组织工程学技术修复关节软骨缺损研究进展

本文从种子细胞、生长因子和基质材料三方面对关节软骨组织工程学和分子组织工程学研究的进展进行了综述,指出了目前存在的问题和今后的研究方向。     

关节软骨缺损生物性修复与再生的研究现状

关节软骨绝大多数属透明软骨,具有低摩擦、富弹性和高渗透性的特性,对维持关节运动具有重要意义。但外伤或骨性关节炎等原因经常造成关节表面缺损，而关节软骨因本身缺乏再生能力而难以修复，所以最终常常导致关节面不完整，需要进行关节融合术或关节置换术，但其寿命有限，不太适宜于青壮年患者。随着医学的发展，关于关节软骨损伤后的再生和生物学愈合，在基础研究和临床探索方面都取得了丰硕的成果，本文着重介绍国内外有关关节软骨缺损的生物性修复及治疗方法在基础和临床研究方面的现状和进展？     

以羊膜为载体培养游离软骨细胞修复兔关节软骨缺损

目的：探讨膜结构为载体培养游离软骨细胞修复软骨缺损的可行性。方法：以兔羊膜为载体将体外培养的同种异体游离软骨细胞植于兔左侧股骨外踝软骨缺损区，分别于4、8、12周处死动物，整个膝关节被解剖，进行大体观察、组织学评价、电镜观察及SRY基因性别鉴定，并以兔体的右膝关节做为对照。结果：术后4、8、12周大体、组织学、电镜观察显示软骨缺损区新生了透明软骨，SRY基因性别鉴定证明新生的软骨来源于移植的同种异体软骨细胞；而对照组则仅见纤维组织样的修复组织。结论：以羊膜为载体进行同种异体软骨细胞移植能够修复关节软骨缺损。     

软骨组织工程学方法修复关节软骨缺损

背景：关节软骨缺损在临床上十分常见,随着分子生物学和组织工程学等学科的不断发展,为关节软骨缺损的修复提供了大量新的方法和思路,利用软骨组织工程学方法修复关节软骨缺损成为目前骨科领域研究的热点。 目的：总结并讨论目前软骨组织工程学方法修复关节软骨缺损的研究现状,综述应用软骨组织工程学方法修复关节软骨缺损的研究进展。 方法：由第一作者应用计算机检索中国期刊全文数据库(CNKI：2000/2010)和Medline(1990/2010)数据库中的相关文章,检索词分别为“关节软骨缺损,软骨组织工程”和“articular cartilage defects (ACD),cartilage tissue engineering”,语言分别设定为中文和英文。共检索得文章786篇,从中选取相关文章44篇,从软骨组织工程学方法修复关节软骨缺损过程中的种子细胞、支架材料和生物因子3个具体方面的研究进展进行归纳及总结。 结果与结论：支架、种子细胞和可调控细胞生长、增殖及分化的生物因子是软骨组织工程的3大要素。软骨组织工程方法主要包括利用体外培养、扩增后的种子细胞,将其种植于支架材料中,在相关调节因素的作用下形成组织工程化软骨,此方法已成为目前治疗关节软骨缺损的重要方法之一,并取得不错的疗效,但迄今为止尚未出现一种法被广泛认可的治疗方案。利用不同新型复合支架材料如修复治疗关节软骨缺损将成为今后研究的主要方向。     

关节软骨损伤临床治疗新进展

关节软骨(articular cartilage,AC)缺乏直接的血液供应、淋巴循环和神经支配,并具有低代谢的生理特点,当外伤或疾病而发生缺损时通常不能自行修复,持续发展会导致骨关节炎。随着全民健身运动普及和社会老龄化等因素,因各种原因导致软骨缺损的发病率逐年递增,寻找行之有效的临床治疗手段迫在眉睫。目前临床修复关节软骨损伤的方法很多,大体分为刺激关节     

组织工程支架材料修复关节软骨缺损

BACKGROUND: Articular cartilage repair has been a difficulty in the clinical setting, which is mainly treated with autologous or allogeneic osteochondral grafts, and cartilage periosteum or periosteum grafts. However, the limited source, secondary lesion and immunological rejection force some researchers to search for a novel treatment strategy, cartilage tissue engineering, that is of great significance for cartilage regeneration and repair. OBJECTIVE: To investigate the tissue-engineered scaffolds for the repair of articular cartilage defects. METHODS: The first author searched the PubMed and WanFang databases for the articles addressing tissue-engineered cartilage for articular cartilage defects published between 1991 and 2015 using the keywords “articular cartilage defect, scaffold, tissue engineered cartilage” in English and Chinese, respectively. The irrelative and repetitive literatures were excluded. RESULTS AND CONCLUSION: Finally 48 eligible literatures were enrolled based on the inclusion and exclusion criteria. Cartilage tissue engineering possesses the advantages of controllability, little damage to tissue itself, and biological repair of injured cartilage. Tissue-engineered scaffold material is a critical factor in tissue engineering construction; therefore, it should hold biodegradability and histocompatibility. The commonly used scaffold materials include natural macromolecule materials (collagen, silk fibroin and chitosan), and synthetic polymer materials (polylactic acid and tricalcium phosphate). It is necessary to prepare composite scaffolds with high bioactivity integrate advantages of each material. The tissue engineering is bound to be a hotspot in the field of articular cartilage repair.      

丹参关节腔注射与针刺修复膝关节全层软骨缺损

背景：研究表明软骨下钻孔可有效修复关节软骨缺损,应用针药结合能有效控制软骨损伤出现的症状。推测针药结合可与软骨下钻孔在修复软骨缺损中产生协同作用。 目的：观察针药结合联合软骨下钻孔对兔膝关节全层软骨缺损的修复效果。 方法：于50只新西兰兔双侧股骨髁关节面造成6 mm×8 mm全层软骨缺损,随机分为5组。模型组造模后不作任何处理;钻孔组只做钻孔处理;丹参组钻孔后1周向膝关节腔注射丹参注射液0.3 mL,1次/周,共5周;针刺组钻孔后1周针刺兔双侧足三里穴,1次/d,30 min/次,6次为1个疗程,共治疗5个疗程;针药组钻孔后1周同时行丹参关节腔注射与针刺治疗,方法和疗程同上。 结果与结论：组织学及透射电镜检查均显示,模型组主要为纤维组织,其余4组缺损修复组织均为类透明软骨、幼稚软骨和纤维软骨,且以针药组修复组织的数量和质量最佳。与模型组比较,其余4组的致密修复组织覆盖缺损的面积明显增大(P < 0.01),以针药组的疗效最优,丹参组、针刺组次之。结果提示丹参关节腔注射、电针足三里和针药结合均可促进软骨下钻孔修复关节软骨缺损,且针药结合优于单纯的丹参关节腔注射及电针足三里治疗。     

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.     

运动性关节软骨缺损支架材料的选择及其生物相容性

背景：骨软骨支架是用于承载细胞,供细胞黏附、生长、增殖、分化的载体。 目的：总结运动性关节软骨缺损支架材料的应用进展及其生物替代材料的生物相容性。 方法：以“关节软骨,生物材料,工程软骨,支架材料,生物相容性”为中文关键词,以“ tissue enginneering ,articular cartilage,scaffold material”为英文关键词,采用计算机检索维普数据库、PubMed数据库1993-01/2010-11相关文章。纳入与有关修复关节软骨损伤、生物材料、支架材料、生物相容性等相关的文章。以20篇文献为重点对运动性关节软骨缺损修复用的生物材料的生物相容性进行了讨论。 结果与结论：天然软骨支架材料因其具有细胞识别信号,故生物相容性好,细胞黏附率高,但力学性能较差。有些人工合成材料生物相容性不理想、亲水性差、对细胞吸附不足,人工合成高分子聚合物生物相容性良好。复合支架利用不同生物材料的优点克制材料的局限性制备理想的复合支架,其混合比例、混合技术还有待进一步研究。目前尚无一种材料完全满足组织工程的要,通过材料制备技术的改进或将几种不同材料的复合,材料的性能会不断的提高。     

组织工程修复关节软骨缺损的力学状态研究

背景：力学状态对软骨的正常生理有重要影响,若应力集中过大将造成人工软骨退变和原宿主软骨退化,影响治疗效果。目前的各种力学手段很难实现活体软骨力学状态测量,而有限元动态分析能有效地模拟修补后软骨的受力情况。 目的：通过有限元仿真研究组织工程修复膝关节软骨缺损后人工软骨和宿主软骨的力学状态。 方法：以人体膝关节软骨受滚压部分为研究对象,建立滚动运动下关节软骨的有限元模型。根据行走过程中股骨与胫骨间的滚压边界条件,对软骨在取不同弹性模量、不同压缩量、不同载荷速度及不同缺损大小的情况进行了滚压受力分析。 结果与结论：在滚压载荷下,植入人工软骨弹性模量和软骨压缩量的不同都使人工软骨和宿主软骨受到的Mises应力值变化,二者对修复缺损处软骨Mises应力分布的影响比较明显,是临床治疗软骨缺损和术后康复阶段值得注意的因素。模拟中使用的载荷速度和缺损大小对软骨应力值的影响不明显。当人工软骨弹性模量取某个值时,人工软骨和宿主软骨的Mises应力差别可以达到很小值,二者趋于吻合。应力差别还和个体宿主软骨的力学性能有关,据此,应针对不同病例选择最佳弹性模量的人工软骨植入。     

组织工程软骨与柚皮苷联合修复兔膝关节软骨缺损的组织学证实

背景：目前临床上虽有多种方法用于治疗软骨缺损,但没有从根本上解决关节软骨缺损修复问题。 目的：通过组织学研究进一步评价柚皮苷结合组织工程软骨修复兔关节软骨缺损的效果。 方法：取兔骨髓间充质干细胞体外增殖后,复合于改建后的脱细胞真皮基质载体上,制成组织工程软骨,植入到兔膝关节软骨缺损,并以柚皮苷汤灌胃,于 4,8周后分别对修复组织进行苏木精-伊红、Masson三色染色、甲苯胺蓝染色、Ⅱ型胶原染色、Ⅹ型胶原染色等组织学检查。 结果与结论：术后8周, 柚皮苷结合干细胞复合体组缺损处修复组织变成乳白色,半透明光滑组织,缺损修复组织与周围正常软骨已基本难区分,表面光滑。组织学检查发现修复缺损处基本为新生软骨填充。结果证实,柚皮苷结合组织工程软骨能提高家兔膝关节软骨缺损的修复质量。 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;组织工程全文链接：     

组织工程化软骨修复关节软骨缺损

背景：目前治疗软骨缺损的方法均有明显缺陷,组织工程软骨修复关节软骨缺损为微创治疗软骨缺损提供了新方法。 目的：总结组织工程软骨应用于修复关节软骨缺损的新进展。 方法：由第一作者用计算机检索万方数据库(2000/2010)和PubMed数据库(2000/2010),检索词分别为“软骨缺损,组织工程软骨”和“articular cartilage defects, tissue-engineered cartilage”,语言分别设定为中文和英文。从组织工程软骨及其新进展2方面进行总结,对组织工程软骨的发展及构建等方面进行介绍。共检索到666篇文献,按纳入和排除标准对文献进行筛选,共纳入23篇文章。 结果与结论：近年来随着细胞支架材料的不断发展和组织构建技术的日趋成熟,结合活性细胞和支架的组织工程软骨为微创修复关节软骨缺损提供了良好的治疗手段及方法,组织工程软骨修复软骨缺损是完全可行的,C-GP凝胶与种子细胞相容性好,可做为组织工程软骨的理想支架,但细胞支架的制备及选择仍然是组织工程软骨的热点和难点。     

组织工程化软骨细胞和骨髓间充质干细胞用于修复同种异体关节软骨缺损

背景：关节软骨几乎没有自身修复的能力,目前临床大多采用自体或异体软骨移植修复、软骨膜或骨膜移植修复、软骨细胞移植修复。由于自体软骨来源有限,异体软骨又存在慢性免疫排斥反应,最终可能导致预后不佳;软骨膜或骨膜移植修复的软骨易于退化,导致修复效果不佳。 目的：总结组织工程化软骨细胞、骨髓间充质干细胞及两者共培养对同种异体软骨缺损修复作用的研究现状。 方法：应用计算机检索PubMed 数据库及中国期刊网全文数据库1994-01/2012-01有关组织工程化软骨细胞和骨髓间充质干细胞用于修复同种异体关节软骨缺损方面的文章,英文检索词为“cartilage defect,allograft,chondrocyte,mesenchymal stem cells,bone marrow mesenchymal stem cells”,中文检索词为“软骨缺损,同种异体移植,软骨细胞,骨髓间充质干细胞”。排除重复性及非中英文语种研究,共保留35篇文献进行综述。 结果与结论：随着体外细胞培养方法的不断改进,现已能够把软骨细胞从坚韧的软骨中分离出来,并获得大量高纯度的软骨细胞并繁殖出新生软骨细胞。软骨细胞培养增殖能力低,传代培养容易引起老化和去分化;而成体骨髓中骨髓间充质干细胞含量少,随传代次数的增多成软骨潜能明显降低。骨髓间充质干细胞和软骨细胞共培养,两种细胞相互促进增殖和分化,作为种子细胞可减少软骨细胞增殖传代次数并节省软骨细胞数量,与组织工程支架材料复合能有效修复关节软骨缺损。     

Hydrogels for the repair of articular cartilage defects

The repair of articular cartilage defects remains a significant challenge in orthopedic medicine. Hydrogels, three-dimensional polymer networks swollen in water, offer a unique opportunity to generate a functional cartilage substitute. Hydrogels can exhibit similar mechanical, swelling, and lubricating behavior to articular cartilage, and promote the chondrogenic phenotype by encapsulated cells. Hydrogels have been prepared from naturally derived and synthetic polymers, as cell-free implants and as tissue engineering scaffolds, and with controlled degradation profiles and release of stimulatory growth factors. Using hydrogels, cartilage tissue has been engineered in vitro that has similar mechanical properties to native cartilage. This review summarizes the advancements that have been made in determining the potential of hydrogels to replace damaged cartilage or support new tissue formation as a function of specific design parameters, such as the type of polymer, degradation profile, mechanical properties and loading regimen, source of cells, cell-seeding density, controlled release of growth factors, and strategies to cause integration with surrounding tissue. Some key challenges for clinical translation remain, including limited information on the mechanical properties of hydrogel implants or engineered tissue that are necessary to restore joint function, and the lack of emphasis on the ability of an implant to integrate in a stable way with the surrounding tissue. Future studies should address the factors that affect these issues, while using clinically relevant cell sources and rigorous models of repair.     

Integrative repair of cartilage with articular and nonarticular chondrocytes

Articular chondrocytes can synthesize new cartilaginous matrix in vivo that forms functional bonds with native cartilage. Other sources of chondrocytes may have a similar ability to form new cartilage with healing capacity. This study evaluates the ability of various chondrocyte sources to produce new cartilaginous matrix in vivo and to form functional bonds with native cartilage. Disks of articular cartilage and articular, auricular, and costal chondrocytes were harvested from swine. Articular, auricular, or costal chondrocytes suspended in fibrin glue (experimental), or fibrin glue alone (control), were placed between disks of articular cartilage, forming trilayer constructs, and implanted subcutaneously into nude mice for 6 and 12 weeks. Specimens were evaluated for neocartilage production and integration into native cartilage with histological and biomechanical analysis. New matrix was formed in all experimental samples, consisting mostly of neocartilage integrating with the cartilage disks. Control samples developed fibrous tissue without evidence of neocartilage. Ultimate tensile strength values for experimental samples were significantly increased (p < 0.05) from 6 to 12 weeks, and at 12 weeks they were significantly greater (p < 0.05) than those of controls. We conclude that articular, auricular, and costal chondrocytes have a similar ability to produce new cartilaginous matrix in vivo that forms mechanically functional bonds with native cartilage.     

关节软骨损伤后体外培养软骨细胞的功能变化

背景：关节软骨损伤可以影响软骨细胞功能,诱发创伤性骨关节炎。 目的：观察关节软骨损伤后体外培养的软骨细胞功能的变化。 方法：通过酶消化法分离培养高能量、低能量撞击后和正常兔膝关节透明软骨细胞,观察创伤能量对软骨细胞生存能力的影响;检测软骨细胞合成蛋白多糖和Ⅱ型胶原能力,检测细胞中白细胞介素1β和核转录因子κB mRNA表达水平,检测细胞合成白细胞介素1β和基质金属蛋白酶1的表达。 结果与结论：高能量和低能量关节软骨损伤后,软骨细胞的存活率下降,原代细胞的贴壁细胞数量减少,贴壁时间延长,生长曲线下移,细胞甲苯胺蓝染色异染反应减弱,Ⅱ型胶原免疫组化染色强度减弱,软骨细胞中白细胞介素1β和核转录因子κB mRNA表达水平上升,细胞培养液中白细胞介素1β和基质金属蛋白酶1的质量浓度升高,其中高能量组效果更为显著(P < 0.05)。说明关节软骨损伤后软骨细胞的功能受到影响,受损程度与创伤强度及炎性细胞因子的表达相关。