带血供肌瓣复合BMP微球修复兔桡骨缺损的实验研究

目的：探讨以带血供肌瓣(VMF)复合骨形态发生蛋白微球(BMPMs)修复兔桡骨缺损的效果。方法：将壳聚糖与海藻酸钠制成复合微球，包裹骨形态发生蛋白，以带血供肌瓣复合BMP微球植入骨缺损处。通过大体观察、X线检查及组织学观察了解其修复效果。结果：带血供肌瓣复合BMP微球植入兔桡骨缺损处，在8周时缺损修复，组织学检查显示成熟骨组织形成，效果明显优于带血供肌瓣复合单纯BMP组及空白微球组。结论：壳聚糖海藻酸钠微球可作为骨形态发生蛋白的良好载体，以带血供肌瓣复合BMP微球可以修复免桡骨15mm的骨缺损.     

注射型rhBMP-2骨修复材料对兔桡骨骨缺损修复的实验研究

探讨以纤维蛋白胶(fibrin sealant,FS)为载体,复合重组BMP-2(rhBMP-2)的注射型骨修复材料,修复兔桡骨节段性骨缺损的能力,为其临床应用提供实验依据。将实验动物分为:实验组(FS rhBMP-2)、对照组(FS)。于28只新西兰白兔右侧桡骨干处造成1.5cm缺损,然后严密缝合皮下组织及皮肤,将各组材料经正常皮肤注射骨缺损处,术后4、8、16、24周进行放射学、组织学和骨密度等方法检查,对其成骨效应进行研究。结果表明:实验组(FS rhBMP-2)骨缺损区在成骨活跃程度、骨密度和再生髓腔结构等方面均显著优于对照组(FS),使骨缺损得到了彻底修复(P<0.01);对照组不能产生骨性愈合。由此说明以纤维蛋白胶为载体复合rhBMP-2的注射型骨修复材料具有高效的骨修复能力。     

复合rhBMP-2的注射型成骨材料修复犬桡骨骨缺损的实验研究

探讨以纤维蛋白胶(fibrin sealant,FS)为载体,复合rhBMP-2\bFGF的注射型骨修复材料,修复犬桡骨节段性缺损的作用,为其临床应用提供实验依据.实验分为:实验组(FS bFGF rhBMP-2)、对照组(FS).于12只成年健康家犬右侧桡骨中上段造成2cm缺损,制成犬桡骨2cm骨缺损实验模型,然后严密缝合皮下组织及皮肤,将实验材料经伤口周围正常皮肤注射到骨缺损处,术后4、8、16、24w进行放射学检查,术后24w,标本进行组织学和骨密度检查,研究其成骨效应.结果表明:实验组骨缺损区在成骨活跃程度、骨密度和再生髓腔结构等方面均显著优于对照组,使骨缺损得到了成功的修复(P＜0.01).以纤维蛋白胶为载体复合rhBMP-2和bFGF的注射型骨修复材料,具有高效的骨修复能力,对犬的骨缺损有很好的修复效果.     

骨形态发生蛋白2基因与重组蛋白缓释方法修复骨缺损的比较研究

为比较骨形态发生蛋白2(BMP-2)基因与重组蛋白缓释方法修复节段性骨缺损的效果,于兔双侧桡骨中段造成1.5cm骨缺损,采用四种方法修复:A组植入转基因骨髓基质干细胞(MSCs)与PLA/PCL(聚乳酸/聚己内酯)支架的复合物;B组植入单纯MSCs与含重组BMP-2的PLA/PCL缓释载体的复合物;C组植入单纯MSCs与PLA/PCL复合物;D组植入单纯PLA/PCL。术后4、8、12周行X线、组织学、生物力学和骨密度等检测。结果表明,A组体内植入4周后,成骨细胞和间质细胞呈BMP-2强阳性表达;其成骨速度及成骨质量均明显优于B组,12周时骨缺损完全修复。C组成骨能力较弱,而D组则无新骨形成,残留骨缺损。BMP-2基因治疗是修复节段性骨缺损的好方法。     

缓释万古霉素三维支架修复兔感染性骨软骨缺损

背景:大量研究证实组织工程支架几乎可完全修复骨软骨缺损,但当骨软骨缺损合并感染时,即使前期经过彻底的清创,单纯骨软骨组织工程支架的修复效果往往不理想。目的:制备盐酸万古霉素缓释微球丝素蛋白/壳聚糖/纳米羟基磷灰石支架,观察其对兔股骨远端感染性骨软骨缺损的修复效果。方法:①采用乳化溶剂挥发法制备盐酸万古霉素缓释微球;将不同质量(7.5,10,12.5 mg)的缓释微球分别与丝素蛋白-壳聚糖-纳米羟基磷灰石溶液混合,利用化学交联法制备盐酸万古霉素缓释微球丝素蛋白/壳聚糖/纳米羟基磷灰石支架,表征支架的孔隙率、吸水膨胀率、热水溶失率及体外药物缓释等。②将45只新西兰大白兔随机分为空白组、对照组、实验组,每组15只,均建立右后肢股骨远端骨软骨缺损并感染模型,空白组不作任何处理,对照组缺损处植入丝素蛋白-壳聚糖-纳米羟基磷灰石支架,实验组缺损处植入盐酸万古霉素缓释微球(10 mg)丝素蛋白/壳聚糖/纳米羟基磷灰石支架。术后1周,检测血液样本C-反应蛋白、白细胞水平;术后4,8,12周取术区组织,分别进行大体观察与病理学观察。结果与结论:①随着缓释微球含量的增加,支架的孔隙率降低,组间比较差异有显著性意义(P<0.05);3组支架的孔径大小、吸水膨胀率、热水溶失率比较差异均无显著性意义(P>0.05);3组支架体外均可持续释放盐酸万古霉素达30 d以上。②实验组兔血液样本C-反应蛋白、白细胞水平均低于空白组、对照组(P<0.05);实验组兔术后各时间点的大体软骨修复情况明显好于空白组、对照组;苏木精-伊红、Masson、阿利新蓝及Ⅱ型胶原免疫组化染色显示,实验组兔术后各时间点的骨软骨修复效果明显优于空白组、对照组。③结果表明,盐酸万古霉素缓释微球丝素蛋白/壳聚糖/纳米羟基磷灰石支架能有效促进开放性骨软骨缺损的修复。     

复合骨形态发生蛋白缓释材料修复兔桡骨缺损

背景：在聚乳酸-聚羟基乙酸中加入β-磷酸三钙可调控其降解速率和强度,加载骨形态发生蛋白可增强材料的诱导成骨能力。 目的：检验β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼/骨形态发生蛋白缓释材料修复兔桡骨节段性骨缺损的效果。 方法：制作兔左侧桡骨12 mm缺损模型,随机分4组,分别于骨缺损处植入β-磷酸三钙/聚乳酸-聚羟基乙酸材料、β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼缓释材料、β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼/骨形态发生蛋白缓释材料,并设置空白对照组(不植入任何材料)。 结果与结论：术后12周时,β-磷酸三钙/聚乳酸-聚羟基乙酸材料组、β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼缓释材料组、β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼/骨形态发生蛋白缓释组骨缺损都得到较好修复,其中β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼/骨形态发生蛋白缓释组骨缺损修复效果最佳(P < 0.05),空白对照组骨缺损未修复。表明β-磷酸三钙/聚乳酸-聚羟基乙酸/异烟肼/骨形态发生蛋白缓释材料可很好修复兔节段性骨缺损。     

羟基磷灰石/凝胶纳米复合物修复兔颅骨缺损的影像学评估

背景:羟基磷灰石/凝胶纳米新型复合物具备与天然骨相同的机械强度,但对骨缺损的修复能力与成骨效果还有待于进一步研究证实。目的:探索羟基磷灰石/凝胶纳米复合物在兔颅骨缺损中的修复效果。方法:建立兔颅顶骨洞型骨缺损模型,缺损区植入羟基磷灰石/凝胶新型复合物,空白缺损设为阴性对照,自体颅骨设为阳性对照。于植入后4,8,12周通过X射线片与CT观察分析骨缺损修复状况。结果与结论:X射线片观察显示,植入后8周,大鼠自体颅骨修复区与周围正常的骨组织形态类似;羟基磷灰石/凝胶纳米复合物修复区中央位置骨质致密,形态接近正常骨组织,边缘略显模糊,植入后12周,羟基磷灰石/凝胶纳米复合物与骨缺损边缘更加模糊,复合物中心呈不连贯状态,空白缺损组可观察到明显的规则的透射阴影。植入后12周CT检测显示,羟基磷灰石/凝胶新型复合物缺损边缘部分紧密融合,与周围的正常骨组织结合为一体。结果说明,羟基磷灰石/凝胶新型仿生复合物在兔颅骨缺损中可取得较好的修复效果。     

负载转基因成肌细胞和骨形态发生蛋白组织工程化骨修复骨缺损

背景：近年来有报道提出了睫状神经营养因子能够抑制成肌细胞体外成肌分化,可保持成肌细胞去分化状态的理论。两者可通过基因工程技术结合为组织工程化骨的构建提供种子细胞,联合骨形态发生蛋白在一定条件下促进骨缺损的修复。 目的：探讨转染睫状神经营养因子基因成肌细胞与骨形态发生蛋白通过透明质酸钠凝胶载体结合修复兔桡骨节段性缺损的效果。 方法：新西兰大白兔制备左侧桡骨中段造成1.0 cm的节段性骨缺损,缺损处以1.4 cm硅胶管桥接固定,管内植入负载骨形态发生蛋白与转睫状神经营养因子基因成肌细胞的透明质酸凝胶(实验组)、仅复合骨形态发生蛋白的透明质酸凝胶(对照组)或仅植入透明质酸凝胶(空白组)。 结果与结论：动物体内实验表明实验组的X射线阻射密度、大体标本以及病理组织学观察情况均优于对照组和空白组,对照组也优于空白组。说明应用透明质酸钠凝胶复合转基因成肌细胞和骨形态发生蛋白因子作为膜内填充物构建组织工程化骨具有可行性,其应用于修复兔桡骨节段性缺损效果理想。     

用复合诱导型骨修复材料修复兔桡骨缺损的实验研究

以不同的载药方式构建4种壳聚糖/聚羟基丁酸酯-羟基戊酸酯(PHBV)复合诱导型骨修复材料,检测并比较4种支架材料对兔桡骨缺损的修复效果,筛选出最佳骨修复材料并确定最佳药物控释方式。以淫羊藿苷为诱导因子,采用两相混合冷冻干燥技术以微球载药、改性药物微球(W/O法制得并表征)、改性药物与材料共价结合等药物添加方式及不加药制得4种支架材料,并对其进行显微结构以及载药支架药物缓释表征,后将4种材料分别植入兔桡骨缺损处,于1、3、6个月进行X射线及三维CT观察支架材料对兔桡骨缺损的修复情况,HE,Masson染色观察其诱导成骨效果。结果表明,支架材料呈网络状串珠状的显微结构,载药微球粒径分布在3～11 μm,载药支架材料有着良好的药物缓释,其中共价结合组药物释放峰值时间较其他组推迟,为72 h,且峰值后药物缓释量迅速平稳为75 μg左右。X射线及三维CT观察显示,最终共价结合组支架材料骨缺损处连通,且骨密度高于其他3组。HE、Masson染色结果显示,共价结合组成骨效果优于其他组。共价结合的药物添加方式能使支架具有良好的药物缓释效果,进而对兔桡骨缺损表现出良好的修复效果。     

人骨形成蛋白7修饰骨髓间充质干细胞复合仿生型支架材料的体内成骨研究

目的 应用携带人骨形成蛋白7(hBMP7)基因的兔骨髓间充质干细胞(BMSC)与仿生型生物玻璃-胶原-透明质酸-磷脂酰丝氨酸(BG-COL-HYA-PS)支架材料复合培养,植入兔桡骨缺损模型中观察其在体内成骨的能力.方法 携带hBMP7基因或增强型绿色荧光蛋白(EGFP)基因的2型重组腺相关病毒(rAAV2)载体在体外分别转染兔BMSC,再将转染后和未转染的兔BMSC分别与BG-COL-HYA-PS支架材料复合培养7 d后植入3组兔桡骨缺损模型,每组6只兔.各组在分别术后8周、12周通过大体标本观察、影像学、组织学等方法观察骨缺损的修复情况.以正常兔桡骨为对照组(n=3),术后12周比较各组骨缺损修复组织生物力学差异.结果 rAAV2-hBMP7转染的兔BMSC与BG-COL-HYA-PS复合支架材料有良好的生物相容性,植入兔桡骨缺损模型内表现出明确的成骨能力和骨修复能力,而形成的新骨最大压力载荷值低于正常桡骨组织[(188.46±12.24)N比(203.25±19.29)N,P＜0.05].结论 用hBMP7修饰BMSC复合仿生型BG-COL-HYA-PS支架材料构建的组织工程骨具有较强的骨修复能力,但形成的新生骨组织与正常骨组织比较仍然有早期生物力学方面的不足.     

Novel Scaffolds Based on Poly(2-hydroxyethyl methacrylate) Superporous Hydrogels for Bone Tissue Engineering

In this study, poly(2-hydroxyethyl methacrylate) (pHEMA)-based superporous hydrogels were synthesized by radical polymerization of 2-hydroxyethyl methacrylate (HEMA) in the presence of a gas blowing agent, sodium bicarbonate. These hydrogels are: pHEMA, pHEMA–gelatin, glycerol phosphate (GP) cross-linked pHEMA–gelatin, glutaraldehyde (GA) cross-linked pHEMA–gelatin superporous hydrogels (SPHs) and pHEMA–hydroxyapatite (HA) superporous hydrogel composites (SPHCs). The hydrogels have a structure of interconnected pores with pore sizes of approx. 500 μm. Although the extent of swelling decreased when gelatin and HA were incorporated to the pHEMA structure, the time to reach the equilibrium swelling (approx. 20 s) was not affected so much. In the presence of gelatin and cross-linkers, mechanical properties significantly improved when compared with pHEMA SPH. Among all the synthesized hydrogels, pHEMA–HA SPHC showed great improvement in mechanical strength and its elastic modulus value was 0.027±0.002 N/mm². Osteogenic activities of pHEMA-based scaffolds were examined by preosteoblastic MC3T3-E1 cell-culture studies. The mitochondrial activity test (MTT) showed that gelatin-containing scaffolds stimulated cell proliferation compared with other scaffolds, while alkaline phosphatase levels (ALP) and mineralization were found highest for the GP cross-linked pHEMA–gelatin SPH. However, pHEMA SPH and pHEMA–HA SPHC did not support cell proliferation and also differentiation. In conclusion, pHEMA–gelatin SPH and GP cross-linked pHEMA–gelatin SPH can be considered as potential scaffolds for bone tissue-engineering applications.     

Insulin-Like Growth Factor (IGF) and Bone

Bone is not only a rich source of a diverse group of growth factors, but is also a very responsive tissue to such growth promoting agents. IGF-I and IGF-I1 are reported to be synthesized and retained in bone. While both IGF-I and IGF-II stimulate DNA, collagen, and noncollagenous protein synthesis in cultured calvariae, these explant cultures have quantitative differential sensitivities to these IGF's. In addition to the observed increase in collagen synthesis, collagen degradation decreased in calvariae treated with IGF-I or IGF-II.     

Bone bonding behavior of MgO-CaO-SiO2-P2O5-CaF2 glass (mother glass of A.W-glass-ceramics)

In this study, it was found that a Ca-P layer and a Si layer were formed on the interface of the mother glass of apatite-wollastonite containing glass-ceramics (designated AW) and bone tissue. The dissolution of Si, Ca, and P from glass (MgO-CaO-SiO2-P2O5-CaF2) is necessary to form a chemical film (a Si layer and a Ca-P layer). The three kinds of glasses used were 1) a mirror surface of the mother glass (MgO 4.6, CaO 44.9, SiO2 34.2, P2O5 16.3, CaF 0.5 weight ratio) of AW (designated G-AW (mirror], 2) an abraded surface of G-AW (designated G-AW (#2000)), 3) a mirror surface SiO2 glass (designated G-Si, 100% SiO2). The glass plates (15 mm x 10 mm x 2 mm) were implanted into the metaphysis of tibia of mature male rabbits for 10 and 25 weeks. The failure load, when an implant detached from the bone or when the bone itself broke, was measured by a detaching test and the interface of glass/bone was observed by SEM-EPMA. Failure loads in G-Si, G-AW (mirror), and G-AW (#2000) 10 weeks after implantation were 0.18 +/- 0.24, 3.06 +/- 1.29, and 2.94 +/- 1.77 kg, respectively. Those in G-Si, G-AW (mirror), and G-AW (#2000) 25 weeks after implantation were 1.30 +/- 1.18, 3.88 +/- 1.06, and 3.55 +/- 1.51, respectively. The failure loads in G-Si vs. G-AW (mirror) and those in G-Si vs. G-AW (#2000) differed significantly (P less than 0.01). There were no significant differences in the failure load according to the surface roughness of G-AW. As shown by SEM-EPMA observation, a Si layer next to G was adjacent to a Ca-P layer next to the bone. The chemical film showed no increase in thickness as time passed. A Ca-P layer did not form on the interface of Si-G and bone.     

The Role of Ellis‐Van Creveld 2(EVC2) in Mice During Cranial Bone Development

EvC syndrome is a type of autosomal‐recessive chondrodysplasia. Previous case studies in patients suggest abnormal craniofacial development, in addition to dwarfism and tooth abnormalities. To investigate how craniofacial development is affected in EvC patients, surface models were generated from micro‐CT scans of control mice, Evc2 global mutant mice and Evc2 neural crest‐specific mutant mice. The anatomic landmarks were placed on the surface model to assess the morphological abnormalities in the Evc2 mutants. Through analyzing the linear and angular measurements between landmarks, we identified a smaller overall skull, shorter nasal bone, shorter frontal bone, and shorter cranial base in the Evc2 global mutants. By comparing neural crest‐specific Evc2 mutants with control mice, we demonstrated that the abnormalities within the mid‐facial regions are not accounted for by the Evc2 mutation within these regions. Additionally, we also identified disproportionate length to width ratios in the Evc2 mutants at all levels from anterior to posterior of the skull. Overall, this study demonstrates a more comprehensive analysis on the craniofacial morphological abnormalities in EvC syndrome and provides the developmental insight to appreciate the impact of Evc2 mutation within the neural crest cells on multiple aspects of skull deformities. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:46–55, 2018. © 2017 Wiley Periodicals, Inc.     

甲亢患者尿DPD、血清BGP水平变化与骨代谢的关系

目的 :探讨甲状腺功能亢进对骨代谢的影响。方法 :本文对 4 1例甲状腺功能亢进症患者及 4 8例正常人对照用化学发光法测定尿脱氧吡啶啉 (DPD)、FT3、FT4,放射免疫法测定血清BGP。结果 :甲亢组尿DPD水平及血清FT3、FT4、骨钙素 (BGP)水平均明显升高 (p <0 0 1)。结论 :表明甲状腺功能亢进与骨代谢密切相关     

Bone Marrow Stromal Cells (BMSCs) in Bone Engineering: Limitations and Recent Advances

Bone marrow stromal cells (BMSCs) have been isolated for the first time by Friedenstein et al. and since then have been considered the progenitor cells for the skeletal tissues. Indeed BMSCs are clonogenic, fibroblastic in shape, and can differentiate along multiple lineages such as osteoblasts, chondrocytes, adipocytes, and hematopoiesis-supportive stroma. When implanted in vivo on a three-dimensional bioceramic scaffold into immunocompromised mice, BMSCs form bone and hematopoiesis-supportive stroma. The ease of harvest from a donor bone marrow together with the ability to form bone in vivo make BMSCs ideal for clinical applications. Thus, ex vivo expanded BMSCs have been employed, first in large animal models, then in human clinical trials, to repair large bone segmental defects. Further investigation of the expanded BMSC population led to the observation that in vitro expansion appears a limiting passage: cells tend to senesce and lose their multidifferentiation potential with time in culture. To overcome these limitations, two approaches have been proposed: (1) identification of the appropriate culture conditions to prevent senescence by possibly selecting a subpopulation with stem cell characteristics, and (2) engineering of the cells by transfection with the telomerase gene to prevent cells from telomere shortening and consequent aging.     

Bone marrow examination in the koala (Phascolarctos cinereus)

A technique for bone marrow aspiration from the iliac crest in the koala (Phascolarctos cinereus) is described. Bone marrow was obtained from ten healthy koalas under general anaesthesia using the combination of tiletamine HCL and zolazepam HCL. Reference ranges were wide. The mean value for M:E was 1.7 (range 0.8–2.7), the mean percentage proliferating erythroid was 11% (range 5%–16%), and the mean percentage proliferating myeloid was 25% (range 17%–35%). In addition, bone marrow aspiration was performed on 14 koalas with various haematological diseases. Results showed aspiration to be useful in the diagnosis of leukaemia, and the investigation of regenerative anaemia and dysplastic anaemia. It was of varying use in the investigation of non-regenerative anaemia.