A review on gold nanoparticles as an innovative therapeutic cue in bone tissue engineering: Prospects and future clinical applications.

Bone damage is a complex orthopedic problem primarily caused by trauma, cancer, or bacterial infection of bone tissue. Clinical care management for bone damage remains a significant clinical challenge and there is a growing need for more advanced bone therapy options. Nanotechnology has been widely explored in the field of orthopedic therapy for the treatment of a severe bone disease. Among nanomaterials, gold nanoparticles (GNPs) along with other biomaterials are emerging as a new paradigm for treatment with excellent potential for bone tissue engineering and regenerative medicine applications. In recent years, a great deal of research has focused on demonstrating the potential for GNPs to provide for enhancement of osteogenesis, reduction of osteoclastogenesis/osteomyelitis, and treatment of bone cancer. This review details the latest understandings in regards to GNPs based therapeutic systems, mechanisms, and the applications of GNPs against various bone disorders. The present review aims to summarize i) the mechanisms of GNPs in bone tissue remodeling, ii) preparation methods of GNPs, and iii) functionalization of GNPs and its decoration on biomaterials as a delivery vehicle in a specific bone tissue engineering for future clinical application.

Development of a Temperature-Responsive Hydrogel Incorporating PVA into NIPAAm for Controllable Drug Release in Skin Regeneration.

Melanoma, a highly malignant and aggressive form of skin cancer, poses a significant global health threat, with limited treatment options and potential side effects. In this study, we developed a temperature-responsive hydrogel for skin regeneration with a controllable drug release. The hydrogel was fabricated using an interpenetrating polymer network (IPN) of N-isopropylacrylamide (NIPAAm) and poly(vinyl alcohol) (PVA). PVA was chosen for its adhesive properties, biocompatibility, and ability to address hydrophobicity issues associated with NIPAAm. The hydrogel was loaded with doxorubicin (DOX), an anticancer drug, for the treatment of melanoma. The NIPAAm-PVA (N-P) hydrogel demonstrated temperature-responsive behavior with a lower critical solution temperature (LCST) around 34 °C. The addition of PVA led to increased porosity and faster drug release. In vitro biocompatibility tests showed nontoxicity and supported cell proliferation. The N-P hydrogel exhibited effective anticancer effects on melanoma cells due to its rapid drug release behavior. This N-P hydrogel system shows great promise for controlled drug delivery and potential applications in skin regeneration and cancer treatment. Further research, including in vivo studies, will be essential to advance this hydrogel system toward clinical translation and impactful advancements in regenerative medicine and cancer therapeutics.

In-situ forming injectable GFOGER-conjugated BMSCs-laden hydrogels for osteochondral regeneration.

The collagen-mimetic peptide GFOGER possesses the chondrogenic potential and has been used as a cell adhesion peptide or chondrogenic inducer. Here, we prepared an injectable in situ forming composite hydrogel system comprising methoxy polyethylene glycol-b-polycaprolactone (MPEG-PCL) and GFOGER-conjugated PEG-PCL (GFOGER-PEG-PCL) with various GFOGER concentrations based on our recently patented technology. The conjugation of GFOGER to PEG-PCL was confirmed by 1H NMR, and the particle size distribution and rheological properties for the sol-gel transition behavior of the samples with respect to the GFOGER content were evaluated systemically. In vitro experiments using rat bone marrow-derived mesenchymal stem cells (BMSCs) revealed that the GFOGER-PEG-PCL hydrogel significantly enhanced expression of integrins (ß1, α2, and α11), increased expression of FAK, and induced downstream signaling of ERK and p38. Overexpression of chondrogenic markers suggested that BMSCs have the potential to differentiate into chondrogenic lineages within GFOGER-PEG-PCL samples. In vivo studies using a rat osteochondral defect model revealed that transplanted BMSCs with GFOGER0.8-PEG-PCL survived at the defect with strong chondrogenic expression after 4 weeks. The stem cell-laden GFOGER0.8-PEG-PCL hydrogel produced remarkable osteochondral regeneration at 8 weeks of transplantation, as determined by histological findings and micro-CT analysis. The histomorphological score in the GFOGER0.8-PEG-PCL + BMSCs group was ~1.7-, 2.6-, and 5.3-fold higher than that in the GFOGER0.8-PEG-PCL, MPEG-PCL, and defect groups, respectively. Taken together, these results provide an important platform for further advanced GFOGER-based stem cell research for osteochondral repair.

Guided cortical and cancellous bone formation using a minimally invasive technique of BMSC- and BMP-2-laden visible light-cured carboxymethyl chitosan hydrogels.

A cavity defect inside the bone is formed by deformed cancellous bone from the fixation of the cortical bone, and consequently, abnormal bone healing occurs. Therefore, repairing cancellous bone defects is a remarkable topic in orthopedic surgery. In this study, we prepared bone marrow-derived stem cell (BMSC)-laden and bone morphogenetic protein-2 (BMP-2)-laden visible light-cured carboxymethyl chitosan (CMCS) hydrogels for cortical and cancellous bone healing. Proton nuclear magnetic resonance (1H NMR) analysis confirmed the methacrylation of CMCS (CMCSMA), resulting in 55 % of substitution. The higher concentration of CMCSMA hydrogel resulted in the lower swelling ratio, the larger viscosity, the slower degradation behavior, and the stronger compressive strength. The 5 w/v% hydrogel exhibited a controlled BMP-2 release for 14 days, while the 7 and 10 w/v% hydrogels displayed a controlled BMP-2 release for 28 days. Results of in vitro cytotoxicity and cell proliferation assays revealed the biocompatibility of the samples. In vivo animal tests demonstrated that BMSC- and BMP-2-laden 7 w/v% CMCSMA (CMCSMA+Cell+BMP-2) improved bone formation in the defected cortical and cancellous bones of the femur, as analyzed by micro-computed tomography (micro-CT) and histological evaluations. Consequently, we suggested that CMCSMA+Cell+BMP-2 can be a valuable scaffold for restoring cortical and cancellous bone defects.

Theranostic potential of biodegradable polymeric nanoparticles with paclitaxel and curcumin against breast carcinoma.

In this study, integrin-mediated targeting and near-infrared fluorescence (NIRF) traceable polyethylene glycol-b-poly(lactic-co-glycolic acid) (PEG-PLGA)-based polymeric nanoparticles (NPs) were prepared to investigate the effects of paclitaxel (PTX) and curcumin (CUR) combination therapy on breast cancer. Cyclic (arginine-glycine-aspartic acid-phenylalanine-lysine) (cRGDfK) was selected as a ligand for breast cancer and conjugated to the end of NPs (cRGDfK-NPs). For fluorescence imaging, sulfo-cyanine 5.5 (Cy5.5) was incorporated into NPs (Cy5.5-NPs). A series of hybrid NPs consisting of NPs, cRGDfK-NPs, and Cy5.5-NPs with drugs encapsulated inside the core (Cy5.5-cRGDfK-NPs/PTX + CUR) were prepared by self-assembly. The efficacy of PTX and CUR combination and the ability of the integrin-mediated targeting of NPs were systemically investigated using a 4T1 mouse breast cancer cell line and a nude mouse xenograft model. We suggested that Cy5.5-cRGDfK-NPs/PTX + CUR has superior theranostic potential against breast carcinoma.

Injectable Glycol Chitosan Hydrogel Containing Folic Acid-Functionalized Cyclodextrin-Paclitaxel Complex for Breast Cancer Therapy.

We prepared a drug carrier which consisted of injectable methacrylated glycol chitosan (MGC) hydrogel, and a conjugate of 6-monodeoxy-6-monoamino-ß-cyclodextrin⋅hydrochloride (6-NH2-ß-CD⋅HCl), polyethylene glycol (PEG), and folic acid (FA) for the local delivery and improved cellular uptake of paclitaxel (PTX) (MGC/CDPF-ic-PTX). CDPF refers to a conjugate of 6-NH2-ß-CD⋅HCl, PEG, and FA. The anti-cancer effect was investigated using a xenograft mouse model. As controls, the animal study on MGC/PTX and MGC/CD-ic-PTX was performed. The swelling ratio of all samples was analyzed for 7 days, and it showed a gradual increase for 3 days and a maintained state afterward. From the release result, the MGC-based samples have an initial burst for 1 day and a sustained release for 7 days. Results of cytotoxicity and animal study showed the biocompatibility and superior anti-cancer effect of MGC/CDPF-ic-PTX against breast cancer. Furthermore, histological results showed the anti-cancer capacity of MGC/CDPF-ic-PTX against breast cancer. These findings suggest that MGC/CDPF-ic-PTX has clinical potential for breast cancer therapy.

Near-infra-red fluorescent chitosan oligosaccharide lactate for targeted cancer imaging and photothermal therapy.

Photothermal therapy (PTT) is a promising approach for effective cancer treatment because of its non-invasive procedure, low toxicity to normal tissues, and high tumour ablation efficiency. Developing a PTT agent with precise tumour imaging capabilities is an essential prerequisite for effective PTT. In this study, we developed a bifunctional near-infra-red (NIR) fluorescent conjugate consisting of chitosan oligosaccharide lactate (COL) and the ZW800-1 NIR fluorophore (COL-ZW). We demonstrate that this conjugate is easy to use and that it is an effective theranostic agent for fluorescence-guided photothermal treatment. The temperature of COL-ZW increased by 62.3 °C after NIR laser irradiation (1.1 W/cm2) for 5 min in HT-29 tumour-bearing mice. The HT-29 tumours targeted by COL-ZW showed a remarkable decrease in tumour volume until a week after photothermal treatment. These in vivo results demonstrate that the bifunctional COL-ZW generates strong fluorescence and light-triggered PTT in tumour sites, indicating successful fluorescence-guided PTT. Importantly, no tumour recurrence or treatment-induced toxicity was observed after a single dose of COL-ZW with laser irradiation. Therefore, a combinatorial treatment with COL-ZW and NIR laser irradiation could serve as a promising strategy for photothermal cancer therapy.

Bioluminescence and near-infrared fluorescence imaging for detection of metastatic bone tumors.

Bioluminescence imaging is being increasingly utilized in biological research. However, since the most commonly used firefly luciferase generates relatively weak bioluminescent signals, detection of low numbers of luciferase-expressing cells in vivo is challenging. The weak signal makes it difficult to detect cells located in deep tissues, which is problematic for preclinical research in tumor metastasis. In this study, three different types of fluorophores such as D-luciferin, AkaLumine-HCl, and P800SO3 were compared to evaluate the progression of bone metastasis induced by MDA-MB-231 breast cancer cells in vivo. The fluorescent signals for D-luciferin, AkaLumine-HCl, and P800SO3 were differently detected in the chest and knee joint. In particular, the fluorescence signal of P800SO3 was clearly observed in a section of the ribs, where it pointed out fractured bone fragments by tumor mass. Moreover, the P800SO3 signal from the left knee joint also showed a small bone fragment in the distal femur and was highlighted in the proximal tibia. Using targeted NIR fluorophores, metastatic bone tumors were monitored under the NIR fluorescence imaging system in real time, which enabled the in vivo diagnosis of bone metastasis by providing the location of the metastatic bone tumors.

Doxorubicin·Hydrochloride/Cisplatin-Loaded Hydrogel/Nanosized (2-Hydroxypropyl)-Beta-Cyclodextrin Local Drug-Delivery System for Osteosarcoma Treatment In Vivo.

Osteosarcoma (OSA) is a difficult cancer to treat due to its tendency for relapse and metastasis; advanced methods are therefore required for OSA treatment. In this study, we prepared a local drug-delivery system for OSA treatment based on doxorubicin·hydrochloride (DOX·HCl)/cisplatin (CP)-loaded visible light-cured glycol chitosan (GC) hydrogel/(2-hydroxypropyl)-beta-cyclodextrin (GDHCP), and compared its therapeutic efficiency with that of DOX·HCl- and CP-loaded GC hydrogels (GD and GHCP). Because of diffusion driven by concentration gradients in the swollen matrix, the three hydrogels showed sustained releases of DOX·HCl and CP over 7 days, along with initial 3-h bursts. Results of in vitro cell viability and in vivo animal testing revealed that GDHCP had a stronger anticancer effect than GD and GHCP even though there were no significant differences. Body weight measurement and histological evaluations demonstrated that the drug-loaded GC hydrogels had biocompatibility without cardiotoxicity or nephrotoxicity. These results suggested that GDHCP could be a good platform as a local drug-delivery system for clinical use in OSA treatment.

Hydrogel-Mediated DOX⋠HCl/PTX Delivery System for Breast Cancer Therapy.

We used a hydrogel-mediated dual drug delivery approach, based on an injectable glycol chitosan (GC) hydrogel, doxorubicin hydrochloride (DOX⋅HCl), and a complex of beta-cyclodextrin (ß-CD) and paclitaxel (PTX) (GDCP) for breast cancer therapy in vitro and in vivo. The hydrogel was swollen over 3 days and remained so thereafter. After an initial burst period of 7 hours, the two drugs were released in a sustained manner for 7 days. The in vitro cell viability test showed that GDCP had a better anticancer effect than well plate and DOX⋅HCl/PTX (DP). In addition, the in vivo tests, which evaluated the anticancer effect, systemic toxicity, and histology, proved the feasibility of GDCP as a clinical therapy for breast cancer.

Near-Infrared Fluorescent Sorbitol Probe for Targeted Photothermal Cancer Therapy.

Abstract: Photothermal therapy (PTT) using a near-infrared (NIR) heptamethine cyanine fluorophore has emerged as an alternative strategy for targeted cancer therapy. NIR fluorophores showing a high molar extinction coefficient and low fluorescence quantum yield have considerable potential applications in photothermal cancer therapy. In this study, a bifunctional sorbitol-ZW800 conjugate was used as an advanced concept of photothermal therapeutic agents for in vivo cancer imaging and therapy owing to the high tumor targetability of the sorbitol moiety and excellent photothermal property of NIR heptamethine cyanine fluorophore. The sorbitol-ZW800 showed an excellent photothermal effect increased by 58.7 °C after NIR laser irradiation (1.1 W/cm2) for 5 min. The HT-29 tumors targeted by sorbitol-ZW800 showed a significant decrease in tumor volumes for 7 days after photothermal treatment. Therefore, combining the bifunctional sorbitol-ZW800 conjugate and NIR laser irradiation is an alternative way for targeted cancer therapy, and this approach holds great promise as a safe and highly efficient NIR photothermal agent for future clinical applications.

Photo-Cured Glycol Chitosan Hydrogel for Ovarian Cancer Drug Delivery.

In this study, we prepared an injectable drug delivery depot system based on a visible light-cured glycol chitosan (GC) hydrogel containing paclitaxel (PTX)-complexed beta-cyclodextrin (ß-CD) (GC/CD/PTX) for ovarian cancer (OC) therapy using a tumor-bearing mouse model. The hydrogel depot system had a 23.8 Pa of storage modulus at 100 rad/s after visible light irradiation for 10 s. In addition, GC was swollen as a function of time. However, GC had no degradation with the time change. Eventually, the swollen GC matrix affected the releases of PTX and CD/PTX. GC/PTX and GC/CD/PTX exhibited a controlled release of PTX for 7 days. In addition, GC/CD/PTX had a rapid PTX release for 7 days due to improved water solubility of PTX through CD/PTX complex. In vitro cell viability tests showed that GC/CD/PTX had a lower cell viability percentage than the free PTX solution and GC/PTX. Additionally, GC/CD/PTX resulted in a superior antitumor effect against OC. Consequently, we suggest that the GC/CD system might have clinical potential for OC therapy by improving the water solubility of PTX, as PTX is included into the cavity of ß-CD.

A local drug delivery system based on visible light-cured glycol chitosan and doxorubicin⋠hydrochloride for thyroid cancer treatment in vitro and in vivo.

Systemic drug delivery systems (SDDSs) for thyroid cancer treatment are associated with serious side effects including nausea, anorexia, and hair loss as a result of damage to normal tissues. In this study, we investigated the feasibility of a local DDS (LDDS) based on visible light-cured glycol chitosan (GC) hydrogel and doxorubicin⋅hydrochloride (DOX⋅HCl), called GC10/DOX, on thyroid cancer treatment in vivo. Visible light irradiation increased the storage modulus and swelling ratio of the GC10/DOX hydrogel precursor. The release of DOX⋅HCl from GC10/DOX exhibited two unique patterns comprising an initial burst within 18 hours, followed by a controlled and sustained release thereafter. In vitro cell viability testing showed that GC10/DOX had a greater antitumor effect than free DOX⋅HCl and GC10 hydrogel controls. In vivo, local injection of GC10/DOX near tumor tissue led to a superior antitumor effect compared with controls consisting of free DOX⋅HCl intravenously injected to the tail vein of thyroid cancer-bearing mouse and GC10 hydrogel subcutaneously injected near the tumor. Altogether, our results suggest that GC10/DOX may have clinical potential for thyroid cancer treatment.

Injectable visible light-cured glycol chitosan hydrogels with controlled release of anticancer drugs for local cancer therapy in vivo: a feasible study.

Currently available chemotherapy is associated with serious side effects, and therefore novel drug delivery systems (DDSs) are required to specifically deliver anticancer drugs to targeted sites. In this study, we evaluated the feasibility of visible light-cured glycol chitosan (GC) hydrogels with controlled release of doxorubicin⋅hydrochloride (DOX⋅HCl) as local DDSs for effective cancer therapy in vivo. The storage modulus of the hydrogel precursor solutions was increased as a function of visible light irradiation time. In addition, the swelling ratio of the hydrogel irradiated for 10 s (GC10/DOX) was greater than in 60 s (GC60/DOX). In vitro release test showed that DOX was rapidly released in GC10/DOX compared with GC60/DOX due to the density of cross-linking. In vitro and in vivo tests including cell viability and measurement of tumor volume showed that the local treatment of GC10/DOX yielded substantially greater antitumor effect compared with that of GC60/DOX. Therefore, the visible light-cured GC hydrogel system may exhibit clinical potential as a local DDS of anticancer drugs with controlled release, by modulating cross-linking density.

Preparation of poly-l-lysine-based nanoparticles with pH-sensitive release of curcumin for targeted imaging and therapy of liver cancer in vitro and in vivo.

Poly-l-lysine (PLL) nanoparticle (NP) system was prepared for the controlled release of curcumin (CUR) by pH stimuli, and its theranostic efficacy on cancer was compared to that of CUR solution in vitro and in vivo. Deoxycholic acid (DOCA), methoxy polyethylene glycol (MPEG) and cyanine 5.5 (cy5.5) were conjugated to the amine group of PLL through condensation reaction (PLL-DOCA-MPEG-cy5.5), followed by encapsulation of CUR by dialysis method (PLL-DOCA-MPEG-cy5.5/CUR NPs). The composition, morphology and size distribution of PLL-DOCA-MPEG-cy5.5 NPs were characterized by proton nuclear magnetic resonance (1H NMR), transmission electron microscopy (TEM) and dynamic light scattering (DLS), respectively. In vitro tests exhibited that changes in the charge and size of the NPs at low pH led to the improved cellular uptake of CUR into human hepatoma Hep3B cell line by electrostatically absorptive endocytosis. PEGylation with MPEG was turn out to be very effective to have a prolonged blood circulation time, in turn increased the EPR effect. In addition, the incorporation of Cy5.5 into NPs provided successful biodistribution images in vivo and ex vivo. Our findings suggest that PLL-DOCA-MPEG-cy5.5/CUR NPs may have promising applications in cancer theranosis.

Real-time in vivo imaging of metastatic bone tumors with a targeted near-infrared fluorophore.

Tumors of the prostate or breast are particularly likely to metastasize to the bone, and early diagnosis of metastatic bone tumors is important for designing an effective treatment strategy. Imaging modalities for the detection of bone metastasis are limited, and radiation-based techniques are commonly used. Here, we investigated the efficacy of selective near-infrared (NIR) fluorescence detection of metastatic bone tumors and its role in the detection of bone metastasis in prostate and breast cancer cell lines and in a xenograft mouse model. A targeted NIR fluorophore was used to monitor metastatic bone tumors using a NIR fluorescence imaging system in real time, enabling the diagnosis of bone metastasis in vivo by providing the location of the metastatic bone tumor. The NIR fluorescence imaging technique using targeted NIR contrast agents is a potential tool for the early diagnosis of bone tumors.

Visible Light-Cured Glycol Chitosan Hydrogel Containing a Beta-Cyclodextrin-Curcumin Inclusion Complex Improves Wound Healing In Vivo.

Scarless wound healing is ideal for patients suffering from soft tissue defects. In this study, we prepared a novel wet dressing (ß-CD-ic-CUR/GC) based on the visible light-cured glycol chitosan (GC) hydrogel and inclusion complex between beta-cyclodextrin (ß-CD) and curcumin (CUR). We also evaluated its efficacy in the acceleration of wound healing as compared to that of CUR-loaded GC (CUR/GC). The conjugation of glycidyl methacrylate (GM) to GC for photo-curing was confirmed by ¹H-NMR measurement, and the photo-cured GC hydrogel was characterized by the analyses of rheology, swelling ratio, SEM and degradation rate. After visible light irradiation, the surface/cross-sectional morphologies and storage (G')/loss (G'') moduli revealed the formation of hydrogel with interconnected porosity. The dressing ß-CD-ic-CUR/GC exhibited a controlled release of 90% CUR in a sustained manner for 30 days. On the other hand, CUR/GC showed CUR release of 16%. ß-CD acted as an excipient in improving the water-solubility of CUR and affected the release behavior of CUR. The in vivo animal tests including measurement of the remaining unhealed wound area and histological analyses showed that ß-CD-ic-CUR/GC may have potential as a wet dressing agent to enhance soft tissue recovery in open fractures.

Preparation and Evaluation of Dexamethasone (DEX)/Growth and Differentiation Factor-5 (GDF-5) Surface-Modified Titanium Using ß-Cyclodextrin-Conjugated Heparin (CD-Hep) for Enhanced Osteogenic Activity In Vitro and In Vivo.

The most ideal implant models in the dental and orthopedic fields to minimize the failure rate of implantation involve the improvement of osseointegration with host bone. Therefore, a focus of this study is the preparation of surface-modified titanium (Ti) samples of disc and screw types using dexamethasone (DEX) and/or growth and differentiation factor-5 (GDF-5), as well as the evaluation of their efficacies on bone formation in vitro and in vivo. X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and contact angle measurement were used to evaluate the surface chemical composition, surface morphology and wettability, respectively. The results showed that implant surfaces were successfully modified with DEX and/or GDF-5, and had rough surfaces along with hydrophilicity. DEX, GDF-5 or DEX/GDF-5 on the surface-modified samples were rapidly released within one day and released for 28 days in a sustained manner. The proliferation and bone formation of MC3T3-E1 cells cultured on pristine and surface-modified implants in vitro were examined by cell counting kit-8 (CCK-8) assay, as well as the measurements of alkaline phosphatase (ALP) activity and calcium deposition, respectively. MC3T3-E1 cells cultured on DEX/GDF-5-Ti showed noticeable ALP activity and calcium deposition in vitro. Active bone formation and strong osseointegration occurred at the interface between DEX/GDF-5-Ti and host bone, as evaluated by micro computed-tomography (micro CT) analysis. Surface modification using DEX/GDF-5 could be a good method for advanced implants for orthopaedic and dental applications.

Effects of Titanium Mesh Surfaces-Coated with Hydroxyapatite/ß-Tricalcium Phosphate Nanotubes on Acetabular Bone Defects in Rabbits.

The management of severe acetabular bone defects in revision reconstructive orthopedic surgery is challenging. In this study, cyclic precalcification (CP) treatment was used on both nanotube-surface Ti-mesh and a bone graft substitute for the acetabular defect model, and its effects were assessed in vitro and in vivo. Nanotube-Ti mesh coated with hydroxyapatite/ß-tricalcium phosphate (HA/ß-TCP) was manufactured by an anodizing and a sintering method, respectively. An 8 mm diameter defect was created on each acetabulum of eight rabbits, then treated by grafting materials and covered by Ti meshes. At four and eight weeks, postoperatively, biopsies were performed for histomorphometric analyses. The newly-formed bone layers under cyclic precalcified anodized Ti (CP-AT) meshes were superior with regard to the mineralized area at both four and eight weeks, as compared with that under untreated Ti meshes. Active bone regeneration at 2-4 weeks was stronger than at 6-8 weeks, particularly with treated biphasic ceramic (p < 0.05). CP improved the bioactivity of Ti meshes and biphasic grafting materials. Moreover, the precalcified nanotubular Ti meshes could enhance early contact bone formation on the mesh and, therefore, may reduce the collapse of Ti meshes into the defect, increasing the sufficiency of acetabular reconstruction. Finally, cyclic precalcification did not affect bone regeneration by biphasic grafting materials in vivo.

Surface modification of titanium with hydroxyapatite-heparin-BMP-2 enhances the efficacy of bone formation and osseointegration in vitro and in vivo.

Surface-modified titanium (Ti) samples with hydroxyapatite (HAp) and heparin (Hep)-bone morphogenetic protein-2 (BMP-2) complex (Ti/HAp/Hep/BMP-2) were prepared, and their efficacies on the enhancements of bone formation and osseointegration in vitro and in vivo were examined as compared to Ti/HAp and Ti/Hep/BMP-2. The modified surfaces were characterized by X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and contact angle goniometry. In vitro studies revealed that MG-63 human osteosarcoma cell lines grown on Ti/HAp/Hep/BMP-2 increased the amounts of alkaline phosphatase (ALP) activity, calcium deposition and the levels of OCN mRNA gene expression as compared to those grown on Ti/HAp, Ti/Hep/BMP-2 or pristine Ti. Moreover, Ti/HAp/Hep/BMP-2 exhibited higher bone volume (BV), bone volume/tissue volume (BV/TV), removal torque value and bone-implant contact (BIC) than Ti/HAp, Ti/Hep/BMP-2 or pristine Ti in vivo. Histological evaluations showed that many desirable features of bone remodelling existed at the interface between Ti/HAp/Hep/BMP-2 and the host bone. Consequently, Ti/HAp/Hep/BMP-2 may have potential for clinical use as dental or orthopaedic implants.