羟基磷灰石表面修饰的人工角膜钛支架体外生物相容性研究

目的采用细胞培养法观察人工角膜纯钛支架经羟基磷灰石(HA)表面修饰后,其生物相容性是否增加。方法采用第4～6代兔角膜基质成纤维细胞直接接种于HA-Ti、Ti及盖玻片表面,培养3、24、48、72h后,用丫啶橙染色法观察材料表面细胞的黏附、伸展和增殖情况,在扫描电子显微镜下观察材料表面的细胞形态及细胞外基质产生情况。结果细胞接种3、24、48、72h后,HA-Ti表面的活细胞数多于其他材料表面(P0.05)。细胞接种3h,细胞扩展面积:HA-Ti盖玻片Ti。48h后扫描电子显微镜观察发现HA-Ti表面的细胞扩展面积最大,细胞张力丝最长。72h后,HA-Ti表面完全被胶原覆盖。结论HA表面修饰增加了人工角膜纯Ti支架的生物活性。     

Injectable nanocrystalline hydroxyapatite paste for bone substitution: in vivo analysis of biocompatibility and vascularization

The nanocrystalline hydroxyapatite paste Ostim represents a fully degradable synthetic bone substitute for the filling of bone defects. Herein, we investigated in vivo the inflammatory and angiogenic host tissue response to this biomaterial after implantation. For this purpose, Ostim was implanted into the dorsal skinfold chambers of Syrian golden hamsters. The hydroxyapatite ceramic Cerabone and isogeneic transplanted cancellous bone served as controls. Angiogenesis, microhemodynamics, microvascular permeability, and leukocyte-endothelial cell interaction of the host tissue were analyzed over 2 weeks using intravital fluorescence microscopy. Ostim exhibited good biocompatibility comparable to that of Cerabone and cancellous bone, as indicated by a lack of venular leukocyte activation after implantation. Cancellous bone induced a more pronounced angiogenic response and an increased microvessel density when compared with the synthetic bone substitutes. In contrast to Cerabone, however, Ostim showed a guided neovascularization directed toward areas of degradation. Histology confirmed the ingrowth of proliferating vascularized tissue into the hydroxyapatite paste at sites of degradation, while the hydroxyapatite ceramic was not pierced by new microvessels. Thus, Ostim represents an injectable synthetic bone substitute, which may optimize the conditions for the formation of new bone at sites of bone defects by supporting a guided vascularization during biodegradation.     

The in vitro corneal biocompatibility of hydroxyapatite coated carbon mesh

The purpose of this study was to consider the use of a hydroxyapatite (HA) coated porous carbon matrix as a synthetic dental laminate substitute in osteo-odonto-keratoprosthetic (OOKP) design. 3 types of carbon meshes were coated with HA by sonoelectrochemical deposition. The materials were characterised by scanning electron microscopy (SEM) and HA deposition was characterised by elemental analysis and X-ray diffractometry (XRD). In vitro assays were carried out to quantify the effects of HA coating on human keratocyte adhesion. Cellular cytokine production was used to assess inflammatory potential. HA coating significantly increased keratocyte adhesion to the carbon matrix (p < 0.01). The materials did not induce excessive cytokine production by the adherent keratocytes. In addition, the matrices themselves adsorbed significant levels of the cytokine IL-8 (p < 0.05). The results indicate that HA coated carbon matrices provide a suitable environment to enhance in-growth of corneal cells without inducing further inflammation. The materials may also suppress excessive inflammation by adsorption of the cytokine IL-8 into the porous, internal carbon structure.     

Growth of osteoblasts on porous calcium phosphate ceramic: an in vitro model for biocompatibility study

Biomaterial implantation in animals is commonly used for biocompatibility studies as well as examination of long-term interaction between tissue and the test material. An in vitro cell culture model is proposed as an alternative which will save animal lives and reduce the pain and discomfort of animals used for such studies. In this study the biomaterial was matched to the cell types typical of the implant site of the particular material: porous calcium phosphate ceramic, used as dental and orthopaedic implants, with periosteal fibroblasts, osteoblasts and chondrocytes. All three cell types attached on to the ceramic and formed multicellular layers. Numbers of periosteal fibroblasts, osteoblasts and chondrocytes increased 29-, 23- and 17-fold, respectively, during the 10 wk period. Osteoblasts retained their phenotypic expression by producing only Type I collagen. Parathyroid hormone (PTH, 50 nM) suppressed the alkaline phosphatase activity of osteoblasts by over 50% and increased cAMP by more than 10-fold over control cultures.     

Dissolution behavior of hydroxyapatite coating by hydrothermal method: an in vitro study

Hydroxyapatite (HA) coated CaO-SiO2-B2O3-Na2O glass composites prepared by a hydrothermal coating method were soaked in four kinds of solutions to evaluate the dissolution behavior and the chemical stability. 0.5 mol/l HCl and 0.5 mol/l HNO3 were used as acidic solution, and 5% physiological saline and sodium lactate compound solution as the neutral physiological solution. It was found that HA coated glass composites had the better chemical stability and the corroding-resistance ability. The dissolution behavior of the HA coating was mainly correlated with the phase composition of the as-received HA coating and its microstructure as well as the pH value of the solution.     

壳聚糖-镁膜体外生物相容性的初步研究

目的： 研究壳聚糖镁膜的制备及其与PC12细胞的体外生物相容性,初步探讨壳聚糖-镁络合后形成的复合物作为组织工程材料的可行性。方法: 合成壳聚糖-镁膜,应用扫描电镜观察复合材料的表面形态,并用X射线能谱仪分析壳聚糖-镁复合膜中镁的含量;测定复合膜的膨胀率;在体外将复合膜与PC12细胞共培养,用MTT方法检测细胞活力。扫描电镜观察PC12细胞在材料上的形态学变化。结果: 单纯壳聚糖与壳聚糖-镁膜各组相比,表面较为光滑,而复合膜表面含有大量的细小孔隙;复合膜中的镁元素的的含量与投入的硫酸镁量呈一定的剂量依赖。形态学观察表明,PC12细胞在CM上生长良好,至7 d时,可见微绒毛丰富并有较长突起,部分细胞之间形成突触状结构;MTT检测结果表明,培养7 d时实验组细胞活力超过对照组。结论: 壳聚糖可以与镁络合形成复合物,但是两者之间的络合率不成正比关系;壳聚糖镁膜与PC12细胞有良好的体外相容性。     

The biocompatibility of hydroxyapatite ceramic: a study of retrieved human middle ear implants

The biocompatibility of 11 hydroxyapatite auditory canal-wall prostheses and 4 hydroxyapatite incus prostheses implanted for 4 to 40 months was evaluated by light microscopy, scanning electron microscopy, transmission electron microscopy, and R?ntgen microanalysis. These 15 prostheses representing 4% of 375 prostheses, has been removed because of unresolved chronic middle ear infection, residual cholesteatoma, or poor fit. The findings confirmed earlier reports on the biocompatibility of hydroxyapatite in vitro, in animals, and in man. An electron-dense layer was found at the interface with bone and fibrous tissue, and a firm bond between the ceramic and bone at the hydroxyapatite ceramic/bone interface developed. Macropores became filled with bone and fibrous tissue, and the tissue in the individual pores was interconnected. Furthermore the incus prostheses were covered with an epithelium similar to that found in the human middle ear. Findings diverging from those made in other studies were the relatively large amount of exudate in the pores, an apparent increase of degradation during infection, and the accumulation of trace elements in one of the canal-wall prostheses. In all likelihood these three phenomena may be attributed to the unfavorable conditions to which these prostheses were exposed during implantation.     

In vitro and in vivo study to the biocompatibility and biodegradation of hydroxyapatite/poly(vinyl alcohol)/gelatin composite

A novel porous composite material composed of hydroxyapatite, poly(vinyl alcohol) (PVA), and gelatin (Gel) was fabricated by emulsification. Scanning electron microscopy showed that the material had a well-interconnected porous structure including many macropores (100-500 microm) and micropores (less than 20 microm) on their walls. The composite had a porosity of 78% and showed high water absorption up to 312.7% indicating a good water-swellable behavior that is a characteristic of hydrogel materials. When immersed in water, the scaffold's weight continuously decreased. After immersion in simulated body fluid, the weight continuously increased because Ca(2+) and PO(3-) (4) ions deposited on the surface and the internal surfaces of the material pores. The deposit was proved to be carbonated hydroxyapatite by thin-film X-ray diffraction, Fourier transform infrared spectroscopy and energy dispersive X-ray analysis. The composite was detected to be non-cytotoxicity by MTT assay. The HA/PVA/Gel material was also implanted subcutaneously in the dorsal region of adult female rats. After 12 weeks of implantation, the porous material adhered tightly with the surrounding tissue, and the ingrowth of fibrous tissue as well as the material's partial degradation was observed, which partly indicated that the composite was biocompatible in vivo. In conclusion, the porous HA/PVA/Gel composite is a promising scaffold for cartilage tissue engineering with more studies.     

In vitro biocompatibility of an ultrafine grained zirconium

We have investigated a novel ultrafine grained (UFG) Zr obtained by severe plastic deformation (SPD) which resulted in a refinement of the grain size by several orders of magnitude. Compared to conventional Zr, higher hardness values were measured on UFG Zr. Polished surfaces having similar topographical features from both materials were prepared, as assessed by atomic force microscopy (AFM). Surface hydrophobicity of Zr, evaluated by measuring water contact angles, was unaffected by grain size reduction. In vitro biocompatibility was addressed on conventional and UFG Zr surfaces and, for comparative purposes, a polished Ti6Al4V alloy was also investigated. Cell attachment and spreading, actin and beta-tubulin cytoskeleton reorganisation, fibronectin secretion and cellular distribution as well as cell viability were evaluated by culturing human osteoblastic Saos-2 cells on the surfaces. The osteoblastic response to conventional Zr was found to be essentially identical to Ti6Al4V and was not affected by grain size reduction. In order to evaluate the ability of the surfaces to promote osteogenic maturation and bone matrix mineralisation, human mesenchymal cells from bone marrow were switched to the osteoblastic phenotype by incubation in osteogenic induction media. Compared to undifferentiated mesenchymal cells, alkaline phosphatase activity and formation of mineralisation nodules were enhanced to the same extent on both Zr surfaces and Ti6Al4V alloy after induction of osteoblastic differentiation. In summary, improved mechanical properties together with excellent in vitro biocompatibility make UFG Zr a promising biomaterial for surgical implants.     

Cellular activity of osteoblasts in the presence of hydroxyapatite: an in vitro experiment.

The use of synthetic calcium phosphate as bone substitute calls for the knowledge of the influence on adjacent cells. Effects on monocytes, macrophages, synovial cells and fibroblasts have been largely described in vivo and in vitro but few data are available as concerns osteoblast responses. The present experiments tested the activity of MC3T3-E1, ROS 17/2.8 and mouse calvaria cells cultured in the presence of hydroxyapatite powders. The three osteoblast-like cells were shown to phagocytoze the calcium phosphate particles. As a consequence, they exhibit reduced cell growth and alkaline phosphatase activity. This response was different when compared with other cell types. The osteogenetic function of osteoblastic cells could be involved in these specific effects of hydroxyapatite.     

Surface characteristics and biocompatibility of sandblasted and acid-etched titanium surface modified by ultraviolet irradiation: an in vitro study

Sandblasting with large grit and acid-etching (SLA) treatment is considered to be a reliable modification to achieve excellent titanium surface. However, contamination of hydrocarbons would make SLA surface hydrophobic and influence its bioactivity. Thus, appropriate methods of preservation or further treatments could be used for improvement. In present study, preservation in deionized water (dH(2)O) and ultraviolet (UV) irradiation were, respectively, applied to achieve modSLA and UV-SLA surfaces. Surface characteristics were assessed by scanning electron microscopy, optical profilometer and x-ray photoelectron spectroscopy as well as wettability by optical contact angle analyzer. Additionally, biocompatibility was evaluated by the response to osteoblast-like MG63 cells. Prevented from further contamination, modSLA surface with fewer hydrocarbons (25.31%) remained hydrophilic and showed better affinity to mineralization of MG63 cells than hydrophobic polluted SLA surface (p < 0.01). Furthermore, with the lowest content of hydrocarbons (14.26%) and super-hydrophilicity, UV-SLA surface, which had the hydrocarbons effectively decomposed by photocatalysis and meanwhile acquired abundant hydroxyl groups, had most greatly promoted the attachment, proliferation, differentiation, and mineralization of MG63 cells (p < 0.05). Therefore, hydrocarbons were found to be an important influencing factor to compatibility of biomaterials. In addition, UV irradiation was recognized as a trustworthy method for surface cleaning without change of topography and roughness and could ever lead to greater biocompatibility of sandblasted and acid-etched titanium surface.     

A PVA/PVP hydrogel for human lens substitution: Synthesis, rheological characterization, and in vitro biocompatibility

To overcome opacification and absence of accommodation of human lens substitutes a new poly(vinyl alcohol) (PVA)/poly(N-vinyl-2- pyrrolidinone) (PVP) based hydrogel (PPS31075) was realised. The Infrared Spectroscopy and the mechanical spectra confirmed the successful occurrence of crosslinking reaction. The rheological analysis pointed out a behavior comparable with that of young human lens in terms of complex shear modulus and accommodation capability. Further analysis in terms of optical properties, water content measurements, diffusion coefficient, cytotoxicity, and human capsular cell adhesion confirmed the applicability of such a hydrogel as potential human lens substitute.     

二价阳离子掺杂羟基磷灰石的基础研究进展

通过离子掺杂到羟基磷灰石(HA)可合成理化和生物性能更优良的无机复合物。本文将从基本特性、体外以及体内的生物学性能概述Mg~(2+)、Zn~(2+)、Sr~(2+)三种二价阳离子掺杂HA骨修复材料的基础研究进展。掺杂Mg~(2+)、Zn~(2+)、Sr~(2+)三种二价阳离子均增强HA的体外生物相容性、生物降解性和骨诱导性。此外,掺杂Zn~(2+)提高HA的抗炎和抗菌性能,掺杂Sr~(2+)提高HA的机械性能。Mg~(2+)、Zn~(2+)、Sr~(2+)三种二价阳离子掺杂HA的动物体内实验均显示出良好的骨修复能力。     

Magnetron co-sputtered silicon-containing hydroxyapatite thin films--an in vitro study

The use of silicon-substituted hydroxyapatite (Si-HA) as a biomaterial has been reported recently. In vivo testing has shown that Si-HA promotes early bonding of the bone/implant interface. In order to extend its usage to major load-bearing applications such as artificial hip replacement implants, it has been proposed that the material could be used in the form of a coating on implant surfaces. This paper reports a preliminary study of the biocompatibility of magnetron co-sputtered silicon-containing hydroxyapatite (Si-HA) coatings on a metallic substrate. Magnetron co-sputtered Si-HA films of thickness 600 nm with a Si content of approximately 0.8 wt% were produced on titanium substrates. X-ray diffraction analysis showed that the as-deposited Si-HA films were either amorphous or made up of very small crystals. The crystallinity of Si-HA films was increased after post-deposition heat treatment at 700 degrees C for 3 h, and the principal peaks were attributable to HA. The formation of nano-scale silicon-calcium phosphate precipitates was noted on the heat-treated films. In vitro cell culture has demonstrated that human osteoblast-like cells attached and grew well on all films, with the highest cell growth and signs of mineralisation observed on the heat-treated Si-HA films. In addition, many focal contacts were produced on the films and the cells had well-defined actin cytoskeletal organisation. This work shows that as-deposited and heat-treated Si-HA films have excellent bioactivity and are good candidates when rapid bone apposition is required. Furthermore, heat-treated Si-HA films have improved biostability compared to as-deposited films under physiological conditions.     

In vitro bioactivity of starch thermoplastic/hydroxyapatite composite biomaterials: an in situ study using atomic force microscopy

The in vitro bioactivity of a composite composed by a biodegradable starch-based polymeric matrix and hydroxyapatite fillers was investigated, in situ, as a function of immersion time in a simulated body fluid (SBF) using atomic force microscopy (AFM). The surface roughness of the composite started to increase after the initial 8h because of both the degradation of the polymer matrix and the nucleation of calcium phosphate. After 24h of immersion the surface of the composite was fully covered with calcium phosphate nuclei with diameters around 126 nm. As the immersion time increased, the nuclei increased both in number and size, and coalesced leading to the formation of a dense and uniform calcium phosphate layer on the surface of the composite only after 126 h of SBF immersion. The results of in situ AFM observation agreed with those of standard in vitro bioactivity tests in combination with scanning electron microscopy observations. Thin-film X-ray diffraction demonstrated that the ratio of apatite to the polymer matrix was higher within the surface layer (40 microm deep from the surface) than that in the bulk after the immersion for 7 days. The water-uptake capability of the polymer contributes to the nucleation and growth of the calcium phosphate layer. These results suggest the great potential of the composite for a range of temporary applications in which bone-bonding ability is a desired property.     

In vitro study on different cell response to spherical hydroxyapatite nanoparticles

Hydroxyapatite (HA) is widely used in filling of bone defects and coating on metal parts of prosthetic implants due to its excellent biocompatibility, bioactivity, and bone-bonding properties. It has been demonstrated that micro-sized HA particles cause inflammatory reaction, especially for the needle shaped particles. However, little effort has been concentrated on the cell responses of the spherical HA nanoparticles. The aim of the present work is to chemically and physically characterize the synthesized HA nanoparticles and to investigate the in vitro cell responses. X-ray diffraction, electron microscopy, nitrogen adsorption, and Fourier transform infrared spectroscopy revealed that the particles consisted of nearly spherical crystallites of carbonate-substituted HA with size of 20-40 nm and specific surface area of 75 m(2)/g. L929 cell proliferation experiments demonstrate that the spherical HA nanoparticles is more biocompatible than commercially available HA. On the other hand, U2-OS cell test results show that the inhibition rate of the spherical HA nanoparticles increases with time and concentration. The half effective inhibitory concentration (IC50) of the nanoparticles was determined to be 50.8 mug/mL at 72 h. All these data indicated that the synthesized spherical nanocrystalline HA particles can function as an effective biomaterial for bone tumorectomy repair, while having little adverse effect.     

Novel silicon-doped hydroxyapatite (Si-HA) for biomedical coatings: an in vitro study using acellular simulated body fluid

Magnetron co-sputtering was used to produce silicon-doped hydroxyapatite (Si-HA) as coatings intended for potential applications such as orthopedic and dental implants. It was found that the crystallinity of the as-sputtered coatings increased after annealing, resulting in a nanocrystalline apatite structure. Subsequently, the bioactivity of the coatings was evaluated in an acellular simulated body fluid (SBF). Physicochemical evaluation demonstrated that a carbonate-containing apatite layer, which is essential for bonding at the bone/implant interface, was formed on the coating surfaces after immersion in SBF between 4 and 7 days. The annealed coatings exhibited enhanced bioactivity and chemical stability under physiological conditions, as compared with the as-sputtered coatings. It is proposed that the rate at which the carbonate-containing apatite layer forms is dependent on the scale factor of the structure. A nanocrystalline structure can provide a higher number of nucleation sites for the formation of apatite crystallites, leading to a more rapid precipitation of carbonate-containing apatite layer. This work shows that Si-HA coatings offer considerable potential for applications in hard tissue replacement, owing to their ability to form a carbonate-containing apatite layer rapidly.     

Flow cytometry analysis of the effects of pre-immersion on the biocompatibility of glass-reinforced hydroxyapatite plasma-sprayed coatings

Multilayered coatings composed of mixtures of HA and P2O5-based bioactive glasses are of potential clinical benefit in orthopaedic and dental surgery. Pre-immersion of these materials has been reported to further enhance their efficacy in vivo, although the precise biological effects of this treatment are not yet known. In this study we have therefore prepared double-layer plasma-sprayed coatings and evaluated the effects of pre-immersion on the growth and function of human osteosarcoma cells in vitro, using the MTT assay and flow cytometry analysis, respectively. The results showed that the increase in numbers of viable cells was the same or elevated following incubation on the pre-immersed HA and glass-reinforced HA coatings compared with the non-immersed materials. In addition, the expression of bone sialoprotein and fibronectin, two key connective tissue antigens, was up-regulated in cultures grown on the pre-immersed surfaces compared with the non-treated materials. Moreover, cell numbers and antigen expression both improved as the proportion of glass increased, particularly in the pre-immersed samples. Our findings thus suggest that the immersion treatment of these materials appeared to improve the response of these bone-like cells.     

Evaluation of the in vitro biocompatibility of various elastomers

A previous study highlighted the superior shock absorption of silicone rubbers compared to other elastomers. We evaluated and compared the in vitro biocompatibility of silicone-based rubbers and propose them as an alternative to conventional products. We used the MTT colorimetric test to assess cell viability and flow cytometry to evaluate cell proliferation. Tests were conducted at 24 and 72 h. Changes in cell morphology were evaluated by scanning electron microscopy. Positive (polyurethane) and negative (polystyrene) toxicity controls were included. The number of viable cells was significantly higher on polystyrene than on polyurethane. A decrease in the total number of cells from 24 to 72 h compared to the negative control was correlated with a lower percentage of S-phase cells. The differences in cell viability noted between the samples and the polystyrene control mainly resulted from an initial lack of adhesion, which was confirmed by scanning electron microscopy. The biocompatibility of the three silicone rubbers was comparable to the best of the three products currently being used. These results, combined with those of the previous study, indicate that silicone rubber could be considered for the manufacture of mouth guards.