Formation of bone-like apatite on poly(L-lactide) to improve osteoblast-like compatibility in vitro and in vivo

The biomimetic apatite coating process was adopted to modify poly(L-lactide) (PLLA) surfaces with osteoblasts-like cell compatibility. The apatite coating was formed on the pre-hydrolyzed PLLA film and scaffold surfaces by incubating in simulated body fluid (SBF). Scanning electron microscopy and energy dispersive X-ray analyzer were utilized to characterize the composition and the structure of the apatite coating. The cytocompatibility of the modified PLLA films was investigated by testing osteoblast-like attachment, proliferation, alkaline phosphatase (ALP) activity, and cell cycle. Subsequently, the modified PLLA scaffolds were co-cultured with the osteoblasts-like in vitro and subcutaneously implanted into nude mice. The experimental results showed that the formed apatite had a nano-sized particle and matrix configuration. The surface modification of PLLA with apatite coating significantly promoted osteoblast-like compatibility. After a four-week culture in vivo, no significant inflammatory signs were observed in the implanted regions and osteoblast-like congeries with bone-like structure began to form in the scaffolds. The positive results of this study suggest a good way to produce desirable PLLA biomaterials for bone tissue engineering.     

In vitro adhesion and biocompatability of osteoblast-like cells to poly(methylmethacrylate) and poly(ethylmethacrylate) bone cements

A bone cement, poly(ethylmethacrylate)/n-butylmethacrylate (PEMA/nBMA) has been developed with lower exotherm and monomer leaching compared to the traditional poly(methylmethacrylate)/methylmethacrylate (PMMA/MMA) cement. This study compares the in vitro biological response to the cements using primary human osteoblast-like cells (HOB). Cell attachment was qualified by immunolocalization of vinculin and actin cytoskeleton, showing more organization on PEMA/nBMA compared to PMMA/MMA. Proliferation was assessed using tritiated thymidine incorporation, and phenotype expression determined by measuring alkaline phosphatase (ALP) activity. An increase in proliferation and ALP activity was observed on PEMA/nBMA compared to PMMA/MMA. The results confirm the biocompatability of PEMA/nBMA, and an enhanced cell attachment and expression of differentiated cell phenotype.     

软骨细胞在聚乳酸支架中的体外生长行为

采用明胶和氯化钠颗粒作为致孔剂,使用溶剂浇铸／颗粒沥滤法制备了高孔隙率、孔间连通和高机械性能的聚乳酸支架,采用软骨细胞体外培养研究了这两种多孔支架对细胞生长性能的影响．结果表明,软骨细胞在以明胶颗粒为致孔剂制备的多孔支架中的相对数量和GAG的分泌量更多,细胞的活性更高。     

高分子量聚L-乳酸的合成和表征

采用丙交酯开环聚合合成聚L-乳酸(PLLA),研究了引发剂用量、聚合时间对聚L-乳酸分子量的影响.采用FTIR、DSC、TG分析方法对聚乳酸的结构和热性能加以表征.研究结果表明通过对聚合时间以及引发剂用量的控制,合成出高分子量聚L-乳酸.FTIR分析结果证实了聚L-乳酸的结构,DSC分析表明合成出的聚L-乳酸玻璃化转变温度为62℃,结晶度达到42.3%;TG分析表明聚L-乳酸热分解温度为299℃.     

Formation of bone-like apatite layer on chitosan fiber mesh scaffolds by a biomimetic spraying process

Bone-like apatite coating of polymeric substrates by means of biomimetic process is a possible way to enhance the bone bonding ability of the materials. The created apatite layer is believed to have an ability to provide a favorable environment for osteoblasts or osteoprogenitor cells. The purpose of this study is to obtain bone-like apatite layer onto chitosan fiber mesh tissue engineering scaffolds, by means of using a simple biomimetic coating process and to determine the influence of this coating on osteoblastic cell responses. Chitosan fiber mesh scaffolds produced by a previously described wet spinning methodology were initially wet with a Bioglass((R))-water suspension by means of a spraying methodology and then immersed in a simulated body fluid (SBF) mimicking physiological conditions for one week. The formation of apatite layer was observed morphologically by scanning electron microscopy (SEM). As a result of the use of the novel spraying methodology, a fine coating could also be observed penetrating into the pores, that is clearly within the bulk of the scaffolds. Fourier Transform Infrared spectroscopy (FTIR-ATR), Electron Dispersive Spectroscopy (EDS) and X-ray diffraction (XRD) analysis also confirmed the presence of apatite-like layer. A human osteoblast-like cell line (SaOs-2) was used for the direct cell contact assays. After 2 weeks of culture, samples were observed under the SEM. When compared to the control samples (unmodified chitosan fiber mesh scaffolds) the cell population was found to be higher in the Ca-P biomimetic coated scaffolds, which indicates that the levels of cell proliferation on this kind of scaffolds could be enhanced. Furthermore, it was also observed that the cells seeded in the Ca-P coated scaffolds have a more spread and flat morphology, which reveals an improvement on the cell adhesion patterns, phenomena that are always important in processes such as osteoconduction.     

The mechanism of biomineralization of bone-like apatite on synthetic hydroxyapatite: an in vitro assessment.

The mechanism of biomineralization of bone-like apatite on synthetic hydroxyapatite (HA) has been investigated in vitro, in which the HA surface was surveyed as a function of soaking time in simulated body fluid (SBF). In terms of surface structure by transmission electron microscopy with energy-dispersive X-ray spectrometry, the HA whose Ca/P atomic ratio was 1.67 revealed three different characteristic soaking periods in SBF, i.e. the first soaking period, in which the HA surface increased the Ca/P ratio up to 1.83 to form an amorphous phase of Ca-rich calcium phosphate; the second soaking period, in which the HA surface decreased the Ca/P ratio up to 1.47 to form an amorphous phase of Ca-poor calcium phosphate; and the third soaking period, in which the HA surface gradually increased the Ca/P ratio up to 1.65 to eventually produce the bone-like nano-cerystallites of apatite, which grew forming complex crystal assemblies with a further increase in immersion time. Analysis using electrophoresis spectroscopy indicated that, immediately after immersion in SBF, the HA revealed a highly negative surface potential, which increased to reach a maximum positive value in the first soaking period. The surface potential then decreased to again reach a negative value in the second soaking period and thereafter converge to a constant negative value in the third soaking period. This implies that the HA induces biomineralization of apatite by smartly varying its surface potential to trigger an electrostatic interaction, first with positive calcium ions and second with negative phosphate ions in the SBF.     

类骨磷灰石的结构分析与形成机制研究

采用在模拟体液(SBF)中浸泡材料的方法,在致密羟基磷灰石(HA)表面形成类骨磷灰石层,利用XRD和FT-IR分析表面相结构,SEM观察表面形貌,X光电子能谱分析表面化学组成及其价态.结果表明,表面新生相为含PO43-、CO32-和OH-的低结晶度的类骨磷灰石微晶.其形成机制属于溶解-沉淀型,HA陶瓷浸泡与SBF液中,首先产生表面溶解,使表面附近Ca2+、PO43-等离子浓度升高,然后通过静电吸引,在其表面吸附、成核、长大,形成类骨磷灰石新相.     

Formation of bone-like apatite on titanium filaments incubated in a simulated body fluid by using an electrochemical method

Apatite (HA) layer can be formed on the surface of titanium using the electrochemical deposit method. In the present study, we found the current and loading time strongly affected the granule size and quantity of HA. When a low current was applied, needle-like HA was formed and irregular granules were seen at a higher current. We also found the main components of both the HA structures were Ca and P, and the percentage Ca/P was 1.69, which is similar to that of a natural bone.     

L-丙交酯和D,L-丙交酯混合制备PLA工艺探讨

以辛酸亚锡为催化剂,采用开环聚合法,对L-丙交酯和D,L-丙交酯混合制备PLA工艺进行探讨,分别考察了D,L-丙交酯含量、催化剂用量、聚合时间、聚合温度和体系真空度对PLA粘均分子量的影响。经过GPC测试,所得PLA的质均分子量达到1.15×105,并采用红外光谱仪和差热-热重分析仪对PLA进行了结构和热性能表征。     

静电纺丝法制备高分子量聚左旋乳酸纳米纤维及其表征

利用静电纺丝法制备了高分子量聚左旋乳酸(PLLA)纳米纤维,研究了溶液浓度、纺丝距离和纺丝电压对纤维形貌的影响.用扫描电子显微镜(SEM)对纤维形貌进行了表征,结果表明,当PLLA溶液浓度为3%,纺丝距离为20.5cm,纺丝电压为13kV时得到的纤维形貌较好.再用差示扫描量热法(DSC)对纤维的热学性能进行了表征,用X...     

羧化左旋聚乳酸膜的表面性能

用低温等离子体接枝聚合丙烯酸获得表面羧基化的PLLA膜并对其表面性能进行表征.结果表明,等离子体处理条件对膜表面羧基接枝密度有较大影响;与空白PLLA膜相比,羧基化后膜的亲水性得到显著提高,表面接触角由75°减少到24°并长时间保持稳定;L929成纤维细胞在羧化膜上的生长状态良好,呈现出细胞的聚集态,其细胞密度大于空白组.材料表面的羧基化能改善其亲水性并使其带上负电荷,从而有效的促进细胞的黏附和生长.     

N-组氨酸壳聚糖/左旋聚乳酸支架的制备及表征

在冷冻诱导相分离制备N-组氨酸壳聚糖支架(NHCS)的基础上进行二次相分离,利用不同取代度的NHCS和不同N-组氨酸壳聚糖/聚乳酸质量比制备一系列N-组氨酸壳聚糖/聚乳酸(NHCS/PLLA)支架。通过红外光谱、广角X射线衍射、热分析和扫描电镜等来表征NHCS/PLLA支架。结果表明,同一种N-组氨酸壳聚糖支架(50kD-NHCS-3),N-组氨酸壳聚糖/聚乳酸质量比减小,支架孔隙率减小,密度增大。支架材料的孔尺寸约在12~25μm,孔隙率均大于92%,抗压强度和弹性模量分别在0.33~0.78 MPa和1.75~5.28 MPa之间,有望适用于软骨组织工程支架。     

聚乳酸熔体中缠结网络的形成

采用冷冻萃取法制备了不同相对分子质量的部分解缠结聚乳酸试样。通过流变学的方法实时检测了聚乳酸熔体中缠结网络的形成和发展演变过程。结果表明,部分解缠结聚乳酸分子链的缠结过程分为快速穿插阶段和慢速穿插阶段。同时考察了在可控外场作用下缠结网络的形成,结果表明,预剪切和快速熔融会促进分子链缠结网络的形成,而检测频率则对缠结形成没有影响。     

Biodegradation and tissue-reaction in a long-term implantation study of poly(L-lactide)

Three poly(L-lactides) with different molecular weights were synthesized as solid blocks from the melt. Two of those were ground and samples were produced by injection moulding. Specimens, cubes (2×3×3 mm) and rods (2×3×25 mm), were machined out of the samples, yielding the amorphous parts. Specimens of the crystalline parts were directly machined out of the third block. All were implanted into the paravertebral muscle of 70 rats to explore the biodegradation of poly(L-lactide) in vivo and the tissue changes at the implantation site. The rats were sacrificed after 1 to 116 weeks and the implants recovered. Histological sections of the cubes including the surrounding tissue were prepared by the cutting-grinding technique according to Donath. The three different materials were incorporated well, forming a collagenous layer. The crystalline poly(L-lactide) (M _vis 429 000) remained almost stable in form and structure over the whole observation period. No signs of inflammation or foreign-body reaction were observed. The amorphous poly(L-lactide) of higher molecular weight (M _vis 203 000) degraded nearly completely, whereas the amorphous poly(L-lactide) of lower molecular weight (M _vis 120 000) was totally resorbed. After about 8 weeks both injection-moulded materials degraded progressively, subsequently accompanied by a mild to moderate foreign-body reaction. The degradation in the inner part of the implants proceeded faster than in the cortex. The final biodegradation appeared accompanied by a resorptive histiocytic inflammation. The degradation rate and velocity of the amorphous poly(L-lactides) did not overtax the absorption capacity of the surrounding tissue. These properties of biodegradation seem to meet the requirements for a biodegradable material in osteosynthesis.     

生物陶瓷及其复合材料表面诱导类骨磷灰石层形成的研究

钙磷生物材料表面类骨磷灰石层的形成对其植入体内诱导新骨生成起非常重要的作用.本实验采用2倍模拟体液(2×SBF)为介质,通过仿生浸泡的方法研究了羟基磷灰石(HA)陶瓷及其复合材料羟基磷灰石/聚乳酸(HA/PLA)、羟基磷灰石/聚己內脂(HA/PCL)、羟基磷灰石/丝素(HA/SF)和羟基磷灰石/壳聚糖(HA/CS)表面诱导类骨磷灰石层形成的差异.实验结果表明,HA陶瓷及其复合材料在2×SBF溶液中仿生浸泡7d后,各样品表面均有一层厚度不同的类骨磷灰石层生成.并且该类骨磷灰石层的结晶度较低,晶粒较细(15～30nm),与人体自然骨无机物的结构非常类似.其中,在这4种复合材料中,HA/CS和HA/SF复合材料中因丝素蛋白和壳聚糖富含多种功能基团,能有效促进类骨磷灰石晶体的形核和生长,因而诱导类骨磷灰石生成的能力最强,显示其良好的生物活性.     

Hybrid coatings of poly(L-lysine) and apatite on micro-arc oxidized titania

Hybrid coatings of poly(L-lysine) and apatite were formed on the micro-arc oxidized titania through a biomimetic process. Phosphorous (P)-containing titania films were prepared by micro-arc oxidation (MAO) of titanium (Ti) substrates in an electrolyte solution containing β-glycerol phosphate disodium salt pentahydrate. The hybrid coatings were grown by immersing MAO titania in the poly(L-lysine)-containing simulated body fluid (SBF) solution. After 72 h immersion, the globular precipitates appeared on the surface of titania films and grew up to ~ 10 µm. These precipitates consisted of 100–200 nm nano-flakes with a distorted (less straight) morphology. XRD and FT-IR confirmed that these precipitates were poly(L-lysine)-containing apatite nanocomposite.     

聚L-丙交酯合成工艺条件研究

在超声波辐射条件下 ,以丙交酯为原料、辛酸亚锡为催化剂 ,通过开环聚合反应制得聚L 丙交酯 (PLLA)。应用红外光谱 (IR)表征聚合物的结构 ,详细讨论了丙交酯纯度、聚合温度、聚合时间和超声辐射对聚合物分子质量的影响。优化聚合条件可获得粘均分子量 >1 80× 1 0 3的聚丙交酯材料 ,聚合结果具有良好的重复性     

Combinatorial design of calcium meta phosphate poly(vinyl alcohol) bone-like biocomposites

The incidence of degenerative diseases and the ageing population have added to the growing demand for bone grafts. Although autologous bone continues to be the gold standard, limited yield and potential morbidity of the donor site pose considerable challenges. Currently, clinically used synthetic grafts based on calcium phosphates are mechanically brittle and not compliant hence composite scaffolds are expected to be provide viable solutions. In this study we report composites of calcium meta phosphate-poly (vinyl alcohol) with tunable mechanical properties, low swelling and excellent biocompatibility. The elastomeric nature of the composites resist brittle fracture and the scaffolds can be easily shaped to the bone defect by the surgeon. Testing on bone plug shaped specimens of the scaffolds, exhibited superior mechanical properties compared to currently commercially available bone plugs with additional advantages being the ability to increase porosity without compromising properties in compression and degree of swelling, which make these composites promising synthetic alternatives for bone grafts and bone tissue engineering.     

Tailoring the morphology and crystallinity of poly(L-lactide acid) electrospun membranes

Biodegradable poly(L-lactic acid) (PLLA) microfibers were prepared by electrospinning by varying the applied potential, solution flow rate and collector conditions. PLLA fibers with smoothly oriented and random morphologies were obtained and characterized by scanning electron microscopy. The optimum fiber orientation was obtained at 1000 rpm using a 20.3 cm diameter collecting drum, while for higher and lower drum rotation speeds, the rapid random motion of the jets resulted in a random fiber distribution. The deformation of the jet with rapid solidification during electrospinning often results in a metastable phase. PLLA electrospun fibers are amorphous but contain numerous crystal nuclei that rapidly grow when the sample is heated to 70–140 °C. In this way, the degree of crystallinity of the fibers can be tailored between 0 and 50% by annealing. Infrared transmission spectra revealed that the processing conditions do not affect the PLLA samples at the molecular level and that the crystallinity of the samples is related to the presence of α-crystals.