Hydroxyapatite particles are capable of inducing osteoclast formation

Hydroxyapatite (HA) coatings have been used to improve implant fixation by promoting bone formation around the prosthesis. A macrophage response to HA particulates has been noted around loosened HA-coated prostheses. As biomaterial wear particle-associated macrophages are known to be capable of differentiating into osteoclasts that are capable of bone resorption, we examined whether particulate HA could similarly induce macrophage-osteoclast differentiation. HA-associated macrophages were isolated from granulomas, formed by subcutaneous implantation of HA, and co-cultured with UMR 106 osteoblast-like cells in the presence of 1,25-dihydroxyvitamin D₃ for up to 14 days on glass coverslips and bone slices. HA-associated macrophage-osteoclast differentiation was evidenced by the formation of numerous multinucleated tartrate resistant acid phosphatase (TRAP)-positive cells which formed lacunar resorption pits on bone slices. Polymethylmethacrylate (PMMA) particle-associated macrophages, isolated from subcutaneous PMMA-containing granulomas, caused significantly more osteoclast formation and bone resorption than HA-associated macrophages. These results indicate that macrophages responding to HA particles are capable of osteoclast differntiation. They also suggest that particles derived from uncemented (HA-coated) implants are likely to induce less osteoclast formation and osteolysis than cemented implants. © 2001 Kluwer Academic Publishers     

Highly porous hydroxyapatite scaffolds with elongated pores using stretched polymeric sponges as novel template

This study reports a simple way of improving the compressive strength of highly porous hydroxyapatite (HA) scaffolds by adopting elongated polymeric sponges as a novel template. In this method, as-received polymeric sponges with isotropic pores were stretched uniaxially to 50% elongation at 200 °C for 2 h, and then coated with a HA slurry. The HA-coated sponges were heat-treated at 800 °C for 3 h to remove the polymeric sponges and at 1250 °C for 3 h to sinter the HA walls. The fabricated samples showed a highly anisotropic pore structure with elongated pores parallel to the direction of the elongation of the polymeric sponge. This simple method allowed a highly porous scaffold to have a high compressive strength of 3.8 ± 0.1 MPa at a porosity of 76% when tested parallel to the direction of pore elongation.     

A histological evaluation of the effect of hydroxyapatite coating on interfacial response

The effect of hydroxyapatite (HA) coating on cortical bone apposition around press-fit inserted implants and implants surrounded by a gap was investigated. Uncoated and HA-coated titanium implants were inserted in burr holes with three different diameters in the tibia of rabbits. Implantation time was four months. The histological evaluation demonstrated that after four months implantation the interfacial bone reaction appeared to be identical for HA-coated and non-coated implants with various degrees of surgical fit. Although after four months the interface showed the same response, there still might be an initial advantage of the HA-interface with bony tissue.     

Preparation of an electrodeposited hydroxyapatite coating on titanium substrate suitable for in-vivo applications

In this paper the porous hydroxyapatite coating on Ti implant materials was prepared by the process of electrodeposition, hydrothermal and sinter. The surface morphology, bond strength and thickness of HA coatings were investigated by SEM, AFM, and its biocompatibility was evaluated by cytotoxicity experiments and implant experiments, respectively. Results showed that (1) The HA coatings was 50 μm thickness and adhered on the Ti substrate strongly, which bond strength reached 38MPa. AFM analysis showed that the HA coating was porous structure, in which the mean pore size was 236.5 μm, (2) Cytotoxicity experiments and implant experiments showed that HA-coated Ti implant materials has little cytotoxicity in vitro and little inflammatory reaction in vivo, and there were no statistically disparity between HA-coated Ti implant and titanium implant materials of clinical application (p > 0.05), which demonstrated that HA-coated Ti has a good biocompatibility.     

Preparation of electrospun siloxane-poly(lactic acid)-vaterite hybrid fibrous membranes for guided bone regeneration

A nonwoven fabric scaffold for guided bone regeneration (GBR) consisting of siloxane-poly(lactic acid) (PLA)-vaterite hybrid material (SiPVH) was prepared by an electrospinning method. To improve the cellular compatibility of the fabric, the fibers were completely coated with hydroxyapatite (HA) by soaking in simulated body fluid. The HA-coated SiPVH nonwoven fabric contained large-sized spaces and showed the ability of releasing soluble silica and calcium species, which have previously been reported to stimulate osteogenic cells at the genetic level. A new type of GBR membrane was prepared by bonding SiPVH fabric with PLA nonwoven fabric with small-sized pores for preventing the intrusion of soft tissue. The resultant bi-layered membrane was expected to be effective not only for having an open structure for bone formation and a barrier to soft tissue, but also for enhancing bone growth by the release of ionic species.     

Monte Carlo determination of age-dependent steady-state dose to red bone marrow and bone from 14C exposure

Monte Carlo simulations were carried out to calculate the age-dependent dose from the beta decay of 14C to marrow and bone on the basis of a steady-state specific-activity model. A model of the trabecular cavity containing spherical fat cells in a square lattice surrounded by haemopoietic tissue was employed. The age-dependent 14C dose to haemopoietic (red) marrow was estimated taking account of the change in the fat cell size with fat fraction. Allowances were made for the change in the percentage cellularity and distribution of active marrow in the whole skeleton as a function of age. Age-dependent changes in trabecular cavity size and bone composition were found to have only a small effect on dose. Dose rates were estimated under steady-state conditions, for food ingested with a 14C specific activity of 1 Bq g(-1) of C. The equivalent dose rate to the haemopoietic tissue of a 20 year-old adult is 77 microSv a(-1), and 39 microSv a(-1) for a 3-month-old infant. Similarly, the equivalent dose rate to the bone surfaces of an adult is 48 microSv a(-1), and 38 microSv a(-1) for an infant. Therefore, the equivalent dose rate to marrow and bone stem cells increases with age under steady state conditions.     

仿生矿化法制备可降解羟基磷灰石/氧化细菌纤维素复合材料

细菌纤维素是具有天然纳米网状结构的支架材料,对其进行氧化改性后可获得可调控的降解性能。通过仿生矿化氧化改性的细菌纤维素支架,制备了可降解羟基磷灰石/氧化细菌纤维素复合骨组织工程支架材料。观察并分析了仿生矿化过程氧化细菌纤维素的降解和羟基磷灰石的形成,并通过SEM、EDS、XRD对羟基磷灰石在可降解氧化细菌纤维素支架上沉积进行了表征,矿化7天的羟基磷灰石/氧化细菌纤维素复合材料表面和内部均有磷灰石形成,测得磷灰石的钙磷比为1.75,主要为羟基磷灰石,伴有少量碳羟磷灰石。结果表明,使用仿生矿化法成功获得了一种新型可降解羟基磷灰石/氧化纤维素复合材料支架。     

2, 3-Dihydrazone cellulose: Prospective material for tissue engineering scaffolds

Cellulose was oxidized by sodium metaperiodate to give rise to 2, 3-dialdehyde cellulose with 92% oxidation ratio, which was further reacted with hydrazine to form 2, 3-dihydrazone cellulose for the incorporation of NH₂ groups. Two forms of matrix, i.e. films and sponges were fabricated. The materials were characterized by FTIR spectroscopy. Scanning electron microscopy revealed its porous architecture with an average pore size of 150 μm. Swelling studies were carried out in phosphate buffer saline (PBS) at physiological pH 7.4. The contact angle of the 2, 3-dihydrazone cellulose surface was determined for assessing its hydrophilicity which came out to be 23° ± 2°. NIH3T3 mice fibroblast cells were used for determining the cytocompatibility of the surfaces. The morphology of the cells was observed through optical inverted microscopy. The results show that 2, 3-dihydrazone cellulose can be used as scaffold material in tissue engineering.     

Functional synergy of anti-mir221 and nanohydroxyapatite scaffold in bone tissue engineering of rat skull

An appropriate cell source, effective cell modification, and proper supportive matrices are the main bases of tissue engineering. The effectiveness of anti-mir221 or hydroxyapatite (HA) in improving the osteogenic differentiation of mesenchymal stem cells (MSCs) has been reported previously. Herein, simultaneous application of these osteogenic inducers was investigated in vivo. The Poly-caprolactone (PCL)/HA nanofibers were characterized using contact angle measurement, tensile test, Fourier transform infrared spectroscopy, and electron microscopy. Rat MSCs were isolated, characterized and transfected with anti-mir221. The rats were divided into 4 groups and an 8 mm defect were created in the mid-calvaria of each rat by trephine bur. Group 1 received (PCL)/HA nanofibers, group 2 received (PCL)/HA nanofibers plus autologous MSCs, group 3 received (PCL)/HA nanofibers plus MSCs transfected with anti-mir221, and group 4 rats were left empty as an additional control group. Histomorphometric and radiomorphometric evaluation after 4 and 8 weeks revealed more new bone formation in the cell-treated groups compared to the scaffold alone group. There was evidence for a combination of increased osteoclasts and osteoblast vascular lake containing red blood cells in the anti-mir221 transfected group. New bone penetration into the scaffolds empirically demonstrated the capability of this combination for efficient osteointegration. Altogether, the co-application of HA and anti-mir221 transfected cells can enhance bone healing of the rat skull.     

Hyaluronic acid hydrogels for biomedical applications

Hyaluronic acid (HA), an immunoneutral polysaccharide that is ubiquitous in the human body, is crucial for many cellular and tissue functions and has been in clinical use for over thirty years. When chemically modified, HA can be transformed into many physical forms-viscoelastic solutions, soft or stiff hydrogels, electrospun fibers, non-woven meshes, macroporous and fibrillar sponges, flexible sheets, and nanoparticulate fluids-for use in a range of preclinical and clinical settings. Many of these forms are derived from the chemical crosslinking of pendant reactive groups by addition/condensation chemistry or by radical polymerization. Clinical products for cell therapy and regenerative medicine require crosslinking chemistry that is compatible with the encapsulation of cells and injection into tissues. Moreover, an injectable clinical biomaterial must meet marketing, regulatory, and financial constraints to provide affordable products that can be approved, deployed to the clinic, and used by physicians. Many HA-derived hydrogels meet these criteria, and can deliver cells and therapeutic agents for tissue repair and regeneration. This progress report covers both basic concepts and recent advances in the development of HA-based hydrogels for biomedical applications.     

Osteogenesis of freeze-dried cancellous bone allograft loaded with autologous marrow-derived mesenchymal cells

In this work, we explored the osteogenic potential of freeze-dried cancellous bone allograft loaded with autologous mesenchymal stem cells (MSCs). Human MSCs were isolated, purified and identified. Flow cytometry revealed that the isolated and enriched cells were 90–97% homogeneous and were negative for reactivity to antigens CD34 and CD45 but positive for reactivity to antigen CD44. The expressions of transformation growth factor-β1 (TGFβ1) and basic fibroblast growth factor (bFGF) were detected as positive in freeze-dried cancellous bone allograft using the immunohistochemistry method. The expression of Alkaline Phosphatase (AKP) and Type I collagen mRNA in MSCs, co-cultured in supernatant of allograft, showed a stronger positive than that in hydroxyapatite (HA). After 1 day of incubation, MSCs grew in both surface and inner pores of bone allograft blocks observed by electron microscopy scan. The composites of allograft loaded with MSCs or HA loaded with MSCs were implanted in nude mice subcutaneously. Osteoid and fluorescence labeling were observed in the allograft by the 4th and 6th weeks, respectively, but not at any time in the HA. This study suggests that the freeze-dried cancellous bone allograft would be an ideal choice for the delivery of mesenchymal stem cells for bone tissue engineering applications.     

Fabrication and characterization of injection molded poly (ε-caprolactone) and poly (ε-caprolactone)/hydroxyapatite scaffolds for tissue engineering

In this study, poly(ε-caprolactone) (PCL)/sodium chloride (NaCl), PCL/poly(ethylene oxide) (PEO)/NaCl and PCL/PEO/NaCl/hydroxyapatite (HA) composites were injection molded and characterized. The water soluble and sacrificial polymer, PEO, and NaCl particulates in the composites were leached by deionized water to produce porous and interconnected microstructures. The effect of leaching time on porosity, and residual contents of NaCl and NaCl/HA, as well as the effect of HA addition on mechanical properties was investigated. In addition, the biocompatibility was observed via seeding human mesenchymal stem cells (hMSCs) on PCL and PCL/HA scaffolds.The results showed that the leaching time depends on the spatial distribution of sacrificial PEO phase and NaCl particulates. The addition of HA has significantly improved the elastic (E′) and loss moduli (E″) of PCL/HA scaffolds. Human MSCs were observed to have attached and proliferated on both PCL and PCL/HA scaffolds. Taken together, the molded PCL and PCL/HA scaffolds could be good candidates as tissue engineering scaffolds. Additionally, injection molding would be a potential and high throughput technology to fabricate tissue scaffolds.     

包埋载药微球的羟基磷灰石/聚氨酯复合组织工程支架的研究

研制具有药物缓释功能的骨组织工程支架, 对载药微球包埋于羟基磷灰石/聚氨酯(HA/PU)支架中的药物缓释体系进行了可行性研究. 首先将盐酸环丙沙星作为模型药物, 包裹于乙基纤维素(EC)微球中, 然后将EC微球与HA/PU材料进行复合, 制备了抗生素药物缓释支架. 结果显示EC微球均匀地分布在HA/PU支架基质中, 未对支架的开孔结构和孔隙形貌构成影响. 与单纯将药物载入HA/PU支架中相比, 复合载药EC微球的HA/PU支架的初期药物暴释明显降低, 药物缓释时间延长. 体外药物释放实验和抑菌实验结果表明, 该载药微球支架具有良好的药物缓释功能和抑菌性能, 是一种集骨修复和治疗于一体的新型组织工程支架材料.     

Fabrication,characterization, and biocompatibility of ethyl cellulose/carbonated hydroxyapatite composite coatings on Ti6Al4V

In order to improve the biocompatibility of metallic implants, bioactive components are often used as coatings so that a real bond with the surrounding bone tissue can be formed. We prepared ethyl cellulose/carbonated hydroxyapatite composite coatings (ECHCs) on Ti6Al4V substrates with carbonated hydroxyapatite coatings (CHACs) without ethyl cellulose as controls. The inorganic constituent on the CHACs and ECHCs is calcium-deficient carbonated hydroxyapatite with a flaky texture and a low degree of crystallinity. The flaky carbonated hydroxyapatite plates aggregate to form macropores with an aperture size of around 0.5–2.0 μm. The presence of ethyl cellulose provides superior morphology, contact angle, and biocompatibility characteristics. In comparison to CHACs, ECHCs exhibit a smoother, crack-free surface because the cracks are filled by ethyl cellulose. Moreover, the contact angle of ECHCs is 37.3°, greater than that of CHACs (13.0°). Surface biocompatibility was investigated by using human bone mesenchymal stem cells (hBMSCs). The attachment, spreadability, viability and proliferation of hBMSCs on ECHCs are superior to those on CHACs. Thus, the crack-free ECHCs have excellent biocompatibility and are appropriate for use as biological implants.     

Cell proliferation,viability, and in vitro differentiation of equine mesenchymal stem cells seeded on bacterial cellulose hydrogel scaffolds

The culture of multipotent mesenchymal stem cells on natural biopolymers holds great promise for treatments of connective tissue disorders such as osteoarthritis. The safety and performance of such therapies relies on the systematic in vitro evaluation of the developed stem cell-biomaterial constructs prior to in vivo implantation. This study evaluates bacterial cellulose (BC), a biocompatible natural polymer, as a scaffold for equine-derived bone marrow mesenchymal stem cells (EqMSCs) for application in bone and cartilage tissue engineering. An equine model was chosen due to similarities in size, load and types of joint injuries suffered by horses and humans. Lyophilized and critical point dried BC hydrogel scaffolds were characterized using scanning electron microscopy (SEM) to confirm nanostructure morphology which demonstrated that critical point drying induces fibre bundling unlike lyophilisation. EqMSCs positively expressed the undifferentiated pluripotent mesenchymal stem cell surface markers CD44 and CD90. The BC scaffolds were shown to be cytocompatible, supporting cellular adhesion and proliferation, and allowed for osteogenic and chondrogenic differentiation of EqMSCs. The cells seeded on the BC hydrogel were shown to be viable and metabolically active. These findings demonstrate that the combination of a BC hydrogel and EqMSCs are promising constructs for musculoskeletal tissue engineering applications.     

Histological aspects in bone regeneration of an association with porous hydroxyapatite and bone marrow cells

The osteogenic potential of an association of two kinds of hydroxyapatite (HA) porous ceramics with autologous bone marrow cells cultured with or without dexamethasone (10^-8M) addition in the culture medium and non-cultured rabbit marrow stromal stem cells (MSCs) was tested after 4 weeks of implantation in the dorsal muscles of spine in rabbit. A significantly higher number of rabbits with implants containing bone tissue inside pores were obtained with 10⁷ cells ml^-1 cultured treated with Dex. In the HA porous ceramic using naphtalen as porogen agent, the bone recolonization remains only at the periphery of implants and in the second row of pores, while in the HA porous ceramic using polymethacrylate (PMMA) microbeads as porogen agent, the bone recolonization is observed in the depth of implants. In the PMMA HA group, the Kru¨skal–Wallis variance analysis between the rabbits is significantly different with the percentage of number of occupied pores and occupied pores with bone tissue is different (p < 0.05).     

Morphology and immersion behavior of plasma-sprayed hydroxyapatite/bioactive glass coatings

A series of hydroxyapatite/bioactive glass (HA/BG) coatings have been plasma-sprayed on Ti6Al-4V substrate using HA/BG powders that were prepared by both sinter-granulation and direct mixing methods. The morphology and immersion behavior of these coatings in a simulated body fluid (SBF) were investigated. The results showed that in-house fabricated BG and sinter-granulated HA powders were irregularly shaped and dense. When 5 wt % or more BG was added in HA, the powder became rough and porous. X-ray diffraction (XRD) patterns showed that the presence of BG enhanced the decomposition of HA structure during fabrication of the powders. Reasonably high bond strengths were obtained from all coatings. The granulated type HA/BG coatings showed no significant differences in bond strength from the mixed type HA/BG coatings. The plasma spray process itself and the presence of BG enhanced the decomposition of apatite. Surface morphology of all sinter-granulated type coatings was similar to that of monolithic HA coating, that was comprised of patches of smooth and shiny glassy film and irregularly-shaped particles on its surface. The dissolution depth of plasma-sprayed coatings immersed in SBF was largely dependent on the type and composition of the coating. Granulated type HA/BG coatings were much less dissolvable than monolithic HA or mixed type HA/BG coatings. It seems that the presently used granulation method for the preparation of HA/BG powders plays a predominant role in determining the dissolution behavior of the plasma-sprayed coatings. ©©2000 Kluwer Academic Publishers     

Effects of PVA sponge containing chitooligosaccharide in the early stage of wound healing

Poly(vinyl alcohol) (PVA) sponges with different chitooligosaccharide (COS) content were prepared for wound-dressing application. The morphological structure of PVA sponges was observed by scanning electron microscopy. As the concentration of COS-loaded PVA sponge increased, the average pore size of sponge decreased and the release rate of COS from the sponge also slightly decreased. The accelerating effect of the COS-loaded PVA sponges on open wound healing in rats was investigated by macroscopic examination and measurement of wound area. The COS-loaded sponges were found to be very effective as a wound-healing accelerator in the early stage of wound healing. The wound treated with the COS-loaded PVA sponge was almost reepithelialized, granulation tissues in the wound were considerably replaced by fibrosis at 8 days after initial wounding. The COS-loaded PVA sponge was considered to be a suitable wound-healing formulation due to its easy preparation and high effectiveness.     

Preparation and characterization of multi-walled carbon nanotube/hydroxyapatite nanocomposite film dip coated on Ti–6Al–4V by sol–gel method for biomedical applications: An in vitro study

In the present research, the introduction of multi-walled carbon nanotubes (MWCNTs) into the hydroxyapatite (HA) matrix and dip coating of nanocomposite on titanium alloy (Ti–6Al–4V) plate was conducted in order to improve the performance of the HA-coated implant via the sol–gel method. The structural characterization and electron microscopy results confirmed well crystallized HA–MWCNT coating and homogenous dispersion of carbon nanotubes in the ceramic matrix at temperatures as low as 500 °C. The evaluation of the mechanical properties of HA and HA/MWCNT composite coatings with different weight percentages of MWCNTs showed that the addition of low concentrations of MWCNTs (0.5 and 1 wt.%) had improved effect on the mechanical properties of nanocomposite coatings. Moreover, this in vitro study ascertained the biocompatibility of the prepared sol–gel-derived HA/MWCNT composite coatings.     

Overcoming drug-resistant lung cancer by paclitaxel-loaded hyaluronic acid-coated liposomes targeted to mitochondria

Abstract

As a major cause for the inefficiency of cancer chemotherapy, multidrug resistance (MDR) has become a major barrier to cancer treatment. Mitochondrion-orientated transportation of smart liposomes has been developed as a promising strategy to deliver anticancer drugs directly to tumor sites and actively target the mitochondria, so that drugs can interfere with mitochondrial function and facilitate cell apoptosis, overcoming MDR. Herein, we report a novel dual-functional paclitaxel (PTX) liposome system possessing both CD44-targeting and mitochondrial-targeting properties to enhance drug accumulation in mitochondria and trigger apoptosis of drug-resistant cancer cells. Mitochondria-targeting PTX-loaded liposomes were prepared by thin-film hydration and then coated with hyaluronic acid (HA) by electrostatic adsorption. We evaluated the characteristics of the PTX liposomes in vitro, and found that their particle size was about 100?nm and increased to ～140?nm after modification by HA. The entrapment efficiency was larger than 85%, and stability data indicated that the liposomes were physically and chemically stable for at least one week at 4?°C. We further evaluated the intake, mitochondrial targeting, ATP levels, caspase-3 activity measurement, and antitumor actives of the liposomes. The results indicated that HA-coated liposomes with mitochondria targeting had significant inhibitory effects against A549 and A549/Taxol cells, showing them to be a promising means of improving therapeutic efficacy and overcoming MDR in cancer treatment.