Effects of cell phenotype and seeding density on the chondrogenic capacity of human osteoarthritic chondrocytes in type I collagen scaffolds.
细胞表型和种植密度对 I 型胶原支架中人骨关节炎软骨细胞成软骨能力的影响。
- 作者列表："Cao C","Zhang Y","Ye Y","Sun T
OBJECTIVE:Matrix-associated autologous chondrocyte implantation (MACI) achieves good clinical efficacy in young patients with focal cartilage injury; however, phenotypic de-differentiation of chondrocytes cultured in monolayer and the treatment of older OA patients are still challenges in the field of cartilage tissue engineering. This study aimed to assess the in vitro re-differentiation potential and in vivo chondrogenic capacity of human OA chondrocytes inoculated into collagen I scaffolds with different cellular phenotypes and seeding densities. METHODS:OA chondrocytes and articular chondrocyte-laden scaffolds were cultured over 2 weeks in in vitro. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and histological staining were used to detect the mRNA expression profiles and extracellular matrix secretion of chondrocyte-specific markers. OA chondrocyte-laden collagen I scaffolds with different cellular phenotypes, and seeding densities were implanted into SCID mice over 4 weeks to evaluate the chondrogenic capacity in vivo. RESULTS:Increased COL2a1, ACAN, COMP, SOX9, and BMP2 expression levels and decreased COL1a1, VCAN, MMP13, and ADAMTS5 amounts were observed in OA chondrocytes seeded in collagen I scaffolds; Implantation of phenotypically superior OA chondrocytes in collagen I scaffolds at high density could improve the chondrogenic capacity of human OA chondrocytes, as confirmed by RT-qPCR assessed gene expression patterns in vitro and histological evaluation in vivo. CONCLUSIONS:Freshly isolated chondrocytes from OA patients could be a source of replacement for articular chondrocytes being commonly used in MACI. Implantation of phenotypically superior OA chondrocytes in collagen I scaffolds at high density could be a promising tool for the treatment of elderly OA patients.
目的: 基质相关的自体软骨细胞植入 (MACI) 在年轻的局灶性软骨损伤患者中取得了良好的临床疗效; 然而,单层培养的软骨细胞的表型去分化和老年 OA 患者的治疗仍然是软骨组织工程领域的挑战。本研究旨在评估接种到不同细胞表型和种植密度的胶原 I 支架中的人 OA 软骨细胞的体外再分化潜能和体内成软骨能力。 方法: 体外培养 OA 软骨细胞和关节软骨细胞负载支架超过 2 周。采用逆转录定量聚合酶链反应 (RT-qPCR) 和组织学染色检测软骨细胞特异性标志物的 mRNA 表达谱和细胞外基质分泌。将不同细胞表型的 OA 软骨细胞负载胶原 I 支架和种植密度植入 SCID 小鼠 4 周，评价体内软骨形成能力。 结果: 种植在胶原 I 支架中的 OA 软骨细胞表达 COL2a1 、 ACAN 、 COMP 、 SOX9 和 BMP2 水平增加，COL1a1 、 VCAN 、 MMP13 和 ADAMTS5 水平降低; 将表型优越的 OA 软骨细胞高密度植入胶原 I 支架可提高人 OA 软骨细胞的成软骨能力,RT-qPCR 证实，体外评估基因表达模式和体内组织学评估。 结论: OA 患者新鲜分离的软骨细胞可能是 MACI 中常用关节软骨细胞的替代来源。将表型优越的 OA 软骨细胞高密度植入胶原 I 支架可能是治疗老年 OA 患者的一种有前途的工具。
METHODS:Purpose To determine outcomes of transphyseal ACL reconstruction using a living parental hamstring tendon allograft in a consecutive series of 100 children. Methods One hundred consecutive juveniles undergoing ACL reconstruction with a living parental hamstring allograft were recruited prospectively and reviewed 2 years after ACL reconstruction with IKDC Knee Ligament Evaluation, and KT1000 instrumented laxity testing. Skeletally immature participants obtained annual radiographs until skeletal maturity, and long leg alignment radiographs at 2 years. Radiographic Posterior tibial slope was recorded. Results Of 100 juveniles, the median age was 14 years (range 8–16) and 68% male. At surgery, 30 juveniles were graded Tanner 1 or 2, 21 were Tanner 3 and 49 were Tanner 4 or 5. There were no cases of iatrogenic physeal injury or leg length discrepancy on long leg radiographs at 2 years, despite a median increase in height of 8 cm. Twelve patients had an ACL graft rupture and 9 had a contralateral ACL injury. Of those without further ACL injury, 82% returned to competitive sports, IKDC ligament evaluation was normal in 52% and nearly normal in 48%. The median side to side difference on manual maximum testing with the KT1000 was 2 mm (range − 1 to 5). A radiographic PTS of 12° or more was observed in 49%. Conclusions ACL reconstruction in the juvenile with living parental hamstring tendon allograft is a viable procedure associated with excellent clinical stability, patient-reported outcomes and return to sport over 2 years. Further ACL injury to the reconstructed and the contralateral knee remains a significant risk, with identical prevalence observed between the reconstructed and contralateral ACL between 12 and 24 months after surgery. Level of evidence III (Cohort Study).
METHODS:Purpose The purpose of the study was to investigate the biomechanics of walking and of the knee joint in the acute phase of ACL injury. Methods We examined 18 patients with acute ACL injuries and 20 healthy adults as controls. The biomechanics of the knee joint and of walking was assessed by 5 inertial sensors fixed with special cuffs to the lower back, the lower third of the thigh, and the lower third of the shank of the right and left legs. The movements and temporal characteristics were recorded while the subject was walking 10 m at a comfortable pace. Based on the results of examination, the patients were divided into two groups: with severe function impairment (6 patients) and with moderate function impairment (12 patients). Results It was found that in the first days post-trauma, not only the knee function was reduced, but the function of the entire lower limb as well. This led to a functional asymmetry. The kinematics of movements in the joints changed in accordance with slower walking. The walking became not only slower, but it was also associated with a decreased impact load in the weight acceptance phase. At later dates, the functional impairments were less pronounced. The total range of flexion motion did not exceed 20 degrees in the first group and 55 degrees in the second one. The injured joint developed functional immobilization within the first days post-injury. This was a guarding response by additional muscle strain to prevent unusual and limit physiological movements in the knee joint. The movements in the knee joint while walking were of small amplitude, rocking, and occurred only under load. The amplitude of the main flexion in the swing phase was reduced. Conclusion The stage of an ACL injury should be assessed not only based on the time post-trauma, but also taking account of the functional parameter—the knee range of flexion while walking. According to our findings, the only factor that had influenced the functional condition of the KJ was the duration of joint immobilization after trauma.
METHODS:Purpose Numerous techniques have been described for the tibial-sided graft preparation in anterior cruciate ligament (ACL) reconstruction. The use of less suture material for graft preparation is thought to improve ingrowth and to reduce the risk for infection. At the same time, the suture construct should be strong enough to resist the surgeon’s pull during tensioning of the transplant. Methods In total, 39 fresh-frozen procine deep flexor tendons were used and prepared as four-strand grafts. In the three-suture group ( n = 19), graft preparation was performed using three tibial-sided sutures, with each tendon end sutured separately. In the one-suture group ( n = 20), a modified graft preparation using only one tibial-sided suture was applied. Each sample underwent load-to-failure testing ( N _max) after cyclic pre-loading. To estimate intraoperative tension forces acting on the tibial-sided suture constructs, the maximal tension force of 26 volunteers on such a construct was measured using a load cell. Results The biomechanical testing of the two different suture constructs showed a significantly higher load-to-failure for the three-suture group (711 N ± 91 N) compared to the one-suture group (347 N ± 24 N) ( p = 0.0001). In both groups, the mode of failure was a tear of the suture in all samples. A failure of the suture–tendon interface was not observed in any case. The median maximal tension force on the construct applied by the 26 volunteers was 134 N (range 73–182 N). Conclusion The presented single-suture tendon graft preparation resisted to smaller failure loads than the conventional three-suture technique. However, no failures in the suture–tendon interface were seen and the failure loads observed were far beyond the tension forces that can be expected intraoperatively. Hence, the single-suture graft preparation technique may be a valuable alternative to the conventional technique.