Rabbit Cartilage Research Articles

Therapeutic Effects of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Combined with Cartilage Acellular Matrix Mediated Via Bone Morphogenic Protein 6 in a Rabbit Model of Articular Cruciate Ligament Transection.

Osteoarthritis (OA) is a general joint disease. Cartilage damage is associated with a decrease in the density of chondrocytes. Mesenchymal stem cells (MSCs) differentiate into adipocytes, osteocytes and chondrocytes, and are an excellent source of cell therapy. Cartilage-derived extracellular matrix (ECM) promotes chondrogenesis of MSCs. However, the role of MSCs stimulated by ECM is not well known in OA. The purpose of this study is to determine the role of specific factors generated by the application of ECM and umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) in managing OA symptoms. Cartilage acellular matrix (CAM), which is a cartilage-derived ECM, was used to promote the chondrogenesis of UCB-MSCs. Induced MSCs were analyzed using chondrogenic markers (aggrecan, collagen type 2, and SOX9) and bone morphogenic protein 6 (BMP6). BMP6 is known to be involved in early chondrogenesis of MSCs. As a result, treatment with CAM significantly increased the expression of chondrogenic markers and BMP6 in UCB-MSCs. Treatment with recombinant human BMP6 also dramatically increased the levels of chondrogenic markers in UCB-MSCs. In addition, UCB-MSCs and CAM were used to evaluate OA symptom improvement in a rabbit articular cruciate ligament transection (ACLT) model. Application of UCB-MSCs and CAM enhanced not only the structure and synthesis of proteoglycan and collagen type 2 but also anti-inflammatory effects in both rabbit joint and synovial fluid. Moreover, the detection of human cells and involvement of BMP6 were confirmed in rabbit cartilage tissues. This study indicates that therapeutic potential of UCB-MSCs with CAM is mediated via BMP6 in OA.

REMODELING OF THE TEMPROMANDIBULAR JOINT FOLLOWING DIFFERENT RATES OF MANDIBULAR DISTRACTION OSTEOGENESIS (AN EXPERIMENTAL STUDY)

Introduction: patients with craniofacial deformities resulting from trauma, infection, tumors and congenital defects usually require skeletal expansion. Tradition treatment methods, involve autogenous bone grafting, which often results in problems including graft resorption, infection and donor site morbidity. Distraction osteogenesis has been adopted more and can help overcome these problems. Aim of the work: the purpose of this study was to compare histologically the effect of bilateral distraction osteogenesis of the mandibular body at different rates on the integrity of the condylar cartilage in rabbits. Material and Methods: This study was performed on 2 groups of twelve healthy rabbits (adult male) : (group 1 and group 2) each group (n=6 rabbits, and 12 joints). Two rabbits (4 joints) in each subgroup according to the post distraction period (1, 2, and 4 weeks). In group 1, the distraction was performed daily at rate of 1 mm for 5 days. In group 2, the distraction was performed twice daily at rate of 1.5 mm (3 mm per day) for 5 days.Results: Animals in both groups (1 and 2) showed changes in condylar head, which seemed to be reversible according to the post distraction period. After the first week, condylar layers were decreased in thickness with irregular fibrous outline, bone trabeculae were decreased in number and invaded by large number of osteoclasts. After the second week, the condylar head showed evidence of regeneration of all layers including the disc. After the fourth week, the subchondral area started to be refilled with new bone with numerous osteoblast. Group 2 (3 mm per day) showed more bone and collagen formation.Conclusion: this experimental study reported that distraction rates of 1 mm per day and 3 mm per day showed degenerative change in the TMJ condylar cartilage in the first post distraction week. At the second week, all the condyles showed adaptive and remodeling signs. Regeneration to the normal histological structure appeared in the fourth post distraction weeks, in group 2 (3 mm per day) more than in group 1 (1 mm per day).

A decellularized scaffold derived from squid cranial cartilage for use in cartilage tissue engineering.

Decellularized cartilage scaffold (DCS) is an emerging substitute for cartilage defect application. However, it is hard to preserve an intact structure of such scaffolds derived from terrestrial animals, because of their dense and compact constitution. In contrast, squid (Dosidicus gigas) cranial cartilage, which possesses a relatively loose structure, could be easily decellularized using mild conditions and it retains the original microstructures of extracellular matrix (ECM). Herein, decellularized squid cranial cartilage scaffold (DSCS) was fabricated successfully after substantially removing the cells, which contained abundant ECM components (proteoglycans and type II collagen). Microscopic structure results showed that the DSCS possesses a relatively smooth and dense surface with a favorable interconnected inner porous structure for cell growth. DSCS exhibited excellent biomechanics and hydrophilicity. In vitro experiments indicated that the scaffold extracts were not toxic to cells, and were amenable to chondrocyte migration. Meanwhile, chondrocytes seeded in DSCS could maintain a favorable viability and present a spreading morphology. Furthermore, the in vivo experiments revealed that both cell-free scaffold and cell-laden scaffold exert promotive effects for the regeneration of cartilage in a full-thickness rabbit cartilage defect model. Taken together, these results suggested that DSCS presents a novel and promising cell-free therapeutic choice for cartilage tissue engineering.

Quantitative µMRI and PLM study of rabbit humeral and femoral head cartilage at sub-10 µm resolutions.

This study aimed to establish the baseline characteristics in humeral and femoral cartilage in rabbit, using quantitative magnetic resonance imaging (MRI) relaxation times (T2, T1ρ, and T1) at 9.75 and 70-82 µm pixel resolutions, and quantitative polarized light microscopy (PLM) measures (retardation, angle) at 1.0 and 4.0 µm pixel resolutions. Five intact (i.e., unopened) shoulder joints (the scapula and humeral heads) and three femoral heads of the hip joints from five healthy rabbits were imaged in MRI at 70-82 µm resolution. Thirteen cartilage-bone specimens were harvested from these joints and imaged in µMRI at 9.75 µm resolution. Subsequently, quantitative PLM study of these specimens enabled the examination of the fibril orientation and organization in both intact joints and individual specimens. Quantitative MRI relaxation data and PLM fibril structural data show distinct features in tissue properties at different depths of cartilage, different in individual histological zones. The thicknesses of the histological zones in µMRI and PLM were successfully obtained. This is the first correlated and quantitative MRI and PLM study of rabbit cartilage at sub-10 µm resolutions, which benefits future investigation of osteoarthritis using the rabbit model. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:1052-1062, 2020.

HMGB2 is associated with pressure loading in chondrocytes of temporomandibular joint: In vitro and in vivo study

High mobility group protein B2 (HMGB2) is an important protein attributed to tissue function and homeostasis in osteoarthritis (OA), however, its roles in the temporomandibular joint with mechanical pressure are unclear. This study investigated the expression patterns of HMGB2 in chondrocytes of TMJ on OA-related cellular and animal models. The transcriptional level and protein expression of HMGB2 as well as Col- II, MMP-13 and β-catenin were examined in primary cultured chondrocytes from TMJs with hydrostatic pressures (HP). The immunohistochemistry of HMGB2, Col- II, MMP13 and β-catenin in rabbits with surgical anterior disc displacement (ADD) were analyzed. The results indicated that HP decreased the mRNA expression of HMGB2, MMP-13, and β-catenin. HMGB2 knockdown attenuated the sensitivity of chondrocytes response to pressure loading. Immunohistochemical results showed that the rabbits with ADD exhibited thinner condylar cartilage with disordered cell arrangement. The distribution of HMGB2 was chiefly in the fibrous layer and the proliferative layer of the condylar cartilage. In rabbit cartilage with ADD, the expression of HMGB2 and β-catenin were elevated at 1 week followed by sustained decrease at 2 and 4 weeks. Our findings suggested that HMGB2 is necessary for TMJ chondrocytes to sense pressure loading, which plays an important role in the pathogenesis of chondrogenesis and TMJOA following mechanical loading.

Dasatinib Promotes Chondrogenic Differentiation of Human Mesenchymal Stem Cells via the Src/Hippo-YAP Signaling Pathway.

Mesenchymal stem cells (MSCs) are progenitors of chondrocytes and could be used as a potential therapy for cartilage defects in diarthrodial joints. However, promoting chondrogenic differentiation of MSCs remains a daunting challenge. As a small molecular drug, dasatinib can promote MSC differentiation, although the exact mechanisms of chondrogenic differentiation are unclear. In this study, the differentiation of MSCs into osteoblasts, adipocytes, and chondrocytes was assessed by the protein and mRNA levels of osteoblast- and chondrocyte-related proteins using western blotting and real-time polymerase chain reaction, respectively. MSCs were induced to differentiate into chondrocytes or osteoblasts with or without dasatinib in vitro. The effects of dasatinib on cartilage regeneration were also assessed in vivo in a rabbit model of full-thickness cartilage defects using methacrylate gelatin hydrogel as scaffolds. Dasatinib promoted chondrogenic differentiation and inhibited osteogenic differentiation of MSCs. Furthermore, dasatinib significantly inhibited the expression of YAP and TAZ and the phosphorylation of Src, but it enhanced serine phosphorylation of YAP during the chondrogenic differentiation of MSCs in vitro. Inhibition of the Hippo pathway using XMU-MP-1 dramatically suppressed the serine phosphorylation of YAP and chondrogenic differentiation of MSCs. Moreover, we confirmed that the sustained release of dasatinib from the hydrogel promoted rabbit cartilage repair. The results demonstrated that dasatinib might promote chondrogenic differentiation of MSCs via the Src/Hippo-YAP signaling pathway and that hydrogel sustained-release dasatinib had a certain effect on the repair of cartilage defects.

Glucosamine-grafted methacrylated gelatin hydrogels as potential biomaterials for cartilage repair.

Glucosamine (GlcN) has been widely used to reduce joint pain and osteoarthritis progression, but the efficacy of GlcN remains controversial because of the low GlcN concentration reaching the articular cavity. The aim of this study is to provide an effective approach of GlcN delivery to a target site using photocrosslinkable methacrylated gelatin (GelMA)-based hydrogels, where GlcN could be gradually released during the degradation of the GelMA hydrogel. Herein, GlcN was acrylated as the acryloyl glucosamine (AGA) and covalently grafted to GelMA, and more than 87.7% of 15% (w/v) GelMA hydrogel was grafted with AGA. Moreover, in vitro outgrowth and apoptosis assay of bone marrow stem cells (BMSCs) demonstrated that the GelMA-AGA hydrogels had better biocompatibility, larger cell attachment, and higher cell viability than pure GlcN and AGA materials. Also, 15% (w/v) GelMA-AGA hydrogel was injected into the defect site for in vivo rabbit cartilage repair. Compared with oral administration of pure GlcN and injection of pure GelMA, the repaired cartilages using GelMA-AGA hydrogels had the smoothest surface of the defect site, filling more than 95% defect bulk. The similar content of glycosaminoglycans to the native tissue and the maximum amount of type II collagen was found in the repaired cartilage using GelMA-AGA hydrogels, indicating the outgrowth of hyaline cartilage.

Hyperbaric oxygen inhibits the HMGB1/RAGE signaling pathway by upregulating Mir-107 expression in human osteoarthritic chondrocytes

MicroRNA (miRNA)107 expression is downregulated but high mobility group box 1 (HMGB-1), Toll-like receptors (TLRs), and receptor for advanced glycation end products (RAGE) are upregulated in osteoarthritic (OA) cartilage. We investigated mir-107/HMGB-1 signaling in OA after hyperbaric oxygen (HBO) treatment. MiR-107 mimic was transfected and the HMGB-1 was analyzed in OA chondrocytes. MiRNA targets were identified using bioinformatics and a luciferase reporter assay. After HBO treatment, the mRNA or protein levels of HMGB-1, RAGE, TLR2, TLR4, and inducible nitric oxide (NO) synthase (iNOS) and phosphorylation of mitogen-activated protein kinase (MAPK) were evaluated. The secreted HMGB-1 and matrix metalloproteases (MMPs) levels were quantified. Finally, we detected the HMGB-1 and iNOS expression in rabbit cartilage defects. Overexpression of miR-107 suppressed HMGB-1 expression in OA chondrocytes. The 3'UTR of HMGB-1 mRNA contained a 'seed-matched-sequence' for miR-107. MiR-107 was induced by HBO and a marked suppression of HMGB-1 was observed simultaneously in OA chondrocytes. Knockdown of miR-107 upregulated HMGB-1 expression in hyperoxic cells. HBO downregulated the mRNA and protein expression of HMGB-1, RAGE, TLR2, TLR4, and iNOS, and the secretion of HMGB-1. HBO decreased the nuclear translocation of nuclear factor (NF)-κB, downregulated the phosphorylation of MAPK, and significantly decreased the secretion of MMPs. Morphological and immunohistochemical observation demonstrated that HBO markedly enhanced cartilage repair and the area stained positive for HMGB-1 and iNOS tended to be lower in the HBO group. HBO inhibits HMGB-1/RAGE signaling related pathways by upregulating miR-107 expression in human OA chondrocytes.

PNNS-Conjugated Chitosan Mediated IGF-1 and miR-140 Overexpression in Articular Chondrocytes Improves Cartilage Repair

The aim of the present study was to investigate the effects of phosphorylatable nucleus localization signal linked nucleic kinase substrate short peptide (pNNS)-conjugated chitosan (pNNS-CS) mediated miR-140 and IGF-1 in both rabbit chondrocytes and cartilage defects model. pNNS-CS was combined with pBudCE4.1-IGF-1, pBudCE4.1-miR-140, and negative control pBudCE4.1 to form pDNA/pNNS-CS complexes. Then these complexes were transfected into chondrocytes or injected intra-articularly into the knee joints. High levels of IGF-1 and miR-140 expression were detected both in vitro and in vivo. Compared with pBudCE4.1 group, in vitro, the transgenic groups significantly promoted chondrocyte proliferation, increased glycosaminoglycan (GAG) synthesis, and ACAN, COL2A1, and TIMP-1 levels, and reduced the levels of nitric oxide (NO), MMP-13, and ADAMTS-5. In vivo, the exogenous genes enhanced COL2A1, ACAN, and TIMP-1 expression in cartilage and reduced cartilage Mankin score and the contents of NO, IL-1β, TNF-α, and GAG contents in synovial fluid of rabbits, MMP-13, ADAMTS-5, COL1A2, and COL10A1 levels in cartilage. Double gene combination showed better results than single gene. This study indicate that pNNS-CS is a better gene delivery vehicle in gene therapy for cartilage defects and that miR-140 combination IGF-1 transfection has better biologic effects on cartilage defects.

3D-bioprinting a genetically inspired cartilage scaffold with GDF5-conjugated BMSC-laden hydrogel and polymer for cartilage repair.

Rationale: Articular cartilage injury is extremely common in congenital joint dysplasia patients. Genetic studies have identified Growth differentiation factor 5 (GDF5) as a shared gene in joint dysplasia and OA progression across different populations. However, few studies have employed GDF5 in biological regeneration for articular cartilage repair.Methods & Results: In the present study, we report identified genetic association between GDF5 loci and hip joint dysplasia with genome-wide association study (GWAS). GWAS and replication studies in separate populations achieved significant signals for GDF5 loci. GDF5 expression was dysregulated with allelic differences in hip cartilage of DDH and upregulated in the repaired cartilage in a rabbit cartilage defect model. GDF5 in vitro enhanced chondrogenesis and migration of bone marrow stem cells (BMSCs), GDF5 was tested in ectopic cartilage generation with BMSCs by GDF5 in nude mice in vivo. Genetically inspired, we further generated functional knee articular cartilage construct for cartilage repair by 3d-bioprinting a GDF5-conjugated BMSC-laden scaffold. GDF5-conjugated scaffold showed better cartilage repairing effects compared to control. Meanwhile, transplantation of the 3D-bioprinted GDF5-conjugated BMSC-laden scaffold in rabbit knees conferred long-term chondroprotection.Conclusions: In conclusion, we report identified genetic association between GDF5 and DDH with combined GWAS and replications, which further inspired us to generate a ready-to-implant GDF5-conjugated BMSC-laden scaffold with one-step 3d-bioprinting for cartilage repair.

Codelivery of Synovium-Derived Mesenchymal Stem Cells and TGF-β by a Hybrid Scaffold for Cartilage Regeneration.

Synovium-derived mesenchymal stem cells (SMSCs) are attractive tissue-specific cells for cartilage regeneration because of their easy availability, higher chondrogenic potential, and joint specificity. In the present study, we established a hybrid scaffold to codeliver SMSCs and transforming growth factor beta (TGF-β), which can integrate the scaffolds, the growth factor, and the autogenous cells within rabbit cartilage defects. A chitosan hydrogel and a decellularized bone matrix were used to fabricate the gel-solid duplex phase biomaterials, which were proven to retain more cells, promote tissue integration, and provide mechanical support. In vitro experiments demonstrated that this hybrid scaffold could release TGF-β in a controlled biphasic pattern, which can promote cell proliferation and chondrogenic differentiation of loaded rabbit SMSCs. For in vivo experiments, we filled cartilage defects with the hybrid materials, delivering autogenous SMSCs and TGF-β simultaneously via chitosan's sol-gel transition. Histological analysis, magnetic resonance imaging, and nanoindentation assessment indicated superior cartilage regeneration using this codelivery system compared with that from routine microfracture or control delivery scaffolds. Beyond cartilage regeneration, the easy preparation, favorable biophysical properties, and controlled release ability make this codelivery system applicable to transport other tissue-specific cells or biofactors for tissue engineering.

IGF-1 and BMP-7 synergistically stimulate articular cartilage repairing in the rabbit knees by improving chondrogenic differentiation of bone-marrow mesenchymal stem cells.

Knee injury is known as a frequently occurred damage related to sports, which may affect the function of cartilage. This study aims to explore whether Insulin-like growth factor 1 (IGF-1) and bone morphogenetic protein-7 (BMP-7)-modified bone-marrow mesenchymal stem cells (BMSCs) affect the repair of cartilage damage found in the knee. Primarily, BMSCs were treated with a series of pEGFP-C1, IGF-1, and BMP-7, followed by determination of IGF-1 and BMP-7 expression. A rabbit cartilage defect model was also established. Afterfward, cell morphology, viability, cartilage damage repair effect, and expression of collagen I and collagen II at the 6th and the 12th week were measured. BMSCs treated with pEGFP-C1/IGF-1, pEGFP-C1/BMP-7, and pEGFP-C1/BMP-7-IGF-1 exhibited elevated expression of BMP-7 and IGF-1. Besides, BMSCs in the P10 generation displayed decreased cell proliferation. Moreover, BMSCs treated with IGF-1, BMP-7, and IGF-1-BMP-7 showed reduced histological score and collagen I expression while elevated collagen II expression, as well as better repair effect, especially in those treated with IGF-1-BMP-7. Collectively, these results demonstrated a synergistic effect of IGF-1 and BMP-7 on the BMSC chondrogenic differentiation on the articular cartilage damage repair in the rabbit knees, highlighting its therapeutic potential for the treatment of articular cartilage damage.

Study on the protective mechanism of autophagy on cartilage by magnesium sulfate

To investigate the mechanism of magnesium sulfate in protecting rabbit cartilage by initiating autophagy. Twenty-four adult female New Zealand rabbits were used to prepare post-traumatic osteoarthritis (PTOA) models by anterior cruciate ligament transection. Then, the PTOA models were randomly divided into PTOA group, distilled water group, and magnesium sulfate group, with 8 rabbits in each group. Immediately after operation, the distilled water group and the magnesium sulfate group were injected with 0.5 mL distilled water and 20 mmol/L magnesium sulfate solution in the joint cavity 3 times a week for 4 weeks, respectively. The PTOA group was not treated. The general condition of the animals was observed after operation. After 4 weeks, the expressions of tumor necrosis factor α (TNF-α) and collagen typeⅡ in the joint fluid and the expression of collagen type Ⅱ in venous blood were detected by ELISA assay. The protein expressions of transient receptor potential channel vanilloid 5 (TRPV5) and microtubule associated protein 1 light chain 3 (LC3; LC3-Ⅱ/LC3-Ⅰ) in femoral cartilage were detected by Western blot. The mRNA expressions of interleukin 1β (IL-1β), TNF-α, matrix metalloproteinases 3 (MMP-3) in synovial tissue and collagen type Ⅱ, Aggrecan (AGN), SOX9 in cartilage tissue were detected by real-time fluorescence quantitative PCR. Cartilage tissue sections were stained with HE staining, Masson staining, and Alcian blue staining and scored according to the modified histological osteoarthritis (OA) score. All animals survived until the experiment was completed. Compared with the other two groups, the expression of TNF-α in joint effusion and collagen type Ⅱ in joint effusion and venous blood were decreased in magnesium sulfate group; the protein expression of TRPV5 decreased, and the ratio of LC3-Ⅱ/LC3-Ⅰ increased significantly; the mRNA expressions of IL-1β, TNF-α, and MMP-3 in synovial tissue were decreased, and the mRNA expressions of collagen type Ⅱ, AGN, and SOX9 in cartilage tissue were increased; OA scores also decreased significantly. All differences were statistically significant ( P<0.05). There was no significant difference in the above indicators between the PTOA group and the distilled water group ( P>0.05). Intra-articular injection of magnesium sulfate can reduce intra-articular inflammation, reduce the loss of collagen type Ⅱ and AGN, and is beneficial to cartilage regeneration in rabbits. The mechanism may be related to the initiation of chondroautophagy by inhibiting the calcium channel TRPV5.

Silk fibroin/collagen/hyaluronic acid scaffold incorporating pilose antler polypeptides microspheres for cartilage tissue engineering

A silk fibroin/collagen/hyaluronic acid (SF/COL/HA) composite scaffold was prepared via admixing, crosslinking, and lyophilizing processes. We studied its physicochemical and biological properties, such as water absorption, porosity, weight loss, and biocompatibility. The optimal ratio of SF/COL/HA scaffold was 3:6.5:0.5. Then, the optimal ratio of scaffold incorporating pilose antler polypeptides (PAPs)–PLGA microspheres was prepared, and their compatibility was studied. PAP-SF/COL/HA scaffold had favorable adhesion and proliferation. A rabbit cartilage defect model was established. The repair effect of cartilage defects was observed and evaluated among PAP-SF/COL/HA, SF/COL/HA, and sham operation groups. The defects were almost completely repaired after 13 weeks in the PAP-SF/COL/HA group, thereby indicating that the PAP-SF/COL/HA composite had a favorable effect on articular cartilage repair.

Effects of press-fit biphasic (collagen and HA/βTCP) scaffold with cell-based therapy on cartilage and subchondral bone repair knee defect in rabbits.

Human spontaneous osteonecrosis of the knee (SPONK) is still challenging as the current treatments do not allow the production of hyaline cartilage tissue. The aim of the present study was to explore the therapeutic potential of cartilage regeneration using a new biphasic scaffold (type I collagen/hydroxyapatite) previously loaded or not with concentrated bone marrow cells. Female rabbits were operated of one knee to create articular lesions of the trochlea (three holes of 4 × 4mm). The holes were left empty in the control group or were filled with the scaffold alone or the scaffold previously loaded with concentrated bone marrow cells. After twomonths, rabbits were sacrificed and the structure of the newly formed tissues were evaluated by macroscopic, MRI, and immunohistochemistry analyses. Macroscopic and MRI evaluation of the knees did not show differences between the three groups (p > 0.05). However, histological analysis demonstrated that a higher O'Driscoll score was obtained in the two groups treated with the scaffold, as compared to the control group (p < 0.05). The number of cells in treated area was higher in scaffold groups compared to the control group (p < 0.05). There was no difference for intensity of collagen type II between the groups (p > 0.05) but subchondral bone repair was significantly thicker in scaffold-treated groups than in the control group (1mm for the control group vs 2.1 and 2.6mm for scaffold groups). Furthermore, we observed that scaffolds previously loaded with concentrated bone marrow were more reabsorbed (p < 0.05). The use of a biphasic scaffold previously loaded with concentrated bone marrow significantly improves cartilage lesion healing.

Expression of endoplasmic reticulum stress protein in rabbit condyle cartilage following anterior disk displacement.

Anterior disk displacement (ADD) is a most common subtype of temporomandibular disorders (TMD), which is promoted by chondrocytes apoptosis. However, the signaling pathways that trigger apoptosis are still unknown. The aim of this study was to investigate the expression of endoplasmic reticulum (ER) stress-related proteins in the condylar cartilage of rabbits following ADD. Sixty healthy adult rabbits were randomly assigned to the experimental and sham-operated control groups (n=12). The experimental rabbits were subjected to surgical ADD in the right temporomandibular joints. The production of ER stress-related proteins C/EBP homologous protein (CHOP), glucose-regulated protein 78 (GRP78), cleaved caspase-3, and caspase-12 in cartilage was evaluated by immunohistochemistry, quantitative real-time PCR, and Western blot analysis. Our results showed that the expression of CHOP, GRP78, cleaved caspase-3, and caspase-12 increased significantly along with degenerative changes in cartilage after ADD. These results indicate that the ER stress pathway is activated in ADD cartilage and might promote the development of TMD.

Calcium Phosphate Bioceramic Material Local Influence on the Bone Biomechanical Properties at Rabbits with Experimental Osteoporosis

Biomechanical characteristics of the rabbit cartilage component from the femur were investigated. The control osteoporosis had been activated by ovariectomy alongside the injections of methylprednisolone. During the experiment defects in the femur’s greater trochanter zone was created and afterwards filled with the fragments of a hydroxyapatite as well as tricalcium phosphate (HAP/TCP 70/30) or HAP/TCP 70/30 mixed with five percent strontium. Three months later animals were euthanazed, squared samples were cut out from the femur’s body and later tested on a bend. The results of the research revealed that the femurs become more stringent after filling off defects in the greater trochanter zone with HAP/TCP 70/30 or HAP/TCP 70/30 mixed with five percent strontium. The ultimate pain for the cartilage component in the control groups was less, than in the test group. The flexure modulus of flexibility of a cartilage component in the test group was statistically reliable less, than the value of the flexure modulus of flexibility of the cartilage component in the control groups. Therefore, local usage of calcium – phosphatic bioceramic component over the greater trochanter part increases biomechanical characteristics of the cartilage component in the femur of animals.

Effects of fibroblast growth factors 2 and low intensity pulsed ultrasound on the repair of knee articular cartilage in rabbits.

To investigate the effect of low intensity ultrasound irradiation combine with fibroblast growth factors (FGF2) on the repair of the knee articular cartilage and to explore its mechanism in rabbit. The model of the rabbit knee joint injury was established. 40 rabbits were divided into four groups, including control group, model group, FGF2 group and FGF2 + low intensity pulsed ultrasound group (FGF2 + LIPU). The knee joints of rabbits were taken at 4 and 8 weeks, respectively. Histopathological changes were detected by Hematoxylin and Eosin stain (HE) and evaluated by Wakitani score. The expression of FGF2 mRNA was detected by Real-time polymerase chain reaction (RT-PCR) and the levels of Collagen I and Collagen II protein were analyzed by Western blotting. In FGF2 group and FGF2 + LIPU group, it was found that the tissues of knee joint were gradually repaired following the change of time. Further, the recovery was better in FGF2 + LIPU group. Cartilage defect areas were filled with cartilage-like cells and the repair surface was fused with surrounding cartilage in FGF2 and FGF2 + LIPU groups. Wakitani scores were consistent with HE results. The expressions of FGF2 mRNA were higher in FGF2 and FGF2 + LIPU group than the model group. Western blotting results showed that the levels of Collagen I and Collagen II protein in FGF2 and FGF2 + LIPU groups were significantly increased compared with that in model group. FGF2 and LIPU combined application on the rabbit knees joint repair is better than FGF2 alone. FGF2 and LIPU combination can promote the synthesis and secretion of collagen in chondrocytes, promote the differentiation and maturation of chondrocytes during the repair of cartilage defects.

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.

ADIPOCHONDROCYTES IN RABBIT AURICULAR CARTILAGE

Chondrocytes are described as one cell population in different cartilage types. The auricular cartilage in mouse and rat contains unique chondrocytes similar in morphology to white adipocytes and known as lipochondrocytes. Lipochondrocytes were not mentioned in other species. The current study aimed to explore the existence of this cell type in rabbits. The auricles of adult male white rabbits were harvested and processed for histological examination with light and electron microscopy. With the light microscopy, the auricular cartilage of adult rabbits contained central large rounded adipocyte-like chondrocytes, termed in the current study “adipochondrocytes” The adipochondrocytes were embedded in relatively wide lacunae and had large lipid droplets with a rim of cytoplasm. This result was confirmed by the scanning electron microscopy. With the transmission electron microscopy, the adipochondrocytes showed dark nucleus and electron-dense cytoplasm with few organelles and cytoplasmic processes. The adipochondrocytes of the auricular cartilage in adult rabbits were unique cell type and different from chondrocytes in other cartilage subtypes. This result should be considered during cartilage transplant. Further studies are suggested to investigate the development and physiological roles of adipochondrocytes in the auricular cartilage in rabbits.