The effects of staged intra-articular injection of cultured autologous mesenchymal stromal cells on the repair of damaged cartilage: a pilot study in caprine model

Introduction

Treatment of chondral injuries remains a major issue despite the many advances made in cartilage repair techniques. Although it has been postulated that the use of marrow stimulation in combination with cell-based therapy may provide superior outcome, this has yet to be demonstrated. A pilot study was thus conducted to determine if bone marrow derived mesenchymal stromal cells (BM-MSCs) have modulatory effects on the repair outcomes of bone marrow stimulation (BMS) techniques.

Methods

Two full-thickness chondral 5 mm diameter defects were created in tandem on the medial condyle of left stifle joints of 18 Boer caprine (N = 18). Goats were then divided equally into three groups. Simultaneously, bone marrow aspirates were taken from the iliac crests from the goats in Group 1 and were sent for BM-MSC isolation and expansion in vitro. Six weeks later, BMS surgery, which involves subchondral drilling at the defect sites, was performed. After two weeks, the knees in Group 1 were given autologous intra-articular BM-MSCs (N = 6). In Group 2, although BMS was performed there were no supplementations provided. In Group 3, no intervention was administered. The caprines were sacrificed after six months. Repairs were evaluated using macroscopic assessment through the International Cartilage Repair Society (ICRS) scoring, histologic grading by O’Driscoll score, biochemical assays for glycosaminoglycans (GAGs) and gene expressions for aggrecan, collagen II and Sox9.

Results

Histological and immunohistochemical analyses demonstrated hyaline-like cartilage regeneration in the transplanted sites particularly in Group 1. In contrast, tissues in Groups 2 and 3 demonstrated mainly fibrocartilage. The highest ICRS and O’Driscoll scorings was also observed in Group 1, while the lowest score was seen in Group 3. Similarly, the total GAG/total protein as well as chondrogenic gene levels were expressed in the same order, that is highest in Group 1 while the lowest in Group three. Significant differences between these 3 groups were observed (P <0.05).

Conclusions

This study suggests that supplementing intra-articular injections of BM-MSCs following BMS knee surgery provides superior cartilage repair outcomes.     

In Vivo Evaluation of a Novel Oriented Scaffold-BMSC Construct for Enhancing Full-Thickness Articular Cartilage Repair in a Rabbit Model

Tissue engineering (TE) has been proven usefulness in cartilage defect repair. For effective cartilage repair, the structural orientation of the cartilage scaffold should mimic that of native articular cartilage, as this orientation is closely linked to cartilage mechanical functions. Using thermal-induced phase separation (TIPS) technology, we have fabricated an oriented cartilage extracellular matrix (ECM)-derived scaffold with a Young''s modulus value 3 times higher than that of a random scaffold. In this study, we test the effectiveness of bone mesenchymal stem cell (BMSC)-scaffold constructs (cell-oriented and random) in repairing full-thickness articular cartilage defects in rabbits. While histological and immunohistochemical analyses revealed efficient cartilage regeneration and cartilaginous matrix secretion at 6 and 12 weeks after transplantation in both groups, the biochemical properties (levels of DNA, GAG, and collagen) and biomechanical values in the oriented scaffold group were higher than that in random group at early time points after implantation. While these differences were not evident at 24 weeks, the biochemical and biomechanical properties of the regenerated cartilage in the oriented scaffold-BMSC construct group were similar to that of native cartilage. These results demonstrate that an oriented scaffold, in combination with differentiated BMSCs can successfully repair full-thickness articular cartilage defects in rabbits, and produce cartilage enhanced biomechanical properties.     

Allogeneic Mesenchymal Stem Cells in Combination with Hyaluronic Acid for the Treatment of Osteoarthritis in Rabbits

Mesenchymal stem cell (MSC)-based therapies may aid in the repair of articular cartilage defects. The purpose of this study was to investigate the effects of intraarticular injection of allogeneic MSCs in an in vivo anterior cruciate ligament transection (ACLT) model of osteoarthritis in rabbits. Allogeneic bone marrow-derived MSCs were isolated and cultured under hypoxia (1% O₂). After 8 weeks following ACLT, MSCs suspended in hyaluronic acid (HA) were injected into the knees, and the contralateral knees were injected with HA alone. Additional controls consisted of a sham operation group as well as an untreated osteoarthritis group. The tissues were analyzed by macroscopic examination as well as histologic and immunohistochemical methods at 6 and 12 weeks post-transplantation. At 6 and 12 weeks, the joint surface showed less cartilage loss and surface abrasion after MSC injection as compared to the tissues receiving HA injection alone. Significantly better histological scores and cartilage content were observed with the MSC transplantation. Furthermore, engraftment of allogenic MSCs were evident in surface cartilage. Thus, injection of the allogeneic MSCs reduced the progression of osteoarthritis in vivo.     

Effects of Notch/p38MAPK signaling pathway on articular cartilage defect recovery by BMSCs tissue based on the rabbit articular cartilage defect models

ObjectiveThe objective is to clarify the effects of Notch/p38MAPK signaling pathway on articular cartilage defect recovery by BMSCs tissue and provide a basis for clinical treatments of articular cartilage defects.MethodsA total of 96 healthy male rabbits (weighed 1.5–2.0 kg) that were fully-grown were selected and grouped as the no-treatment group, the model group, and the treatment group in a random manner. Each group included 32 rabbits in total. The no-treatment group was fed without any interventions. The model group and the treatment group were constructed into rabbit knee-joint articular cartilage defect models. In addition, rabbits in the treatment group were given intervention treatments with Notch inhibitor (DAPT) combined with p38MAPK inhibitor (SB203580). The general conditions of rabbits in each group and the conditions of the stained articular cartilage tissue samples were observed, the proliferation of chondrocytes of rabbits in each group was compared.Results(1) After drug interventions, in contrast to the rabbits in the model group, the general conditions and the chondrocyte recovering situations of rabbits in the treatment group were obviously improved; (2) 8 weeks after model construction, the articular cartilage empty bone lacuna rate of rabbits in the treatment group was (12.13 ± 1.81)%, which was obviously lower than the synchronous (21.55 ± 3.07)% articular cartilage empty bone lacuna rate of rabbits in the model group, and there was a statistical significance in the differences (P < 0.05); (3) the absorbance value (OD value) of chondrocytes in the treatment group was (0.34 ± 0.015), which was obviously higher than the (0.10 ± 0.020) OD value of chondrocytes in the model group, and there was a statistical significance in the differences (P < 0.05).ConclusionThe inhibition of Notch/p38MAPK signaling pathway can promote the recovery of articular cartilage by BMSCs tissue, accelerate the proliferation of chondrocytes, and contribute to the recovery of knee-joint injuries in rabbits, which provides a reliable basis for clinical treatments of articular cartilage defects.     

骨碎补结合组织工程软骨促进软骨再生的实验研究

目的:评估骨碎补结合组织工程软骨治疗对实验兔软骨缺陷模型软骨再生的疗效。方法:将h IGF-1基因转染MSCs,并与脱细胞真皮基质(ADM)构建组织工程软骨。24只新西兰白兔随机分为A、B、C、D四组,A、C组进行自体软骨移植,B、D组进行改建的细胞-ADM移植。C、D组用40%骨碎补汤喂养4周,150 m L/d。第12周处死实验动物,分离缺损关节软骨部位,蜡块包埋染色,通过总体形态评价软骨再生组织。采用组织学评分评估再生软骨质量。采用甲苯胺蓝染色评价缺损部位产生软骨糖胺聚糖的情况。结果:与B组比较,C组和D组的新生软骨覆盖度、新骨髓的颜色、缺损边缘和表面粗糙度均显著提高(P0.05);再生软骨的组织学评分软骨表面评分显著改善(P0.05)。C组与D组具有比其他组更好的基质、细胞分布和表面指数。并且有较厚的透明样软骨组织,具有正常的糖胺聚糖产生。表明该治疗方法可以通过再生透明样软骨且没有不良事件来减少软骨缺陷。结论:工程软骨结合骨碎补治疗可显著改善兔膝关节软骨缺损修复的质量,为临床治疗软骨病变提供重要理论依据。     

Intra-articular delivery of chondroitin sulfate for the treatment of joint defects in rabbit model

Chondroitin sulfate (CS) is considered as a possible candidate for the treatment of joint defect. This study is to evaluate the efficacy of intra-articular injection of CS carried by hydrogel in the treatment of chondral defects in adult rabbit models. Inclusion of CS (0–50 μg/ml) in in vitro chondrocyte culture exerts a dose-dependent increase in cell proliferation. To select for optimal carrier for in vivo study, the release kinetic of CS embedded in five types of hydrogel was studied using fluorescence technique and their biocompatibilities in vivo were investigated by injecting the CS-hydrogel into rabbit knees. α-CD-EG 4400 hydrogel was chosen as the carrier based on progressively released CS from the hydrogel, with 80% released by in one week while the remaining 20% was retained for 30 days. In vivo studies showed high biocompatibility of CS-hydrogel. To evaluate the efficacy of CS in the treatment of cartilage injury, chondral defects were created in femoral medial condyle (punch diameter 2.7 mm) or trochlea (punch diameter 3.5 mm) of the rabbits without damaging subchondral bone. CS (100 mg/ml) in 0.5 ml α-CD-EG 4400 hydrogel was then injected into the knee joint. Hydrogel and saline served as controls. On day 50 the chondral defect in the saline group showed no signs of healing and defect treated with hydrogel alone was covered with a thin and slightly irregular layer of fibrous tissue. The CS-hydrogel group showed a thicker layer composed of both hyaline and fibrocartilage. The modulus of elasticity was the highest in the CS-hydrogel group and lowest in the group injected with saline only. Our results suggest that intra-articular delivery of CS by α-CD-EG 4400 improved the biomechanical and histological properties of the repaired cartilage. It may be an effective treatment for cartilage injury. Paper presented at ICRS and OARSI.     

Early cartilage abnormalities at the hip are associated with obesity and body composition measures – a 3.0T MRI community-based study

IntroductionAlthough obesity is a risk factor for hip osteoarthritis (OA), the role of body composition, if any, is unclear. This study examines whether the body mass index (BMI) and body composition are associated with hip cartilage changes using magnetic resonance imaging (MRI) in community-based adults.Methods141 community-based participants with no clinical hip disease, including OA, had BMI and body composition (fat mass and fat free mass) measured at baseline (1990 to 1994), and BMI measured and 3.0 T MRI performed at follow-up (2009–2010). Femoral head cartilage volume was measured and femoral head cartilage defects were scored in the different hip regions.ResultsFor females, baseline BMI (β = −26 mm³, 95% Confidence interval (CI) -47 to −6 mm³, p = 0.01) and fat mass (β = −11 mm³, 95% CI −21 to −1 mm³, p = 0.03) were negatively associated with femoral head cartilage volume. Also, while increased baseline fat mass was associated with an increased risk of cartilage defects in the central superolateral region of the femoral head (Odds Ratio (OR) = 1.08, 95% CI 1.00–1.15, p = 0.04), increased baseline fat free mass was associated with a reduced risk of cartilage defects in this region (OR = 0.82, 95% CI 0.67–0.99; p = 0.04). For males, baseline fat free mass was associated with increased femoral head cartilage volume (β = 40 mm³, 95% CI 6 to 74 mm³, p = 0.02).ConclusionsIncreased fat mass was associated with adverse hip cartilage changes for females, while increased fat free mass was associated with beneficial cartilage changes for both genders. Further work is required to determine whether modifying body composition alters the development of hip OA.     

Requirement of fibroblast growth factor signaling for regeneration of epiphyseal morphology in rabbit full-thickness defects of articular cartilage

The involvement of fibroblast growth factor-2 (FGF-2) during the repair process in rabbit full-thickness defects of articular cartilage was studied. Fibroblast growth factor-2 (50 pg/h) was administered for 2 weeks in a 5mm defect of articular cartilage, which is large enough not to repair spontaneously. The administration of FGF-2 resulted in the regeneration of the articular cartilage and the subchondral bone within 8 weeks. In these defects, undifferentiated mesenchymal cells initiated chondrogenic differentiation coupled with replacement by subchondral bone, resulting in the resurfacing of the defects with hyaline cartilage and the recovery of subchondral bone up to the original bone–articular cartilage junction. In rabbits, full-thickness defects are capable of regenerating articular cartilage as long as the defect size is limited to ≤3 mm in diameter. In the defects, strong immunoreactivity for FGF-2 was observed in the granulation tissue filling the defects in the early stage of repair, in association with the expression of FGF-2 mRNA shown by in situ hybridization. Once the undifferentiated mesenchymal cells had differentiated into chondrocytes, both the immunoreactivity and the in situ hybridization signal declined significantly. Upon the local administration of a monoclonal antibody against FGF-2 (bFM-1, 50ng/h), the defects were filled with fibrous tissue and no resurfacing hyaline cartilage was formed. Compared to the non-treated defects, there were marked increases in FGF-2 immunoreactivity and the overexpression of FGF-2 mRNA in the reparative tissue in the bFM-1 -treated defects. This rebound phenomenon indicates that the autocrine FGF-2 signaling is critically important for the regeneration of articular cartilage.     

Local intra-articular injection of rapamycin delays articular cartilage degeneration in a murine model of osteoarthritis

Introduction

Recent studies have revealed that rapamycin activates autophagy in human chondrocytes preventing the development of osteoarthritis (OA) like changes in vitro, while the systemic injection of rapamycin reduces the severity of experimental osteoarthritis in a murine model of OA in vivo. Since the systemic use of rapamycin is associated with numerous side effects, the goal of the current study was to examine the beneficial effect of local intra-articular injection of rapamycin in a murine model of OA and to elucidate the mechanism of action of rapamycin on articular cartilage.

Methods

Destabilization of the medial meniscus (DMM) was performed on 10-week-old male mice to induce OA. Intra-articular injections of 10 μl of rapamycin (10 μM) were administered twice weekly for 8 weeks. Articular cartilage damage was analyzed by histology using a semi-quantitative scoring system at 8 and 12 weeks after surgery. Mammalian target of rapamycin (mTOR), light chain 3 (LC3), vascular endothelial growth factor (VEGF), collagen, type X alpha 1 (COL10A1), and matrix metallopeptidase 13 (MMP13) expressions were analyzed by immunohistochemistry. VEGF, COL10A1, and MMP13 expressions were further examined via quantitative RT-PCR (qPCR).

Results

Intra-articular injection of rapamycin significantly reduced the severity of articular cartilage degradation at 8 and 12 weeks after DMM surgery. A reduction in mTOR expression and the activation of LC3 (an autophagy marker) in the chondrocytes was observed in the rapamycin treated mice. Rapamycin treatment also reduced VEGF, COL10A1, and MMP13 expressions at 8 and 12 weeks after DMM surgery.

Conclusion

These results demonstrate that the intra-articular injection of rapamycin could reduce mTOR expression, leading to a delay in articular cartilage degradation in our OA murine model. Our observations suggest that local intra-articular injection of rapamycin could represent a potential therapeutic approach to prevent OA.     

Icariin accelerates cartilage defect repair by promoting chondrogenic differentiation of BMSCs under conditions of oxygen‐glucose deprivation

This study explored the role played by combined ICA and bone mesenchymal stem cells (BMSCs) in repairing rabbit knee cartilage defects. Firstly, rabbit BMSCs were isolated and used to construct an in vitro cellular model of oxygen‐glucose deprivation/reoxygenation (OGD/R). Subsequently, ICA processing, Alcian blue staining, immunofluorescence and Western blot studies were performed to evaluate the ability of BMSCs to display signs of chondrogenic differentiation. Furthermore, a rabbit knee cartilage injury model was established in vivo. International Cartilage Repair Society (ICRS) macroscopic evaluations, H&E, Alcian blue and EdU staining, as well as immunohistochemistry, were analysed cartilage repair and pathological condition of the knee cartilage tissue. Our in vitro results showed that ICA promoted the chondrogenic differentiation of BMSCs, as well as aggrecan (AGR), bone morphogenetic protein 2 (BMP2) and COL2A1 protein expression in BMSCs. In vivo experiments showed that rabbits in the BMSCs or ICA treatment group had higher ICRS scores and displayed a better restoration of cartilage‐like tissue and chondrocyte expression on the surface of their cartilage defects. In conclusion, ICA or BMSCs alone could repair rabbit knee cartilage damage, and combined treatment with ICA and BMSCs showed a better ability to repair rabbit knee cartilage damage.     

Operative Treatment of Terrible Triad of the Elbow via Posterolateral and Anteromedial Approaches

The aim of the study was to explore the clinical outcome of posterolateral and anteromedial approaches in treatment of terrible triad of the elbow. The study involved 12 patients with closed terrible triad of the elbow treated by posterolateral and anteromedial approaches between January 2010 and June 2012. The mechanism of injury included fall from height in 9 patients and traffic accident in 3. According to O’Driscoll classification for fractures of the ulnar coronoid, there were 11 patients with type Ⅰ and 1 with type Ⅱ fractures. According to Mason classification for fractures of the radial head, there were 3 patients with type Ⅰ, 7 with type Ⅱ and 2 with type Ⅲ fractures. All patients were followed up for 12-27 months (average 15.5 months), which showed no pain or severe pain in all patients except for 2 patients with mild pain. At the last follow-up, the mean flexion was for 125°(range, 90°-140°), the mean extension loss for 20°(range, 0°-70°), the mean pronation for 66°(range, 20°-85°) and the mean supination for 60°(range, 30°-85°). The bony union time was 8-14 weeks (average 11 weeks) and the elbows were stable in flexion-extension and varus-valgus in all patients. The elbows maintained a concentric reduction of both the ulnotrochlear and the radiocapitellar articulation. Mild heterotopic ossification of the elbow occurred in 3 patients at 6 months after operation and mild degenerative change in 1 patient at 18 months after operation. The Broberg and Morrey elbow performance score was 82 points (range, 58-98 points). The results were excellent in 6 patients, good in 4, fair in 1 and poor in 1, with excellence rate of 83.3%. The results showed that the combined posterolateral and anteromedial approaches can facilitate the reduction and fixation of terrible triad of the elbow. Repair of radial head, coronoid, medial and lateral collateral ligaments can sufficiently restore the elbow stability, allow early postoperative motion and enhance the functional recovery.     

Xenogeneic transplantation of articular chondrocytes into full-thickness articular cartilage defects in minipigs: fate of cells and the role of macrophages

Xenogeneic or allogeneic chondrocytes hold great potential to build up new cartilage in vivo. However, immune rejection is a major concern for the utility of universal donor-derived cells. In order to verify the reported immune privilege of chondrocytes in vivo, the aim of this study was to assess engraftment of human articular chondrocytes (HAC) in minipig knee cartilage defects and their contribution to cartilage regeneration. HAC were transplanted matrix-assisted within two hydrogels into full-thickness cartilage defects of minipigs or implanted ectopically into immune deficient mice to assess redifferentiation capacity. At 2 and 4 weeks after surgery, cell-persistence and host cell invasion were monitored by species-specific in situ hybridization and RT-PCR. Early tissue regeneration was evaluated by histomorphometry and a modified O’Driscoll score. HAC capable of successful in vivo chondrogenic redifferentiation persisted at ectopic sites for 4 weeks in both carrier materials. Early defect regeneration involved extensive host cell invasion and a decline of HAC to less than 5 % of initial cell numbers in 6/12 defects within 2 weeks. Few clusters of persisting HAC within collagen type II-rich tissue were surrounded by porcine macrophages. Four weeks after cell transplantation, most of the defects contained well-integrated cell-rich tissue free of human cells with no apparent difference between hydrogel carriers. In summary, HAC failed to engraft in porcine articular cartilage defects despite their ability for successful in vivo redifferentiation. The co-localization of macrophages to hydrogel-implanted HAC suggests active graft rejection without evidence for an immune-privileged status of xenogeneic chondrocytes in a large animal joint.     

Occupational risk factors for hip osteoarthritis are associated with early hip structural abnormalities: a 3.0?T magnetic resonance imaging study of community-based adults

IntroductionOccupational exposure to heavy lifting and stair climbing are associated with radiographic hip osteoarthritis (OA). This study examined whether these activities are associated with early structural hip joint changes in a community-based population.MethodsIn total, 198 community-based people with no history of hip disease, including OA, had 3.0 T-magnetic resonance imaging (MRI) to assess hip cartilage volume, defects and bone marrow lesions (BMLs). Recall of occupational exposure to heavy lifting and stair climbing aged 18 to 30 years and in the previous 10 years were collected. A persistence score was defined as exposure at neither time point (0), at one time point (1) or at both time points (2).ResultsExposure to heavy lifting when aged 18 to 30 years was associated with BMLs of the central superolateral femoroacetabular region (odds ratio (OR) 3.9, 95% confidence interval (CI) 1.6 to 9.8, P <0.01), with persistence score associated with cartilage defects in the central superolateral region of the femoral head (OR 1.6, 95% CI 1.0 to 2.5, P = 0.04). Exposure to stair climbing aged 18 to 30 years and persistence score were associated with an increased risk of cartilage defects in the central superolateral femoral head and BMLs in the central superolateral and posterior femoroacetabular regions (OR range 2.1 to 3.2, all P ≤0.03).ConclusionsOccupational exposure to heavy lifting and stair climbing are associated with hip structural abnormalities. If confirmed by longitudinal data, such associations may explain how occupational activities affect the hip joint and may identify new targets for the prevention of hip OA.     

Prospective evaluation of serum biomarker levels and cartilage repair by autologous chondrocyte transplantation and subchondral drilling in a canine model

Introduction

The purpose of this study was to evaluate serum chondroitin sulfate (CS) and hyaluronic acid (HA) levels and the capability of cartilage repair of full-thickness cartilage defects after treatment with two different fundamental surgical techniques: autologous chondrocyte transplantation (AC) and subchondral drilling (SD).

Methods

A 4-mm-diameter full-thickness cartilage defect was created in each of 10 skeletally mature male outbred dogs. The dogs were randomly separated into two groups. Groups A and B were treated with AC and SD, respectively. An evaluation was made at the 24th week of the experiment. Serum was analyzed prospectively – preoperatively and at 6-week intervals – for CS and HA levels by enzyme-linked immunosorbent assay (ELISA) and ELISA-based assays, respectively.

Results

The cartilage repair assessment score (median ± standard deviation) of group A (9.5 ± 2.5) was significantly higher than that of group B (2.5 ± 1.3) (P < 0.05). Group A also demonstrated a better quality of hyaline-like cartilage repair. Prospective analysis of serum WF6 and HA levels between the two groups did not show any significant difference. Serum WF6 levels at the 24th week of the experiment had a negative correlation (r = -0.69, P < 0.05) with the cartilage repair assessment score, whereas serum HA levels tended to correlate positively (r = 0.46, 0.1 <P < 0.05).

Conclusions

AC treatment provides superior results to SD treatment, according to morphology, histology, and cartilage marker levels. AC treatment demonstrated a smoother surface, less fissure, better border integration, and a more reliable outcome of repairing cartilage. Moreover, a decreasing level of serum WF6, which correlated with good quality of the repairing tissue at the end of the follow-up period, was found predominantly in the AC group. Serum WF6 therefore should be further explored as a sensitive marker for the noninvasive therapeutic evaluation of cartilage repair procedures.     

Osteochondral regeneration by a bilayered construct in a cell-free or cell-based approach

A bilayered construct with or without adipose-derived stem cells (ASCs) was applied to repair full-thickness defects in the patellar groove of 18 rabbits. Non-treated and treated defects were divided into three groups: a control group (n = 12), a cell-free group (n = 12) and a cell-based group (n = 12). Histological appearance and grading were evaluated at 8 and 12 weeks. At 12 weeks, osteochondral-like tissues completely filled in the defects and integrated with host tissues in the cell-based group. The semi-quantitative score of the cell-based group (4.2 ± 1.2), which is a total score ranging from 0 (best) to 20 (worst), was significantly better than that of the other two groups (cell-free: 13.8 ± 2.5; control: 10.3 ± 2.4). This finding indicated that the bilayered constructs combined with ASCs could be an effective way to enhance osteochondral regeneration.     

Effect of Transplanting Various Concentrations of a Composite of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells and Hyaluronic Acid Hydrogel on Articular Cartilage Repair in a Rabbit Model

BackgroundMesenchymal stem cells (MSCs) are known to have therapeutic potential for cartilage repair. However, the optimal concentration of MSCs for cartilage repair remains unclear. Therefore, we aimed to explore the feasibility of cartilage repair by human umbilical cord blood-derived MSCs (hUCB-MSCs) and to determine the optimal concentrations of the MSCs in a rabbit model.MethodsOsteochondral defects were created in the trochlear groove of femur in 55 rabbits. Four experimental groups (11 rabbits/group) were treated by transplanting the composite of hUCB-MSCs and HA with various MSCs concentrations (0.1, 0.5, 1.0, and 1.5 x 10⁷ cells/ml). One control group was left untreated. At 4, 8, and 16 weeks post-transplantation, the degree of cartilage repair was evaluated grossly and histologically.FindingsOverall, transplanting hUCB-MSCs and HA hydrogel resulted in cartilage repair tissue with better quality than the control without transplantation (P = 0.015 in 0.1, P = 0.004 in 0.5, P = 0.004 in 1.0, P = 0.132 in 1.5 x 10⁷ cells/ml). Interestingly, high cell concentration of hUCB-MSCs (1.5×10⁷ cells/ml) was inferior to low cell concentrations (0.1, 0.5, and 1.0 x 10⁷ cells/ml) in cartilage repair (P = 0.394,P = 0.041, P = 0.699, respectively). The 0.5 x 10⁷ cells/ml group showed the highest cartilage repair score at 4, 8 and 16 weeks post transplantation, and followed by 0.1x10⁷ cells/ml group or 1.0 x 10⁷ cell/ml group.ConclusionsThe results of this study suggest that transplantation of the composite of hUCB-MSCs and HA is beneficial for cartilage repair. In addition, this study shows that optimal MSC concentration needs to be determined for better cartilage repair.     

Kartogenin prevents cartilage degradation and alleviates osteoarthritis progression in mice via the miR-146a/NRF2 axis

Osteoarthritis (OA) is a common articular degenerative disease characterized by loss of cartilage matrix and subchondral bone sclerosis. Kartogenin (KGN) has been reported to improve chondrogenic differentiation of mesenchymal stem cells. However, the therapeutic effect of KGN on OA-induced cartilage degeneration was still unclear. This study aimed to explore the protective effects and underlying mechanisms of KGN on articular cartilage degradation using mice with post-traumatic OA. To mimic the in vivo arthritic environment, in vitro cultured chondrocytes were exposed to interleukin-1β (IL-1β). We found that KGN barely affected the cell proliferation of chondrocytes; however, KGN significantly enhanced the synthesis of cartilage matrix components such as type II collagen and aggrecan in a dose-dependent manner. Meanwhile, KGN markedly suppressed the expression of matrix degradation enzymes such as MMP13 and ADAMTS5. In vivo experiments showed that intra-articular administration of KGN ameliorated cartilage degeneration and inhibited subchondral bone sclerosis in an experimental OA mouse model. Molecular biology experiments revealed that KGN modulated intracellular reactive oxygen species in IL-1β-stimulated chondrocytes by up-regulating nuclear factor erythroid 2-related factor 2 (NRF2), while barely affecting its mRNA expression. Microarray analysis further revealed that IL-1β significantly up-regulated miR-146a that played a critical role in regulating the protein levels of NRF2. KGN treatment showed a strong inhibitory effect on the expression of miR-146a in IL-1β-stimulated chondrocytes. Over-expression of miR-146a abolished the anti-arthritic effects of KGN not only by down-regulating the protein levels of NRF2 but also by up-regulating the expression of matrix degradation enzymes. Our findings demonstrate, for the first time, that KGN exerts anti-arthritic effects via activation of the miR-146a-NRF2 axis and KGN is a promising heterocyclic molecule to prevent OA-induced cartilage degeneration.Subject terms: Osteoarthritis, Drug development     

深低温冻存组织工程化软骨在关节软骨缺损修复的应用

目的：探讨经深低温冻存组织工程化软骨修复关节软骨缺损的可行性。方法：分离收集3周龄新西兰大白兔关节软骨细胞进行体外培养,接种于PGA三维支架材料上,复合物体外培养1周后冻存,冻存1个月后解冻、复苏及体外培养,1周后接种于已建立的双侧兔膝关节软骨缺损模型的膝关节软骨缺损处,并设对照组。分别于手术后4周、8周、12周行大体标本及组织观察。结果：大体观察结果表明,实验组与对照组缺损处均由软骨组织修复;组织学观察可以见到实验组和对照组关节软骨缺损处有密集的软骨细胞,均有软骨生成及基质分泌,两组差异无统计学意义。结论：应用深低温冻存组织工程化软骨修复关节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实验依据。     

Increased physical activity severely induces osteoarthritic changes in knee joints with papain induced sulfate-glycosaminoglycan depleted cartilage

Introduction

Articular cartilage needs sulfated-glycosaminoglycans (sGAGs) to withstand high pressures while mechanically loaded. Chondrocyte sGAG synthesis is regulated by exposure to compressive forces. Moderate physical exercise is known to improve cartilage sGAG content and might protect against osteoarthritis (OA). This study investigated whether rat knee joints with sGAG depleted articular cartilage through papain injections might benefit from moderate exercise, or whether this increases the susceptibility for cartilage degeneration.

Methods

sGAGs were depleted from cartilage through intraarticular papain injections in the left knee joints of 40 Wistar rats; their contralateral joints served as healthy controls. Of the 40 rats included in the study, 20 rats remained sedentary, and the other 20 were subjected to a moderately intense running protocol. Animals were longitudinally monitored for 12 weeks with in vivo micro-computed tomography (μCT) to measure subchondral bone changes and single-photon emission computed tomography (SPECT)/CT to determine synovial macrophage activation. Articular cartilage was analyzed at 6 and 12 weeks with ex vivo contrast-enhanced μCT and histology to measure sGAG content and cartilage thickness.

Results

All outcome measures were unaffected by moderate exercise in healthy control joints of running animals compared with healthy control joints of sedentary animals. Papain injections in sedentary animals resulted in severe sGAG-depleted cartilage, slight loss of subchondral cortical bone, increased macrophage activation, and osteophyte formation. In running animals, papain-induced sGAG-depleted cartilage showed increased cartilage matrix degradation, sclerotic bone formation, increased macrophage activation, and more osteophyte formation.

Conclusions

Moderate exercise enhanced OA progression in papain-injected joints and did not protect against development of the disease. This was not restricted to more-extensive cartilage damage, but also resulted in pronounced subchondral sclerosis, synovial macrophage activation, and osteophyte formation.