Growth of chondrocytes on different biomaterials

Summary Chondrocytes can be cultured on different three-dimensional culture systems suitable for transplantation to enhance the repair of localized cartilage defects. Articular cartilage chondrocytes from adult rabbit knees and from bovine calf metacarpophalangeal joints were isolated by enzymatic digestion and cultured in a monolayer system to amplify cell count. After amplification the cells were seeded on different biocompatible materials. We investigated two types of bioresorbable polymer fleece matrices (a composite fleece of polydioxanon and polyglactin and a resorbable poly-L-lactic acid fleece) and lyophilized dura as a biological carrier. On all three types of transport media the phenotypic and morphological appearance of cultured chondrocytes could be observed. The production of glycosaminoglycans was revealed by Alcian blue staining and immunohistochemical detection of Chondroitin-4 and 6-sulfate in the created constructs. The material properties of the carriers allow for transplantation of the artificial cartilage-like products into full thickness articular cartilage defects and could therefore improve the minor intrinsic healing capacity of cartilage tissue. Bioartificial cartilage may become a future perspective in the treatment options of orthopaedic and plastic surgery.      

组织工程化软骨修复兔膝关节软骨缺损的实验研究

目的 评估同种异体组织工程软骨修复兔膝关节全层软骨缺损的有效性。方法分离收集成年新西兰大白兔软骨细胞进行体外培养。建立双侧兔膝关节软骨缺损模型，用去端肽胶原（atelocollagen）凝胶与所培养的异体兔关节软骨细胞共同植入兔膝关节软骨缺损处，并设对照组。分别于手术后4周、8周观察大体标本以及组织学修复结果，并进行Wakitan的评分，评估此方法的有效性。结果大体观察结果表明，与对照组相比，实验组缺损处由软骨组织修复而对照组缺损处由纤维样组织填充。组织学观察可以见到实验组关节软骨缺损处有密集的软骨细胞而对照组关节缺损处只有纤维细胞无软骨细胞。结论通过短期观察表明以同种异体软骨细胞-去端肽胶原复合物修复全层软骨缺损的方法是有效可行的，为其进一步临床应用提供了参考。     

同种异体软骨细胞移植术后关节软骨蛋白多糖的测定

目的 应用Pluronic F-127负载同种异体软骨细胞移植修复兔全厚关节软骨损伤,对于新生的修复组织进行基质蛋白多糖含量测定,以探讨此方法修复全厚关节软骨损伤的可行性.方法 取3个月龄新西兰大白兔关节软骨细胞体外培养扩增,与20%Plurortic F-127凝胶混合.选27只健康同种成年大白兔,人为造成双侧膝关节软骨缺损.实验组软骨缺损处植入培养的软骨细胞/Pluronic F-127混合物,对照组缺损处单纯注入Pluronic F-127凝胶和空白对照.然后,对修复组织进行大体观察及蛋白多糖含量测定.结果 移植的软骨细胞-载体复合物中的软骨细胞能良好地生长,12周时再生组织与周围正常软骨组织外观相似,界限模糊.实验组与对照组各时期蛋白多糖含量均有非常显著性差异,实验组不同时期的蛋白多糖含量之间均有显著性差异,实验组12周时蛋白多糖含量与正常软骨组织无显著性差异.结论 Pluronic F-127负载同种异体软骨细胞移植是治疗关节软骨缺损的有效方法.     

Allograft of cultured chondrocytes into articular cartilage defects in rabbits--experimental study of the repair of articular cartilage injuries

Articular cartilage defects were created by dill holes, 2 mm wide and 3 mm deep, through the articular cartilage into the subchondral bone in the patellar groove of the femur in mature rabbits. The defects received graft of cultured chondrocytes and the matrix obtained from the primary culture of chondrocytes isolated from the articular cartilage or auricular cartilage in immature rabbits. The isolated cells were cultured for 10 to 14 days. For graft, the cultured chondrocytes together with the matrix were detached from the culture chamber using rubber policemen and centrifuged. The repair of the grafted defects or defects without graft (control) was histologically studied 2 to 12 weeks after operation. The defects without the graft were progressively filled with fibrous tissue containing spindle shaped cells, fibers perpendicular to the surface, and matrix showing weak metachromasia with toluidin blue at 8 weeks. The defects received articular cartilage cell graft were occupied by new cartilage tissue consisting colonylike crumps of chondrocytes 2 weeks after operation. The crumps showed strong metachromasia with toluidin blue and strong stainability for safranin-O. By 4-8 weeks, the defects were filled with homogeneous cartilage. At 12 weeks, arrangement of the chondrocytes of the superficial layer of the new cartilage became columnar as seen in the normal articular cartilage. The defects received elastic cartilage cell graft were filled by reformed cartilage with chondrocytes surrounded by elastic fibers 2-12 weeks after operation. The results indicate that allograft of cultured chondrocytes with matrix into the articular cartilage defects accerated the repair process of the defects by formation of the new cartilage derived from the grafted chondrocytes.     

Restoration of arthritic cartilage defects using autologous chondrocytes transplantation is superior to cartilage-paste graft in rabbits

This study compared the articular cartilage repair potential of cultured chondrocytes transplantation with bone-cartilage paste-graft in the resurfacing of full-thickness defects without breaching of the subchondral bone plate in rabbit knees. A 5 x 5-mm articular cartilage defect was created in the patellar groove of the femur. Three months following creation, the defect was filled with cultured autologous chondrocytes (group 1) or bone-cartilage paste (group 2). A control group of untreated defects was followed for 1 year. The reparative tissue was analyzed macroscopically, histologically, and by immunohistochemistry 3-12 months post-transplantation. The surfaces of the reparative tissue in group 1 were smooth, and the defects were filled with reparative tissue that resembled hyaline cartilage. The composition of the repair tissue more closely resembled cartilage, as demonstrated by cartilage-specific stains. In contrast, the reparative tissue in group 2 was fibrous and exhibited markers of mesenchymal stem cells and bone formation. Transplantation of cultured chondrocytes into a full-thickness defect in the rabbit generates a biologic substitute tissue that resembles native articular cartilage with living cells capable of synthesizing the surrounding cartilage matrix. In contrast, analysis of the healing response to the paste-graft technique failed to show cartilage-like characteristics. This information may be clinically applicable to direct the use of these treatments in chondral injuries.     

Chondrocyte transplantation in PGLA/polydioxanone fleece

The transplantation of chondrogenic cells in a supportive carrier structure proved to be a promising alternative for the treatment of cartilage defects. In the study presented we focused on the transplantation of allogeneic chondrocytes in a biodegradable polymer scaffold (PGLA/Polydioxanon) in articular cartilage defects in a rabbit defect model. Isolated allogeneic chondrocytes embedded in a PGLA polymer scaffold were transplanted into osteochondrogenic defects of the patellar groove and compared with empty defects and transplants of polymer scaffolds without cells. The histological and histochemical analysis was performed after 4 and 12 weeks. The transplant integration and the architecture of the newly formed cartilage were evaluated with a semiquantitative score. After 4 weeks the development of a hyaline-like cartilage tissue of the cell-polymer-transplants was observed, after 12 weeks the defects were nearly completely filled with hyaline-like cartilage. The biodegradation of the polymer construct did not affect the histological structure of the transplant area. Defects of the groups with empty defect and polymer transplants without cells revealed no or insufficient healing indices. The study demonstrated that biodegradable polymers served as suitable carriers for the chondrocyte transplantation, which is due to the in-vitro establishment of a semi-solid cartilage transplant and the resulting effective transplant fixation into the defect. In-vivo the polymer cell transplants seem to provide a supportive microenvironment for the development of hyaline cartilage. The controlled release of morphogenic factors or bioactive molecules and the use of pluripotent mesenchymal progenitor cells opens new perspectives for the optimization of cartilage repair procedures.     

胶原/羟基磷灰石支架负载软骨细胞修复兔膝关节骨软骨缺损

目的 探讨胶原复合梯度羟基磷灰石(Col/HA)双相支架负载软骨细胞修复兔膝关节骨软骨缺损的可行性及疗效.方法 构建Col/HA双相支架,将软骨细胞种植于支架培养1周,再将软骨细胞-支架复合体移植修复兔膝关节股骨髁的骨软骨缺损,并对骨软骨缺损的修复进行检测.结果 光镜及扫描电镜观察显示软骨细胞在Col/HA支架中贴附良好,表型维持稳定,分泌胞外基质.大体观察和组织学检测显示,植入体内16周后实验组软骨层呈透明软骨样修复,软骨下骨缺损有新骨构建;对照组骨软骨缺损修复不良,组织学检测以纤维性组织或纤维软骨组织形成.Wakitani评分显示实验组修复组织优于对照组,差异有统计学意义(P＜0.05).结论 双相Col/HA复合支架可作为骨软骨组织工程支架,负载软骨细胞可修复兔膝关节骨软骨缺损,重建关节软骨的结构和功能.     

Rabbit articular cartilage defects treated by allogenic chondrocyte transplantation

Articular cartilage defects have a poor capacity for repair. Most of the current treatment options result in the formation of fibro-cartilage, which is functionally inferior to normal hyaline articular cartilage. We studied the effectiveness of allogenic chondrocyte transplantation for focal articular cartilage defects in rabbits. Chondrocytes were cultured in vitro from cartilage harvested from the knee joints of a New Zealand White rabbit. A 3 mm defect was created in the articular cartilage of both knees in other rabbits. The cultured allogenic chondrocytes were transplanted into the defect in the right knees and closed with a periosteal flap, while the defects in the left knees served as controls and were closed with a periosteal flap alone, without chondrocytes. Healing of the defects was assessed at 12 weeks by histological studies. Allogenic chondrocyte transplantation significantly increased the amount of newly formed repair tissue (P=0.04) compared with that found in the control knees. The histological quality score of the repair tissue was significantly better (P=0.05), with more hyaline characteristics in the knees treated with allogenic chondrocytes than in the control knees. Articular cartilage defects treated with allogenic chondrocyte transplantation result in better repair tissue formation with hyaline characteristics than those in control knees.     

Repairing articular cartilage defects with tissueengineering cartilage in rabbits

Objective: To investigate the effect of cancellous bone matrix gelatin ( BMG ) engineered with allogeneic chondrocytes in repairing articular cartilage defects in rabbits. Methods: Chondrocytes were seeded onto three-dimensional cancellous BMG and cultured in vitro for 12 days to prepare BMG-chondrocyte complexes. Under anesthesia with 2.5% pentobarbital sodium (1ml/kg body weight), articular cartilage defects were made on the right knee joints of 38 healthy New Zealand white rabbits (regardless of sex, aged 4-5 months and weighing 2. 5-3 kg) and the defects were then treated with 2. 5% trypsin. Then BMG-chondrocyte complex ( Group A, n = 18 ), BMG (Group B, n = 10), and nothing (Group C, n = 10) were implanted into the cartilage defects, respectively. The repairing effects were assessed by macroscopic, histologic, transmission electron microscopic ( TEM ) observation, immunohistochemical examination and in situ hybridization detection, respectively, at 2, 4, 8, 12 and 24 weeks after operation. Results: Cancellous BMG was degraded within 8 weeks after operation. In Group A, lymphocyte infiltration was observed around the graft. At 24 weeks after operation, the cartilage defects were repaired by cartilage tissues and the articular cartilage and subchondral bone were soundly healed. Proteoglycan and type II collagen were detected in the matrix of the repaired tissues by Safranin-O staining and immunohistochemical staining, respectively. In situ hybridization proved gene expression of type II collagen in the cytoplasm of chondrocytes in the repaired tissues. TEM observation showed that chondrocytes and cartilage matrix in repaired tissues were almost same as those in the normal articular cartilage. In Group B, the defects were repaired by cartilage-fibrous tissues. In Group C, the defects were repaired only by fibrous tissues. Conclusions: Cancellous BMG can be regarded as the natural cell scaffolds for cartilage tissue engineering. Articular cartilage defects can be repaired by cancellous BMG engineered with allogeneic chondrocytes. The nature of repaired tissues is closest to the normal cartilage. Local administration of trypsin can promote the adherence of repaired tissues to host tissues. Transplantation of allogeneic chondrocytes has immunogenicity, but the immune reaction is weak.     

骨髓基质细胞源性软骨细胞修复兔全层关节软骨缺损

目的观察体外诱导骨髓基质细胞(MSCs)源性软骨细胞在兔股骨滑车关节面全层软骨缺损修复中的作用. 方法高密度传代培养第3代诱导MSCs分化为软骨细胞,以酸溶性Ⅰ型胶原为载体,两者混合后形成凝胶样植入物(细胞浓度为5×106/ml).于36只新西兰大耳白兔一侧股骨滑车关节面造成3 mm×5 mm全层关节软骨缺损,凝胶样植入为实验侧;另一侧分别为单纯胶原植入组(18个膝关节)和空白对照组(18个膝关节).术后4、8、12、24、32和48周取材观察缺损修复情况及新生组织的类型.参照Pineda标准对新生组织评分. 结果实验侧术后4周,植入细胞类似软骨细胞,周围有异染基质,形成透明软骨样组织;8周,深层有软骨下骨形成,软骨细胞层较正常关节软骨厚;12周,新生软骨厚度减小,与正常软骨相近,细胞呈柱状排列,结构与正常关节软骨相似,软骨下骨形成,潮线恢复;24周,新生软骨厚度较正常薄,约占55%,表面平整,潮线附近仍有肥大的软骨细胞;32周,潮线附近无肥大软骨细胞;48周,组织结构与32周时基本相同,为类透明软骨.Pineda评分24、32和48周间无差异,与4周比较有统计学意义(P＜0.05).实验组2～48周期间关节功能良好.单纯胶原组与空白对照组缺损无修复,48周时软骨下骨外露,关节退变;关节功能逐渐减退,动度受限. 结论 MSCs源性软骨细胞移植体内可形成透明样软骨组织,24周后新生软骨特性稳定,48周时为透明样软骨,能维持良好的关节功能.     

PLGA和胶原海绵复合BMP修复兔关节软骨缺损的对比研究

目的以PLGA和胶原海绵为载体，分别复合rhBMP-2，比较两者修复兔关节软骨缺损的效果。方法将PLGA和胶原海绵制成直径4mm、厚3mm的圆柱形，分别与0．5mgrhBMP-2复合。2月龄新西兰兔24只，雌雄不拘，体重1．8～2．3kg，平均2．0kg。于24只兔双侧股骨髁部制造直径4mm的缺损，其中18只动物右侧缺损处植入PLGA／rhBMP-2复合物（实验1组），左侧缺损处植入胶原海绵／rhBMP-2复合物（实验2组）；余6只动物（12个膝关节）缺损不作任何处理，作为空白对照组。术后4、12和24周取材，行大体及组织学观察，并根据关节软骨半定量评分标准进行组织学评分。结果大体及组织学观察：4周，实验1组缺损内被半透明组织填充；实验2组缺损未被新生组织填满，两组形成的软骨组织与周围界限明显，软骨细胞分布较均一，排列无方向性；对照组中形成少量纤维组织。12周，实验1组缺损内完全充填白色半透明新生软骨组织，与正常软骨界限不清；实验2组缺损内形成白色半透明软骨组织，与周围正常软骨界限仍可辨认；两组新生软骨厚度逐渐接近正常软骨厚度，表面细胞平行排列，深层细胞排列较紊乱，基质异染，软骨下骨及潮线恢复，与正常软骨连接良好；对照组缺损边缘及底部形成少量软骨细胞，分布不均匀，底部纤维组织不连续。24周，实验1组缺损内形成半透明新生软骨组织，与正常软骨界限消失；实验2组形成的新生软骨组织颜色、质地与12周接近；两组新生软骨与正常软骨厚度相近，表面平整，细胞排列均一，但较紊乱，基质异染，软骨下骨及潮线恢复，与正常软骨连接良好；对照组缺损底部形成一层纤维组织，不连续。组织学评分：实验1组、实验2组与对照组在各时间点比较，差异均有统计学意义（P〈0．01）；两个实验组12、24周与4周比较差异有统计学意义（P〈0．01），但12周和24周比较差异无统计学意义（P〉0．05）；同一时间点两实验组间比较差异无统计学意义（P〉0．05）。结论PLGA和胶原海绵与rhBMP-2复合均可修复关节软骨缺损。     

Repair of articular cartilage defects with cultured chondrocytes in Atelocollagen gel

We attempted to repair full-thickness articular cartilage defects in rabbit knee joints with allogeneic cultured chondrocytes embedded in Atelocollagen gel. An articular cartilage defect was created on the patellar groove of the femur. The defect was filled with chondrocytes cultured in the collagen gel and covered with periosteal flap (G group). In three other experimental groups, the same defects were transplanted with chondrocytes in monolayer culture with periosteal flap (M group), periosteal graft only (P group), or left empty (E group). At 4, 12, and 24 weeks after operation, the reparative tissue was analyzed macroscopically and histologically. At 4 weeks after operation, the surfaces of the reparative tissue were smooth, and the defects were filled with reparative tissues that resembled hyaline cartilage in all four groups. However, the reparative tissues degenerated gradually with time in the M, P, and E groups. In contrast, in the G group, the reparative tissue retained its thickness, and there was a steady integration of the grafted tissue into the adjacent normal cartilage at 24 weeks after operation. The results suggest that transplantation of allogeneic chondrocytes cultured in Atelocollagen gel is effective in repairing an articular cartilage defect.     

Repair of rabbit articular surfaces with allografts of chondrocytes embedded in collagen gels

In an attempt to restore articular cartilage, allogeneic articular chondrocytes embedded in collagen gels were transplanted onto full-thickness defects in rabbit articular cartilage. Within 24 weeks after the transplantation, the defects were filled with hyaline cartilage, specifically synthesizing type II collagen. These chondrocytes were autoradiographically proven to be originated from the originally transplanted chondrocytes. As histologically assessed, success rate was about 80%, a marked improvement over the results (40% success rate) in previous studies reporting chondrocyte transplantation without collagen gels. On the other hand, the defects without chondrocyte transplantation healed with fibrocartilaginous tissue more than 24 weeks after treatment. Immunological enhancement induced by transplanted allogeneic chondrocytes or collagen was not significant for eight weeks after treatment, so far as shown by both direct and indirect blastformation reactions. Thus, allogeneic transplantation of isolated chondrocytes embedded in collagen gels appears to be one of the most promising methods for the restoration of articular cartilage.     

A new biotechnology for articular cartilage repair: subchondral implantation of a composite of interconnected porous hydroxyapatite, synthetic polymer (PLA-PEG), and bone morphogenetic protein-2 (rhBMP-2)

OBJECTIVE: Articular cartilage repair remains a major obstacle in tissue engineering. We recently developed a novel tool for articular cartilage repair, consisting of a triple composite of an interconnected porous hydroxyapatite (IP-CHA), recombinant human bone morphogenetic protein-2 (rhBMP-2), and a synthetic biodegradable polymer [poly-d,l-lactic acid/polyethylene glycol (PLA-PEG)] as a carrier for rhBMP-2. In the present study, we evaluated the capacity of the triple composite to induce the regeneration of articular cartilage. METHODS: Full-thickness cartilage defects were created in the trochlear groove of 52 New Zealand White rabbits. Sixteen defects were filled with the bone morphogenetic protein (BMP)/PLA-PEG/IP-CHA composite (group I), 12 with PLA-PEG/IP-CHA (group II), 12 with IP-CHA alone (group III), and 12 were left empty (group IV). The animals were killed 1, 3, and 6 weeks after surgery, and the gross appearance of the defect sites was assessed. The harvested tissues were examined radiographically and histologically. RESULTS: One week after implantation with the BMP/PLA-PEG/IP-CHA composite (group I), vigorous repair had occurred in the subchondral defect. It contained an agglomeration of mesenchymal cells which had migrated from the surrounding bone marrow either directly, or indirectly via the interconnecting pores of the IP-CHA scaffold. At 6 weeks, these defects were completely repaired. The regenerated cartilage manifested a hyaline-like appearance, with a mature matrix and a columnar organization of chondrocytes. CONCLUSIONS: The triple composite of rhBMP-2, PLA-PEG, and IP-CHA promotes the repair of full-thickness articular cartilage defects within as short a period as 3 weeks in the rabbit model. Hence, this novel cell-free implant biotechnology could mark a new development in the field of articular cartilage repair.     

Repair of rabbit articular surfaces with allograft chondrocytes embedded in collagen gel

In an attempt to repair articular cartilage, allograft articular chondrocytes embedded in collagen gel, were transplanted into full-thickness defects in rabbit articular cartilage. Twenty-four weeks after the transplantation, the defects were filled with hyaline cartilage, specifically synthesising Type II collagen. These chondrocytes were autoradiographically proven to have originated from the transplanted grafts. Assessed histologically the success rate was about 80%, a marked improvement over the results reported in previous studies on chondrocyte transplantation without collagen gel. By contrast, the defects without chondrocyte transplantation healed with fibrocartilage. Immunological enhancement induced by transplanted allogenic chondrocytes or collagen was not significant at eight weeks after treatment, so far as shown by both direct and indirect blastformation reactions. Thus, allogenic transplantation of isolated chondrocytes embedded in collagen gel appears to be one of the most promising methods for the restoration of articular cartilage.     

Synthesis of articular cartilage-like tissue in vitro

Defects in mature articular cartilage do not heal without residues, and therefore they remain a challenging problem in orthopaedic surgery. Modern tissue culture techniques facilitate the synthesis of cartilage-like tissue. A requirement of retaining the phenotypic characteristics of chondrocytes in vitro is the use of three-dimensional culture techniques. Articular chondrocytes of adult rabbits were isolated and cultured on different transplantable media for several weeks. A resorbable fleece, a non-absorbable net and lyophilized dura were used. Viability was tested by immunohistochemical techniques. Deposition of extracellular matrix could be observed by electron microscopy. The phenotypical and morphological appearance of cultured chondrocytes was preserved on the resorbable polymer fleece and the lyophilized dura. Cells cultured on the non-absorbable net had a more fibroblastic appearance. The resorbable fleece is apparently most suitable in terms of viability of the cultured chondrocytes and biocompatibility. The cultured three-dimensional artificial cartilage constructs reveal a future possibility for autologous cartilage transplantation into mature cartilage defects. Received: 21 March 1997     

增强型绿色荧光蛋白表达在同种异体组织工程软骨构建中的示踪作用

目的 观察重组逆转录病毒载体转染的软骨细胞能否形成同种异体组织工程化软骨并示踪构建过程，方法 利用重组逆转录病毒载体转染的软骨细胞和组织工程材料pluronicF-127复合物构建兔同种异体组织工程软骨，荧光显微镜和激光共聚焦显微镜观察其构建过程。结果 重组逆转录病毒载体转染的软骨细胞能形成组织工程软骨，增强型绿色荧光蛋白（EGFP）在3周和6周后均得到了明显表达，且未影响软骨组织的形成过程，结论 重组逆转录病毒载体转染的软骨细胞能形成组织工程软骨，EGFP是组织工程化软骨形成过程的良好示踪剂，且不影响组织的形成。     

In vivo gene delivery to articular chondrocytes mediated by an adeno-associated virus vector.

PURPOSES: (1) To investigate the efficiency of direct in vivo adeno-associated virus (AAV) vector-mediated gene transduction to chondrocytes in relation to normal and injured articular cartilage. (2) To evaluate the effects of ultra-violet light-activated gene transduction (LAGT) in chondrocytes in vivo. (3) To determine dissemination of active rAAV vector after intra-articular administration. METHODS: Rabbit knees with either normal or injured cartilage received an intra-articular injection with 1.5x10(12) infectious rAAV-eGFP particles. The right knees received rAAV-eGFP alone, whereas the left knees were given LAGT-treatment. The transduction efficiencies were determined at 1 and 3 weeks after infection by fluorescence-activated cell scanning. The occurrence of active shedding was monitored in serum and various tissues. RESULTS: After 1 week, 7% of the chondrocytes in normal cartilage were transduced by direct rAAV transduction technique. Chondrocytes in cartilage defects demonstrated higher transduction rates compared to chondrocytes in normal cartilage. LAGT increased the cellular eGFP expression in the internal zones to 12%, but did not have any effect in the external zones in defects. Finally, infectious particles were not detected in either serum or tissue samples. CONCLUSIONS: Direct rAAV-mediated gene transfer in vivo to articular chondrocytes is possible. LAGT improves rAAV transduction of chondrocytes in vivo but appears to have a very limited range of effect induction. Expression of eGFP was not determined in other tissues than synovium and cartilage in the treated joints.     

Sizable Scaffold‐Free Tissue‐Engineered Articular Cartilage Construct for Cartilage Defect Repair

This study introduces an implantable scaffold‐free cartilage tissue construct (SF) that is composed of chondrocytes and their self‐produced extracellular matrix (ECM). Chondrocytes were grown in vitro for up to 5 weeks and subjected to various assays at different time points (1, 7, 21, and 35 days). For in vivo implantation, full‐thickness defects (n = 5) were manually created on the trochlear groove of the both knees of rabbits (16‐week old) and 3 week‐cultured SF construct was implanted as an allograft for a month. The left knee defects were implanted with 1, 7, and 21 days in vitro cultured scaffold‐free engineered cartilages. (group 2, 3, and 4, respectively). The maturity of the engineered cartilages was evaluated by histological, chemical and mechanical assays. The repair of damaged cartilages was also evaluated by gross images and histological observations at 4, 8, and 12 weeks postsurgery. Although defect of groups 1, 2, and 3 were repaired with fibrocartilage tissues, group 4 (21 days) showed hyaline cartilage in the histological observation. In particular, mature matrix and columnar organization of chondrocytes and highly expressed type II collagen were observed only in 21 days in vitro cultured SF cartilage (group 4) at 12 weeks. As a conclusion, cartilage repair with maturation was recapitulated when implanted the 21 day in vitro cultured scaffold‐free engineered cartilage. When implanting tissue‐engineered cartilage, the maturity of the cartilage tissue along with the cultivation period can affect the cartilage repair.     

Cell-based treatment of osteochondral defects in the rabbit knee with natural and synthetic matrices: cellular seeding determines the outcome

Introduction: Matrix-associated transplantation of cartilage constructs is an appealing method in cartilage repair. Three different matrices seeded with allogenic chondrocytes were compared in an osteochondral defect model in the rabbit. An investigation was conducted to identify the best matrix for cell-based treatment of osteochondral defects in the rabbit knee joint. Materials and methods: Osteochondral defects (diameter 3 mm) were created in the trochlea and the femoral condyles of 33 New Zealand White rabbits, which were then treated with bioartificial cartilage constructs. The cartilage constructs were created in vitro using three different resorbable carrier materials (two fleece matrices: one of PLLA, and one composite of polydioxanon/polyglactin, as well as one consisting of lyophilized dura) cultured with isolated allogenic chondrocytes. The defects were evaluated macroscopically, by histological and immunhistological techniques, and by scanning electron microscopy after 6 weeks, 6 months, and 12 months. The chondrocyte-seeded constructs were compared to defects treated with carrier material alone as well as to untreated control defects. Results: There was a significant improvement in defect repair quality in the transport materials, which were cultured with chondrocytes prior to implantation (P<0.0005). No significant differences were observed between the three carrier matrices, and no significant differences were seen between the unseeded matrices and the untreated control defects. Conclusion: There is no difference in the outcome between the three tested matrices in the treatment of osteochondral defects in the rabbit knee. The results of this in vitro experiment are promising and with refinement may lead to useful clinical therapies.