羟基磷灰石陶瓷诱导成骨细胞的研究

通过定量检测在羟基磷灰石(HA)表面生长的成骨细胞骨涎蛋白和骨钙蛋白的表达,研究了羟基磷灰石陶瓷对成骨细胞的影响．研究结果表明,随着成骨细胞培养时间的延长,在羟基磷灰石表面生长的成骨细胞骨钙蛋白和骨涎蛋白的表达量相对于对照组有显著增加．由于骨钙蛋白和骨涎蛋白是成骨细胞分化晚期高度表达的两种标志性蛋白质,因此其表达量的显著提高意味着羟基磷灰石陶瓷可以诱导成骨细胞的成熟。这一结果可进一步为理解HA陶瓷的生物相容性和生物活性提供分子生物学依据。     

纳米相陶瓷支架与人成骨细胞生物相容性的体外实验研究

研究了2种新型多孔纳米陶瓷支架材料(hydroxyapatite,HA和tricalcium phosphate,TCP)的生物相容性.从人4月龄胚胎骨膜组织中分离培养出骨膜来源的成骨细胞(periosteal-derived osteoblast,POB),采用人胚骨膜成骨细胞与2种陶瓷支架的体外复合培养,观察细胞在材料上的生长及生理功能表达情况.发现人POB在HA和TCP表面生长繁殖良好,并发挥成骨功能.材料对细胞的增殖还有一定的促进作用(p<0.05).实验表明: 2种新型多孔陶瓷支架材料均有良好的生物相容性.     

Preparation of bioactive glasses with controllable degradation behavior and their bioactive characterization

Bioactive glasses and ceramics have been widely investigated for bone repair because of their excel-lent bioactive characteristics. However, these biomaterials undergo incomplete conversion into a bone-like material, which severely limits their biomedical application. In this paper, borosilicate bioac-tive glasses were prepared by traditional melting process. The results showed that borosilicate glasses possessed high biocompatibility and bioactivity. In addition, when immersed in a 0.02 mol/L K2HPO4 solution, particles of a borate glass were fully converted to HA. The desirable conversion rate to HA may be achieved through the adjustment of the B2O3/SiO2 ratio. The results of XRD and FTIR analysis indicated that the degradation product was carbonate-substituted hydroxyapatite, which was similar to the inorganic component of bone.     

兔成骨细胞的体外培养及生物学行为观察

以一月龄幼兔的颅顶骨为材料 ,采用组织块法 ,进行细胞的原代培养并传代之 .结果显示 :(1)原代细胞具有成骨细胞的形态特征 ,经 6次传代后细胞形态变化不明显 ;(2 )细胞具有较强的碱性磷酸酶 (AL P)活性且稳中有升 ;(3)细胞培养液和细胞内均含有骨钙素 ;(4)连续培养 1个月后 ,成骨细胞有矿化结节形成 ;(5 )骨片经 4次贴壁仍有细胞爬出且细胞形态相同 .该结果表明所培养的细胞具有成骨细胞的生物学行为 ,为骨细胞生物学和骨组织工程的研究提供了一种客观而有效的手段     

Osteogenic differentiation of mesenchymal stem cells promoted by overexpression of connective tissue growth factor

Objective: Large segmental bone defect repair remains a clinical and scientific challenge with increasing interest focusing on combining gene transfection with tissue engineering techniques. The aim of this study is to investigate the effect of connective tissue growth factor (CTGF) on the proliferation and osteogenic differentiation of the bone marrow mesenchymal stem cells (MSCs). Methods: A CTGF-expressing plasmid (pCTGF) was constructed and transfected into MSCs. Then expressions of bone morphogenesis-related genes, proliferation rate, alkaline phosphatase activity, and mineralization were examined to evaluate the osteogenic potential of the CTGF gene-modified MSCs. Results: Overexpression of CTGF was confirmed in pCTGF-MSCs. pCTGF transfection significantly enhanced the proliferation rates of pCTGF-MSCs (P<0.05). CTGF induced a 7.5-fold increase in cell migration over control (P<0.05). pCTGF transfection enhanced the expression of bone matrix proteins, such as bone sialoprotein, osteocalcin, and collagen type I in MSCs. The levels of alkaline phosphatase (ALP) activities of pCTGF-MSCs at the 1st and 2nd weeks were 4.0- and 3.0-fold higher than those of MSCs cultured in OS-medium, significantly higher than those of mock-MSCs and normal control MSCs (P<0.05). Overexpression of CTGF in MSCs enhanced the capability to form mineralized nodules. Conclusion: Overexpression of CTGF could improve the osteogenic differentiation ability of MSCs, and the CTGF gene-modified MSCs are potential as novel cell resources of bone tissue engineering.     

兔骨髓间充质干细胞复合脱细胞牛松质骨体外相容性研究

探讨脱细胞牛松质骨（Acellular Bovince Cancellous Bone，ABcB）生物衍生材料与骨髓间充质干细胞（bone marrow mesenchymal stem cells，BMSCs）的生物相容性，为组织工程方法修复骨缺损提供依据。将兔骨髓间充质干细胞与ABCB复合并体外培养，然后进行形态学、细胞增殖、蛋白质含量测定。骨髓间充质干细胞能够贴附在ABCB孔隙内生长，细胞生长及蛋白合成功能不受ABCB的影响。ABCB生物相容性良好，可作为组织工程的支架材料应用于骨缺损的修复；BMSCs可以作为骨组织工程种子细胞。     

钛合金/HA涂层新型医用复合材料的制备及研究现状

钛合金/HA涂层材料是一种新型的生物医用复合材料,HA涂层与钛合金基体间的结合强度是影响该材料长期应用效果的重要因素;对其生物相容性的研究,是该复合材料能广泛应用于临床的关键.对此从制备技术、涂层与基体间的结合强度和材料的生物相容性三方面研究了该材料的现状.并指出对医用钛合金/HA涂层复合材料的研究,不仅应从制备工艺出发,探索出能制备涂层与基体间具有足够结合强度、符合临床应用、具有长期应用价值植入材料的制备技术,而且还应加强植入物生物相容性的基础理论研究,确切理解材料表面性能影响生物相容性的原理.通过合适的工艺参数提高钛合金/HA复合材料的生物相容性与生物活性,为该材料在临床中的广泛应用打下坚实的基础.     

大豆苷元对体外培养大鼠成骨细胞增殖与分化的影响

目的探讨大豆苷元对体外培养大鼠成骨细胞的增殖与分化能力的影响.方法从新生大鼠颅骨中分离培养大鼠成骨细胞,加入不同浓度的大豆苷元溶液进行培养,48,72 h后用MTT法测定成骨细胞的增殖,PNPP法测定碱性磷酸酶活性及RIA法测定骨钙素含量.结果不同浓度的大豆苷元作用48,72 h后,测定OD值均高于阴性对照组;成骨细胞在大豆苷元的作用下,与阴性对照相比,碱性磷酸酶活性显著增高;成骨细胞BGP含量显著增加.结论大豆苷元具有促进体外大鼠成骨细胞的增殖以及分化的作用.     

体外培养大鼠颅骨成骨细胞的生物学特性

应用机械和多步酶解的方法，从新生大鼠颅骨分离成骨细胞。偶氮染色法和钙钴法证明细胞的碱性磷酸酶活性呈阳性反应；Ｖｏｎ Ｋｏｓｓａ和钙分子探针Ｆｌｕｏ－３染色显示细胞节内有钙的沉积；电镜和倒置相差显微镜观察可见矿化结节及其形成过程；碱性磷酸酶性活性和骨钙素的动态分析提示这两种蛋白因子与成骨细胞分化与成熟的相关性。这一体外培养方法的建立，为骨的发育和成骨细胞生物学的研究提供了模型。     

Cytoskeleton reorganization and FAK phosphorylation are involved in lanthanum （Ⅲ）-promoted proliferation and differentiation in rat osteoblasts

The effect of lanthanum ion (La³⁺) on osteoblast function and cytoskeleton is assessed in vitro. Osteoblasts were isolated from Sprague-Dawley fetal neonatal rats. Cell proliferation and gene expression levels of cbfa-1, alkaline phosphatase (ALP), osteocalcin (OC), bone sialoprotein (BSP) and osteopontin (OPN) were examined by cell counting and RT-PCR. Cytoskeleton F-actin was stained with rhodamine-conjugated phalloidin and was visualized by a confocal microscope. As the results, 10⁻⁸-10⁻⁴ M La³⁺-induced osteoblast proliferation on day 2. Data from the RT-PCR assay revealed that 10⁻⁶ M La³⁺ up-regulated the expression levels of ALP, BSP, and cbfa-1 on day 4, while it enhanced the expressions of OC and OPN on day 21. The F-actin cytoskeleton was strengthened and reorganized under the exposure of La³⁺. In addition, the phosphorylation of focal adhesion kinase (FAK) was significantly promoted in 24 h evaluated by Western blot analysis. These findings indicate that La³⁺ promotes osteoblast activity through the phosphorylation of FAK and reorganization of the cytoskeleton.     

HMBA对人成骨肉瘤MG-63细胞形态结构和终末分化指标表达的影响

应用环六亚甲基双乙酰胺处理人成骨肉瘤MG-63细胞,观察MG-63细胞处理前后形态与超微结构及其相关终末分化指标的表达变化.实验结果显示,经5 mmol/L HMBA处理后,MG-63细胞体积增大,趋于扁平铺展状态,细胞大小较为一致,排列较为规则,细胞核形态规则,核质比例减小,核仁减少,核内异染色质减少,常染色质增多,细胞核内的线粒体和高尔基体较为发达,内质网数量增多,细胞表面的微绒毛减少,在成熟细胞中可见钙化糖原颗粒,细胞表面出现钙化小泡沉积,并且形成典型的骨结节.常规细胞化学和免疫细胞化学检测显示,HMBA处理后的细胞中Ⅰ型胶原纤维、骨钙素、骨粘素的表达显著增加,实验结果表明HMBA能够有效诱导人成骨肉瘤细胞的分化,并促进其终末分化指标的表达.     

钛表面微弧氧化羟基磷灰石陶瓷膜的结构及其生物活性

采用微弧氧化技术(MAO), 以纯钛(TA2)为基体, 在醋酸钙和磷酸二氢钠电解液体系中, 制备含羟基磷灰石(HA)的生物活性陶瓷膜, 并利用扫描电子显微镜(SEM)、 X射线衍射(XRD)、 X射线[KG*8]能谱(EDS)和红外光谱(FT IR)对膜层进行表征, 通过体外模拟体液浸泡实验检测膜层的生物活性. 结果表明, 纯钛经微弧氧化处理10 min后, 在其表面能生成一层含羟基磷灰石成分的多孔陶瓷膜, 该膜层经模拟体液浸泡48 h后, 其表面覆盖一层含有CO^2-₃的羟基磷灰石（类骨磷灰石）, 即该陶瓷膜层具有良好的生物活性.     

磷酸钙生物陶瓷涂层制备及其研究进展

羟基磷灰石是哺乳动物体内硬组织的主要无机成分 ,人工合成的羟基磷灰石具有优良的生物相容性和生物活性 但是纯的羟基磷灰石脆性大 ,强度较低 钛及钛合金等医用金属材料具有较好的强度、韧性和优良的加工性能 ,但是生物相容性差 金属基体羟基磷灰石涂层材料兼备金属材料优良的力学性能和生物陶瓷材料的生物特性 ,备受材料领域和医学领域学者的关注 涂层与基体的界面结合强度及涂层材料植入后的稳定性 ,是目前研究的热点和难点 简要评述了羟基磷灰石涂层的制备技术 ,介绍了近年来的研究进展 ,对提高结合强度和植入稳定性的方法进行了归纳     

羟基磷灰石/聚己内酯生物复合材料的制备及其生物活性

以聚己内酯(PCL)为基底、羟基磷灰石(HA)为增强剂,采用溶液浇铸法制备HA/PCL复合材料,考察了浸渍于模拟体液(SBF)中的复合材料生物活性,并用X衍射(XRD)和傅立叶变换红外光谱(FT IR)对材料的结构和组成进行表征。结果表明:HA/PCL(wHA=0.30)复合材料在SBF中浸渍14 d后,即在其表面形成一层弱结晶的碳磷灰石(CHA)覆盖层,显示良好的生物活性。     

Cytoskeleton reorganization and FAK phosphorylation are involved in lanthanum(Ⅲ)-promoted proliferation and differentiation in rat osteoblasts

The effect of lanthanum ion(La3+)on osteoblast function and cytoskeleton is assessed in vitro. Osteoblasts were isolated from Snraeuo-tjawiey tetai neonatal rats.Cell proliferation and gene expression levels of cbfa-1,alkaline phosphatase(ALP),osteocalcin(OC),bone sialo-protein(BSP)and osteopontin(OPN)were examined by cell counting and RT-PCR.Cytoskeleton F-actin was stained with rhodamine-con-jugated phalloidin and was visualized by a confocal microscope.As the results,10-8-10-4M La3+-induced osteoblast proliferation on day2.Data from the RT-PCR assay revealed that 10-6 M La3+ up-regulated the expression levels of ALP,BSP,and cbfa-lon day 4,while it enhanced the expressions of OC and OPN on day 21.The F-actin cytoskeleton was strengthened and reorganized under the exposure of La3+.In addition,the phosphorylation of focal adhesion kinase(FAK)was significantly promoted in 24 h evaluated by Western blot analysis.These findings indi-sate that La 3+ promotes osteoblast activity through the phosphorylation of F AK and reorganization of the cytoskeleton.     

碳酸钠液相改性硅-磷酸钙复合骨水泥研究

以质量分数70%的硅酸三钙(Ca3SiO5,C3S)和30%磷酸氢钙(CaHPO4·2H2O,DCPD)复合得到的DCP30粉体材料为固相,以不同浓度碳酸钠溶液为液相,得到碳酸钠改性骨水泥材料。使用X射线衍射(XRD)、扫描电镜(SEM)、万能材料试验机等手段对不同浓度改性材料进行表征。结果显示:添加碳酸钠液相,骨水泥初、终凝时间分别缩短至16和55min;调控碳酸钠液相浓度,可以实现短期抗压强度优化;使用碳酸钠后,固化自发生成羟基磷灰石(HA)。浸泡模拟体液(SBF)7天,材料表面覆盖HA沉积层,生物活性优越。碳酸钠液相改性硅-磷酸钙复合骨水泥体系的水化性能、短期力学性能以及生物活性均优于Ca3SiO5水泥和未改性硅-磷酸钙复合骨水泥,是一种良好的生物活性骨修复材料。     

Bone recognition mechanism of porcine osteocalcin from crystal structure

Osteocalcin is the most abundant noncollagenous protein in bone, and its concentration in serum is closely linked to bone metabolism and serves as a biological marker for the clinical assessment of bone disease. Although its precise mechanism of action is unclear, osteocalcin influences bone mineralization, in part through its ability to bind with high affinity to the mineral component of bone, hydroxyapatite. In addition to binding to hydroxyapatite, osteocalcin functions in cell signalling and the recruitment of osteoclasts and osteoblasts, which have active roles in bone resorption and deposition, respectively. Here we present the X-ray crystal structure of porcine osteocalcin at 2.0 A resolution, which reveals a negatively charged protein surface that coordinates five calcium ions in a spatial orientation that is complementary to calcium ions in a hydroxyapatite crystal lattice. On the basis of our findings, we propose a model of osteocalcin binding to hydroxyapatite and draw parallels with other proteins that engage crystal lattices.     

细胞周期检测作为生物相容性评价指标的研究

应用体外细胞培养法,观察不同质量分数羟基磷灰石浸提液对L-929细胞的细胞学形态的影响,同时采用MTT比色法评价羟基磷灰石对L-929生长和增殖的影响,流式细胞仪检测羟基磷灰石浸提液对L-929 细胞生长周期及凋亡的影响.结果表明,羟基磷灰石浸提液对体外培养的细胞形态无明显影响,对细胞生长和增殖无明显抑制作用;不同质量分数材料浸提液的细胞毒性为0～1级;随羟基磷灰石浸提液质量分数的升高,细胞凋亡率逐渐上升;50%、75%、100%羟基磷灰石浸提液组能明显降低G0/G1期细胞比例,增加S,G2/M期细胞比例,能增加L-929细胞DNA的合成,促进细胞生长和组织修复.细胞周期检测是生物材料生物相容性评价的一种可靠方法和指标.     

磷酸钙生物陶瓷

磷酸钙生物陶瓷材料包括磷酸三钙（β－TCP）和羟基磷灰石（HA）,具有较好的生物相容性和生物活性,本文总结了磷酸钙生物陶瓷材料的制备方法和力学性能研究的最新进展,指出溶液沉淀法和溶胶凝胶法是目前优先使用的精细磷酸钙陶瓷粉末制备工艺,由于磷酸陶瓷的韧性较低,必须对它进行补强增韧,以扩大在临床中的应用范围。