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.     

神经干细胞在新型复合支架中的生长和分化

BACKGROUND:Neural stem cells with self-proliferation and differentiation potential are the ideal seed cells for central nervous tissue engineering. Although collagen and silk fibroin as biological scaffold materials have been widely used, both of them used alone have certain shortcomings. Is it possible to combine the two materials to build a novel neural tissue-engineered scaffold? What is the effect of this novel scaffold on the growth and differentiation of neural stem cells? OBJECTIVE:To observe the growth and differentiation of neural stem cells seeded onto the novel composite scaffold. METHODS:The rat embryonic neural stem cells were inoculated onto new composite scaffolds, and then, their growth and differentiation were observed by light microscopy and scanning electron microscopy. Neural stem cells were cultured in conventional suspension culture as control group. Cell counting kit-8 assay was used to detect viability of neural stem cells in the two groups. Three-dimensional composite scaffolds carrying neural stem cells were sliced into paraffin sections to observe the growth and differentiation of neural stem cells by hematoxylin-eosin staining and immunofluorescence staining. RESULTS AND CONCLUSION: Neural stem cells cultured on the new composite scaffold grew and differentiated well, and interconnected synapses were observed. Cell counting kit-8 assay showed that neural stem cells on the scaffold grew well, and the cell viability was significantly higher in the composite scaffold group than that in the control group (P < 0.05). Hematoxylin-eosin staining and immunofluorescence staining of paraffin sections further provided evidence for good growth and differentiation of neural stem cells on the scaffold. These results indicate that the novel composite scaffold with good biocompatibility benefits the growth and differentiation of neural stem cells, promising a favorable application prospect.     

Knitted poly-lactide-co-glycolide scaffold loaded with bone marrow stromal cells in repair and regeneration of rabbit Achilles tendon

The objectives of this study were to evaluate the morphology and biomechanical function of Achilles tendons regenerated using knitted poly-lactide-co-glycolide (PLGA) loaded with bone marrow stromal cells (bMSCs). The animal model used was that of an adult female New Zealand White rabbit with a 10-mm gap defect of the Achilles tendon. In group I, 19 hind legs with the created defects were treated with allogeneic bMSCs seeded on knitted PLGA scaffold. In group II, the Achilles tendon defects in 19 hind legs were repaired using the knitted PLGA scaffold alone, and in group III, 6 hind legs were used as normal control. The tendon-implant constructs of groups I and II were evaluated postoperatively at 2, 4, 8, and 12 weeks using macroscopic, histological, and immunohistochemical techniques. In addition, specimens from group I (n = 7), group II (n = 7), and group III (n = 6) were harvested for biomechanical test 12 weeks after surgery. Postoperatively, at 2 and 4 weeks, the histology of group I specimens exhibited a higher rate of tissue formation and remodeling as compared with group II, whereas at 8 and 12 weeks postoperation, the histology of both group I and group II was similar to that of native tendon tissue. The wound sites of group I healed well and there was no apparent lymphocyte infiltration. Immunohistochemical analysis showed that the regenerated tendons were composed of collagen types I and type III fibers. The tensile stiffness and modulus of group I were 87 and 62.6% of normal tendon, respectively, whereas those of group II were about 56.4 and 52.9% of normal tendon, respectively. These results suggest that the knitted PLGA biodegradable scaffold loaded with allogeneic bone marrow stromal cells has the potential to regenerate and repair gap defect of Achilles tendon and to effectively restore structure and function.     

骨形态发生蛋白12诱导骨髓间充质干细胞构建组织工程肌腱

背景：以往肌腱损伤的修复方法有端端吻合、自体肌腱移植、同种异体肌腱或人工肌腱移植等,但均有其各自缺点。 目的：探讨以家兔骨髓间充质干细胞为种子细胞,骨形态发生蛋白12诱导,聚羟基乙酸复合材料作为支架材料,预构组织工程化肌腱重建家兔跟腱缺损的可能性。 方法：家兔抽取股骨近端骨髓,分离所得细胞传代至第2代,加入10 μg/L骨形态发生蛋白12诱导分化,并与Ⅰ型胶原溶液按一定比例移植于预张的聚羟基乙酸缝线上预制组织工程化肌腱备用。将家兔制备成跟腱缺损模型,分别采用不同方法修复跟腱缺损：组织工程化肌腱修复,Ⅰ型胶原-聚羟基乙酸缝线修复,丝线行端端修复。修复12周对各组肌腱行形态学、力学及组织病理学观察。 结果与结论：修复12周家兔肌腱组织病理切片：组织工程化肌腱组可见大量梭形纤维母细胞顺应力学方向排列均匀分布于胶原中,纤维细胞明显增多,胶原致密;Ⅰ型胶原-聚羟基乙酸缝线组可见部分纤维组织增生伴少许肉芽组织形成,胶原纤维呈松散网丝状,细胞排列紊乱分布不均;丝线组可见纤维组织旁见大量肉芽组织形成。修复12周家兔肌腱生物力学强度：骨形态发生蛋白12+聚羟基乙酸重建肌腱的力学强度明显优于Ⅰ型胶原-聚羟基乙酸组,与丝线缝合组差异无显著性意义;但骨形态发生蛋白12+聚羟基乙酸重建肌腱的力学强度低于正常肌腱。提示以自体骨髓间充质干细胞作为种子细胞,骨形态发生蛋白12诱导,以聚羟基乙酸为支架,可望构建组织工程化肌腱。构建的组织工程肌腱具有一定的生物力学特性,能用于修复跟腱缺损。     

聚己二酸丁二醇酯-对苯二甲酸丁二醇酯/Ⅰ型胶原取向纤维促进前交叉韧带断裂后腱-骨愈合

背景:材料表面的微/纳米结构对细胞的行为具有调控作用,肌腱组织主要由平行的胶原纤维组成,因此取向纤维结构对腱-骨愈合具有一定的促进作用.目的:探索聚己二酸丁二醇酯-对苯二甲酸丁二醇酯[poly(butylene adipate-co-terephthalate),PBAT]/Ⅰ型胶原取向纤维支架的生物相容性和成骨诱导活...     

基于丝素/胶原/羟基磷灰石仿生骨材料的多维结构及其性能

目的 体外构建丝素蛋白（silk fibroin,SF）、I型胶原(type I collagen,Col-I)和羟基磷灰石(hydroxyapatite, HA)共混体系制备二维复合膜和三维仿生支架,研究其理化性质和生物相容性,探讨其在组织工程支架材料中应用的可行性。方法 通过在细胞培养小室底部共混SF/Col-I/HA以及低温3D打印结合真空冷冻干燥法制备二维复合膜及三维支架。通过机械性能测试、电子显微镜和Micro-CT检测材料的理化性质,检测细胞的增殖评估其生物相容性。结果 通过共混和低温3D打印获得稳定的二维复合膜及三维多孔结构支架;力学性能具有较好的一致性,孔径、吸水率、孔隙率和弹性模量均符合构建组织工程骨的要求;支架为网格状的白色立方体,内部孔隙连通性较好; HA均匀分布在复合膜中,细胞黏附在复合膜上,呈扁平状;细胞分布在支架孔壁周围,呈梭形状,生长及增殖良好。结论 利用SF/Col-I/HA共混体系成功制备复合膜及三维支架,具有较好的孔连通性与孔结构,有利于细胞和组织的生长以及营养输送,其理化性能以及生物相容性符合骨组织工程生物材料的要求。     

丝素蛋白-壳聚糖支架复合骨髓间充质干细胞体内构建组织工程化软骨的生物相容性

文题释义： 生物相容性：是指生命体组织对非活性材料产生的一种性能,一般是指材料与宿主之间的相容性,包括组织相容性和血液相容性。 检测相容性的方法：是将支架材料与种子细胞在体外共培养,检测支架毒性、细胞活性、细胞增殖及细胞与支架的黏附情况等指标,该方法具有客观性强、可重复性强、影响因素相对简单及敏感性高等特点。 背景：课题组前期的研究中发现,丝素蛋白-壳聚糖支架材料复合诱导后骨髓间充质干细胞在兔体内能修复缺损的软骨组织,但对于该组织工程化软骨组织的生物相容性还未进一步研究。 目的：研究丝素蛋白-壳聚糖支架材料复合骨髓间充质干细胞在体内构建组织工程化软骨的生物相容性。 方法：使用丝素蛋白-壳聚糖按1∶1比例混合制备三维支架材料,提取兔骨髓间充质干细胞,将诱导后的骨髓间充质干细胞与丝素蛋白-壳聚糖支架构建修复体,再将修复体移植到兔关节软骨缺损模型中修复软骨组织。实验分为3组,实验组植入诱导后骨髓间充质干细胞+丝素蛋白-壳聚糖支架,对照组植入丝素蛋白-壳聚糖支架干预,空白组未植入修复体。 结果与结论：①实验成功制备丝素蛋白-壳聚糖三维支架材料及提取骨髓间充质干细胞,并构建软骨缺损的修复体,将修复体植入兔体内能成功修复缺损的软骨组织;②建模后2,4,8,12周,3组血常规、降钙素原、血沉、C-反应蛋白结果提示无明显的全身感染征象,3组血常规及肝肾功能各时间段比较差异无显著性意义(P > 0.05);③一般观察、苏木精-伊红染色及扫描电镜观察：建模后12周,相比其他两组,实验组软骨缺损已修复,支架材料已吸收,修复组织周围未见炎性细胞,修复组织已正常组织整合良好;④结果证实,丝素蛋白-壳聚糖支架复合骨髓间充质干细胞在体内构建的组织工程化软骨具有良好的生物相容性。 ORCID: 0000-0002-8139-1175(佘荣峰) 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

不同来源蚕丝蛋白修复骨软骨组织缺损的效果比较

背景：目前还没有研究比较不同种属来源蚕丝蛋白修复骨软骨的效果。 目的：观察桑蚕和柞蚕来源蚕丝蛋白支架材料修复骨软骨缺损的效果差异。 方法：取新西兰兔20只,制备单侧膝关节骨软骨缺损模型,随机分为5组：其中1组不植入任何材料,作为空白对照;实验1组将3层桑蚕丝蛋白支架粘在一起,充填于缺损处;实验2组将1个包被转化生长因子β3的桑蚕丝蛋白支架与2个包被骨形态发生蛋白2的桑蚕丝蛋白支架粘在一起,充填于缺损处;实验3组将3层柞蚕丝蛋白支架粘在一起,充填于缺损处;实验4组将1个包被转化生长因子β3的柞蚕丝蛋白支架与2个包被骨形态发生蛋白2的柞蚕丝蛋白支架粘在一起,充填于缺损处。术后8周,取关节骨软骨修复区行组织病理学观察,检测Ⅰ型和Ⅱ型胶原蛋白表达。 结果与结论：实验1组胶原蛋白广泛分布于缺损区全层;实验2组胶原纤维在修复区表面呈平行分布,在中间和底部呈垂直顶部的方向分布;实验3组在修复区表面可见胶原;实验4组在支架表面和底部可见软骨样细胞有成团分布。4个实验组修复区Ⅰ型胶原蛋白呈强阳性表达;实验1组、实验2组修复区Ⅱ型胶原蛋白呈微弱表达,实验3组、实验4组修复区Ⅱ型胶原蛋白呈强阳性表达。表明两种蚕丝蛋白均能修复骨软骨缺损,桑蚕丝蛋白倾向于形成骨组织,柞蚕丝蛋白倾向于形成软骨组织。 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      

Repair of Achilles tendon defect with autologous ASCs engineered tendon in a rabbit model

Adipose derived stem cells (ASCs) are an important cell source for tissue regeneration and have been demonstrated the potential of tenogenic differentiation in vitro. This study explored the feasibility of using ASCs for engineered tendon repair in vivo in a rabbit Achilles tendon model. Total 30 rabbits were involved in this study. A composite tendon scaffold composed of an inner part of polyglycolic acid (PGA) unwoven fibers and an outer part of a net knitted with PGA/PLA (polylactic acid) fibers was used to provide mechanical strength. Autologous ASCs were harvested from nuchal subcutaneous adipose tissues and in vitro expanded. The expanded ASCs were harvested and resuspended in culture medium and evenly seeded onto the scaffold in the experimental group, whereas cell-free scaffolds served as the control group. The constructs of both groups were cultured inside a bioreactor under dynamic stretch for 5 weeks. In each of 30 rabbits, a 2 cm defect was created on right side of Achilles tendon followed by the transplantation of a 3 cm cell-seeded scaffold in the experimental group of 15 rabbits, or by the transplantation of a 3 cm cell-free scaffold in the control group of 15 rabbits. Animals were sacrificed at 12, 21 and 45 weeks post-surgery for gross view, histology, and mechanical analysis. The results showed that short term in vitro culture enabled ASCs to produce matrix on the PGA fibers and the constructs showed tensile strength around 50 MPa in both groups (p > 0.05). With the increase of implantation time, cell-seeded constructs gradually form neo-tendon and became more mature at 45 weeks with histological structure similar to that of native tendon and with the presence of bipolar pattern and D-periodic structure of formed collagen fibrils. Additionally, both collagen fibril diameters and tensile strength increased continuously with significant difference among different time points (p < 0.05). In contrast, cell-free constructs failed to form good quality tendon tissue with fibril structure observable only at 45 weeks. There were significant differences in both collagen fibril diameter and tensile strength between two groups at all examined time points (p < 0.05). The results of this study support that ASCs are likely to be a potential cell source for in vivo tendon engineering and regeneration.     

Carbon Nanotube Reinforced Collagen/Hydroxyapatite Scaffolds Improve Bone Tissue Formation <Emphasis Type="Italic">In Vitro</Emphasis> and <Emphasis Type="Italic">In Vivo</Emphasis>

Current bone regeneration strategies faced major challenges in fabricating the bionic scaffolds with nano-structure, constituents and mechanical features of native bone. In this study, we developed a new porous scaffold by adding the multi-walled carbon nanotube (MWCNT) into collagen (Col)/hydroxyapatite (HA) composites. Data showed that 0.5%CNT/Col/HA (0.5%CNT) group was approximately tenfolds stiffer than Col–HA, and it was superior in promoting bone marrow mesenchymal stem proliferation and spreading, mRNA and protein expressions of bone sialoprotein (BSP) and osteocalcin (OCN) than Col–HA group. Moreover, we utilized 0.5%CNT composite to repair the rat calvarial defects (8 mm diameter) in vivo, and observed the new bone formation by 3D reconstruction of micro CT, HE and Masson staining, and BSP, OCN by immunohistochemical analysis. Results showed that newly formed bone in 0.5%CNT group was significantly higher than that in Col–HA group at 12 weeks. These findings highlighted a promising strategy in healing of large area bone defect with MWCNT added into the Col–HA scaffold as they possessed the combined effects of mechanical strength and osteogenicity.     

结合3D打印技术构建聚己内酯-骨细胞外基质复合支架及体外骨诱导性能

BACKGROUND: Scholars are still looking for ideal bone tissue-engineered scaffolds, and  three-dimensional (3D) printing technology is a novel construction method. In the meanwhile, bone extracellular matrix is becoming a hotspot in osteogenic induction. OBJECTIVE: To construct the polycaprolactone/bone extracellular matrix scaffold using 3D printing technology and co-culture method, and to detect its osteogenic property. METHODS: 216 3D-printed polycaprolactone scaffolds were divided into group A (96 pores, n=72) and group B(48 pores, n=144). Passage 5 bone marrow mesenchymal stem cells from Sprague-Dawley rats were seeded onto the two kinds of polycaprolactone scaffolds, and the group A was used for alizarin red staining and Masson staining, while the group B for collagen and glycosaminoglycan detection at 1, 2 and 3 weeks of incubation. Afterwards, the scaffolds at 1, 2 and 3 weeks of culture were decellularized and labeled as groups AE1, AE2, AE3, BE1, BE2 and BE3. Then passage 5 bone marrow mesenchymal stem cells from Sprague-Dawley rats were seeded onto each scaffold again, and the former three groups underwent alizarin red staining, and the latter three were used for calcium, alkaline phosphatase activity and DNA quantitative analysis at 1, 2 and 3 weeks of culture. RESULTS AND CONCLUSION: Masson staining, glycosaminoglycan and hydroxyproline quantitative analysis showed that the extracellular matrix on the composite scaffold increased with time. Alkaline phosphatase activity revealed that the composite scaffold had a significantly stronger osteogenic differentiation than the normal polycaprolactone scaffold (P < 0.05). Alizarin red staining and calcium quantitative analysis showed that the mineralization of the composite scaffold was more obvious than that of the normal polycaprolactone scaffold (P < 0.05), but the total DNA analysis did not differ significantly between scaffolds. These results suggest that the composite scaffold with extracellular matrix is constructed successfully using the 3D technology and co-culture method and exhibits a better osteoinductivity.     

Optimized conditions for mesenchymal stem cells to differentiate into osteoblasts on a collagen/hydroxyapatite matrix

Collagen/hydroxyapatite (HA) composite scaffolds are known to be suitable scaffolds for seeding with mesenchymal stem cells (MSCs) differentiated into osteoblasts and for the in vitro production of artificial bones. However, the optimal collagen/HA ratio remains unclear. Our study confirmed that a higher collagen content increased scaffold stiffness but that a greater stiffness was not sufficient for bone tissue formation, a complex process evidently also dependent on scaffold porosity. We found that the scaffold pore diameter was dependent on the concentration of collagen and HA and that it could play a key role in cell seeding. In conclusion, the optimal scaffold for new bone formation and cell proliferation was found to be a composite scaffold formed from 50 wt % HA in 0.5 wt % collagen I solution.     

胶原/丝素蛋白支架联合神经干细胞治疗创伤性脊髓损伤北大核心

背景:随着交通事故、坠落伤、运动损伤等不断增多,创伤性脊髓损伤已成为危及脊髓健康的一个重要问题。目的:探讨胶原/丝素蛋白支架联合神经干细胞治疗创伤性脊髓损伤的疗效。方法:①分别提取胶原和丝素蛋白原料,将二者以质量比2∶4混合,采用真空冷冻干燥法制备胶原/丝素蛋白支架;将第3代GFP小鼠神经干细胞接种至胶原/丝素蛋白支架上,光学显微镜和扫描电镜下观察神经干细胞生长情况。②采用随机数字表达将40只成年SD大鼠分5组,正常组不进行任何处理,模型组建立T10段脊髓缺损模型,干细胞组脊髓缺损处注射神经干细胞,支架组脊髓缺损处植入胶原/丝素蛋白支架,联合组脊髓缺损处植入接种神经干细胞的胶原/丝素蛋白支架,每组8只。术后每周进行旷场实验BBB评分和斜坡实验,术后第8周行神经诱发电位检测、苏木精-伊红和免疫荧光染色,评价创伤性脊髓损伤大鼠恢复情况。结果与结论:①光学显微镜和扫描电镜下观察显示,胶原/丝素蛋白支架上有利于神经干细胞的黏附、伸展与分化。②旷场实验BBB评分和斜坡实验结果显示,脊髓损伤各组大鼠的运动功能随时间的延长均有不同程度的恢复,各治疗组大鼠的运动功能的恢复速度与程度均优于模型组,其中以联合组大鼠运动功能恢复最好。术后第8周,脊髓损伤后各治疗组大鼠的运动诱发电位和体感诱发电位的潜伏期、振幅检测结果均优于模型组(P<0.05),联合组检测结果优于干细胞组、支架组(P<0.05)。术后第8周苏木精-伊红染色显示,模型组大鼠脊髓损伤部位修复效果最差,联合组修复效果最好;免疫荧光染色显示,模型组大鼠脊髓损伤部位神经丝蛋白阳性细胞最少,各治疗组神经丝蛋白阳性细胞均多于模型组,其中以联合组最多。③胶原/丝素蛋白支架联合神经干细胞对创伤性脊髓损伤大鼠双下肢功能改善和脊髓组织修复具有一定效果。     

Preparation and characterization of collagen-hydroxyapatite composite used for bone tissue engineering scaffold

In this study, highly porous collagen-HA scaffolds were prepared by solid-liquid phase separation method. Microstructure of the composites was characterized by SEM, TEM and XRD. The results show that collagen-HA scaffolds are porous with three-dimension interconnected fiber microstructure, pore sizes are 50-150 microm, and HA particles are dispersed evenly among collagen fiber. Compared with pure collagen, the mechanical property of collagen-HA composite improves significantly. To gain further insight into cell growth throughout 3D scaffolds, the cell proliferation and attachment on the scaffold in vitro was investigated. The collagen-HA composite has good biocompatibility, and adding HA does not affect the histocompatibility of the scaffold materials. The porous collagen-HA composite is suitable as scaffold used for bone tissue engineering.     

复合骨碎补总黄酮的丝素蛋白/壳聚糖支架在兔关节软骨损伤中的应用研究

目的 以丝素蛋白/壳聚糖支架为载体将骨碎补总黄酮应用于兔软骨损伤局部,观察修复效果,为临床提供实验数据。方法 制备丝素蛋白/壳聚糖支架、骨碎补总黄酮缓释微球与负载骨碎补总黄酮缓释微球的丝素蛋白/壳聚糖支架,扫描电子显微镜下观察支架形貌,同时检测该支架的体外缓释能力。24只新西兰大白兔随机分3组,利用电钻在股骨滑车部位构建直径3.5 mm、深1.5 mm的软骨损伤模型,空白组软骨缺损处不植入任何材料,对照组植入单纯的丝素蛋白/壳聚糖支架,实验组植入负载骨碎补总黄酮缓释微球的丝素蛋白/壳聚糖支架,术后12周、24周行标本大体与组织学观察,RT-PCR检测修复组织Sox-9、II型胶原与聚集蛋白聚糖mRNA的表达量,Western blot检测软骨缺损部位II型胶原蛋白表达,分析软骨修复效果。结果 丝素蛋白/壳聚糖支架具有良好的三维孔隙结构,孔洞之间相互联通;制备的载药微球表面较光滑,为较规则的圆球形;载药微球均匀分散于丝素蛋白/壳聚糖支架基质中。丝素蛋白/壳聚糖支架可在体外持续稳定地释放骨碎补总黄酮,实验组软骨损伤修复效果优于对照组,对应的ICRS评分与Wakitani组织学评分高于对照组...     

Effect of self-assembled nanofibrous silk/polycaprolactone layer on the osteoconductivity and mechanical properties of biphasic calcium phosphate scaffolds

We here present the first successful report on combining nanostructured silk and poly(ε-caprolactone) (PCL) with a ceramic scaffold to produce a composite scaffold that is highly porous (porosity ∼85%, pore size ∼500 μm, ∼100% interconnectivity), strong and non-brittle with a surface that resembles extracellular matrix (ECM). The ECM-like surface was developed by self-assembly of nanofibrous structured silk (20-80 nm diameter, similar to native collagen found in ECM) over a thin PCL layer which is coated on biphasic calcium phosphate (BCP) scaffolds. The effects of different concentrations of silk solution on the mechanical and physical properties of the scaffolds were also comprehensively examined. Our results showed that using silk only (irrespective of concentration) for the modification of ceramic scaffolds could drastically reduce the compressive strength of the modified scaffolds in aqueous media, and the modification made a limited contribution to improving scaffold toughness. Using PCL/nanostructured silk the compressive strength and modulus of the modified scaffolds reached 0.42 MPa (compared with 0.07 MPa for BCP) and ∼25 MPa (compared with 5 MPa for BCP), respectively. The failure strain of the modified scaffold increased more than 6% compared with a BCP scaffold (failure strain of less than 1%), indicating a transformation from brittle to elastic behavior. The cytocompatibility of ECM-like composite scaffolds was investigated by studying the attachment, morphology, proliferation and bone-related gene expression of primary human bone-derived cells. Cells cultured on the developed scaffolds for 7 days had significant up-regulation of cell proliferation (∼1.6-fold higher, P < 0.001) and osteogenic gene expression levels (collagen type I, osteocalcin and bone sialoprotein) compared with the other groups tested.     

Hybrid coating of hydroxyapatite and collagen within poly(D,L-lactic-co-glycolic acid) scaffold

To improve the cell-affinity of biodegradable polymer scaffold, coating hydroxyapatite (HA) or collagen on the surface of polymer materials seemed to be a strategy to combine both advantages of them. The objective of this study was to develop a novel method to introduce HA and collagen inside polymer scaffold uniformly. HA and collagen suspension was mixed with paraffin microspheres, and molded to form a composite sample. After the sample was dried, HA/collagen composite was left among and on the surface of paraffin microspheres. Poly(D,L-lactic-co-glycolic acid) (PLGA) (50/50) solution was cast into the inter-space of the paraffin microspheres and dried. Afterwards, the paraffin was dissolved and removed, HA/collagen was transferred to the surface and even inside of the pore wall of PLGA scaffolds. Collagen fibers and HA particles which were inlaid inside the PLGA pore wall could help to enhance the coating strength between HA/Col coating and the pore wall surface of the PLGA scaffold. The scaffolds with HA/Col coating were expected to exhibit desirable properties in bone tissue engineering.     

三维环境下软骨细胞诱导滑膜间充质干细胞向软骨样细胞的分化

BACKGROUND: The co-culture of chondrocytes and synovial mesenchymal stem cells can induce the cartilage differentiation of synovial mesenchymal stem cells in vitro, but the cell differentiation induced by co-culture in vivo is rarely reported. OBJECTIVE: To investigate the chondrogenic differentiation of synovial mesenchymal stem cells co-cultured with chondrocytes on the chitosan/type I collagen composite scaffolds after being transplanted into the subcutaneous layer of Sprague-Dawley rats. METHODS: The synovial mesenchymal stem cells and chondrocytes harvested from the synovial membrane and articular cartilage of Sprague-Dawley rats were obtained by enzyme digestion method and cultured respectively. Passage 3 synovial mesenchymal stem cells and passage 2 chondrocytes, which were divided into four groups: group A (chondrocytes alone), group B (synovial mesenchymal stem cells alone), group C (ratio of synovial mesenchymal stem cells:chondrocytes=1:2) and group D (scaffold material without cells), were cultured on chitosan/type I collagen composite scaffolds and transplanted into the subcutaneous layer of rats followed by morphological observation and immunohistochemical staining at 4 and 8 weeks.   . RESULTS AND CONCLUSION: After 4 and 8 weeks, the discoid-like scaffold was visible. The immunohistochemical staining of type II collagen and the toluidine blue staining of aggrecan were significantly positive in groups A and C. These results show that the co-culture of synovial mesenchymal stem cells and chondrocytes on the scaffold in vivo can form cartilage-like tissues.