改性纳米羟基磷灰石/聚乳酸-聚羟乙酸复合骨髓基质干细胞修复兔桡骨缺损*

背景：前期试验证实骨髓基质干细胞能够在改性纳米羟基磷灰石/聚乳酸-聚羟乙酸材料表面黏附、增殖,该材料具有良好的生物安全性。 目的：观察骨髓基质干细胞与改性纳米羟基磷灰石/聚乳酸-聚羟乙酸材料复合修复兔桡骨缺损的效果。 方法：建立兔15 mm桡骨缺损模型,随机分为3组：空白对照组不进行任何处理,实验组植入改性纳米羟基磷灰石/聚乳酸-聚羟乙酸+骨髓基质干细胞组织工程化骨,对照组植入单纯改性纳米羟基磷灰石/聚乳酸-聚羟乙酸支架材料。 结果与结论：①X射线评价：术后1~12周,实验组骨缺损修复程度及速度明显优于空白对照组与对照组(P < 0.05)。②组织学检测：实验组术后4周即可观察到新生骨和纤维组织长入材料空隙,局部形成陷窝结构;8周时新生骨组织增多,部分可观察到成熟的骨小梁结构;12周时可见大量成熟骨细胞,骨小梁排列紧密,移植材料逐步被新生骨取代,与正常骨组织形态基本一致,且骨小梁出现时间早于空白对照组与对照组。说明骨髓基质干细胞复合改性纳米羟基磷灰石/聚乳酸-聚羟乙酸构建的组织工程化骨能够促进骨缺损处新骨的生成,较单纯支架材料具有明显优势。     

Non-viral bone morphogenetic protein 2 transfection of rat dental pulp stem cells using calcium phosphate nanoparticles as carriers

Calcium phosphate nanoparticles have shown potential as non-viral vectors for gene delivery. The aim of this study was to induce bone morphogenetic protein (Bmp)2 transfection in rat dental pulp stem cells using calcium phosphate nanoparticles as a gene vector and then to evaluate the efficiency and bioactivity of the transfection. We also intended to investigate the behavior of transfected cells when seeded on 3-dimensional titanium fiber mesh scaffolds. Nanoparticles of calcium phosphate encapsulating plasmid deoxyribonucleic acid (DNA) (plasmid enhanced green fluorescent protein-BMP2) were prepared. Then, STRO-1-selected rat dental pulp stem cells were transfected using these nanoparticles. Transfection and bioactivity of the secreted BMP2 were examined. Thereafter, the transfected cells were cultured on a fibrous titanium mesh. The cultures were investigated using scanning electron microscipy and evaluated for cell proliferation, alkaline phosphatase activity and calcium content. Finally, real-time polymerase chain reaction was performed for odontogenesis-related gene expression. The results showed that the size of the DNA-loaded particles was approximately 100 nm in diameter. Nanoparticles could protect the DNA encapsulated inside from external DNase and release the loaded DNA in a low-acid environment (pH 3.0). In vitro, nanoparticle transfection was shown to be effective and to accelerate or promote the odontogenic differentiation of rat dental pulp stem cells when cultured in the 3-dimensional scaffolds. Based on our results, plasmid DNA-loaded calcium phosphate nanoparticles appear to be an effective non-viral vector for gene delivery and functioned well for odontogenic differentiation through Bmp2 transfection.     

牙髓干细胞修复种植体周围骨缺损的研究

目的 建立兔下颌骨前牙即刻种植种植体周围骨缺损的动物模型,考察牙髓干细胞(DPSCs)在种植体周围骨缺损中的骨再生能力.方法 将8只新西兰白兔随机分为两组,分别拔除兔双侧下颌前牙,在拔牙窝颊侧建立2 mm×3 mm骨缺损区,即刻植入种植体,对照组植入Bio-oss骨粉和磷酸盐缓冲液,实验组植入Bio-oss骨粉和DPSCs.通过大体观察和组织学观察(苏木精-伊红染色、Goldner's三色染色和扫描电镜观察)评价植入后4周骨缺损区的骨再生能力.结果 所有实验兔种植体植入顺利,且种植体植入后均稳固,无明显差异.苏木精-伊红和Goldner's三色染色观察均可见实验组有新生骨组织形成,且部分新生骨组织为编织骨,骨细胞体积大、数量多,呈编织状排列,骨细胞分化成熟;而对照组可见少量新生骨样组织形成,骨细胞分化较成熟.扫描电镜结果显示,实验组新生骨与种植体间的骨结合较对照组多.结论 与单独使用Bio-oss骨粉比较,DPSCs与Bio-oss骨粉联用的成骨能力相对较强,新生骨组织与种植体的结合能力更好.     

纳米晶胶原基骨材料在腰椎后外侧植骨融合中的应用：安全性及效果

背景：腰椎后外侧植骨融合中需要进行植骨,但骨量常不够,需再取患者部分自体髂骨补充。 目的：探讨纳米晶胶原基骨材料在腰椎后外侧植骨融合中的应用安全性及效果。 方法：回顾性分析四川省眉山骨科医院2013年2月至2014年2月收治的52例腰椎退行性疾病患者的临床资料,均实施腰椎后外侧植骨融合治疗,术中所使用的植骨材料为患者自体骨颗粒混合纳米晶胶原基骨材料。 结果与结论：52例患者中,内固定后伤口一期愈合的有51例,患者局部皮肤均未出现皮温上升以及红肿、疼痛、流液等现象。患者内固定后功能和疼痛症状均得到不同程度的改善,对患者的内固定后植骨融合情况进行分析,可得内固定后3个月完全融合率达13%,内固定后6个月达54%,内固定后12个月达84%,且未出现假关节,随访12个月,所有患者均未出现螺钉松动或者移位等现象,对固定节段相临椎体情况进行检查,均未出现滑脱或者移位等情况。表明在腰椎后外侧植骨融合中应用纳米晶胶原基骨材料可以获得良好的效果,材料具有生物相容性及一定的安全性。 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     

人骨髓间充质干细胞接种纳米晶胶原骨构建组织工程骨

目的观察体外培养的人骨髓间充质干细胞（mesenchymal stem cells,MSCs）与纳米晶羟基磷灰石/胶原骨（nano-hydroxyapatite/collagen,nHAC）的生物相容性及细胞在nHAC上的生长情况。方法全骨髓法体外培养骨髓间充质干细胞,应用成骨诱导剂诱导向成骨细胞表型转化,通过细胞活性、免疫组化鉴定诱导培养的成骨细胞的细胞学特性。通过倒置显微镜、扫描电镜观察细胞生长及其在nHAC上的生长情况。结果原代培养的骨髓细胞增殖迅速,10～12d左右即可稳定传代,传代细胞7～9d即可传代。经诱导培养的细胞的ALP染色阳性,Von Kossa染色阳性,可见钙化的基质沉积,呈现典型的成骨细胞形态和生物学特征。构建的MSCs与nHAC共培养的模型中,细胞可在nHAC表面良好贴壁。复合培养8天,分布于支架材料上的细胞大量增殖、分泌细胞外基质。第14天,大量细胞在材料表面和孔隙中生长。细胞之间广泛存在突起连接。结论nHAC适合种子细胞的贴附、生长和增殖,是组织工程良好的载体材料。     

STRO-1 selected rat dental pulp stem cells transfected with adenoviral-mediated human bone morphogenetic protein 2 gene show enhanced odontogenic differentiation

Dental pulp stem cells harbor great potential for tissue-engineering purposes. However, previous studies have shown variable results, and some have reported only limited osteogenic and odontogenic potential.Because bone morphogenetic proteins (BMPs) are well-established agents to induce bone and dentin formation,in this study STRO-1-selected rat dental pulp-derived stem cells were transfected with the adenoviral mediated human BMP-2 gene. Subsequently, the cells were evaluated for their odontogenic differentiation ability in medium not containing dexamethasone or other stimuli. Cultures were investigated using light microscopy and scanning electron microscopy (SEM) and evaluated for cell proliferation, alkaline phosphatase(ALP) activity, and calcium content. Real-time polymerase chain reaction (PCR) was performed for gene expression of Alp, osteocalcin, collagen type I, bone sialoprotein, dentin sialophosphoprotein, and dentin matrix acidic phosphoprotein 1. Finally, an oligo-microarray was used to profile the expression of odontogenesis-related genes. Results of ALP activity, calcium content, and real-time PCR showed that only BMP2-transfected cells had the ability to differentiate into the odontoblast phenotype and to produce a calcified extracellular matrix. SEM and oligo-microarray confirmed these results. In contrast, the non-transfected cells represented a less differentiated cell phenotype. Based on our results, we concluded that the adenovirus can transfect STRO-1 selected cells with high efficacy. After BMP2 gene transfection, these cells had the ability to differentiate into odontoblast phenotype, even without the addition of odontogenic supplements to the medium.     

骨髓间充质干细胞在纳米晶胶原基骨内生长以及成骨潜能

背景：纳米晶胶原基骨修复材料是根据仿生原理制备的纳米骨框架材料,其微结构和成分两方面都与天然骨有相似性 ,具有良好的生物相容性。 目的：研究兔骨髓间充质干细胞与纳米晶胶原基骨修复材料体外复合培养的结合程度,以及构建组织工程骨的可行性。 方法：分离兔骨髓间充质干细胞,体外培养、纯化,取第3代骨髓间充质干细胞与纳米晶胶原基骨修复材料体外复合培养,第3,7,20天后激光共聚焦和扫描电镜观察二者复合程度。 结果与结论：骨髓间充质干细胞和纳米晶胶原基骨修复材料复合良好,共聚焦显微镜和环境扫描电镜观察均可见细胞生长;纳米晶胶原基骨修复材料能够作为良好的支架,它能使骨髓间充质干细胞在其内稳定生长。提示骨髓间充质干细胞在纳米晶胶原基骨修复材料内能很好的生长,并且具有成骨潜能。     

骨形态发生蛋白2和7共转染骨髓间充质干细胞的成骨分化

背景：骨形态发生蛋白最主要的作用是诱导骨形成,但因提取困难、代谢速度快、难以准确控制其使用浓度、价格昂贵等限制了其在体外和体内相关研究中的应用。 目的：构建含特异细胞生长因子基因骨形态发生蛋白2,7的腺病毒载体,观察骨形态发生蛋白2和骨形态发生蛋白7基因共转染对兔骨髓间充质干细胞成骨分化的促进作用。 方法：将全骨髓贴壁法分离得到兔骨髓间充质干细胞传至第3代,分为5组,空白组和常规诱导组分别以常规培养基和成骨诱导分化培养基培养;骨形态发生蛋白2, 7腺病毒转染组：分别单独以骨形态发生蛋白2、骨形态发生蛋白7腺病毒进行转染;联合转染组：以2种骨形态发生蛋白腺病毒联合转染。 结果与结论：转染第7天,联合转染组成骨相关基因Runx-2、Osx、Ⅰ型胶原和碱性磷酸酶mRNA表达均较其他各组增高(P < 0.05);骨形态发生蛋白2,7腺病毒转染组上述指标较空白组和常规诱导组增高(P < 0.05)。转染第14天,联合转染组4个指标均较其他各组明显增高(P < 0.05);同时,4个指标表达均高于第7天(P < 0.05)。病毒转染后第7天,联合转染组Ⅰ型胶原和骨钙素蛋白表达均较其他组明显增高(P < 0.05)。提示对骨髓间充质干细胞进行骨形态发生蛋白2腺病毒和骨形态发生蛋白7腺病毒共转染后,较单基因转染和成骨诱导液诱导更能促进成骨相关基因和蛋白的表达。     

骨形态发生蛋白2基因转染犬牙髓细胞

背景：研究发现骨形态发生蛋白2具有诱导间充质细胞向成骨细胞表型分化以及诱导新骨及修复性牙本质形成的能力。 目的：通过对细胞超微结构的分析,探讨骨形态发生蛋白2基因转染的犬牙髓细胞,向成牙本质细胞转化的过程。 方法：将培养的性状稳定的第4代犬牙髓细胞分为3组：基因转染组转染pEGFP-N1-BMP-2基因,空白载体转染组转染EGFP-N1空白荧光载体,未转染组不转染。 结果与结论：成功构建pEGFP-N1-BMP-2真核表达质粒,并成功将其转染犬牙髓细胞, pEGFP-N1-BMP-2质粒转染促进犬牙髓细胞分泌骨形态发生蛋白2。pEGFP-N1-BMP-2质粒转染有促进牙髓细胞碱性磷酸酶活性的作用。透射电镜观察结果显示：转染后的犬牙髓细胞具有成牙本质细胞的形态特点。说明pEGFP-N1-BMP-2 真核表达质粒转染后的牙髓细胞,具有成牙本质细胞的特征。     

骨髓间充质干细胞成骨分化中血管内皮生长因子和骨形态发生蛋白2的表达

背景：有研究表明血管内皮生长因子可促进内皮细胞增殖、分化和迁移,而骨髓间充质干细胞经诱导分化后细胞自身的血管内皮细胞生长因子及骨形态发生蛋白2如何表达,却鲜见报道。 目的：观察兔骨髓间充质干细胞成骨诱导分化过程中血管内皮细胞生长因子和骨形态发生蛋白2的表达。 方法：取第3代新西兰大白兔骨髓间充质干细胞,1 d后将细胞分为两组：对照组不更换培养基;诱导组加入成骨诱导培养基进行成骨诱导,流式细胞仪检测其表面抗原。RT-PCR和Western-Blot检测细胞成骨分化过程中血管内皮细胞生长因子和骨形态发生蛋白2的表达。 结果与结论：兔骨髓间充质干细胞成骨诱导后,细胞形态由长梭形逐渐变为短梭形,多角形,最后成集落层叠生长;在诱导过程中,碱性磷酸酶活性逐渐增高,与对照组比较有显著性差异(P < 0.05);诱导7 d时Ⅰ型胶原免疫细胞化学染色、14 时骨钙素免疫细胞化学染色、21 d时钙化结节染色阳性;RT-PCR和Western-Blot结果显示两组均有骨形态发生蛋白2和血管内皮细胞生长因子表达,诱导后第7,14,21天表达比较,差异有显著性意义(P < 0.05),第7天,两者表达最强。说明兔骨髓间充质干细胞分离、提取操作简便易行,且有较强的生长增殖能力,可诱导分化为成骨细胞。其在成骨诱导过程中,骨形态发生蛋白2和血管内皮生长因子表达均先增强后减弱,血管内皮细胞生长因子表达可能对血管生成有促进作用。     

Reconstruction of calvarial bone defects using poly(amino acid)/hydroxyapatite/calcium sulfate composite

The aim of this study was to evaluate the thermal properties and in vivo interface performance of poly(amino acid) (PAA) and a composite containing PAA, hydroxyapatite (HA), and calcium sulfate (CS), with respect to their suitability for skull repair. Biocompatibility was evaluated by implantation of materials into muscles of rabbits for eight weeks. Skull repair was assessed by implanting PAA, the compact PAA/HA/CS composite (c-PAA/HA/CS), and a one-side-porous PAA/HA/CS composite (p-PAA/HA/CS) into rabbit calvarial defects. The results showed that the PAA/HA/CS composite possessed good heat resistance and possessed excellent biocompatibility and osteoconductivity. Guided bone regeneration and calvarial repair were observed, with excellent integration between calvarial tissue and implant. The p-PAA/HA/CS composite performed best in terms of stability and bone bonding between implant and host bone tissue. Thus, the present work provides new information for the potential use of osteoconductive PAA/HA/CS composites with a macrostructure in calvarial bone repair.     

Osteogenic differentiation of human mesenchymal stem cells in collagen matrices: effect of uniaxial cyclic tensile strain on bone morphogenetic protein (BMP-2) mRNA expression

Human mesenchymal stem cells (hMSCs) differentiate down an osteogenic pathway with appropriate mechanical and/or chemical stimuli. This study describes the successful culture of hMSCs in 3D collagen matrices under mechanical strain. Bone marrow-derived hMSCs were seeded in linear 3D type I collagen matrices and subjected to 0%, 10%, or 12% uniaxial cyclic tensile strain at 1 Hz for 4 h/day for 7 or 14 days. Cell viability studies indicated that hMSCs remained viable throughout the culture period irrespective of the applied strain level. Real-time RT-PCR studies indicated a significant increase in BMP-2 mRNA expression levels in hMSCs strained at 10% compared to the same day unstrained controls after both 7 and 14 days. An increase in BMP-2 was also observed in hMSCs subjected to 12% strain, but the increase was significant only in the 14-day sample. This is the first report of the culture of bone marrow-derived hMSCs in 3D collagen matrices under cyclic strain, and the first demonstration that strain alone can induce osteogenic differentiation without the addition of osteogenic supplements. Induction of bone differentiation in 3D culture is a critical step in the creation of bioengineered bone constructs.     

人骨形态发生蛋白2基因重组腺病毒表达载体转染兔骨髓间充质干细胞*

背景：采用GatewayTM技术构建腺病毒载体,显著弥补了外源性细胞因子局部应用的缺陷。 目的：观察人骨形态发生蛋白2重组腺病毒表达载体(Ad-BMP-2/GFP)转染兔骨髓间充质干细胞后骨形态发生蛋白2的表达情况。 方法：密度梯度离心联合贴壁培养法,分离、培养、纯化兔骨髓间充质干细胞;GatewayTM技术构建的Ad-BMP-2/GFP腺病毒载体转染兔骨髓间充质干细胞。 结果与结论：RT-PCR检测转染诱导后的细胞表达成骨细胞特异性产物骨钙素;转染后兔骨髓间充质干细胞在mRNA水平和蛋白水平均有人骨形态发生蛋白2的表达。结果表明构建的Ad-BMP-2/GFP可高效转染兔骨髓间充质干细胞,骨形态发生蛋白2在细胞中能高效表达。     

骨髓间充质干细胞与复合I型胶原和骨形态发生蛋白2的聚乳酸乙醇酸构建组织工程骨

目的探讨骨髓间充质干细胞(BMSC)体外分离培养后种植到复合Ⅰ型胶原和重组人类骨形态发生蛋白2(rhBMP-2)的聚乳酸乙醇酸(PLGA)生物支架上,构建组织工程骨的可行性.方法密度梯度离心法提取分离BMSC,倒置显微镜观察细胞形态,流式细胞分析法对细胞表面抗原进行鉴定.相分离法制备多孔三维PLGA生物支架,支架材料上复合Ⅰ型胶原和rhBMP-2,扫描电镜观察其超微结构.将第3代的BMSC接种于复合支架上,扫描电镜观察材料的细胞黏附性,将培养6 h 后的细胞-支架复合体植入SD大鼠肌袋内,于2个月后取材进行HE染色,观察其构建组织工程骨的情况.结果 BMSC可在体外分离扩增,表达CD29、CD44,不表达CD34和CD45.制备的PLGA支架孔隙率为90%,平均孔径为100 μm,与BMSC有较好的黏附性.2个月后动物体内细胞-支架复合体的大体观察和HE染色显示,BMSC种植到复合Ⅰ型胶原和rhBMP-2的PLGA生物支架上可构建骨组织.结论 BMSC可在体外长期、稳定培养,是理想的组织工程种子细胞.PLGA与干细胞有较好的黏附性,可用来做组织工程生物材料.BMSC种植到复合Ⅰ型胶原和rhBMP-2的PLGA生物支架上后,在动物体内可构建组织工程骨.     

骨形态发生蛋白4和骨形态发生蛋白4/7基因转染对骨髓基质干细胞成骨活性差异的比较

背景：有文献报道骨形态发生蛋白(bone morphogenetic proteins,BMPs)异源二聚体比同源二聚体的活性高,目前国内外尚无BMP-4/7异源二聚体与BMP-4同源二聚体间诱导成骨活性比较的报道。 目的：观察不同基因BMP-4、BMP-4/7对骨髓基质干细胞(bone marrow mesenchymal stem cells,BMSCs)成骨活性的差异,探讨融合基因BMP-4/7的生物学活性。 方法：经脂质体转染分离培养的兔BMSCs,G418筛选出稳定表达株,利用RT-PCR法证明目的基因在BMSCs中的表达情况;以BMP-4和BMP4/7融合基因修饰的AAV载体,转染兔BMSCs,MTT法检测不同基因转染细胞的增殖能力;以BMP-4单基因和BMP-4/7融合基因修饰的AAV载体转染兔BMSCs 7 d后,测定两组细胞内碱性磷酸酶及骨钙素水平,观察成骨活性的变化。 结果与结论：实验成功构建高滴度的分别携带BMP-4单基因、BMP-4/7融合基因的重组腺相关病毒载体AAV-BMP-4、AAV-BMP-4/7;RT-PCR结果证明目的基因能够在BMSCs中得到稳定表达。AAV-BMP-4及AAV-BMP-4/7载体转染效率分别为68.20%、72.18%;转染BMSCs后,细胞增殖能力均明显提高,BMP-4/7融合基因的细胞增殖能力活性高于BMP-4单基因。以BMP-4和BMP-4/7融合基因修饰的AAV转染细胞7 d后,细胞内碱性磷酸酶活性明显上调,骨钙素水平升高,成骨活性均增强;两种基因比较,BMP-4/7融合基因成骨活性强于BMP-4单基因(P < 0.01)。提示BMP-4、BMP-4/7修饰的AAV载体转染效率高,对BMSCs均有成骨活性,其中BMP-4/7融合基因成骨活性强于BMP-4单基因。     

深低温冻存羟基磷灰石/骨髓间充质干细胞修复兔桡骨缺损

背景：以往研究表明羟基磷灰石复合骨髓间充质干细胞具有良好的骨缺损修复效果,但这种复合材料在冻存后是否具有骨缺损修复效果还不清楚。 目的：观察深低温冻存羟基磷灰石/骨髓间充质干细胞复合修复骨缺损的效果。 方法：制备27只日本大耳白兔桡骨10 mm缺损模型,随机分为冻存复合材料组、新鲜复合材料组与羟基磷灰石组,3组分别于骨缺损处植入-80 ℃保存3个月的羟基磷灰石/同种异体骨髓间充质干细胞复合物、新鲜制备的羟基磷灰石/同种异体骨髓间充质干细胞复合物及单纯羟基磷灰石,植入后8,12周行大体观察、X射线观察及苏木精-伊红染色等组织学观察,并于12周行生物力学检测。 结果与结论：术后12周,冻存复合材料组、新鲜复合材料组骨缺损大部分愈合,有成熟骨小梁通过,有的可见髓腔通畅,塑形较好;羟基磷灰石组骨痂生成少,骨缺损部分愈合,塑形欠佳,新骨生成少于冻存复合材料组、新鲜复合材料组(P < 0.05)。冻存复合材料组、新鲜复合材料组最大载荷明显大于羟基磷灰石组(P < 0.05)。表明低温冻存羟基磷灰石/骨髓间充质干细胞复合植骨材料的骨缺损修复能力与新鲜制作的复合材料几乎一致,未因冻存受到影响。     

振动应力刺激骨髓间充质干细胞修复兔骨缺损

背景：研究证实,力学因素可调控、诱导骨髓间充质干细胞定向分化为骨细胞,提高分化效率。 目的：观察振动应力刺激对兔骨缺损微环境中骨髓间充质干细胞移植修复肱骨骨缺损成骨分化能力的影响。 方法：24只兔按随机数字表法分为非振动单纯骨基质明胶组、非振动骨基质明胶+骨髓间充质干细胞复合植入组、振动骨基质明胶+骨髓间充质干细胞复合植入组,每组8只,建立兔肱骨骨缺损模型。振动组兔置于振动平台,以0.3 G的加速度,25 Hz,正弦波型,1次/d,30 min/次,持续4周施加振动刺激。 结果与结论：造模4周后,大体观察结果显示,振动组骨痂生长良好,组织学切片显示其新生骨量较多,可见大量成骨细胞,骨缺损与断端形成骨性连接;振动组Ⅰ型胶原蛋白、RUNX2 mRNA表达水平明显高于非振动组。提示振动应力刺激可促进骨缺损微环境中骨髓间充质干细胞的成骨分化能力,提高Ⅰ型胶原蛋白、RUNX2 mRNA表达水平,从而加速骨缺损修复的进程。     

NAC-PCCs载骨形态发生蛋白2基因纳米微球促大鼠骨髓间充质干细胞成骨分化

目的　构建半胱氨酸及磷酸胆碱共接枝的仿生壳聚糖载体（NAC-PCCs）,并包封骨形态发生蛋白2（BMP2）基因进行诱导骨髓间充质干细胞成骨分化的研究。 方法　制备pBMP2/NAC-PCCs材料,检测其粒径、形态,绘制DNA缓释曲线,研究微球抗DNA酶降解能力。在骨髓间充质干细胞与材料共培养过程中,检测材料的转染效能、ROS清除能力、BMP2蛋白分泌水平、成骨相关基因RUNX2、OC表达、碱性磷酸酶活性等指标,以研究其对骨髓间充质干细胞成骨分化的影响。 结果　微球材料能有效避免DNA被生物酶降解,其转染效率为23.1%,ROS清除率为(36.13±0.47)%。同时实验结果显示与NAC-PCCs共培养的细胞, 其细胞内BMP2表达水平高于其余各组且成骨分化效果最佳。 结论　NAC-PCCs包封BMP2基因形成的纳米微球材料具有促进BMSC成骨分化作用。     

Weft-knitted silk-poly(lactide-co-glycolide) mesh scaffold combined with collagen matrix and seeded with mesenchymal stem cells for rabbit Achilles tendon repair

Abstract

Natural silk fibroin fiber scaffolds have excellent mechanical properties, but degrade slowly. In this study, we used poly(lactide-co-glycolide) (PLGA, 10:90) fibers to adjust the overall degradation rate of the scaffolds and filled them with collagen to reserve space for cell growth. Silk fibroin-PLGA (36:64) mesh scaffolds were prepared using weft-knitting, filled with type I collagen, and incubated with rabbit autologous bone marrow-derived mesenchymal stem cells (MSCs). These scaffold–cells composites were implanted into rabbit Achilles tendon defects. At 16 weeks after implantation, morphological and histological observations showed formation of tendon-like tissues that expressed type I collagen mRNA and a uniformly dense distribution of collagen fibers. The maximum load of the regenerated Achilles tendon was 58.32% of normal Achilles tendon, which was significantly higher than control group without MSCs. These findings suggest that it is feasible to construct tissue engineered tendon using weft-knitted silk fibroin-PLGA fiber mesh/collagen matrix seeded with MSCs for rabbit Achilles tendon defect repair.     

Examination of bone marrow mesenchymal stem cells seeded onto poly(3-hydroxybutyrate-co-4-hydroxybutyrate) biological materials for myocardial patch

The implantation of bone marrow mesenchymal stem cells (BMSCs) into the heart has been reported to be effective for myocardial infarction; however, it is unknown what methods are most suitable for supporting stem cell growth in a myocardial patch. This study used a new polymer material composed of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] co-cultured with BMSCs to create a myocardial patch. Bone marrow mesenchymal stem cells were obtained from healthy male BSL-C57 mice. The cells were treated with 5-azacytidine to investigate their differentiation into cardiomyocytes. The cells were seeded for 24-hours onto P(3HB-co-4HB) biological material films (n?=?8). Cell-biomaterial constructs were fixed and analyzed using different methods. Bone marrow mesenchymal stem cells were CD34, CD45^??, CD90⁺ (low), and CD73⁺. The cells were stained with anti-cardiac troponin T (anti-cTnT) and anti-connexin 43 (anti-CX43) antibodies after 5-azacytidine treatment. Scanning electron microscopy showed that BMSC morphology was normal, and cell numbers were more abundant on the P(3HB-co-4HB) material surfaces. The growth curve of BMSCs on the biomaterial patches showed that the P(3HB-co-4HB) material permitted good stem cell growth. Owing to its excellent biocompatibility and biodegradability properties, in particular its porosity, P(3HB-co-4HB) is highlighted as an optimal material to support myocardial cell growth and myocardial patch formation in patients with myocardial infarction.