Cartilage Defects In Rabbits Research Articles

Tsmu solution improves rabbit osteochondral allograft preservation and transplantation outcome.

To compare the effects of Tsmu solution with vitrification on chondrocyte viability and examine histological and biomechanical properties of osteochondral allografts (OCAs) after storage, OCAs from femoral condyles of New Zealand rabbits were harvested, stored for 35days in Tsmu solution or by in vitro vitrification, and subjected to in vivo and in vitro assays. Stored OCAs were transplanted into knee femoral condyle cartilage defects in recipient rabbits. Chondrocyte viability and histological changes of cartilage grafts were assessed in vitro. Gross assessment, chondrocyte viability, histological assessment, OCA biomechanics, and immunological markers were evaluated in vivo 6months after transplantation. Fresh OCAs served as in vitro and in vivo controls. Chondrocyte viability and scores for cartilage surface and histological quantitative assessment were superior for Tsmu solution compared with vitrification, but inferior compared with fresh OCAs in vitro and in vivo. With the exception of interleukin 6 content, biomechanical features of samples stored in Tsmu solution were superior to vitrification, and inferior to fresh OCAs in vivo. Thus, Tsmu solution provided suitable storage that improved chondrocyte viability, intact OCA cartilage matrix architecture, and transplantation outcomes.

HWJECM-Derived Oriented Scaffolds with Autologous Chondrocytes for Rabbit Cartilage Defect Repairing.

Previously, we synthesized an articular cartilage extracellular matrix (ECM)-derived oriented scaffold for cartilage tissue engineering, which was biomimetic in terms of structure and biochemical composition. However, the limit resource of the cartilage-derived ECM is a hindrance for its application. In this study, we developed a new material for cartilage tissue engineering-human umbilical cord Wharton's jelly-derived ECM (hWJECM). The hWJECM has an abundant resource and similar biochemistry with cartilage ECM, and the use of it is not associated with ethical controversy. We adopted the method previously used in cartilage ECM-derived oriented scaffold preparation to generate the oriented hWJECM-derived scaffold, and the scaffold properties were tested in vitro and in vivo. The three-dimensional scaffold has a porous and well-oriented structure, with a mean pore diameter of ∼104 μm. Scanning electron microscopy and cell viability staining results demonstrated that the oriented scaffold has good biocompatibility and cell alignment. In addition, we used functional autologous chondrocytes to seed the hWJECM-derived oriented scaffold and tested the efficacy of the cell-scaffold constructs to repair the full-thickness articular cartilage defect in a rabbit model. Defects of 4 mm diameter were generated in the patellar grooves of the femurs of both knees and were implanted with chondrocyte-scaffold constructs (group A) or scaffolds alone (group B); rabbits with untreated defects were used as a control (group C). Six months after surgery, all defects in group A were filled completely with repaired tissue, and most of which were hyaline cartilage. In contrast, the defects in group B were filled partially with repaired tissue, and approximately half of these repaired tissues were hyaline cartilage. The defects in group C were only filled with fibrotic tissue. Histological grading score of group A was lower than those of groups B and C. Quantification of glycosaminoglycan indicated that newly formed cartilage in group A rabbits was comparable with normal cartilage. In conclusion, hWJECM-derived oriented scaffolds loaded with autologous chondrocytes induced cartilage repair in rabbit knees, which was comparable with native cartilage in terms of macroscopic view, microstructure, and biochemical composition.

The experimental research of calcium alginate gel loaded transforming growth factor beta 3 and compounded with adipose mesenchymal stem cells to repair rabbit articular cartilage defect

Objective To investigate the effect of tissue engineered cartilage on the repair of articular cartilage defects in rabbits with calcium alginate gel (CAG) loaded transforming growth factor beta 3 (TGF-β3) and compounded with adipose mesenchymal stem cells (ADSCs). Methods The ADSCs were separated and cultured in subcutaneous fat of New Zealand white rabbits. The full-thickness articular cartilage defect model was made at the patellar groove by exposure of the femoral ankle joint. ADSCs were implanted into calcium alginate scaffolds loaded with TGF-β3 to construct tissue-engineered cartilage and transplanted into rabbit articular cartilage defects. The animals were divided into four groups: control group (injected with sterile isotonic saline), CAG group (injected with CAG), ADSCs+ CAG group (injected with CAG loaded with ADSCs), TGF-β3+ CAG group (injected with CAG loaded with TGF-β3) and TGF-β3+ ADSCs+ CAG group (injected with CAG loaded with TGF-β3 and ADSCs). At the end of 12 weeks, the repair of articular cartilage defects was observed by gross observation, hematoxylin-eosin (HE) staining, immunohistochemical staining of safranin-O and type Ⅱ collagen, and the scoring method developed by International Association of Cartilage Repair (ICRS) Histological score. Results The cartilage repair effect of TGF-β3+ ADSCs+ CAG group was better than that of other groups, not only the neonatal tissue and the surrounding normal tissue closely, but also the secretion of extracellular matrix and normal tissue similar. The ICRS scores of each group were (7.06±0.18)score, (7.15±0.23)score, (7.45±0.25)score, (7.47±0.24)score and (15.78±0.24)score. That of TGF-β3+ ADSCs+ CAG group was better than that of other groups, the difference was significant (P<0.05). Conclusions CAG loaded with TGF-β3 and combined with ADSCs has a good effect in repairing articular cartilage defects in rabbits, and is a structural repair. Key words: Transforming growth factor beta3; Alginates/AD; Mesenchymal stem cell transplantation; Adipose tissue; Cartilage, articular/PA; Cartilage diseases/TH; Gels

Examine the effect of intra- articular injection of Bone Marrow Mesenchymal Stem Cells (BM-MSCs) and Chondrogenic Differentiated Mesenchymal Stem Cells (CD- MSCs) on the repair of articular cartilage defects in rabbits

Examine the effect of intra- articular injection of Bone Marrow Mesenchymal Stem Cells (BM-MSCs) and Chondrogenic Differentiated Mesenchymal Stem Cells (CD- MSCs) on the repair of articular cartilage defects in rabbits

The Neutrophil Response to Rabbit Antimicrobial Extract After Implantation of Biomaterial into a Bone/Cartilage Defect.

In this study, the neutrophil response to an antimicrobial extract was evaluated as a potential marker of inflammatory process after implantation of alginate and carbon fiber biomaterials into a bone or cartilage defect in rabbits. The response to biomaterials used was assessed based on enzyme release, generation of reactive oxygen species (ROS) and survival of neutrophils isolated from the rabbit's blood after implantation. The implantation of alginate biomaterial increased elastase and alkaline phosphatase release, whereas carbon fibers did not evoke increased elastase release. The implantation of both biomaterials resulted in a higher myeloperoxidase (MPO) release after 30-min incubation. Stimulation with different fractions of antimicrobial peptide (AMP) extract diminished MPO release and nitric oxide generation, as well as slightly reducing neutrophil survival. Our study permits the assessment of the neutrophil intravital response to an implant without the need for preparation of histological sections. Additionally, AMP extract restricted some manifestations of the pro-inflammatory response and may be considered a regulator of neutrophil activity in the early inflammatory phase, preventing rejection of the implant.

Non-invasive monitoring of in vivo hydrogel degradation and cartilage regeneration by multiparametric MR imaging.

Numerous biodegradable hydrogels for cartilage regeneration have been widely used in the field of tissue engineering. However, to non-invasively monitor hydrogel degradation and efficiently evaluate cartilage restoration in situ is still challenging.Methods: A ultrasmall superparamagnetic iron oxide (USPIO)-labeled cellulose nanocrystal (CNC)/silk fibroin (SF)-blended hydrogel system was developed to monitor hydrogel degradation during cartilage regeneration. The physicochemical characterization and biocompatibility of the hydrogel were evaluated in vitro. The in vivo hydrogel degradation and cartilage regeneration of different implants were assessed using multiparametric magnetic resonance imaging (MRI) and further confirmed by histological analysis in a rabbit cartilage defect model for 3 months.Results: USPIO-labeled hydrogels showed sufficient MR contrast enhancement and retained stability without loss of the relaxation rate. Neither the mechanical properties of the hydrogels nor the proliferation of bone-marrow mesenchymal stem cells (BMSCs) were affected by USPIO labeling in vitro. CNC/SF hydrogels with BMSCs degraded more quickly than the acellular hydrogels as reflected by the MR relaxation rate trends in vivo. The morphology of neocartilage was noninvasively visualized by the three-dimensional water-selective cartilage MRI scan sequence, and the cartilage repair was further demonstrated by macroscopic and histological observations.Conclusion: This USPIO-labeled CNC/SF hydrogel system provides a new perspective on image-guided tissue engineering for cartilage regeneration.

Development of chondrocyte-seeded electrosprayed nanoparticles for repair of articular cartilage defects in rabbits

Due to limited self-healing capacity in cartilages, there is a rising demand for an innovative therapy that promotes chondrocyte proliferation while maintaining its biofunctionality for transplantation. Chondrocyte transplantation has received notable attention; however, the tendencies of cell de-differentiation and de-activation of biofunctionality have been major hurdles in its development, delaying this therapy from reaching the clinic. We believe it is due to the non-stimulative environment in the injured cartilage, which is unable to provide sustainable physical and biological supports to the newly grafted chondrocytes. Therefore, we evaluated whether providing an appropriate matrix to the transplanted chondrocytes could manipulate cell fate and recovery outcomes. Here, we proposed the development of electrosprayed nanoparticles composed of cartilage specific proteins, namely collagen type II and hyaluronic acid, for implantation with pre-seeded chondrocytes into articular cartilage defects. The fabricated nanoparticles were pre-cultured with chondrocytes before implantation into injured articular cartilage. The study revealed a significant potential for nanoparticles to support pre-seeded chondrocytes in cartilage repair, serving as a protein delivery system while improving the survival and biofunctionality of transplanted chondrocytes for prolonged period of time.

Biostable scaffolds of polyacrylate polymers implanted in the articular cartilage induce hyaline-like cartilage regeneration in rabbits.

PurposeTo study the influence of scaffold properties on the organization of in vivo cartilage regeneration. Our hypothesis was that stress transmission to the cells seeded inside the pores of the scaffold or surrounding it, which is highly dependent on the scaffold properties, determines the differentiation of both mesenchymal cells and dedifferentiated autologous chondrocytes.Methods4 series of porous scaffolds made of different polyacrylate polymers, previously seeded with cultured rabbit chondrocytes or without cells, were implanted in cartilage defects in rabbits. Subchondral bone was injured during the surgery to allow blood to reach the implantation site and fill the scaffold pores.ResultsAt 3 months after implantation, excellent tissue regeneration was obtained, with a well-organized layer of hyaline-like cartilage at the condylar surface in most cases of the hydrophobic or slightly hydrophilic series. The most hydrophilic material induced the poorest regeneration. However, no statistically significant difference was observed between preseeded and non-preseeded scaffolds. All of the materials used were biocompatible, biostable polymers, so, in contrast to some other studies, our results were not perturbed by possible effects attributable to material degradation products or to the loss of scaffold mechanical properties over time due to degradation.ConclusionsCartilage regeneration depends mainly on the properties of the scaffold, such as stiffness and hydrophilicity, whereas little difference was observed between preseeded and non-preseeded scaffolds.

Effect of polycaprolactone-ascobic acid scaffold in repairing articular cartilage defects in rabbits

To investigate the effect of polycaprolactone-ascobic acid (PCL-AA) scaffolds in promoting repair of articular cartilage defects in a rabbit model. The cartilage defects (3.5 mm in diameter and 3.0 mm in depth) were created in the trochlear groove of the bilateral knees of eight 6-month-old male New Zealand white rabbits. The rabbit models were then randomized into 3 groups to receive implantation of PCL-AA scaffolds (group A, n=8), implantation of PCL scaffolds without AA (group B, n=5), or no treatment (group C, n=3). In groups A and B, the mixture of fibrin gel (10 µg) and thrombinogen (10 µg) was injected into the defects to fix the scaffolds during the surgery. Histological analyses and quantitative assessments of defect repair were conducted at 6 and 12 weeks after implantation of the scaffold. At 6 weeks after scaffold implantation, macroscopic observation showed better filling of the cartilage defects in group A than in group B, while no obvious defect repair was observed in group C. The rabbits in group A showed a significant improvement of the Wakitani score than those in group B (4.05∓1.11 vs 7.05∓0.98, P<0.05). HE staining revealed the presence of newly generated cells in and around the PCL-AA scaffolds without inflammatory cells. Safranin O staining showed a significantly greater ECM of the newly regenerated tissue in groups A and B than in group C (P<0.05), and the volume of the regenerated cartilage and cells was significantly greater in group A than in group B (P<0.05). Samples harvested at 12 weeks showed more hyalione-like cartilage formation than that at 6 weeks in group A. PCL-AA scaffolds have a good biocompatibility and promotes the healing of articular cartilage defects. Adding ascorbic acid into PCL scaffolds better promotes cartilage formation in terms of both quantity and quality of the regenerated tissues. PCL-AA scaffolds can serve as a promising biomaterial to promote the regeneration of articular cartilage using tissue engineering techniques.

Hyaluronan thiomer gel/matrix mediated healing of articular cartilage defects in New Zealand White rabbits\u2014a pilot study

BackgroundArticular cartilage defects are limited to their regenerative potential in human adults. Our current study evaluates tissue regeneration in a surgically induced empty defect site with hyaluronan thiomer as a provisional scaffold in a gel/matrix combination without cells on rabbit models to restore tissue formation.MethodsAn osteochondral defect of 4 mm in diameter and 5 mm in depth was induced by mechanical drilling in the femoral center of the trochlea in 18 New Zealand White rabbits. Previously evaluated from an in vitro study hyaluronan thiomer matrix, and a hyaluronan thiomer gel was used to treat the defect. As a control, the defect was left untreated. During the whole study, rabbits were clinically examined and after 4 (n = 3) or 12 (n = 3) weeks, the rabbits were sacrificed. Joints were evaluated macroscopically (Brittberg score) and by histology (O’Driscoll score). Synovial cells from the synovial fluid smear were histopathologically evaluated.ResultsThe healing of the defects varied intra-group wise at the first observation period. After 12 weeks the results concerning the cartilage repair score were inhomogeneous within each group, while the macroscopic analysis was more homogenous. In the synovial fluid smear, the mean score of infiltrated synovial and non-synovial cells was slightly increased after 4 weeks and slightly decreased after 12 weeks in both the treatment groups in comparison to the untreated control.ConclusionsTaken together with results from the in vivo study indicated that implantation of hyaluronan thiomer as a combination of gel and matrix might enhance articular cartilage regeneration in an empty defect. Despite their benefits, the intrinsic healing capacity of New Zealand rabbits is a limitation for comparative test subject in pre-clinical models of cartilage defects.

Microfracture combined with functional pig peritoneum-derived acellular matrix for cartilage repair in rabbit models.

We report the one-step transplantation of functional acellular peritoneum matrix (APM-E7) with specific mesenchymal stem cell recruitment to repair rabbit cartilage injury. The experimental results illustrated that the APM-E7 scaffold was successfully fabricated, which could specifically recruit MSCs and fill the cartilage defects in the femoral trochlear of rabbits at 24weeks post-surgery. The repaired tissue was hyaline cartilage, which exhibited ideal mechanical stability. The APM-E7 biomaterial could provide scaffold for MSCs and improve cell homing, which are two key factors required for cartilage tissue engineering, thereby providing new insights into cartilage tissue engineering.

Repair of Osteochondral Defects Using Human Umbilical Cord Wharton's Jelly-Derived Mesenchymal Stem Cells in a Rabbit Model.

Umbilical cord Wharton's jelly-derived mesenchymal stem cell (WJMSC) is a new-found mesenchymal stem cell in recent years with multiple lineage potential. Due to its abundant resources, no damage procurement, and lower immunogenicity than other adult MSCs, WJMSC promises to be a good xenogenous cell candidate for tissue engineering. This in vivo pilot study explored the use of human umbilical cord Wharton's jelly mesenchymal stem cells (hWJMSCs) containing a tissue engineering construct xenotransplant in rabbits to repair full-thickness cartilage defects in the femoral patellar groove. We observed orderly spatial-temporal remodeling of hWJMSCs into cartilage tissues during repair over 16 months, with characteristic architectural features, including a hyaline-like neocartilage layer with good surface regularity, complete integration with adjacent host cartilage, and regenerated subchondral bone. No immune rejection was detected when xenograft hWJMSCs were implanted into rabbit cartilage defects. The repair results using hWJMSCs were superior to those of chondrogenically induced hWJMSCs after assessing gross appearance and histological grading scores. These preliminary results suggest that using novel undifferentiated hWJMSCs as seed cells might be a better approach than using transforming growth factor-β-induced differentiated hWJMSCs for in vivo tissue engineering treatment of cartilage defects. hWJMSC allografts may be promising for clinical applications.

Matrigel scaffold combined with Ad-hBMP7-transfected chondrocytes improves the repair of rabbit cartilage defect.

The aim of this study was to explore an effective method for the repair of cartilage defects using chitosan/glycerophosphate (C/GP) gel- and Matrigel-engineered human bone morphogenetic protein 7 (hBMP7)-expressing chondrocytes. Rabbit chondrocytes were obtained, cultured in vitro and transfected with an adenovirus containing hBMP7 and green fluorescent protein (Ad-hBMP7-GFP). The expression of hBMP7 in the transfected cells was tested by reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. The phenotype of the transfected cells was evaluated by detecting the yields of collagen II and hyaluronic acid using RT-PCR and enzyme-linked immunosorbent assay (ELISA). The growth of chondrocytes in the C/GP gel and Matrigel was accessed by measuring the cell growth rate, hematoxylin and eosin (H&E) staining and observation under a scanning microscope. Twelve adult male New Zealand white rabbits were randomly divided into three groups. Two cartilage defects were created in the rabbits' knees by aseptic surgery. Group A (n=4) did not receive any treatment, group B (n=4) were treated with C/GP gel and Matrigel-engineered Ad-mock-GFP-transfected chondrocytes, and group C (n=4) were treated with C/GP gel and Matrigel-engineered Ad-hBMP7-GFP-transfected chondrocytes. Rabbits were sacrificed at 4 weeks after transplantation, and the repair effect was measured by the Wakitani scoring method. On the basis of the RT-PCR and western blot results, hBMP7 was efficiently overexpressed in the Ad-hBMP7-GFP-transfected chondrocytes. The ELISA results showed that the yields of collagen II and hyaluronic acid in Ad-hBMP7-GFP-transfected chondrocytes were significantly higher than those in Ad-mock-GFP-transfected chondrocytes. Chondrocytes have a better morphology and arrangement in a Matrigel scaffold than in C/GP, as assessed by H&E staining and scanning microscopy. According to the Wakitani score, Matrigel combined with Ad-hBMP7-GFP-transfected chondrocytes successfully promoted the repair of cartilage defects in rabbit knees.

Dual Function of Glucosamine in Gelatin/Hyaluronic Acid Cryogel to Modulate Scaffold Mechanical Properties and to Maintain Chondrogenic Phenotype for Cartilage Tissue Engineering.

Glucosamine (GlcN) fulfills many of the requirements as an ideal component in scaffolds used in cartilage tissue engineering. The incorporation of GlcN in a gelatin/hyaluronic acid (GH) cryogel scaffold could provide biological cues in maintaining the phenotype of chondrocytes. Nonetheless, substituting gelatin with GlcN may also decrease the crosslinking density and modulate the mechanical properties of the cryogel scaffold, which may be beneficial as physical cues for chondrocytes in the scaffold. Thus, we prepared cryogel scaffolds containing 9% GlcN (GH-GlcN9) and 16% GlcN (GH-GlcN16) by carbodiimide-mediated crosslinking reactions at −16 °C. The crosslinking density and the mechanical properties of the cryogel matrix could be tuned by adjusting the content of GlcN used during cryogel preparation. In general, incorporation of GlcN did not influence scaffold pore size and ultimate compressive strain but increased porosity. The GH-GlcN16 cryogel showed the highest swelling ratio and degradation rate in hyaluronidase and collagenase solutions. On the contrary, the Young’s modulus, storage modulus, ultimate compressive stress, energy dissipation level, and rate of stress relaxation decreased by increasing the GlcN content in the cryogel. The release of GlcN from the scaffolds in the culture medium of chondrocytes could be sustained for 21 days for GH-GlcN16 in contrast to only 7 days for GH-GlcN9. In vitro cell culture experiments using rabbit articular chondrocytes revealed that GlcN incorporation affected cell proliferation, morphology, and maintenance of chondrogenic phenotype. Overall, GH-GlcN16 showed the best performance in maintaining chondrogenic phenotype with reduced cell proliferation rate but enhanced glycosaminoglycans (GAGs) and type II collagen (COL II) secretion. Quantitative real-time polymerase chain reaction also showed time-dependent up-regulation of cartilage-specific marker genes (COL II, aggrecan and Sox9) for GH-GlcN16. Implantation of chondrocytes/GH-GlcN16 constructs into full-thickness articular cartilage defects of rabbits could regenerate neocartilage with positive staining for GAGs and COL II. The GH-GlcN16 cryogel will be suitable as a scaffold for the treatment of articular cartilage defects.

The effects of different doses of IGF-1 on cartilage and subchondral bone during the repair of full-thickness articular cartilage defects in rabbits

To investigate the effects of different doses of insulin-like growth factor 1 (IGF-1) on the cartilage layer and subchondral bone (SB) during repair of full-thickness articular cartilage (AC) defects. IGF-1-loaded collagen membrane was implanted into full-thickness AC defects in rabbits. The effects of two different doses of IGF-1 on cartilage layer and SB adjacent to the defect, the cartilage structure, formation and integration, and the new SB formation were evaluated at the 1st, 4th and 8th week postoperation. Meanwhile, after 1 week treatment, the relative mRNA expressions in tissues adjacent to the defect, including cartilage and SB were determined by quantitative real-time RT-PCR (qRT-PCR), respectively. Different doses of IGF-1 induced different gene expression profiles in tissues adjacent to the defect and resulted in different repair outcomes. Particularly, at high dose IGF-1 aided cell survival, regulated the gene expressions in cartilage layer adjacent defect and altered ECM composition more effectively, improved the formation and integrity of neo-cartilage. While, at low dose IGF-1 regulated the gene expressions in SB more efficaciously and subsequently promoted the SB remodeling and reconstruction. Different doses of IGF-1 induced different responses of cartilage or SB during the repair of full-thickness AC defects. Particularly, high dose of IGF-1 was more beneficial to the neo-cartilage formation and integration, while low dose of it was more effective for the SB formation.

Repair of rabbit cartilage defect based on the fusion of rabbit bone marrow stromal cells and Nano-HA/PLLA composite material

Objective To assess the effect of the fusion of rabbit bone marrow stromal cells (rBMSCs) and Nano-hydroxyapatite/poly (l-lactic acid) (Nano-HA/PLLA) in repairing the rabbit knee joint with full-thickness cartilage defect. Method The rBMSCs were isolated and cultured in vitro, and the third generation of rBMSCs was co-cultured with the Nano-HA/PLLA to construct the tissue-engineered cartilage (TEC). Eighteen New Zealand white rabbits were selected and randomly divided into three groups, namely, TEC group, Nano-HA/PLLA group, and control group. A cartilage defect model with the diameter of 4.5 mm and depth of 5 mm was constructed on the articular surface of medial malleolus of rabbit femur. General observation, histological observation, and Wakitani’s histological scoring were conducted in the 12th and 24th week postoperatively. Results The results of TEC group indicated that new cartilage tissue was formed on the defect site and subchondral bone achieved physiological integration basically. Histological and immunohistochemical analyses indicated the generation of massive extracellular matrix. In contrast, limited regeneration and reconstruction of cartilage was achieved in the Nano-HA/PLLA group and control group, with a significant difference from the TEC group (p < 0.05). Moreover, the effect of cartilage repair was positively correlated with time. Conclusion The porous Nano-HA/PLLA combined with BMSCs promoted the repair of weight-bearing bone of adult rabbit’s knee joint with cartilage defect.

Collagen/silk fibroin composite scaffold incorporated with PLGA microsphere for cartilage repair

For cartilage repair, ideal scaffolds should mimic natural extracellular matrix (ECM) exhibiting excellent characteristics, such as biocompatibility, suitable porosity, and good cell affinity. This study aimed to prepare a collagen/silk fibroin composite scaffold incorporated with poly-lactic-co-glycolic acid (PLGA) microsphere that can be applied in repairing cartilage. To obtain optimum conditions for manufacturing a composite scaffold, a scaffold composed of different collagen-to-silk fibroin ratios was evaluated by determining porosity, water absorption, loss rate in hot water, and cell proliferation. Results suggested that the optimal ratio of collagen and silk fibroin composite scaffold was 7:3. The microstructure and morphological characteristics of the obtained scaffold were also examined through scanning electron microscopy and Fourier transform infrared spectroscopy. The results of in vitro fluorescence staining of bone marrow stromal cells revealed that collagen/silk fibroin composite scaffold enhanced cell proliferation without eliciting side effects. The prepared composite scaffold incorporated with PLGA microsphere was implanted in fully thick articular cartilage defects in rabbits. Collagen/silk fibroin composite scaffold with PLGA microspheres could enhance articular cartilage regeneration and integration between the repaired cartilage and the surrounding cartilage. Therefore, this composite will be a promising material for cartilage repair and regeneration.

Cell-free macro-porous fibrin scaffolds for in situ inductive regeneration of full-thickness cartilage defects.

Macro-porous fibrin scaffolds with regular and adjustable inter-connective pores were fabricated through a porogen-leaching method for the in situ inductive regeneration of full-thickness cartilage defects in vivo. In vitro tests proved the survival and proliferation of BMSCs in the scaffolds. In vivo repair experiment was conducted by implantation of the cell-free macro-porous fibrin scaffolds into full-thickness cartilage defects (4 mm in diameter and 4 mm in depth with bone marrow blood effusion) of New Zealand white rabbits for 6 and 12 w. The neo-cartilage integrated well with the surrounding cartilage as well as the subchondral bone. Immunochemical and GAG staining revealed the abundant deposition of type II collagen and GAGs in the neo-cartilage after regeneration for 12 w. Histological score of the regenerated tissues was 2.6. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blotting (WB) revealed that the cartilage-related genes and proteins were significantly up-regulated compared with those of the normal cartilage. With the cell-free advantage and positive restoration of full-thickness cartilage defect in vivo, the fibrin scaffold is shelf-ready and is expected to be conveniently used in clinics.

Effects of cartilage-derived morphogenetic protein 1 (CDMP1) transgenic mesenchymal stem cell sheets in repairing rabbit cartilage defects.

The aim of this study was to investigate the abilities of cartilage-derived morphogenetic protein 1 (CDMP1) transgenic cell sheets in repairing rabbit cartilage defects. Rabbit CDMP1 transgenic bone marrow mesenchymal stem cell (BMSC) sheets (CDMP1-BMSCs) were cultured on temperature-sensitive culture dishes, and CDMP1 expression and type II collagen protein in the cell sheets were detected. Tissue-engineered cell sheets were constructed and transplanted into defect rabbit thyroid cartilage, to investigate the expression of engineered cartilage collagen protein and proteoglycan (GAG). The experiment was divided into three groups; A) BMSC sheet, B) Ad-CMV-eGFP-transfected cell sheet, and C) Ad-CMV-hCDMP1-IRES-eGFP-transfected cell sheet. The expression of CDMP1 was detected in the transgenic cell sheets. The engineered cartilage exhibited positive immunohistochemical and Alcian blue staining. The expression levels of type II collagen protein and GAG in group A were positive, whereas those in group B and group C were negative (P < 0.05). The CDMP1-BMSC sheets had a good cartilage differentiation activity, and could effectively repair rabbit laryngeal cartilage defects.

Repair of articular cartilage defects in rabbits through tissue-engineered cartilage constructed with chitosan hydrogel and chondrocytes.

In our previous work, we prepared a type of chitosan hydrogel with excellent biocompatibility. In this study, tissue-engineered cartilage constructed with this chitosan hydrogel and costal chondrocytes was used to repair the articular cartilage defects. Chitosan hydrogels were prepared with a crosslinker formed by combining 1,6-diisocyanatohexane and polyethylene glycol. Chitosan hydrogel scaffold was seeded with rabbit chondrocytes that had been cultured for one week in vitro to form the preliminary tissue-engineered cartilage. This preliminary tissue-engineered cartilage was then transplanted into the defective rabbit articular cartilage. There were three treatment groups: the experimental group received preliminary tissue-engineered cartilage; the blank group received pure chitosan hydrogels; and, the control group had received no implantation. The knee joints were harvested at predetermined time. The repaired cartilage was analyzed through gross morphology, histologically and immunohistochemically. The repairs were scored according to the international cartilage repair society (ICRS) standard. The gross morphology results suggested that the defects were repaired completely in the experimental group after twelve weeks. The regenerated tissue connected closely with subchondral bone and the boundary with normal tissue was fuzzy. The cartilage lacuna in the regenerated tissue was similar to normal cartilage lacuna. The results of ICRS gross and histological grading showed that there were significant differences among the three groups (P<0.05). Chondrocytes implanted in the scaffold can adhere, proliferate, and secrete extracellular matrix. The novel tissue-engineered cartilage constructed in our research can completely repair the structure of damaged articular cartilage.