Repair of osteochondral defects in the knee by resorbable bioimplants in a rabbit model

Background?The intrinsic healing capacity of articular cartilage remains poor, despite various attempts that have been undertaken to treat cartilage defects. This study describes the experimental use of a double-layer bioimplant consisting of a bone-substitute layer and a cartilage-substitute layer.

Animals and methods?In group A, 12 implants were placed into osteochondral defects of the load-bearing area of rabbit femoral condyles. In group B, 12 implantations were done in the same manner, with a separating membrane consisting of cement between both layers to investigate ingrowth of mesenchymal stem cells from the subchondral marrow space. Group C, with 12 similar defects but without treatment, served as control. Investigations by microscopy and immunohistochemistry were done after 12 weeks.

Results?All bioimplants showed coverage of the defect with a regeneration tissue that contained cartilage-like regions. Implants with a cement layer showed less cartilage and more fibrous tissue. The bioimplant group showed more cartilage-like regeneration tissue than the control groups, which only showed incomplete fibrous filling of the defects. Results from the second group supported the hypothesis that the subchondral space must be opened for adequate regeneration. Additional examinations were done using an established semiquantitative score. The bioimplant group showed a significant improvement in results compared to the group with the separating layer and the control group.

Interpretation?Our findings indicate that cartilage repair by resorbable bioimplants seems to be a promising new approach, especially if mesenchymal stem cells are present and can differentiate under mechanical load.     

Repair of osteochondral defects in the rabbit knee with Gore-Tex (expanded polytetrafluoroethylene). An experimental study

In 28 knee joints in 14 rabbits 4 mm circular osteochondral defects were created in each medial femoral condyle. In 24 of the knee joints 4 mm Gore-Tex (E-PTFE) patches were glued into the defects with fibrin glue. Four joints were left without implants and served as controls. In 16 joints the membrane showed good macroscopic incorporation into the joint surface. In four joints the E-PTFE patches were lying loose. In the controls the defects were covered by thin irregular layers of reparative tissue. On histological examination at 12 weeks, cells were seen proliferating through the membrane and overlying its joint facing surface with the morphological appearance of the outer layers of the normal articular surface. We conclude that Gore-Tex might be of potential value in restoring the architecture of a damaged articular surface.     

Repair of osteochondral defects in articular weightbearing areas in the rabbit's knee

Summary Repair of osteochondral defects in articular weightbearing areas presents its own particular problems because of the low potential of hyaline cartilage for regeneration.Our first group of experiments on the knee of the rabbit confirms that the new regenerated cartilage comes from bone marrow which degenerates before developing into true hyaline cartilage. The second group of experiments shows that autologous grafts from the non-weightbearing articular area suitable for the repair of defects in weightbearing areas. In an third group, autologous mensical fibro-cartilage was used as a graft for the repair of osteochondral defects.

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Résumé La réparation des pertes de substance ostéo-cartilagineuses au niveau des zones d'appui pose des problèmes particuliers en raison du faible pouvoir de régénération du cartilage hyalin.Notre premier groupe d'expériences sur le genou du lapin montre que le cartilage régénéré provient de la moelle osseuse qui dégénère avant de se transformer en véritable cartilage hyalin. Le deuxième groupe d'expériences démontre que des autogreffes, prélevées sur la partie non portante du cartilage articulaire, sont susceptibles de combler les pertes de substance en zone d'appui. Dans un troisième groupe on a utilisé comme greffe le fibro-cartilage ménsial autologue.

     

动物源性骨软骨支架修复兔膝关节复合缺损

目的探讨软骨细胞-动物源性骨软骨支架复合体修复兔膝关节骨软骨复合缺损的可行性和影响因素。方法将改良贴壁离心法获取的骨髓间充质干细胞（bone marrowm esenchymal stem cells,BMSCs）/诱导分化的软骨细胞共培养后与经深低温冷冻、脱脂、脱钙、真空冷冻干燥和辐照消毒的动物源性骨软骨支架复合,构建共培养细胞＋软骨-骨一体化复合支架。27只新西兰大白兔随机分为实验组（A组）、对照组（B组）和空白组（C组）,每组9只。于兔股骨髁间窝处钻一深6mm的骨软骨复合缺损,A组植入共培养细胞＋骨软骨复合支架,B组植入骨软骨复合支架,C组不植入任何支架材料和细胞,分别于术后4周、8周和12周取材,行大体观察、苏木精—伊红染色和甲苯胺蓝染色,并对各标本的软骨切片进行组织学评分。结果随着时间的延长,A组大体观察见复合缺损区已完全修复,局部无凹陷,新生组织和周围组织融合;B组新生组织仍不能完全填充缺损;C组缺损区仍明显。苏木精—伊红染色和甲苯胺蓝染色见A组软骨缺损区由新生的透明软骨样组织修复,细胞呈柱状排列,极性好,软骨陷窝明显,骨缺损区由骨样组织修复,新生软骨和软骨下骨以及宿主骨界面耦合良好;B组新生软骨细胞无软骨陷窝,排列混乱,各界面藕合欠理想;C组可见陈旧性肉芽组织生长并突出于缺损区表面。甲苯胺蓝染色阳性率和组织学评分结果表明,A组与B、C两组之间的差异具有统计学意义（P〈0.05）。结论 BMSCs/诱导分化的软骨细胞共培养细胞复合动物源性骨软骨支架对兔膝关节软骨和软骨下骨的复合缺损具有修复作用。     

Repair of experimentally induced large osteochondral defects in rabbit knee with various concentrations of Escherichia coli-derived recombinant human bone morphogenetic protein-2

Effective therapies for the regeneration of large osteochondral defects are still lacking; however, various approaches have been used. We evaluated the efficacy of Escherichia coli-derived dimeric recombinant human BMP-2 (E-rhBMP-2) for the repair of large osteochondral defects in a rabbit model. Osteochondral defects made in the femoral patellar groove of the knee were treated by transplanting gelatin sponges onto which no or various doses of E-rhBMP-2 were loaded. The outcomes were compared with those of an untreated control group four, 12 and 24 weeks after transplantation. At early time points, the cartilage tissue was repaired in a dose-dependent manner, and bone repair was accelerated in the defects treated with high doses of E-rhBMP-2. At 24 weeks, the repair of cartilage tissue was better with E-rhBMP-2 treatment, even at low doses, than without E-rhBMP-2 treatment. Our findings suggest that the use of E-rhBMP-2 improves and accelerates the repair of osteochondral defects in a rabbit model.     

Blocking of tumor necrosis factor activity promotes natural repair of osteochondral defects in rabbit knee

Background and purpose Osteochondral defects have a limited capacity for repair. We therefore investigated the effects of tumor necrosis factor (TNF) signal blockade by etanercept (human recombinant soluble TNF receptor) on the repair of osteochondral defects in rabbit knees.Material and methods Osteochondral defects (5 mm in diameter) were created in the femoral patellar groove in rabbits. Soon after the procedure, a first subcutaneous injection of etanercept was performed. This single injection or, alternatively, 4 injections in total (twice a week for 2 weeks) were given. Each of these 2 groups was divided further into 3 subgroups: a low-dose group (0.05 μg/kg), an intermediate-dose group (0.4 μ g/kg), and a high-dose group (1.6 μ g /kg) with 19 rabbits in each. As a control, 19 rabbits were injected with water alone. The rabbits in each subgroup were killed 4 weeks (6 rabbits), 8 weeks (6 rabbits), or 24 weeks (7 rabbits) after surgery and repair was assessed histologically.Results Histological examination revealed that the natural process of repair of the osteochondral defects was promoted by 4 subcutaneous injections of intermediate-dose etanercept and by 1 or 4 injections of high-dose etanercept at the various time points examined postoperatively (4, 8, and 24 weeks). Western blot showed that rabbit TNFα had a high affinity for etanercept.Interpretation Blocking of TNF by etanercept enabled repair of osteochondral defects in rabbit knee. Anti-TNF therapy could be a strategy for the use of tissue engineering for bone and cartilage repair.     

Treatment of articular defects with meniscal allografts in a rabbit knee model

Deep-frozen allogeneic meniscal grafts for the treatment of articular cartilage defects were performed experimentally. Osteochondral defects 3 mm in diameter were created bilaterally on the medial femoral condyles of 50 Japanese white rabbits. A meniscus was then grafted into the defect in the left knee, and the right knee was left untreated. At various periods from 2 to 24 weeks postoperatively, the rabbits were killed and macroscopic and histologic examinations were performed. Two weeks after operation, the grafted meniscus was bonded to the floor of the defect. After 12 weeks, chondrocytes producing matrix granules was shown by electron microscopy. After 24 weeks, a congruous articular surface was formed. With time, cellular elements infiltrated into the graft from the surrounding tissues, and gradually increased in penetration. Weight bearing in the early stage after operation did not degrade the grafted menisci, and no changes were shown in the opposing cartilage of the tibia. Deep-frozen allogeneic menisci may be useful as a biological implant to repair articular cartilage defects in this model.     

Repair of cartilage defects in the knee

The factors to be considered in selecting a technique are the diameter of the chondral defect, the depth of the bone defect, and the knee alignment. As a rough guide, I suggest the following guide to treatment. Chondral defects (without bone involvement) <3 cm in diameter can be treated with microfracture, autologous chondrocyte transplantation, osteochondral autografts, or periosteal grafts. Osteochondral defects <3 cm in diameter and 1 cm in bone depth can be treated with autologous chondrocyte transplantation, osteochondral autografts, or periosteal grafts. Articular defects >3 cm in diameter and 1 cm in bone depth require osteochondral allografts. This is a rough guide to treatment and only the opinion of the author. The greater the bone involvement and the less contained the defect, the greater the need for allograft tissue. Allograft tissue should, however, only be used when the size of the lesion is beyond the other techniques. For all of these techniques, realignment osteotomy should be performed as an adjunct procedure if the lesion is in a compartment under more than physiological compression.     

Repair of osteochondral defects with autogenous callo-osseous grafts

A new method of biological repair of osteochondral defects is presented. An osteochondral defect in the rabbit knee was reconstructed with an autogenous callo-osseous graft made of a superficial sheet of medullary fracture callus attached to a base of cancellous bone. The reparative tissues were evaluated for 24 weeks by histology, analysis of uronic acid contents, and immunohistochemical staining of collagen constituents. The callo-osseous graft provided significantly faster and better repair of the articular surface than an untreated defect or a callo-osseous graft in which the cells had been devitalized by irradiation prior to transplantation. The results indicate that the callo-osseous graft contributes to the repair process via providing both favorable extracellular matrices and pluripotential mesenchymal cells.     

Chondral defects of the knee treated by transplantation of autogenous osteochondral plugs

Objective  Complete, lasting coverage of chondral defects of femoral condyles with hyaline cartilage. Indications  Pain, disturbance of function due to chondral defects of weight-bearing joint sections grade III and IV according to Outerbridge and osteochondritis dissecans stage 4 and 5 according to Rodegerdts & Gleissner. Contraindications  Arthritis, infection, preexisting and persisting instability, axial malalignment. Surgical Technique  Anteromedial or anterolateral arthrotomy to evaluate the size of the chondral defect, removal of the cartilage defect by cutting out cartilage-bone cylinders with the Twin Diamond Transplantation Technique (TDTT). Harvesting of appropriately sized cartilage-bone cylinders with healthy hyaline cartilage from non-weight-bearing areas of the posterior or anterior articular surface with the TDTT. Transplantation of the cartilage-bone cylinder “press-fit” into the prepared defect without further fixation. Refilling of the defects caused by harvesting with corresponding periosteum-covered bone cylinders from the iliac crest or with bone substitute materials. Results  In 1996 and 1997, 20 patients with cartilage damage of a femoral condyle were treated by osteochondral cylinder transplantation. All 20 patients were questioned as to their subjective assessment of the outcome and reexamined 6, 12, and 24 months postoperatively. The evaluation of the results was performed using Meyers and Lysholm scores and the Tegner activity scale 3, 6, 12, and 24 months postoperatively. Five women and 15 men had cartilage defects measuring from 3.22 to 4.25 cm². 24 months postoperatively, 17 patients felt significant and three patients slight improvement. According to the evaluation criteria of the Meyers score, results in five patients were considered excellent, in 13 good, and in two moderate. According to the Lysholm score, 15 patients were judged as being excellent and five as being good, and according to the Tegner activity scale, nine were considered excellent, nine good, and two moderate. Reasons for the two moderate results were remaining activity-related pain and knee flexion limited to 120°. In both patients an additional osteochondral cylinder had been harvested from the posterior aspect of a femoral condyle.     

Repair of osteochondral defects with a new porous synthetic polymer scaffold

We developed a new porous scaffold made from a synthetic polymer, poly(DL-lactide-co-glycolide) (PLG), and evaluated its use in the repair of cartilage. Osteochondral defects made on the femoral trochlear of rabbits were treated by transplantation of the PLG scaffold, examined histologically and compared with an untreated control group. Fibrous tissue was initially organised in an arcade array with poor cellularity at the articular surface of the scaffold. The tissue regenerated to cartilage at the articular surface. In the subchondral area, new bone formed and the scaffold was absorbed. The histological scores were significantly higher in the defects treated by the scaffold than in the control group (p<0.05). Our findings suggest that in an animal model the new porous PLG scaffold is effective for repairing full-thickness osteochondral defects without cultured cells and growth factors.     

Repair of osteochondral defects with hyaluronan- and polyester-based scaffolds

OBJECTIVE: The natural repair of osteochondral defects can be enhanced with biocompatible, biodegradable materials that support the repair process. It is our hypothesis that hyaluronan-based scaffolds are superior to synthetic scaffolds because they provide biological cues. We tested this thesis by comparing two hyaluronan-based scaffolds [auto cross-linked polysaccharide polymer (ACP) and HYAFF-11] to polyester-based scaffolds [poly(DL-lactic-co-glycolic acid) (PLGA) and poly(L-lactic acid) (PLLA)] with similar pore size, porosity and degradation times. DESIGN: Fifty-four rabbits received bilateral osteochondral defects. One defect received a hyaluronan-based scaffold and the contralateral defect received the corresponding polyester-based scaffold. Rabbits were euthanized 4, 12 and 20 weeks after surgery and the condyles dissected and processed for histology. RESULTS: Only ACP-treated defects presented bone at the base of the defect at 4 weeks. At 12 weeks, only defects treated with rapidly dissolving implants (ACP and PLGA) presented bone reconstitution consistently, while bone was present in only one third of those treated with slowly dissolving scaffolds (HYAFF-11 and PLLA). After 20 weeks, the articular surface of PLGA-treated defects presented fibrillation more frequently than in ACP-treated defects. The surface of defects treated with slowly dissolving scaffolds presented more cracks and fissures. CONCLUSIONS: The degradation rate of the scaffolds is critical for the repair process. Slowly dissolving scaffolds sustain thicker cartilage at the surface but, it frequently presents cracks and discontinuities. These scaffolds also delay bone formation at the base of the defects. Hyaluronan-based scaffolds appear to allow faster cell infiltration leading to faster tissue formation. The degradation of ACP leads to rapid bone formation while the slow degradation of HYAFF-11 prolongs the presence of cartilage and delays endochondral bone formation.     

组织工程骨-软骨复合体修复犬股骨头负重区大面积骨软骨缺损的实验研究

目的 探讨采用犬股骨头负重区骨和天然软骨制备的骨-软骨双层支架复合成软骨诱导的骨髓间质干细胞(bone marrow mesenchymal stem cells,BMSCs)修复犬股骨头负重区大面积骨软骨缺损的疗效.方法 利用软骨细胞外基质作为软骨支架部分,犬股骨头负重区骨柱经脱细胞处理后作为骨支架部分,采用相分离技术制备骨-软骨双层支架.将成软骨诱导的BMSCs种植到双层支架上体外构建组织工程骨-软骨复合体,并以此修复犬股骨头负重区大面积骨软骨缺损(直径11 mm,高10 mm),第3、6个月时分别取材,行大体、X线片、组织学、Micro-CT和生物力学等检测.结果 X线片及大体观察:3个月时可见股骨头负重区出现轻度塌陷;6个月时出现严重塌陷,呈重度骨关节炎改变.组织学观察:第3、6个月时软骨缺损部分均以纤维组织或纤维软骨充填,周围软骨退变,骨缺损部分不同程度塌陷,与宿主骨质结合紧密.第3、6个月时骨软骨缺损的骨体积分数均低于正常股骨头,差异有统计学意义.6个月时重建软骨下骨的刚度明显低于正常股骨头,差异有统计学意义.结论 结构性骨-软骨双层支架复合成软骨诱导的BMSCs修复犬股骨头负重区骨软骨缺损效果不佳,易导致股骨头塌陷.

Abstract：

Objective To investigate the effects of the novel scaffold on repairing large,high-loadbearing osteochondral defects of femoral head in a canine model.Methods The biphasic scaffolds were fabricated using cartilage extracellular matrix (ECM)-derived scaffold (cartilage layer) and acellular bone matrix (bone layer) by phase separation technique.Articular high-load-bearing osteochondral defects with a diameter of 11-mm and the depth of 10-mm were created in femoral heads.The defects were treated with constructs of a biphasic scaffold seeded with chondrogenically induced bone marrow-derived mesenehymal stem cells (BMSCs).The outcomes were evaluated for gross morphology,histological,biomechanical and micro-CT analysis at the third and sixth month after implantation.Results The gross and X-ray results showed femoral head slightly collapsed at the third month and severely collapse at the sixth month.Histological analysis showed cartilage defects were repaired with fibrous tissue or fibrocartilage with severe osteoarthritis and the varied degrees of the collapse of femoral heads were presented.Micro-CT showed that the values of bone volume fraction in defect area were always lower than those of the normal area in the femoral heads.Biomechanical analysis showed rigidity of the subchondral bone in defect area was significantly lower than that in normal area in the femoral heads at the sixth month.Conclusion The ECM-derived,integrated biphasic scaffold seeded with chondrogenically induced BMSCs could not successfully repair the large high-load-bearing osteochondral defects of the femoral head.     

Techniques for cartilage repair in chondral and osteochondral defects of the knee

Many techniques are currently used in an attempt to regenerate cartilage surfaces in the presence of a chondral or osteochondral defect. Clinical results have been mixed and no single treatment has emerged as being superior. This article reviews the techniques previously and currently being used and evidence to support their use.     

Treatment of cartilage defects with autologous osteochondral grafts in the knee joint

. The treatment of full-thickness cartilage defects still poses problems. One of the currently applied methods to cover defects in the knee joint is osteochondral autografting. The purpose of this prospective study was to assess the outcome of this surgical treatment. Between August 1996 and July 1999, osteochondral transplantation was performed in 38 patients (mean age 37 years). The size of the lesion varied from 1.5 cm² to 6.5 cm². Before the operation the patients had suffered symptoms for an average of 15 months. Osteochondral grafts were harvested from the non-weight-bearing area of the femoral condyle. Patients were evaluated at a mean follow-up of 36 (range 23-58) months. The review protocol included clinical examination, X-ray, and magnetic resonance imaging (MRI). For comparative evaluation the HSS (Hospital for Special Surgery), Lysholm and Tegner activity scores were used. In 86% of the patients studied results were rated good or excellent. There were no infection, thrombophlebitis, or evidence of graft loosening. We conclude autologous osteochondral transplantation is an efficacious and valid one-step repair of the localized damaged articular surface of the knee. Nevertheless, in the future, studies with longer follow-ups and comparison with other forms of biologic resurfacing will be necessary. Résumé. Le traitement des lésions du cartilage articulaire pose encore des problèmes thérapeutiques. Une des méthodes actuellement employées pour réparer des lésions dans l'articulation du genou est l'autogreffe ostéochondrale. Le but de cette étude est d'évaluer les résultats de ce traitement chirugical. Dans la période du ao?t 1996 jusqu'au juillet 1999, 38 patients (age moyen: 37 ans) ont éte traités par une autogreffe ostéochondrale. La taille de la lésion variait de 1.5 cm² à 6.5 cm². Avant d'être opérés, les patients montraient des sympt?mes pour une moyenne de 15 mois. Les greffons ostéochondraux avaient été prélevés dans une zone non portante du condyle fémoral. Les patients ont été surveillès pendant une periode moyenne de 36 mois (de 23 à 58 mois). Le protocole du contr?le incluait des examens cliniques, des radiographies et du MRI. L'évaluation comparative a été executée par les scores d'activités HSS (Hospitale for Special Surgery), Lysholm et Tegner. Dans les 86%, les résultats etaient bons ou excellents. Il n'y a pas eu d'infection ni de thrombophlébite ni de signe que les greffons se détachaient. Nous concluons donc que l'autogreffe ostéochondrale est un moyen efficace, qui peut être exécuté par une seule intervention, pour réparer les lésions localisées de la surface articulaire du genou. Néanmoins, dans l'avenir d'autres études assurant des suivis plus longs et des comparaisons avec d'autres formes de la reconstitution biologique de la surface seront nécessaires.     

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.     

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.     

Transplantation of tissue-engineered osteochondral plug using cultured chondrocytes and interconnected porous calcium hydroxyapatite ceramic cylindrical plugs to treat osteochondral defects in a rabbit model

Abstract:  The purpose of this study was to evaluate the macroscopic and histological results of transplanting a tissue-engineered composite plug made of tissue-engineered cartilage and interconnected porous calcium hydroxyapatite ceramics (IP-CHA) with a very high porosity of 94.9% to treat osteochondral defects. Twelve 12-week-old male Japanese white rabbits were used. Fresh articular cartilage slices were taken, and isolated chondrocytes (2 × 10⁶ cells) were embedded in atelocollagen gel. They were seeded on the top of IP-CHA plugs and cultured for 2 weeks. These tissue-engineered composite plugs were transplanted into the osteochondral defects in the patellar grooves (the experimental group). In the control group, the defects were treated with composite plugs without chondroytes. Twelve weeks after transplantation in the experimental group, the defects were repaired with cartilage-like tissue with good subchondral bone formation histologically. Histological scores in the experimental group were significantly better than those in the control group. This study clearly showed the defects that had been treated with tissue-engineered composite plugs.