医用聚醚醚酮材料骨整合改性的研究进展

聚醚醚酮（PEEK）材料具有良好的生物相容性、优异的机械性能、与骨组织相似的力学性能等优点,目前已广泛应用于临床各个领域。由于固有的生物惰性,使种植体与骨组织不相整合,从而限制了其作为骨科植入材料的应用。提高PEEK骨整合性成为目前研究的热点。研究者将具有促进成骨细胞增殖和分化的材料（如生物陶瓷、金属材料、生物因子等）通过物理或化学方式修饰于材料表面,或者采用表面物理修饰（如粗糙度、孔隙率、纳米结构等）提高其骨整合性。随着研究的逐步深入,PEEK的成骨活性得到增强,在临床上有了更大的应用前景。本文主要对上述提到的近些年用于提高PEEK生物活性的方法作一综述,旨在为研究者提供参考,并对未来发展提出展望。     

Polyetheretherketone/nano-fluorohydroxyapatite composite with antimicrobial activity and osseointegration properties

Lack of antibacterial activity and binding ability to natural bone tissue has significantly limited polyetheretherketone (PEEK) for many challenging dental implant applications. Here, we have developed a polyetheretherketone/nano-fluorohydroxyapatite (PEEK/nano-FHA) biocomposite with enhanced antibacterial activity and osseointegration through blending method. Smooth and rough surfaces of PEEK/nano-FHA biocomposites were also prepared. Our results showed that in vitro initial cell adhesion and proliferation on the nano-FHA reinforced PEEK composite were improved. In addition, higher alkaline phosphatase activity and cell mineralization were also detected in cells cultured on PEEK/nano-FHA biocomposites, especially for rough PEEK/nano-FHA surfaces. More importantly, the as-prepared PEEK/nano-FHA biocomposite could effectively prevent the proliferation and biofilm formation of bacterial. For in vivo test, the newly formed bone volume of PEEK/nano-FHA group was higher than that of bare PEEK group based on 3D microcomputed tomography and 2D histomorphometric analysis. These reports demonstrate that the developed PEEK/nano-FHA biocomposite has increased biocompatibility and antibacterial activity in vitro, and promoted osseointegration in vivo, which suggests that it holds potential to be applied as dental implant material in dental tissue engineering applications.     

多孔钽的理化/生物特性及其骨整合能力研究进展

近年来,以钛合金为主的骨骼材料得到了广泛的应用。临床研究中,植入体的腐蚀磨损和骨整合能力不足是造成其无菌性松动的主要原因,最终可导致手术失败。因此制备综合性能优越的植入体材料是骨组织工程研究的热点。多孔钽拥有良好的理化性质,耐腐蚀和抗磨损性能出色,其良好的生物相容性和多孔结构可以促进新骨的长入和成骨细胞的增殖,从而提高骨整合能力。并且表面改性技术的发展赋予了多孔钽更加优良的性能和功能,扩展了其临床应用。本文将针对多孔钽的理化/生物特性及其骨整合能力的研究进展进行综述。     

Bone and metal: An orthopaedic perspective on osseointegration of metals

The area of implant osseointegration is of major importance, given the predicted significant rise in the number of orthopaedic procedures and an increasingly ageing population. Osseointegration is a complex process involving a number of distinct mechanisms affected by the implant bulk properties and surface characteristics. Our understanding and ability to modify these mechanisms through alterations in implant design is continuously expanding. The following review considers the main aspects of material and surface alterations in metal implants, and the extent of their subsequent influence on osseointegration. Clinically, osseointegration results in asymptomatic stable durable fixation of orthopaedic implants. The complexity of achieving this outcome through incorporation and balance of contributory factors is highlighted through a clinical case report.     

光固定明胶修饰的聚醚醚酮(PEEK)表面促进细胞增殖及成骨性研究

目的聚醚醚酮(PEEK)具有与骨骼相似的弹性模量,但PEEK作为骨修复材料的应用受到其表面生物惰性及缺乏促成骨性的限制。为了解决这个问题,我们通过表面修饰光固定明胶增强PEEK生物活性。方法我科研团队合成的光固定明胶可通过紫外照射粘附于多种材料表面,并可增强材料的生物相容性。进行SEM、静态水接触角、细胞增殖、细胞形态、碱性磷酸酶分化等表征学及细胞学系统研究。结果研究表明光固定明胶可固定于PEEK表面,改变了PEEK等高分子材料的表面性质。在细胞学研究中,光固定明胶改性后相对于普通PEEK,成骨细胞增殖、伸展、基质分泌及分化能力明显增强。结论通过光固定明胶对PEEK表面进行改性,可明显增强其生物活性,是一种有潜力的骨科内植入物及医疗器械材料。     

医用钛合金材料表面改性对骨整合影响的研究进展

医用钛合金材料是临床应用中较为常见的骨科植入物材料之一,作为一种具备良好的生物相容性、耐腐蚀性以及机械强度的生物材料受到了学者的广泛关注.本文专注于PubMed和Web of Science数据库中的相关文献,着重论述4种不同的表面改性设计对医用钛合金材料骨整合性能的影响.在材料学结构设计不断创新和深入研究的背景下,针...     

Osseointegration of machined,injection moulded and oxygen plasma modified PEEK implants in a sheep model

Machined and injection moulded polyetheretherketone (PEEK) implants with and without an oxygen plasma modification were prepared and implanted in sheep cancellous and cortical bone. After 4, 12 and 26 weeks, osseointegration was evaluated through mechanical push-out tests and histomorphometry. In the cancellous bone, push-out force increased with time, a trend toward higher force was observed for machined compared to moulded, and oxygen plasma modified compared to unmodified. On-going remodelling of the bone was detected in the periphery of the implants at 4 weeks. Minimal or no inflammation was observed with all the implants at all locations and time-points. Bone-implant contact (BIC) was quantified at all-time points and locations for all the four PEEK implant surfaces. The BIC values ranged from 15 to 75% with an average of 29 ± 13% in the cancellous bone and 25–65% with an average of 50 ± 12% in the cortical bone. In the cortical bone the BIC increased significantly from 4 to 26 weeks. This in vivo study has identified that surface topography of PEEK implants influences osseointegration. In addition, oxygen plasma has the potential to increase bone-implant interface stability. This study provides a unique reference for further modifications and in vivo assessment of PEEK implants.     

Enhanced antibacterial property and osteo-differentiation activity on plasma treated porous polyetheretherketone with hierarchical micro/nano-topography

Implantable polyetheretherketone (PEEK) has great biomedical potential as hard tissue substitute in orthopedic application due to its outstanding mechanical properties and excellent biological stability. However, the poor osseointegration and bacteriostatic ability of implantable PEEK become the major barrier for its wide clinic application. In this study, a hierarchically micro/nano-topographic PEEK with specific functional groups (amino and COOH/COOR) has been fabricated using facile sulfonation combined with argon plasma treatment. The new developed hierarchically micro/nano-topographic PEEK have enhanced hydrophilicity, surface roughness, as well as the high ability of apatite-layer forming. Antibacterial assessment shows that as-treated samples exhibit better antibacterial activity. The cellular responses in osteoblast-like MG-63 cells culturing experiment reveal that the micro/nano-topography accompanied with specific functional groups improves the cell adhesion at the initial stage, further ameliorates proliferation and osteogenic differentiation of MG-63. This study proposes a promising approach to increase osteo-differentiation activity and bacteriostasis of PEEK via synergistic effects involving surface topologic structure and chemical modification, which shows great potential in developing advanced implantable materials.     

纳米银与纳米羟基磷灰石混合填塞对即刻种植骨结合的作用

背景：纳米银具有显著的抗菌性能,纳米羟基磷灰石具有良好的生物活性及机械强度,将两者混合既可促进骨形成又具有抗菌性能。 目的：观察纳米银与纳米羟基磷灰石混合填塞在即刻种植骨结合过程中的作用。 方法：拔除9只新西兰大白兔上下颌4个切牙后,随机分为实验组(n=6)与对照组(n=3),实验组在拔牙窝内填塞纳米银与纳米羟基磷灰石混合物,对照组在拔牙窝内填塞纳米羟基磷灰石,两组均即刻植入钛螺纹钉。植入后4,8,12周取完整上下颌骨标本,进行大体观察、扭矩实验测试、X射线骨密度分析及组织学观察。 结果与结论：实验组不同时间点的新生骨灰度值、最大扭矩值均高于对照组(P < 0.05)。植入12周内,实验组成骨速度及新生骨组织成熟度均高于对照组,且始终无炎细胞浸润;对照组植入4周时可见大量炎细胞浸润,植入8周时仍可见少量炎细胞。表明纳米银与纳米羟基磷灰石混合植骨有更好的抗菌作用、生物相容性和骨诱导能力,能够加快骨结合,促进成骨,较单纯使用纳米羟基磷灰石效果好。 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

Coating with genetic engineered hydrophobin promotes growth of fibroblasts on a hydrophobic solid

Class I Hydrophobins self-assemble at hydrophilic–hydrophobic interfaces into a highly insoluble amphipathic film. Upon self-assembly of these fungal proteins hydrophobic solids turn hydrophilic, while hydrophilic materials can be made hydrophobic. Hydrophobins thus change the nature of a surface. This property makes them interesting candidates to improve physio- and physico-chemical properties of implant surfaces. We here show that growth of fibroblasts on Teflon can be improved by coating the solid with genetically engineered SC3 hydrophobin. Either deleting a stretch of 25 amino acids at the N-terminus of the mature hydrophobin (TrSC3) or fusing the RGD peptide to this end (RGD-SC3) improved growth of fibroblasts on the solid surface. In addition, we have shown that assembled SC3 and TrSC3 are not toxic when added to the medium of a cell culture of fibroblasts in amounts up to 125 μg ml⁻¹.     

聚醚醚酮表面多孔羟基化改性对MC3T3-E1细胞黏附、增殖的影响

目的对聚醚醚酮(polyetheretherketone,PEEK)薄片表面进行多孔化和羟基化改性,观察PEEK表面形貌和生物活性的变化,并探讨该改性方法对前成骨MC3T3-E1细胞黏附、增殖的影响。方法超声波环境下浓硫酸处理PEEK表面,在其表面形成大量微孔结构;经湿化学法将PEEK表面的酮类基团还原成羟基基团,改善其表面化学活性,提升PEEK薄片的生物相容性。利用扫描电子显微镜(SEM)、傅里叶变换红外光谱仪(FTIR)及静态水接触角检测改性前后材料表面形貌、化学基团及亲水性的变化。未处理PEEK、多孔化PEEK、羟基化PEEK、多孔羟基化PEEK与MC3T3-E1细胞共培养,评价表面改性后PEEK薄片对细胞黏附、增殖的影响。结果 SEM结果显示浓硫酸处理后的PEEK薄片表面形成密集的空隙大小均匀的微孔结构,FT-IR结果证实羟基化改性成功地在PEEK表面还原出了大量羟基基团。同时,表面多孔化和羟基化改性均可有效提升PEEK材料表面的亲水性能。在体外细胞实验中,不同改性的PEEK材料与MC3T3-E1细胞共培养后结果显示,多孔化、羟基化和多孔羟基化改性均可显著促进细胞黏附和伸展,同时随着时间的延长,其促进细胞增殖的功能也逐步增强。结论表面多孔羟基化改性能有效提高PEEK材料表面的生物学活性和亲水性能,进而显著促进细胞的黏附和增殖。     

l-Cysteine: A biocompatible,breathable and beneficial coating for graphene oxide

Graphene oxide (GO) has been employed in various fields, and its ecological and health risks have attracted much attention. A small and inexpensive biomolecule, l-cysteine, was covalently immobilized onto GO to form l-cysteine-GO (CysGO) as a thio-functionalized nanosheet of 1.4 nm in thickness. Both the d-spacing and crystallinity of CysGO were observed to be lower than those of GO, whereas the D and G peaks remained similar to those of GO. CysGO exhibited remarkable uptake in vivo: no tissue defects, malformation, death or significant hatching delay were observed in zebrafish embryos. Significant DNA damage, decreased Na⁺/K⁺–ATPase activity and decreased mitochondrial membrane potential were not observed for CysGO. As a nonspecific activity linked to nanotoxicology, the unpaired electron spinning intensity of CysGO was approximately two orders of magnitude lower than that of GO. Oxygen microsensor analysis showed that the hypoxic and normoxic environments resulting from the presence of GO and CysGO envelopment, respectively, contributed to the difference in biocompatibility. CysGO also protected embryos from arsenic poisoning. Thus, CysGO has the advantageous properties of GO, exhibits excellent biocompatibility, acts as a breathable coating and antidote, and is suitable for various applications.     

LIGHT基因的克隆及其可溶性表达

目的　克隆LIGHT基因 ,构建含有人LIGHT基因的表达载体 ,诱导其在大肠杆菌中可溶性表达 ,并对表达的LIGHT蛋白的生物学活性进行检测。方法　从人的外周血单个核细胞中克隆LIGHT全长cDNA及其胞外区片段 ,并将其胞外区片段亚克隆至原核表达载体pET 11a中 ,筛选阳性重组质粒pET LIGHT ,以IPTG诱导其可溶性表达 ,并以SDS PAGE和Westernblot检测进行分析。表达的蛋白初步纯化后 ,进行生物学活性分析。结果　RT PCR扩增出了LIGHT全长 72 3bp的cDNA。SDS PAGE和Westernblot分析证实重组pET LIGHT质粒可表达出相对分子质量 (Mr)为 19× 10 3的蛋白。可溶性LIGHT重组蛋白可共刺激T细胞的增殖及诱导IFN γ的产生。结论　本实验成功地将LIGHT胞外区片段在大肠杆菌中进行表达 ,表达的蛋白具有生物学功能 ,这为进一步的LIGHT基因的功能研究打下了基础     

Characterization of surface modification on microelectrode arrays for in vitro cell culture

This study aims to investigate surface-modified microelectrodes on the microelectrode arrays (MEAs) for neuronal interfaces with in vitro cell culture. The polyimide (PI) MEA was fabricated by using micro-electro-mechanical systems (MEMS) techniques. Self-assembled monolayers (SAMs) of 11-mercaptoundecanoic acid (MUA) were utilized to modify the microelectrode surface of the MEA. The SAMs’ modified surface of microelectrodes offered a reliable interface to immobilize biological ligands through covalent bonding. To increase biocompatibility, the poly-d-lysine (PDL) was immobilized on the SAMs’ modified microelectrodes. Several analytical techniques were used to define the physical structure and functional groups of surface-modified gold microelectrodes on the MEA. Spectra of the Fourier transform infrared reflection (FTIR) were applied to characterize the molecular structure of MUA-SAMs and PDL on the microelectrodes. The spectra, two peaks of amide I (at 1,613 cm⁻¹) and amide II (at 1,548 cm⁻¹), revealed that covalent amide bonding existed in PDL-MUA-SAMs modified surfaces. The thickness and formation of the MUA and PDL were also observed and quantified by using an atomic force microscope (AFM). The impedance measurement of PDL-MUA-SAMs modified MEA only increased slightly to an average of 524.6 ± 55.8 kΩ from 352.9 ± 34.4 kΩ of bare gold microelectrode (p < 0.05, N = 20). In addition, the time-course changes of total impedance resulting from cell sealing resistance and gap reactance were recorded for 7 days for inferring the growth of cell lines on the electrode contact of modified MEA. The experiment of 3T3 fibroblasts, PC12 cells, primary glial cells, and primary cortical neurons cultured on the modified MEAs displayed a good adhesion rate. These biocompatibility assays demonstrated that the neuronal cells are able to grow in a proximity to PDL-MUA-SAMs modified microelectrodes of the MEAs for effective electrophysiological stimulation/sensing schemes and for future implantation purposes.     

Topography and biocompatibility of patterned hydrophobic/hydrophilic zein layers

The topography and biocompatibility of zein layers adsorbed on patterned templates containing hydrophilic and hydrophobic regions were investigated. Nanopatterned templates consisting of hydrophilic lines on a hydrophobic background were drawn by dip-pen nanolithography (DPN) on gold-coated surfaces. 16-Mercaptohexadecanoic acid (COOH(CH(2))(15)SH, MHA) was used as primary ink to generate hydrophilic lines. Unpatterned surfaces were backfilled with 18-octadecanethiol (CH(3)(CH(2))(17)SH, ODT), which generated hydrophobic regions. Zein was allowed to adsorb on patterned surfaces from alcohol-water solutions. The topography of zein deposits was observed by atomic force microscopy (AFM). Height profiles from AFM measurements revealed that zein deposits followed closely the nanopatterned templates. The biocompatibility of zein layers assembled over hydrophilic/hydrophobic micropatterned templates was investigated. Templates containing MHA lines and ODT regions were generated by micro-contact printing (microCP). Mouse fibroblasts seeded on patterned zein layers proliferated on zein deposited over MHA lines, but not on zein over ODT. The experiment indicated that fibroblast cells were able to respond to variations in the underlying surface chemistry, transmitted by the different orientation adopted by zein on the different substrates. This property may be useful in controlling the spatial distribution of cells on patterned protein layers.     

增殖诱导配体基因的克隆、表达及生物学活性分析

目的克隆人增殖诱导配体(APRIL)基因,分析APRIL蛋白的生物学活性。方法采用RT2PCR技术,从肿瘤细胞株总RNA中克隆了APRIL全长编码基因;构建了APRIL胞外可溶性片段(sAPRIL)的原核表达载体,对表达的蛋白初步纯化后进行生物学活性检测;构建了APRIL编码区的真核表达载体,经脂质体转染转化细胞株,流式细胞仪和MTT法分析表达产物对细胞周期及细胞生长的影响。结果sAPRIL原核表达载体经IPTG诱导后,发现在相对分子质量(Mr)20×103处有一明显的表达条带,纯化蛋白进行初步活性测定显示其可以剂量依赖方式促进细胞的生长;细胞转染实验证实APRIL能促进多种转化细胞的增殖,但对细胞周期无明显影响。结论成功克隆了人APRIL基因,并将其可溶性胞外区片段在大肠杆菌进行了表达;构建了APRIL真核表达载体,初步分析了APRIL的生物学活性,提示APRIL在肿瘤细胞增殖过程中可能起了重要作用,为进一步进行APRIL基因的功能研究及其临床应用奠定了实验基础。     

A review on the wettability of dental implant surfaces II: Biological and clinical aspects

Dental and orthopedic implants have been under continuous advancement to improve their interactions with bone and ensure a successful outcome for patients. Surface characteristics such as surface topography and surface chemistry can serve as design tools to enhance the biological response around the implant, with in vitro, in vivo and clinical studies confirming their effects. However, the comprehensive design of implants to promote early and long-term osseointegration requires a better understanding of the role of surface wettability and the mechanisms by which it affects the surrounding biological environment. This review provides a general overview of the available information about the contact angle values of experimental and of marketed implant surfaces, some of the techniques used to modify surface wettability of implants, and results from in vitro and clinical studies. We aim to expand the current understanding on the role of wettability of metallic implants at their interface with blood and the biological milieu, as well as with bacteria, and hard and soft tissues.     

Whispering gallery mode biosensor quantification of fibronectin adsorption kinetics onto alkylsilane monolayers and interpretation of resultant cellular response

A Whispering Gallery Mode (WGM) biosensor was constructed to measure the adsorption of protein onto alkysilane self-assembled monolayers (SAMs) at solution concentrations unattainable with other techniques. The high sensitivity was provided by a WGM resonance excited in a silica microsphere that was functionalized with alkylsilane SAMs and integrated in a microfluidic flow cell under laminar flow conditions. It was found that FN adsorbed at biologically relevant surface densities, however, the adsorption kinetics and concentration dependent saturation values varied significantly from work published utilizing alkanethiol SAMs. Mathematical models were applied to the experimental results to interpret the observed kinetics of FN adsorption. Embryonic hippocampal neurons and skeletal myoblasts were cultured on the modified surfaces, a live-dead assay was used to determine the viability of the FN surfaces for cell culture, and major differences were noted in the biological response to the different SAMs. The high sensitivity and simplicity of the WGM biosensor, combined with its ability to quantify the adsorption of any dilute protein in a label-free assay, establishes the importance of this technology for the study of surface accretion and its effect on cellular function, which can affect biomaterials for both in vivo and in vitro applications.     

The combined effects of matrix stiffness and growth factor immobilization on the bioactivity and differentiation capabilities of adipose-derived stem cells

Biomaterial designs are increasingly incorporating multiple instructive signals to induce a desired cell response. However, many approaches do not allow orthogonal manipulation of immobilized growth factor signals and matrix stiffness. Further, few methods support patterning of biomolecular signals across a biomaterial in a spatially-selective manner. Here, we report a sequential approach employing carbodiimide crosslinking and benzophenone photoimmobilization chemistries to orthogonally modify the stiffness and immobilized growth factor content of a model collagen-GAG (CG) biomaterial. We subsequently examined the singular and combined effects of bone morphogenetic protein (BMP-2), platelet derived growth factor (PDGF-BB), and CG membrane stiffness on the bioactivity and osteogenic/adipogenic lineage-specific gene expression of adipose derived stem cells, an increasingly popular cell source for regenerative medicine studies. We found that the stiffest substrates direct osteogenic lineage commitment of ASCs regardless of the presence or absence of growth factors, while softer substrates require biochemical cues to direct cell fate. We subsequently describe the use of this approach to create overlapping patterns of growth factors across a single substrate. These results highlight the need for versatile approaches to selectively manipulate the biomaterial microenvironment to identify synergies between biochemical and mechanical cues for a range of regenerative medicine applications.     

Zn/Ag micro-galvanic couples formed on titanium and osseointegration effects in the presence of S. aureus

Titanium implants possessing simultaneous osseointegration and antibacterial ability are desirable. In this work, three types of Zn/Ag micro-galvanic couples are fabricated on titanium by plasma immersion ion implantation to investigate the osseointegration and antibacterial effects as well as the involved mechanisms. The in vitro findings disclose enhanced proliferation, osteogenic differentiation, and gene expressions of the rat bone mesenchymal stem cells (rBMSCs), as well as good antibacterial ability on all three micro-galvanic couples. Excellent antimicrobial ability is also observed in vivo and the micro-CT and histological results reveal notable osseointegration in vivo despite the presence of bacteria. The Zn/Ag micro-galvanic couple formed on Zn/Ag dual-ion co-implanted titanium shows the best osseointegration as well as good antibacterial properties in vivo obtained from a rabbit tibia model. The difference among the three Zn/Ag micro-galvanic couples can be ascribed to the contact between the Ag NPs and Zn film, which affects the corrosion process. Our results indicate that the biological behavior can be controlled by the corrosion process of the Zn/Ag micro-galvanic couples.