Platelet-rich plasma interferes with changes in articular cartilage and synovium of rabbits with knee osteoarthritis / 中国组织工程研究

BACKGROUND: Studies have shown that platelet-rich plasma has a strong role in promoting chondrocyte repair and proliferation. OBJECTIVE: To investigate the efficacy of platelet-rich plasma in chondrocyte repair and synovial inflammation inhibition in osteoarthritis. METHODS: The blood samples were extracted from the central artery of the ear in 40 New Zealand white rabbits and platelet-rich plasma was prepared by the method of Hokugo. The platelet, platelet-derived growth factor, transforming growth factor-3, vascular endothelial growth factor in the peripheral blood and platelet-rich plasma were detected simultaneously. Animal models were made by resection of anterior cruciate ligament and divided into experimental group and control group. In the experimental group, 0.3 mL of platelet-rich plasma was injected once a week for 10 weeks; in the control group, 0.3 mL of sterile saline was injected once a week for 10 weeks. Gross observation and histological observation of knee joints were performed; articular cartilage type II collagen and matrix metalloproteinase 13 levels were measured; and Mankin score of the cartilage tissue was performed at 2, 4, 6, 8, and 10 weeks after injection. The study protocol was approved by the Animal Experiment Ethics Committee of Chongqing Medical University. RESULTS AND CONCLUSION: (1) The concentrations of platelet, platelet-derived growth factor, transforming growth factor-3, vascular endothelial growth factor in platelet-rich plasma were 5.5, 4.8, 7.7, and 6.2 times those of the peripheral blood, respectively. And there were significant differences between the platelet-rich plasma and peripheral blood (P<0.05). (2) From the 6th week, the Mankin score of the experimental group was significantly lower than that of the control group (P<0.05). (3) The expression of type II collagen protein in the experimental group was significantly higher than that in the control group at the 4th, 6th, 8th and 10th weeks after injection (P<0.05). The expression of matrix metalloproteinase-13 was significantly lower than that in the control group at the 2nd, 4th, 6th, 8th and 10th week (P<0.05). All these findings indicate that intra-articular injection of platelet-rich plasma can inhibit the development of osteoarthritis by alleviating synovial inflammation and delaying or blocking the damage of chondrocytes.

Bone marrow mesenchymal stem cells combined with acellular dermal matrix repair beagle dog articular cartilage defects / 中国组织工程研究

BACKGROUND: Dermis-derived extracellular matrix, as a cartilage repair scaffold, provides a space for the growth of cartilage tissue, and promotes cell adhesion and proliferation. Bone marrow mesenchymal stem cells have the potential to differentiate into chondrocytes. Both of them used alone have disadvantages. OBJECTIVE: To evaluate the feasibility of bone marrow mesenchymal stem cells combined with calf acellular dermal matrix to repair beagle dog articular cartilage defects. METHODS: Beagle dog bone marrow blood was extracted from Beagle dogs. Bone marrow mesenchymal stem cells were obtained by density gradient centrifugation and passaged. Acellular dermal matrix was prepared from the dorsal dermis of neonatal calves by ultrasonic oscillation, freeze-drying and pepsin. 0. 2 mL of cell suspension was added to the surface of acellular dermal matrix until covered, then which was placed in a 5% CO2 incubator at 37 °C for 48 hours. Twelve adult beagle dogs were used to establish knee joint cartilage defect models, and then randomized into three groups: In the acellular dermal matrix combined with bone marrow mesenchymal stem cells group (combination group), cartilage defects were repaired with acellular dermal matrix combined with bone marrow mesenchymal stem cells. In the single acellular dermal matrix group, cartilage defects were repaired with acellular dermal matrix. The model control group received no treatment. At 12 weeks after surgery, the right knee joints were observed by stereomicroscope, hematoxylin-eosin staining and type II collagen immunohistochemical staining. RESULTS AND CONCLUSION: (1) Scanning electron microscopy showed that bone marrow mesenchymal stem cells adhered to and grew well in the acellular dermal matrix. (2) Hematoxylin-eosin staining revealed that the repaired surface in the combination group was slightly lower than that of the surrounding normal tissues, and the repaired tissues integrated well with the surrounding cartilages. The defects in the single acellular dermal matrix group were filled with fibrous tissues. Few surrounding tissues of defect were repaired in the model control group. (3) Type II collagen immunohistochemical staining showed that in the combination group, articular cartilage defects were filled with chondrocyte-like tissues. In the single acellular dermal matrix group, the defect was filled with fibrous tissues. No tissue was found in the model control group. (4) These results indicate that the new calf acellular dermal matrix has good biocompatibility and can promote the proliferation of bone marrow mesenchymal stem cells. Autologous bone marrow mesenchymal stem cells combined with acellular dermal matrix can effectively repair beagle dog knee joint cartilage defects.

Intra-articular injection of optimal concentrationof bonemarrowmesenchymal stem cells for treating rabbit cartilage defects / 中国组织工程研究

BACKGROUND: Bone marrow mesenchymal stem cells have been extensively applied in animal experiments and clinical studies. The cell concentration, treatment times and results in each study are different, and there is no standard for optimal cell concentration. OBJECTIVE: To investigate the optimal concentration of bone mesenchymal stem cells injected into articular cavity in the treatment of rabbit cartilage defects. METHODS: Thirty 6-month-old New Zealand white rabbits were selected and randomly divided into control, 1×108, 1×109, 1×1010, and 1×1011/L groups. Cartilage defect models with diameter of 3 mm and depth of 2 mm were established in femoral trochlea in each group. One week after modeling, 1 mL of normal saline was injected into the rabbit’s knee of the control group. The other groups were injected with bone marrow mesenchymal stem cells at corresponding concentrations. After 6 and 12 weeks, gross observation, hematoxylin-eosin staining, Safranin-O-fast green-staining, type I and II collagen staining were performed to assess the cartilage regeneration. RESULTS AND CONCLUSION: In the control group, the defect area was obvious with no cartilage regeneration. The 1×108, 1×109, and 1×1010/L groups showed cartilage regeneration. The repairing effect was increased with the cell concentration increasing. The effect of cartilage repair in the 1×1011/L group was similar to that in the 1×1010/L group (P > 0.05). Therefore, 1×1010/L is the optimal concentration for intra-articular injection of bone marrow mesenchymal stem cells for treating cartilage defects, and higher concentration cannot enhance the repairing effect.

Treatment and Research Progress of Articular cartilage injury in HT3SS / 中国综合临床

Objective Articular cartilage injury is one of the most common orthopedic diseases with high morbidity and morbidity,especially in the elderly. Articular cartilage injury causes degenerative changes of articular cartilage, such as osteoarthritis, which can lead to disability, pain during joint movement and deformation of bone and joint. The prevalence of osteoarthritis accounts for 10% ～12% of the total population in the world. It is a common disease. The prevalence of osteoarthritis has increased to 49. 7% for the elderly aged over 65 years old ( Statistics of the World Health Organization ( who) in 2010 show that with the development of social aging and obesity and other adverse factors,these figures will continue to rise. It is known that osteoarthritis is related to aging,trauma,genetic susceptibility,obesity and inflammation,but the specific cause of osteoarthritis has not been fully identified, which leads to many obstacles in clinical treatment of osteoarthritis. At present,most of the clinical and research work in this field is focused on the restoration of cartilage trauma. In this review, we summarize and discuss the methods of cartilage defect repair,as well as the hot spots and directions of future research work.

Subchondral drilling method combined with gum-bletilla complex to repair articular cartilage defects / 中国中药杂志

Two types(A model and B model) of articular cartilage defect models were prepared by using adult New Zealand white rabbits. A model group was applied by drilling without through subchondral bone, whose right joint was repaired by composite scaffolds made by seed cell, gum-bletilla as well as Pluronic F-127, and left side was blank control. B model group was applied by subchondral drilling method, whose right joint was repaired by using composite scaffolds made by gum-bletilla and Pluronic F-127 without seed cells, and left side was blank control. Autogenous contrast was used in both model types. In addition, another group was applied with B model type rabbits, which was repaired with artificial complex material of Pluronic F-127 in both joint sides. 4, 12 and 24 weeks after operation, the animals were sacrificed and the samples were collected from repaired area for staining with HE, typeⅡcollagen immunohistochemical method, Alcian blue, and toluidine blue, and then were observed with optical microscope. Semi-quantitative scores were graded by referring to Wakitanis histological scoring standard to investigate the histomorphology of repaired tissue. Hyaline cartilage repairing was achieved in both Group A and Group B, with satisfactory results. There were no significant differences on repairing effects for articular cartilage defects between composite scaffolds made by seed cell, gum-bletilla and Pluronic F-127, and the composite scaffolds made by gum-bletilla and Pluronic F-127 without seed cell. Better repairing effects for articular cartilage defects were observed in groups with use of gum-bletilla, indicating that gum-bletilla is a vital part in composite scaffolds material.

Repair of articular cartilage defects by autologous bone mesenchymal stem cells and allogeneic costal chondrocytes in the knee of Wuzhishan miniature pigs / 中南大学学报(医学版)

Objective:To investigate the feasibility of construction of tissue engineered cartilage by co-culture of bone marrow mesenchymal stem cells (BMSCs) and costal chondrocytes (CCs),and to provide theoretical basis and experimental basis for clinical repair of articular cartilage defects by Wuzhishan miniature pig knee cartilage defects with co-cultured cells.Methods:Density gradient centrifugation method was used to isolate BMSCs from Wuzhishan miniature pig.The double enzyme digestion method was used to isolate CCs.The passage 3 generation of BMSCs and passage 2 generation of CCs were randomly divided into 3 groups:a co-culture group of BMSCs∶CCs for 1∶2 (Group A),a simple CCs (Group B),and a simple BMSCs (Group C).The cell growth curve was drawn,and the content of glycosaminoglycan (GAG) of external separation in chondrocytes was determined.The 12 Wuzhishan miniature pigs were randomly divided into a co-culture cells/collagen membrane experimental group,a collagen membrane control group and the blank group.In the co-culture cells/collagen membrane experimental group,the co-cultured cells/collagen membrane were implanted into the cartilage defects of the mandibular condyle;in the collagen membrane control group,only collagen membrane was implanted;while in the blank group,nothing was implanted.Six animals were sacrificed at 8 and 16 weeks after surgery respectively (2 animals in each group).General observation,cartilage histological score and histopathological examination were carried out.Results:The BMSCs and co-culture cells grew well.The biological activity of CCs was good.After 16 weeks of operation,the repair tissues in the co-cultured cells/collagen membrane experimental group showed hyaline cartilage features:smooth,flat,and integrated well with the surrounding cartilage and subchondral bone.The collagen membrane in the collagen membrane control group was fibrously repaired.Repair tissue gross score in the co-culture cells/collagen membrane experimental group was significantly better than that in the collagen membrane control group and the blank group (both P0.05).Conclusion:BMSCs,CCs and co-cultured cells can function as the seed cells for cartilage tissue engineering,and the co-culture cells (BMSCs∶CCs=1∶2) possess more advantages;the short-term effect of co-culture cells with collagen membrane on repairing cartilage defects is satisfied.

The feasibility study of repairment of articular cartilage defects with bone marrow-derived mesenchymal stem cells seeded on acellular amniotic membrane / 中华显微外科杂志

Objective To study the effect of repairment of articular cartilage defects in non weight-bearing area of rabbit with bone marrow-derived mesenchymal stem cells (BMSCs) seeded on human acellular amniotic membrane (HAAM).Methods From July 2012 to March 2013,bone marrow-derived mesenchymal stem cells were isolated and purified from rabbit in vitro.The cells were seeded on human acellular anniotic membrane at the concentration of 1.63 × l05/cm2.From 7 days to 8 days after cultured,the complexes of BMSC and HAAM were examined under electronmicroscope,light microscope and by HE stain.Full thickness empty defects measuring 4 mm in diameter by 3 mm depth were prepared in femoral intercondylar fossa of 24 rabbits.The rabbits were randomized into two groups:group A and group B with 12 each group.The defects of right knees were served as control and the left as experimental group.BMSCs/HAAM composite was cultured and then transplanted into the defect of left knee joint in group A as group BMSCs/HAAM and HAAM into group B as group HAAM.These rabbits were killed at 4 and 12 weeks after surgery in each group and the newly cartilage samples were evaluated grossly,histologically are graded.Results In the 4th and 12th week after the operation,the regenerated tissue were white,soft and smooth.Chondrocytes were found in the tissue In the 12th week,the morphology,distribution and arrangement of the regenerated tissues were similar to normal cartilage in the knees with HAAM-BMSCs transplantation.The regenerated tissues grew to be integrated with the surrounding normal cartilage with obscure boundary between them.Chondrocytes were found in all layer of the tissue,surrounding normal cartilage with obscure boundary between them.In the HAAM transplantation,the rough surface of regenerated tissue sunk obviously and the fibmblasts in all layer were found.While there were no regenerated tissue in the control side.Conclusion BMSCs seeded on HAAM could repair the articular cartilage defects of femoral intercondylar fossa from rabbits.

Experimental study of cartilage defect repair using bone mesenchymal stem cells transfer of BMP-2 and TGF-β3 gene mediated by adenovirus vector combining with deminerized bone matrix in pig / 实用医学杂志

Objective To explore the repair result of full-thickness cartilage defects in diannan small-ear pig by bone mesenchymal stem cells (BMSCs) transferred with both transforming growth factor-β3(TGF-β3) and bone morphogenetic protein-2(BMP-2) gene mediated by adenovirus vector and combined with deminerized bone matrix (DBM). Methods 32 full-thickness defects from 16 knees of 8 pigs were randomly divided into 4 groups in the experiments. In group A, the animals′ lateral femoral condyle of right knee joint was repaired with DBM and BMSC infected with both Ad-TGF-β3 and Ad-BMP-2. In group B, the medial femoral condyle of right knee joint was repaired with DBM and BMSC without infection. In group C, the lateral femoral condyle of left knee joint was repaired with DBM. And the group D is control group. Morphology and histology were observed 2, 4, 8 and 12 weeks after operation. Results 12 weeks after operation, the whole defects were repaired in group A, HE staining showed typical cartilaginous structure in the repaired area. In group D, defects were not repaired but filled with fibrous tissue. The O′driscoll scores were 15.65 ± 0.11 (group A), 11.33 ± 0.22 (group B), 6.13 ± 0.15 (group C) and 5.08 ± 0.15 (group D). There was significant difference among the groups (P < 0.05). Conclusions The new type of tissue engineering scaffold that DBM combined with BMSCs transfected with both Ad-BMP-2 and Ad-TGF-β3 could induce cartilage regeneration and repair the defects.

Experimental study of free periosteal autograft: repair of major defects of cartilage of joint in rabbits of different ages / 中华创伤骨科杂志

Objective To investigate a periosteal autograft method to repair major defects of joint cartilage in rabbits of different ages. Methods Major cartilage defects of 52 rabbits aged 4 to 52 weeks were repaired with free periosteal grafts and the results were compared in the study. The regenerated cartilage by the cambium of periosteum growing up to the cavity of joint and that growing down to the cancellus of femur were also compared. Results The macroscopical and histological observations at different intervals showed that in young and adult rabbits the grafts could develop into cartilages to repair the major cartilage defects. In adult rabbits the newly formed cartilage was the same as the normal cartilage in thickness and histology. There was no difference between the cambium growing up to the cavity and that growing down to the cancellus. Conclusions Periosteum possesses chondrogenic potential and can be used to repair the cartilage defects. Different directions of chondrogenesis leads to no significantly different outcomes. In adult rabbits the newly formed cartilage can develop into a normal one.

Repair of articular cartilage defects by transplantation of PLGA compounded with mesenchymal stem cells modified by hIGF-1 gene / 重庆医科大学学报

Objective: To invest the repair effect of articular cartilage defects by transplantation of PLGA compounded with mesenchymal stem cells modified by hIGF-1 gene.Methods: After the amplification by polymerase chain reaction(PCR) from pcDNA3.1-hIGF-1,the human insulin-like growth factor-1(hIGF-1) fragment was inserted to plasmid pIRES2-EGFP to get recombinated vector pIRES2-EGFP-hIGF-1,which followed by transfection of restructured vectors to mesenchymal stem cells and determination the expression of hIGF-1 gene.The gene-modified MSCs were compounded with PLGA and then cultured in standard condition.Then,the transplantation of graft to the defect positon of articular cartilage was performed.After the transplantation,the specimens was obtained in different time and the repair was inspected.Results: The reconstructed vector contained hIGF-1 and GFP genes was constructed successfully and high efficiency expression of hIGF-1 was detected in target cells.Compared to the control groups,the group of transplantation of graft compounded with MSCs modified hIGF-1 greatly improved the effect of articular cartilage defects.Conclusions: IGF-1 gene is one of the ideal candidate genes for the gene modification of MSCs,the seed cells of cartilage tissue engineering and the gene-modified MSCs can improved effective the repair of articular cartilage defects.