days of healing. Therefore, the results demonstrate that the local HA combined with extracted S. hermanni collagen could potentially enhance osteoblast formation in promoting bone healing and regeneration.

Hybrid micro/nanostructural surface offering improved stress distribution and enhanced osseointegration properties of the biomedical titanium implant.

OBJECTIVES: The aim of the present study was to investigate the surface characteristic, biomechanical behavior, hemocompatibility, bone tissue response and osseointegration of the optimal micro-arc oxidation surface-treated titanium (MST-Ti) dental implant. MATERIALS AND METHODS: The surface characteristic, biomechanical behavior and hemocompatibility of the MST-Ti dental implant were performed using scanning electron microscope, finite element method, blood dripping and immersion tests. The mini-pig model was utilized to evaluate the bone tissue response and osseointegration of the MST-Ti dental implant in vivo. Data were analyzed by analysis of variance using the Student's t-test (P ≤ 0.05). RESULTS: The hybrid volcano-like micro/nanoporous structure was formed on the surface of the MST-Ti dental implant. The hybrid volcano-like micro/nanoporous surface played an important role to improve the stress transfer between fixture, cortical bone and cancellous bone for the MST-Ti dental implant. Moreover, the MST-Ti implant was considered to have the outstanding hemocompatibility. In vivo testing results showed that the bone-to-implant contact (BIC) ratio significantly altered as the implant with micro/nanoporous surface. After 12 weeks of implantation, the MST-Ti dental implant group exhibited significantly higher BIC ratio than the untreated dental implant group. In addition, the MST-Ti dental implant group also presented an enhancing osseointegration, particularly in the early stages of bone healing. CONCLUSION: It can be concluded that the micro-arc oxidation approach induced the formation of micro/nanoporous surface is a promising and reliable alternative surface modification for Ti dental implant applications.

Micro/nanostructured surface modification using femtosecond laser pulses on minimally invasive electrosurgical devices.

The purpose of the present study was to examine thermal damage and a sticking problem in the tissue after the use of a minimally invasive electrosurgical device with a nanostructured surface treatment that uses a femtosecond laser pulse (FLP) technique. To safely use an electrosurgical device in clinical surgery, it is important to decrease thermal damage to surrounding tissues. The surface characteristics and morphology of the FLP layer were evaluated using optical microscopy, scanning electron microscopy, and transmission electron microscopy; element analysis was performed using energy-dispersive X-ray spectroscopy, grazing incidence X-ray diffraction, and X-ray photoelectron spectroscopy. In the animal model, monopolar electrosurgical devices were used to create lesions in the legs of 30 adult rats. Animals were sacrificed for investigations at 0, 3, 7, 14, and 28 days postoperatively. Results indicated that the thermal damage and sticking situations were reduced significantly when a minimally invasive electrosurgical instrument with an FLP layer was used. Temperatures decreased while film thickness increased. Thermographic data revealed that surgical temperatures in an animal model were significantly lower in the FLP electrosurgical device compared with that in the untreated one. Furthermore, the FLP device created a relatively small area of thermal damage. As already mentioned, the biomedical nanostructured layer reduced thermal damage and promoted the antisticking property with the use of a minimally invasive electrosurgical device. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 865-873, 2017.

Effect of nanostructured thin film on minimally invasive surgery devices applications: characterization, cell cytotoxicity evaluation and an animal study in rat.

BACKGROUND: Minimally invasive surgery is performed using an endoscope and other instruments including the electrosurgical units. However, concerns including surgical smoke, tissue sticking and thermal injury are remaining in electrosurgery. AIMS: Accordingly, a newly developed electrosurgical electrode coating with hydrogenated Cu-incorporated diamond-like carbon (DLC-Cu) film is purposed to improve the instrument performance. METHODS: The morphologies of DLC-Cu surfaces were characterized using transmission electron microscopy, scanning electron microscopy and atomic force microscopy. In this study, lesions were made on the liver lobes of adult rats, using a monopolar electrosurgical unit equipped with untreated stainless steel electrodes or treated-electrodes. Animals were killed for evaluations at 0, 3, 7 and 28 days postoperatively. RESULTS: Treated-electrodes generate less sticking tissues and adhesive blood cells. Thermography revealed that the surgical temperature in liver tissue from the treated-electrode was significantly lower than the untreated-electrode. Total injury area of livers treated with treated-electrodes was significantly smaller than the untreated-electrodes treatment. Moreover, treated-electrodes caused a relatively smaller area of lateral thermal injury, a smaller area of fibrotic tissue and a faster process of remodeling than the untreated-electrodes. Western blot analysis showed that rats treated with treated-electrode expressed lower levels of NF-κB, caspase-3 and MMP-9 than untreated-electrode. Immunofluorescence staining for caspase-3 revealed that the untreated-electrode caused more serious injury. CONCLUSIONS: This study reveals that the plating of electrodes with hydrogenated Cu-incorporated diamond-like carbon film is an efficient method for improving the performance of electrosurgical units, and should benefit wound remodeling. However, more tests must be carried out to confirm these promising findings in human patients.

Research of StemBios Cell Therapy on Dental Implants Containing Nanostructured Surfaces: Biomechanical Behaviors, Microstructural Characteristics, and Clinical Trial.

PURPOSE: The aim of the present study was to examine the osseointegration in low-density bone tissue for SLAffinity-treated implants with StemBios (SB) cell therapy. MATERIALS AND METHODS: The morphologies of SLAffinity-treated surfaces were characterized using scanning electron microscopy. In the animal model, implants were installed in the mandibular canine-premolar area of 12 miniature pigs. Each pig received 3 implants of machine, sand blasted, large grit, and acid etched, and SLAffinity-treated implants. In the clinical trial, 10 patients received 1 SLAffinity-treated implant in the maxilla in the posterior area and 1 patient with low bone tissue density received 2 SLAffinity-treated implants with SB cell therapy. Resonance frequency analysis and computed tomography were assessed monthly over the first 3 months after implant placement. RESULTS: The results demonstrated that surface treatment significantly affected early osseointegration in patients who received SB cell therapy. SB cell therapy transferred the stress caused by the implant more uniformly, and the stress decreased with healing time. SLAffinity-treated implants also proved clinically successful after the 3 months. CONCLUSION: The SLAffinity treatments enhanced osseointegration significantly, especially at early stages of bone tissue healing with SB cell therapy.

Early bone response to machined, sandblasting acid etching (SLA) and novel surface-functionalization (SLAffinity) titanium implants: characterization, biomechanical analysis and histological evaluation in pigs.

The purpose of the present study was to examine early tissue response and osseointegration in the animal model. The surface morphologies of SLAffinity were characterized using scanning electron microscopy and atomic force microscopy. The microstructures were examined by X-ray diffraction, and hardness was measured by nanoindentation. Moreover, the safety and toxicity properties were evaluated using computer-aided programs and cell cytotoxicity assays. In the animal model, implants were installed in the mandibular canine-premolar area of 12 miniature pigs. Each pig received three implants: machine, sandblasted, large grit, acid-etched, and SLAffinity-treated implants. The results showed that surface treatment did affect bone-to-implant contact (BIC) significantly. At 3 weeks, the SLAffinity-treated implants were found to present significantly higher BIC values than the untreated implants. The SLAffinity treatments enhanced osseointegration significantly, especially at early stages of bone tissue healing. As described above, the results of the present study demonstrate that the SLAffinity treatment is a reliable surface modification method.

Effect of exogenous nitric oxide on the proliferation of a human osteoblast (HOS) cell line induced by hydroxyapatite.

AIM: The aim of this study was to test the hypothesis that the proliferation of hydroxyapatite (HA)-induced human osteoblast cell line (HOS cells) may be up-regulated by exogenous nitric oxide (NO). METHODS: HOS cells were cultured on the surface of HA with or without the presence of a NO donor, S-nitroso acetyl penicillamine (SNAP) or nitroso acetyl penicillamine (NAP). 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide) known as carboxy PTIO, a NO scavenger, was added in the cell cultures with or without the presence of SNAP. The cells were also pre-treated with L-N5-(1-iminoethyl)orthinine hydrochloride (L-NIO), an endothelial nitric oxide synthase (eNOS) inhibitor, or anti-integrin alphaV antibody before culturing on HA surfaces with or without the presence of SNAP. Medium, cells alone or cells pretreated with these inhibitors or antibodies was used as the controls. Cell proliferation was assessed by colorimetric assay. RESULTS: The results showed that SNAP, but not NAP, augmented HA-induced HOS cell proliferation. This modulatory effect of SNAP on HA-induced HOS cell proliferation was abolished by carboxy PTIO or anti-integrin alphaV antibody, but only partially reduced by L-NIO. CONCLUSION: Therefore, the results of this study suggest that exogenous NO alone may up-regulate the proliferation of HOS cells attached on HA surfaces via integrin alphaV molecules.

The role of nitric oxide on the proliferation of a human osteoblast cell line stimulated with hydroxyapatite.

The aim of the present study was to test the hypothesis that the proliferation of a human osteoblast cell line (HOS cells) stimulated with hydroxyapatite (HA) may be regulated by nitric oxide (NO). The cells were cultured on the surface of HA. Medium or cells alone were used as controls. L-arginine, D-arginine, 7-NI (an nNOS inhibitor), L-NIL (an iNOS inhibitor), L-NIO (an eNOS inhibitor) or carboxy PTIO, a NO scavenger, was added in the HA-exposed cell cultures. The cells were also precoated with anti-human integrin alphaV antibody. The levels of nitrite were determined spectrophotometrically. Cell proliferation was assessed by colorimetric assay. The results showed increased nitrite production and cell proliferation by HA-stimulated HOS cells up to day 3 of cultures. Anti-integrin alphaV antibody, L-NIO, or carboxy PTIO suppressed, but L-arginine enhanced, nitrite production and cell proliferation of HA-stimulated HOS cells. The results of the present study suggest, therefore, that interaction between HA and HOS cell surface integrin alphaV molecule may activate eNOS to catalyze NO production which, in turn, may regulate the cell proliferation in an autocrine fashion.

Cyclic-AMP-dependent proliferation of a human osteoblast cell line (HOS cells) induced by hydroxyapatite: effect of exogenous nitric oxide.

UNLABELLED: BACKGROUND AND AIMS OF THE WORK: Nitric oxide (NO) has been reported to enhance the production of cAMP by hydroxyapatite (HA)-induced a human osteoblast cell line (HOS cells). The aim of the present study was to test the hypothesis that exogenous NO may up-regulate the proliferation of hydroxyapatite (HA)-induced HOS cells via the cyclic-AMP-protein kinase A (PKA) pathway. METHODS: HOS cells were pre-incubated with ODQ (guanylyl cyclase inhibitor), SQ22536 (adenylyl cyclase inhibitor), forskolin (adenylyl cyclase activator), IBMX [phosphodiesterase (PDE) inhibitor], siguazodan (PDE3 inhibitor), rolipram (PDE4 inhibitor), or KT5720 (PKA inhibitor), and then, cultured on the surface of HA with or without the presence of SNAP (NO donor). The HOS cell cultures on the HA surface were added with br-cGMP (cGMP analogue), db-cAMP (cAMP analogue) with or without SNAP. The cell proliferation was assessed by a colorimetric assay. RESULTS: The up-regulatory effect of SNAP on HA-induced HOS cell proliferation was suppressed by SQ22536 and KT5720, but enhanced by db-cAMP, IBMX, and rolipram. The HA-induced HOS cell proliferation with or without the presence of SNAP was unaltered by ODQ br-cGMP and siguazodan. CONCLUSION: These results suggest, therefore, that HA-induced HOS cell proliferation may be mediated by the cAMP-PKA pathway regulated by PDE4 and that exogenous NO may amplify this cyclic nucleotide pathway, thereby augmenting HA-induced HOS cell proliferation.

The effect of nitric oxide on the production of cyclic AMP by a human osteoblast (HOS) cell line stimulated with hydroxyapatite.

The aim of the present study was to determine the effect of nitric oxide (NO) on the production of cyclic AMP (cAMP) by a human osteoblast cell line (HOS cells) stimulated with hydroxyapatite. Cells were cultured on the HA surfaces with or without the presence of NO donors (SNAP and NAP) for 3 days. The effect of adenylyl cyclase inhibitor (SQ22536), NO scavenger (carboxy PTIO) or endothelial nitric oxide synthase (eNOS) inhibitor (L-NIO), was assessed by adding these to the cultures of HA-stimulated HOS cells with or without the presence of SNAP. Furthermore, HOS cells were pre-treated with anti-human integrin alphaV antibody prior to culturing on HA surfaces with or without the presence of SNAP. The levels of cAMP and cGMP were determined from the 3-day culture supernatants. The results showed that the production of cAMP but not cGMP by HA-stimulated HOS cells was augmented by SNAP. SQ22536 and carboxy PTIO suppressed but L-NIO only partially inhibited the production of cAMP by HA-stimulated HOS cells with or without the presence of exogenous NO. Pre-treatment of the cells with anti-human integrin alphaV antibody suppressed the production of cAMP by HA-stimulated HOS cells with or without the presence of NO. Therefore, the results of the present study suggest that NO may up-regulate the production of cAMP, perhaps, by augmenting adenylyl cyclase activity initiated by the binding between HOS cell-derived integrin alphaV and HA surface.

The effect of nitric oxide on the production of prostaglandin E2 by hydroxyapatite-stimulated a human osteoblast (HOS) cell line.

The aim of the present study was to determine the effect of nitric oxide (NO) on the production of prostaglandin E2 (PGE2) by a human osteoblast cell line (HOS cells) stimulated with hydroxyapatite. Cells were cultured on the HA surfaces with or without the presence of NO donors (SNAP and NAP) for 3 days. The effect of NO scavenger, carboxy PTIO, or endothelial nitric oxide synthase (eNOS) inhibitor, L-NIO, was assessed by adding this scavenger in the cultures of HA-stimulated HOS cells with or without the presence of SNAP. Furthermore, HOS cells were pre-treated with anti-human integrin alphaV antibody, indomethacin, a non-specific inhibitor, aspirin, a COX-1 inhibitor, or nimesulide, a COX-2 inhibitor, prior to culturing on HA surfaces with or without the presence of SNAP. The levels of PGE2 were determined from the 3 day culture supernatants. The results showed that the production of PGE2 by HA-stimulated HOS cells was augmented by SNAP. Carboxy PTIO suppressed but L-NIO only partially inhibited the production of PGE2 by HA-stimulated HOS cells with or without the presence of exogenous NO. Pre-treatment of the cells with anti-human integrin alphaV antibody, indomethacin or nimesulide but not aspirin suppressed the production of PGE2 by HA-stimulated HOS cells with or without the presence of NO. Therefore, the results of the present study suggest that NO may up-regulate the production of PGE2 by augmenting the COX-2 pathway initiated by the binding between HOS cell-derived integrin alphaV and HA surface.