腹壁修补材料的研究及应用进展

巨大腹壁缺损的修补是外科医生经常面临的难题,寻找理想的修补材料以修复缺损至关重要。用于修复腹壁缺损的材料一般分为生物性和非生物性两种。前者的力学性能较好,而后者的生物相容性更佳。目前公认尚没找到一种完全理想的腹壁替代材料.作者对常用生物材料和非生物材料的特性、应用、优缺点等做一综述。     

胶原蛋白作为医用生物材料的应用

胶原蛋白是一类具有活跃生物功能的细胞外间质成分,占人体总蛋白质总量的1/3。胶原蛋白是组织的支持物和填充物,也主动参与了细胞迁移、分化及增殖,并与创伤修复及胚胎发育有关。以胶原蛋白为原料制备的生物替代物(如人造皮肤等)必然大大优于非生物材料的同类产品。目前,大量不同性能及用途的胶原蛋白医用材料不断涌现,有些已结束实验室及动物实验阶段,大批量投入临床应用。     

肿瘤细胞三维培养支架、血管及细胞间的相互作用

背景：肿瘤组织工程通过构建综合的培养模型,充分模拟肿瘤在体内生长的微环境,可以较好地研究肿瘤发生发展的动力学及相关治疗策略。 目的：综述肿瘤工程技术中的肿瘤细胞三维培养。 方法：以“tumor engineering; 3D culture; biological materials; dynamic”为关键词,检索PubMed数据库1992年1月至2013年3月相关文献,纳入有关肿瘤工程、肿瘤细胞三维培养、生物支架材料及肿瘤微环境的相关文章。 结果与结论：三维培养因其可再现组织细胞的体内生长情况,已成为研究肿瘤耐药性、侵袭性和肿瘤微环境的重要平台,在许多领域表现出逐步取代平面培养技术的趋势,为肿瘤研究提供了一个非常接近于体内真实情况的研究平台。近年来,随着肿瘤工程学的发展,多种新型高分子聚合材料被应用于肿瘤细胞的三维立体培养,三维培养技术逐渐成为肿瘤生物学领域研究的热点,其利用各种方法及材料使细胞呈空间立体方式生长,形成类似体内生长环境的生物支撑或基质,建立细胞间及细胞与胞外基质间的相互联系,并形成特定的类似组织样的三维空间结构。生物材料就是种子细胞生长的土壤,在肿瘤工程中起着替代细胞外基质或组织、器官的基质的作用。而随着三维细胞培养技术在肿瘤研究中的广泛应用,其已成为肿瘤耐药、血管形成、细胞间相互作用、信号转导、干细胞等方面研究不可或缺的有力工具。     

羟基乙酸负载软骨细胞组织工程软骨修复喉软骨缺损

BACKGROUND: A three-dimensional biodegradable scaffold is important for tissue-engineered cartilage construction, and it that can provide conditions for cell attachment and proliferation. OBJECTIVE: To observe the treatment outcomes of glycolic acid loaded with chondrocytes in laryngeal cartilage repair. METHODS: Sixty New Zealand white rabbits were enrolled and randomly divided into control and experimental groups. Laryngeal cartilage defect models were established in each group, followed by implanted with glycolic acid loaded with chondrocytes and glycolic acid, respectively. Gross and histological observations were conducted at 4 and 8 weeks after implantation. RESULTS AND CONCLUSION: Gross observation showed that at 4 weeks after implantation, a deep red wound with an obvious boundary was seen in the control group; the dark red and smooth defect parallel to the surrounding tissue was found in the experimental group. Toluidine blue staining revealed that at 8 weeks after implantation, the laryngeal defect site showed no obvious inflammation and cartilage collapse, with numerous newly-formed chondrocytes in the experimental group; in contrast, mild inflammation and cartilage collapse were found in the defect region of the control group, and few newly-formed chondrocytes appeared. The positive areas of glycosaminoglycan and type II collagen in the experimental group were significantly larger than those in the control group at 4 and 8 weeks after implantation (P < 0.05). These results indicate that glycolic acid loaded with chondrocytes contributes to the repair of laryngeal cartilage defects.     

丝素蛋白/羟基磷灰石材料复合骨髓间充质干细胞构建组织工程化软骨

背景：随着组织工程的兴起,软骨损伤的修复可能性显著地提高,但单一的支架材料均不能符合理想支架,有一定的局限性。 目的：观察骨髓间充质干细胞复合丝素蛋白/羟基磷灰石构建组织工程化软骨的可行性。 方法：体外分离培养骨髓间充质干细胞,并定向诱导成软骨细胞,与丝素蛋白/羟基磷灰石复合培养,构建膝关节胫骨平台全层关节软骨缺损。54只大白兔单侧膝关节全层软骨缺损模型后随机抽签法分为3组,复合组植入细胞-丝素蛋白/羟基磷灰石复合物;材料组植入单纯丝素蛋白/羟基磷灰石,对照组不行任何植入。植入后8,12周CT检查及组织学检查观察软骨缺损修复情况。 结果与结论：植入后8周,复合组关节面不平整,关节间隙增大,形成新生类软骨细胞,基质丰富。材料组关节面塌陷,软骨细胞少量增殖。植入后12周,复合组关节面平整,关节间隙如常。大量软骨细胞出现,与周边软骨色泽一样,支架材料完全降解。材料组关节面不平整,软骨细胞不完全充填,支架材料部分降解。对照组未见修复。提示用骨髓间充质干细胞复合丝素蛋白/羟基磷灰石可形成透明软骨修复动物膝关节全层软骨缺损,显示了丝素蛋白/羟基磷灰石材料作为关节软骨组织工程支架材料的良好生物相容性。     

脱钙骨基质与骨髓间充质干细胞共培养关节腔内的成软骨活性

背景：将骨髓间充质干细胞附着到支架材料上再植入关节软骨缺损处,细胞不但不消失,而且可形成新的软骨。 目的：观察同种异体脱钙骨基质与骨髓间充质干细胞共培养在关节内的成软骨活性。 方法：在54只青紫蓝兔单侧膝关节制作关节软骨全层缺损模型,随机分组：实验组在缺损处植入自体骨髓间充质干细胞与同种异体脱钙骨基质复合物,对照组缺损处仅植入同种异体脱钙骨基质,空白对照组未植入任何物质。 结果与结论：植入后12周,实验组缺损处修复组织呈软骨样,表面光滑平坦,与周围软骨整合的软骨细胞更为成熟,修复组织与软骨下骨结合牢固;修复组织的细胞为透明软骨样细胞,柱状排列,Ⅱ型胶原染色阳性,与周围软骨及软骨下骨整合良好,且实验组组织学评分优于对照组和空白对照组 (P < 0.01)。对照组缺损处修复组织呈纤维样,与周围软骨未结合,空白对照组缺损区无修复组织,两组均无Ⅱ型胶原染色阳性表达。表明同种异体脱钙骨基质与骨髓间充质干细胞共培养后植入膝关节可形成软骨样组织,有效修复关节软骨缺损。     

骨形态发生蛋白复合物结合CAD/CAM技术原位修复喉甲状软骨缺损

背景：临床修复喉甲状软骨缺损时使用一定的支架材料。骨形态发生蛋白具有成骨诱导活性,与其他形式的载体联合起来进行复合应用可以更好的发挥出其诱导成骨作用。联合使用计算机辅助设计/计算机辅助制造技术能提高修复效果。 目的：进一步验证骨形态发生蛋白复合物结合计算机辅助设计/计算机辅助制造技术在原位修复喉甲状软骨缺损中的应用效果。 方法：纳入18只新西兰大白兔,对动物喉部进行三维扫描,建立兔甲状软骨三维数字模型。利用泡沫凝胶注模法制备羟基磷灰石支架,再制备骨形态发生蛋白复合物。18只动物随机分为观察组和对照组。两组兔行一侧甲状软骨切除后,观察组植入珊瑚羟基磷灰石支架;对照组填充明胶海绵。处理后 4,8,12周,进行大体观察和评分;处理后12周,对动物行喉镜检查,喉组织常规切片后进行苏木精-伊红染色,进行组织学观察。 结果与结论：处理后12周,对照组兔的喉腔出现明显的变窄现象,且存在轻微充血;观察组喉腔黏膜光滑,且通畅、宽敞,未出现肉芽生长情况。处理后4,8,12周,两组喉甲状软骨评分均呈现出不断下降的情况,且不同时间点,观察组评分均显著低于对照组(均P < 0.05)。处理后12周,对照组未出现成熟骨细胞和软骨细胞的生成;观察组则可见植入材料完全降解,存在大量多核样组织细胞增生和吞噬,手术区域软骨母细胞出现增生现象,并伴有成软骨区域形成情况,另外还以观察到软骨骨质形成以及软骨细胞增生现象。结果表明,利用骨形态发生蛋白复合物结合计算机辅助设计/计算机辅助制造技术进行原位喉甲状软骨缺损修复可以获得良好的效果,实验过程中所使用的复合物具有良好的骨诱导作用以及组织相容性。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程      

自体脂肪间充质干细胞复合人脐带Wharton胶支架修复兔膝关节软骨缺损****◇

背景：脐带Wharton胶富含透明质酸,糖胺多糖及胶原等,成分与天然软骨细胞外基质类似,因此由人脐带提取的Wharton胶很可能是一种较为理想的软骨组织工程支架材料。 目的：评价自体脂肪间充质干细胞复合人脐带Wharton胶支架修复兔膝关节软骨缺损的效果。 方法：将终浓度为1010 L -1、成软骨方向诱导后的兔自体脂肪间充质干细胞与人脐带Wharton胶支架复合,继续培养1周构建组织工程软骨,对兔膝关节全层软骨缺损进行修复(实验组),并与单纯支架修复的对照组及空白组进行比较。术后3个月对修复组织行大体观察、组织学检测、糖胺多糖、总胶原定量检测及生物力学测定。 结果与结论：实验组的缺损多为透明软骨修复,对照组以纤维组织修复为主,空白组无明显组织修复。提示脂肪间充质干细胞作为软骨组织工程种子细胞具有可行性;实验构建的组织工程软骨能有效的修复关节软骨缺损,人脐带Wharton胶可作为软骨组织工程良好的支架材料。     

羟基丁酸-羟基辛酸共聚体/胶原骨软骨一体化支架修复膝关节软骨损伤

BACKGROUND: Due to the complex physiological characteristics of the osteochondral tissue, the clinical repair of knee cartilage injury often has dissatisfied outcomes. Tissue engineering methods and tools provide a new idea for osteochondral repair. OBJECTIVE: To observe the effect of poly(hydroxybutyrate-co-hydroxyoctanoate/collagen) osteochondral tissue-engineered scaffold on the repair of articular cartilage injury in a rabbit. METHODS: The poly(hydroxybutyrate-co-hydroxyoctanoate/collagen) osteochondral tissue-engineered scaffold was prepared by solvent casting/particle leaching method. Then, seed cells were isolated and cultured on the scaffold. Twenty-four healthy New Zealand white rabbits, 4 weeks of age, were used for the study. Under balanced anesthesia, an articular cartilage defect (4.5 mm in diameter, 5 mm in depth) was created on the rabbit’s femoral condyle using a bone drill. After modeling, rabbits were randomized into three groups and given direct suture in blank group, pure scaffold implantation in control group and implantation of the scaffold-cell complex in experimental group. Femoral condyle of each rabbit was taken out for gross and histological observations at 8, 20 weeks after surgery. RESULTS AND CONCLUSION: At 8 weeks after surgery, transparent film-covered defects and small/irregular cells were found in the experimental group; the defects were filled with fibrous tissues in the control group; while there was no repair in the blank group. Until the 20th week, the defects were covered with hyaline cartilage-like tissues, accompanied by regular cell arrangement in the experimental group; in the control group, the defects were covered with white membranous tissues, and many chondrocytes were found at the basement and edge; in the blank group, some newborn tissues were visible at the defect region. These findings suggest that the poly (hydroxybutyrate-co- hydroxyoctanoate/collagen) osteochondral tissue-engineered scaffold carrying seed cells contributes to articular cartilage repair.     

同种异体和异种来源骨髓间充质干细胞诱导成软骨修复喉软骨缺损

BACKGROUND: Because chondrocytes have no regeneration ability, to select suitable seed cells is the primary problem to repair cartilage defects. OBJECTIVE: To investigate the effect of allogeneic versus heterologous bone marrow mesenchymal stem cells (BMSCs) in repairing laryngeal cartilage defects after chondrogenic induction. METHODS: BMSCs from human and rabbits were isolated and cultured. Passage 3 cells were cultured in chondrogenic induction medium containing transforming transforming growth factor beta 1 and bone morphogenetic protein, and then were dropped onto a poly(lactic-co-glycolic acid) (PLGA) scaffold. Thirty New Zealand rabbits were randomly assigned into three groups: blank control group, human BMSCs group, rabbit BMSCs group. Animal models of laryngeal cartilage defects were made in the three groups. After modeling, saline-soaked PLGA scaffold, PLAG scaffold with human BMSCs or with rabbit BMSCs were implanted respectively into the rabbits in the normal blank, human BMSCs and rabbit BMSCs groups. The expression of type II collagen in the larynx and its surrounding tissues was detected by immunohistochemistry at 4 and 8 weeks postoperatively. RESULTS AND CONCLUSION: The animals in each group breathed normally with no presence of wheezing, and their eating and activity were good. Moreover, there was no purulency or infection in the three groups. At 4 and 8 weeks after operation, the positive rates of type II collagen in the two BMSCs groups were significantly higher than that in the blank control group (P < 0.05). There was no significant difference between two BMSCs groups (P > 0.05). These results show that both allogeneic and heterologous BMSCs have good therapeutic effects on the repair of laryngeal cartilage defects in rabbits.      

聚羟基烷酸酯聚合物负载软骨细胞修复同种异体喉软骨缺损

背景：软骨组织工程基础研究相当深入,但在耳鼻咽喉科实际应用研究颇少,探索组织工程技术简便实用的喉软骨修复方法是值得研究的课题。 目的：比较多孔海绵状聚羟基丁酸酯与聚羟基己酸酯共聚物生物材料负载软骨细胞体外培养形成的初期组织工程软骨组织与体内植入一定时期形成的较成熟组织工程软骨组织修复同种异体甲状软骨缺损的效果。 方法：收集体外培养第3代乳兔(3 d龄)软骨细胞,以多孔海绵状聚羟基丁酸酯与聚羟基己酸酯共聚物生物材料为细胞外基质,采用组织工程技术制备细胞-材料复合物,共同体外培养形成初级组织工程软骨组织后直接应用于成兔甲状软骨缺损的修复(实验组A,n=5)或将初级组织工程软骨组织体内植入一定时期形成较成熟组织工程软骨再应用于甲状软骨缺损的修复(实验组B,n=5)。设单纯聚羟基丁酸酯与聚羟基己酸酯共聚物材料修复组(对照A组,n=4)和单纯软骨细胞修复组(对照B组,n=4)作为对照。分别于术后4周(实验B组)和8周(实验A组、对照A组、对照B组)取材,对甲状软骨缺损修复效果进行大体和组织学评价。 结果与结论：两者大体支架形态基本一致,修复区与原有软骨均相续平坦,无凹陷及缺损。但实验A组存在界面无细胞区,修复区基质分泌不丰富;实验B组界面区有细胞生长,基质分泌良好。两者炎细胞浸润均不明显。对照组修复区凹陷,呈暗红色软组织充填,组织学及特殊染色检查未发现软骨样结构及其分泌的基质成分。结果表明在有免疫力的动物体内,初级组织工程软骨组织直接应用与体内植入后再应用均能有效修复同种异体甲状软骨缺损,无明显免疫反应;相同时期内,应用较成熟组织工程软骨组织修复效果优于应用初级组织工程软骨组织。然而,直接应用初级组织工程软骨组织可节省时间、成本、工作量及操作环节,避免二次皮下手术的痛苦,是比较实用的方法之一。     

The use of poly(lactic-co-glycolic acid) microspheres as injectable cell carriers for cartilage regeneration in rabbit knees

The use of injectable scaffolding materials for in vivo tissue regeneration has raised great interest because it allows cell implantation through minimally invasive surgical procedures. Previously, we showed that poly(lactic-co-glycolic acid) (PLGA) microspheres can be used as an injectable scaffold to engineer cartilage in the subcutaneous space of athymic mice. The purpose of this study was to determine whether PLGA microspheres can be used as an injectable scaffold to regenerate hyaline cartilage in the osteochondral defects of rabbit knees. A full-thickness wound to the patellar groove of the articular cartilage was made in the knees of rabbits. Rabbit chondrocytes were mixed with PLGA microspheres and injected immediately into these osteochondral wounds. Both chondrocyte transplantations without PLGA microspheres and culture medium injections without chondrocytes served as controls. Sixteen weeks after implantation, chondrocytes implanted using the PLGA microspheres formed white cartilaginous tissues. Histological scores indicating the extent of the cartilaginous tissue repair and the absence of degenerative changes were significantly higher in the experimental group than in the control groups (P < 0.05). Histological analysis by a hematoxylin and eosin stain of the group transplanted with microspheres showed thicker and better-formed cartilage compared to the control groups. Alcian blue staining and Masson's trichrome staining indicated a higher content of the major extracellular matrices of cartilage, sulfated glycosaminoglycans and collagen in the group transplanted with microspheres than in the control groups. In addition, immunohistochemical analysis showed a higher content of collagen type II, the major collagen type in cartilage, in the microsphere transplanted group compared to the control groups. In the group transplanted without microspheres, the wounds were repaired with fibro-cartilaginous tissues. This study demonstrates the feasibility of using PLGA microspheres as an injectable scaffold for cartilage regeneration in a rabbit model of osteochondral wound repair.     

The use of poly(lactic-co-glycolic acid) microspheres as injectable cell carriers for cartilage regeneration in rabbit knees

The use of injectable scaffolding materials for in vivo tissue regeneration has raised great interest because it allows cell implantation through minimally invasive surgical procedures. Previously, we showed that poly(lactic-co-glycolic acid) (PLGA) microspheres can be used as an injectable scaffold to engineer cartilage in the subcutaneous space of athymic mice. The purpose of this study was to determine whether PLGA microspheres can be used as an injectable scaffold to regenerate hyaline cartilage in the osteochondral defects of rabbit knees. A full-thickness wound to the patellar groove of the articular cartilage was made in the knees of rabbits. Rabbit chondrocytes were mixed with PLGA microspheres and injected immediately into these osteochondral wounds. Both chondrocyte transplantations without PLGA microspheres and culture medium injections without chondrocytes served as controls. Sixteen weeks after implantation, chondrocytes implanted using the PLGA microspheres formed white cartilaginous tissues. Histological scores indicating the extent of the cartilaginous tissue repair and the absence of degenerative changes were significantly higher in the experimental group than in the control groups (P < 0.05). Histological analysis by a hematoxylin and eosin stain of the group transplanted with microspheres showed thicker and better-formed cartilage compared to the control groups. Alcian blue staining and Masson's trichrome staining indicated a higher content of the major extracellular matrices of cartilage, sulfated glycosaminoglycans and collagen in the group transplanted with microspheres than in the control groups. In addition, immunohistochemical analysis showed a higher content of collagen type II, the major collagen type in cartilage, in the microsphere transplanted group compared to the control groups. In the group transplanted without microspheres, the wounds were repaired with fibro-cartilaginous tissues. This study demonstrates the feasibility of using PLGA microspheres as an injectable scaffold for cartilage regeneration in a rabbit model of osteochondral wound repair.