同种异体组织工程化软骨修复关节软骨缺损

目的　探讨应用同种异体组织工程化软骨修复软骨缺损的可行性。方法　取新西兰大白兔双膝关节软骨细胞 ,经体外培养扩增 ,与PlruonicF12 7混合 ,植入人为造成的异体兔膝关节软骨缺损。结果　空白对照组和材料对照组只见少许纤维组织修复 ,缺损凹陷 ;实验组 8周后关节软骨缺损区由部分白色透明样软骨组织充填 ,Masson三色染色见胶原分布较均匀 ,软骨陷窝多见 ,未见明显炎症现象。 16周后缺损完全修复 ,缺损表面较光滑 ,部分颜色呈淡兰色 ,软骨陷窝清晰 ,细胞与基质分布均匀 ,未见炎症和退变现象。结论　同种异体组织工程化软骨可用于修复关节软骨缺损。     

组织工程技术修复同种异体兔关节软骨缺损实验研究

目的：探讨关节软骨缺损治疗的新途径。方法：把几丁糖作为软骨细胞培养的支架。将几丁糖与软骨细胞一起体外培养,然后移植修复同种异体兔的膝关节软骨缺损,并对关节软骨的修复过程进行术后16周大体、组织学、电镜观察及修复组织厚度测定。结果：几丁糖无纺网在术后2周开始降解吸收,术后10-12周完全吸收;术后第16周在实验侧关节软骨缺损处可见成熟的透明软骨,软骨缺损得到完全修复。结论：几丁糖泊生物学特性符合组织工程中对细胞培养支架的要求;几个糖负载软骨细胞移植修复同种异体兔膝关节软骨缺损,兔膝关节全层软骨缺损得到成功修复。为临床上关节软骨缺损的治疗提供了可能的途径。     

Treatment of a Focal Articular Cartilage Defect of the Talus with Polymer-Based Autologous Chondrocyte Implantation: A 12-Year Follow-Up Period

Autologous chondrocyte implantation (ACI) is a first-line treatment option for large articular cartilage defects. Although well-established for cartilage defects in the knee, studies of the long-term outcomes of matrix-assisted ACI to treat cartilage defects in the ankle are rare. In the present report, we describe for the first time the long-term clinical and radiologic results 12 years after polymer-based matrix-assisted ACI treat a full-thickness talar cartilage defect in a 25-year-old male patient. The clinical outcome was assessed using the visual analog scale and Freiburg ankle score, magnetic resonance imaging evaluation using the Henderson-Kreuz scoring system and T2 mapping. Clinical assessment revealed improved visual analog scale and Freiburg ankle scores. The radiologic analysis and T2 relaxation time values indicated the formation of hyaline-like repair tissue. Polymer-based autologous chondrocytes has been shown to be a safe and clinically effective long-term treatment of articular cartilage defects in the talus.     

Cartilage repair: A review of Stanmore experience in the treatment of osteochondral defects in the knee with various surgical techniques

Articular cartilage damage in the young adult knee, if left untreated, it may proceed to degenerative osteoarthritis and is a serious cause of disability and loss of function. Surgical cartilage repair of an osteochondral defect can give the patient significant relief from symptoms and preserve the functional life of the joint. Several techniques including bone marrow stimulation, cartilage tissue based therapy, cartilage cell seeded therapies and osteotomies have been described in the literature with varying results. Established techniques rely mainly on the formation of fibro-cartilage, which has been shown to degenerate over time due to shear forces. The implantation of autologous cultured chondrocytes into an osteochondral defect, may replace damaged cartilage with hyaline or hyaline-like cartilage. This clinical review assesses current surgical techniques and makes recommendations on the most appropriate method of cartilage repair when managing symptomatic osteochondral defects of the knee. We also discuss the experience with the technique of autologous chondrocyte implantation at our institution over the past 11 years.     

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.     

Assessment of Cartilage Repair Quality With the International Cartilage Repair Society Score and the Oswestry Arthroscopy Score

The International Cartilage Repair Society (ICRS) score and the Oswestry Arthroscopic Score (OAS) have been validated to evaluate repair tissue quality. However, the performance of these scores has not been studied in typical patients undergoing cartilage repair and who have lesions of varying sizes. In this study, we compared the performance of the ICRS and the OAS scores and analyzed the effect of lesion characteristics on the performance of these two scores. Cartilage repair quality was assessed in a total of 104 arthroscopic observations of cartilage repair sites of the knee in 62 patients after autologous chondrocyte implantation. Two observers scored the repair areas independently with the ICRS and the OAS scores. The performance of both scores was evaluated according to internal consistency and inter-rater reliability and correlation between the scores. The frequency and proportion of disagreements were analyzed according to the repair site area and the given score. The correlation between the scores was good (r = 0.91, 95% confidence interval [CI]: 0.87–0.94). Both scores showed moderate internal consistency and inter-rater reliability. Cronbach's α was 0.88 (95% CI: 0.80–0.92) for the ICRS score and 0.79 (95% CI: 0.70–0.86) for the OAS score. The intraclass correlation coefficient was 0.89 (95% CI: 0.84–0.92) for the ICRS and 0.81 (95% CI: 0.74–0.87) for the OAS scores. The frequency and proportion of disagreements were higher in larger repair sites. In arthroscopic use, both ICRS and OAS scores perform similarly, however, their reliability deteriorates as the lesion size increases. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:555–562, 2020     

组织工程软骨生物反应器的设计

目的 根据流体力学原理优化反应器内腔结构,并设计一套组织工程软骨生物反应器.方法 采用计算机仿真的方法对反应器内腔流场进行仿真计算,通过流场分析确定内腔结构,最终构建一套完整的生物反应器系统.结果 确定了生物反应器的内腔结构,生物反应器由控制系统和细胞培养室两部分构成,能置于培养箱中对软骨细胞材料复合物进行动态培养.结论 反应器内腔的结构是合理的.整个生物反应器系统运行可靠.     

组织工程软骨生物反应器的设计

目的 根据流体力学原理优化反应器内腔结构,并设计一套组织工程软骨生物反应器.方法 采用计算机仿真的方法对反应器内腔流场进行仿真计算,通过流场分析确定内腔结构,最终构建一套完整的生物反应器系统.结果 确定了生物反应器的内腔结构,生物反应器由控制系统和细胞培养室两部分构成,能置于培养箱中对软骨细胞材料复合物进行动态培养.结论 反应器内腔的结构是合理的.整个生物反应器系统运行可靠.     

注射型软骨组织工程活性材料修复关节软骨缺损的实验研究

目的 利用注射型活性材料修复兔关节软骨,比较碱性成纤维细胞生长因子(Basic fibroblastic growth factor,bFGF)和骨形态发生蛋白(Bone morphogenic protein,BMP)的治疗效果。方法 取18只14周龄成年大白兔随机分为3组,每只动物于双侧膝关节股骨髌股关节面制备直径4mm全层软骨缺损模型,钻通骨髓腔。准备注射型bFGF和BMP生物蛋白胶材料,三组分别接受如下治疗。A组注射型碱性成纤维细胞生长因子,B组注射型骨形态发生蛋白,C组单纯生物蛋白胶治疗。于8周,12周,24周观察大体和形态学特征,组织学评分。结果 A组于8,12周时,软骨缺损被白色半透明坚硬组织平滑修复,富有光泽,与正常软骨边界清楚,24周时修复组织与正常软骨边界模糊,质地坚硬,表面平滑光润。B组于8,12,24周时候均未将缺损表面修复平滑,为疏松组织缺损底部,凹陷明显残留。C组全程均未见软骨修复。组织评分A组明显优于B组C组(P<0.05),而B组和C组间无差异(P>0.05)。结论 注射型碱性成纤维细胞生长因子可以有效修复兔关节软骨的缺损,效果明显优于BMP治疗组,为关节镜等微创治疗方法提供了实验参考。     

人脂肪来源干细胞与胶原支架共培养的实验研究

目的观察评价人脂肪来源干细胞(Human adipose derived stem cells,hADSCs)在胶原支架中的生长情况,为进一步体内组织修复研究提供依据。方法取人抽脂术后脂肪,经胶原酶解、过滤、离心获得hADSCs,代传代扩增后,接种到胶原支架上。细胞-材料复合物分别体外培养1周、2周,裸鼠体内培养2周、4周后,HE染色观察细胞在支架上的生长情况,免疫组化HLA-Ⅰ检测经裸鼠体内培养后的复合物上的细胞的种属来源。结果原代培养的hADSCs呈"梭形"或"成纤维细胞"样,并以克隆团形式生长。免疫荧光Vimentin染色阳性。第3代hADSCs经流式细胞鉴定,CD29、CD44、CD105表达阳性,CD45、CD34表达阴性。胶原支架复合hADSCs经过体外、体内培养,hADSCs均能长入胶原支架的空隙内,且体内培养比体外培养有更多的细胞长入支架。体外培养1周,已有细胞粘附生长在胶原支架的边缘,体外培养2周后,更多的细胞渗透到材料内部。体内培养2周后,大量细胞占据材料的边缘,有部分细胞能渗透到材料内部甚至材料全层。体内培养4周后,大量细胞渗透支架全层。HLA-Ⅰ抗体检测发现,支架材料内部细胞阳性表达,说明胶原支架内部的细胞来源于hADSCs。结论胶原支架与hADSCs具有较好的相容性,可作为hADSCs的载体材料,用于组织工程缺损修复的研究。     

改良纤维蛋白胶软骨膜块在模拟微重力培养下修复关节软骨缺损

目的观察在模拟微重力条件下改良纤维蛋白胶支架软骨膜块修复关节软骨缺损的影响。方法将兔软骨细胞种植到经抑肽酶和氨甲环酸改良的纤维蛋白胶支架材料上,然后分别在旋转培养仪和培养板中培养,体外培养2周。然后将培养出的软骨膜块植入于动物关节软骨缺损模型中,并设立对照组定期进行大体、组织形态学观察、生化指标检测及组织工程学评分。结果①采用改良支架的复合物经体外培养2周,基本保持了原有塑形;②修复关节软骨效果由好至差的顺序为旋转培养组、普通培养组、单纯改良支架组、空白对照组。组织工程学评分通过统计学处理各组之间存在显著性差异(P=0.01)。结论①经过改良的纤维蛋白胶支架,其降解速率与软骨组织基质形成同步,能长时间支持体内、外软骨组织的形成,并适合旋转培养,是软骨组织工程中适宜的支架;②旋转培养仪提供的模拟微重力环境,可以提高工程化软骨的质量,为实现关节软骨损伤的修复提供有效途径。     

Pluronic F-127负载同种异体软骨细胞修复兔全厚关节软骨缺损的实验研究

目的:探讨Pluronic F-127作为软骨细胞移植载体的可行性。方法:将培养的软骨细胞与20％的Pluronic F-127的混合,移植到兔膝关节软骨缺损处。在移植后4、8、12周对缺损的修复情况进行大体、光镜组织学评估和电镜观察。结果:移植的软骨细胞在载体中生长良好,形成了透明软骨。扫描电镜下可见多数成熟的透明软骨细胞。Pluronic F-127平均降解时间为6～8周,与正常软骨的再生速度基本一致。根据Wakitani制定的评分标准,采用盲法对修复质量作出评价。Pluronic F-127组和对照组的组织学评分在各个时期无显著的差异(P>0.001),而软骨细胞-载体复合体组和对照组相比在各个时期均存在显著的差异(P<0.001)。结论:Pluronic F-127可作为工程化软骨细胞安全有效的载体。     

Transplantation of Autologous Chondrocytes Seeded on a Fibrin/Hyaluronan Composite Gel Into Tracheal Cartilage Defects in Rabbits: Preliminary Results

Reconstruction of tracheal defects is one of the most difficult procedures in head and neck surgery. To date, various reconstructing techniques have been used with no consensus on the best approach. This study investigated the feasibility of using a fibrin/hyaluronic acid (HA) composite gel with autologous chondrocytes for tracheal reconstruction. Chondrocytes from autologous rabbit auricular cartilages were expanded and seeded into a culture dish at high density to form stable tracheal cartilages mechanically using a fibrin/HA composite gel. A 1‐cm long by 0.5‐cm wide defect was created by a scalpel on the cervical tracheae of six rabbits. Tissue‐engineered cartilages using fibrin/HA composite were trimmed and fixed to the defect boundaries with tissuecol. Postoperatively, the site was evaluated endoscopically, histologically, radiologically, and functionally. None of the six rabbits showed signs of respiratory distress. Postoperatively, in all cases, rigid telescopic examination showed that the implanted scaffolds were completely covered with regenerated mucosa without granulation or stenosis. Histologically, the grafts showed no signs of inflammatory reaction and were covered with ciliated epithelium. Even when grafts were broken and migrated from their original insertion site, the implanted cartilages were well preserved. However, the grafts did show signs of mechanical failure at the implantation site. The beat frequency of ciliated epithelium on implants was very similar to that of normal respiratory mucosa. In conclusion, implants with autologous chondrocytes cultured with fibrin/HA showed good tracheal luminal contour, functional epithelial regeneration, and preservation of neocartilage without inflammation but lacked adequate mechanical stability.     

戊二醛交联提高胶原/壳聚糖真皮支架抗细胞收缩能力的研究

目的 应用交联处理的胶原支架,来减缓其降解过多、易收缩变形、机械性能不足等缺点.方法 采用冷冻-冻干法构建胶原/壳聚糖多孔支架,并通过戊二醛交联构建具有良好抗细胞收缩稳定性和细胞相容性的胶原支架.通过测定不同培养时间各类支架的尺寸变化,研究了戊二醛交联对支架抗细胞收缩能力的影响.结果 戊二醛交联后的胶原/壳聚糖支架未见明显收缩,其细胞活性也始终高于干热交联支架.结论 戊二醛能有效地提高胶原基支架的抗细胞收缩能力,并保持其良好的生物相容性.