BACKGROUND:Presently,cartilage defect is mainly treated by autologous cartilage transplantation,which cannot be accepted by patients.With development of tissue engineering,mesenchymal stem cells have been a hot focus in research.Most scholars believed that dynamic stress,hypoxia and some growth factors are advantageous factors for differentiation of bone marrow mesencymal stem cells into chondrocytes,which suggest microenvironment of the joint cavity.OBJECTIVE:To establish a method for isolating bone marrow mesenchymal stem cells,to found a microenvironment of the joint cavity,and to observe outcome of injure repair by transplanting autologous bone marrow mesenchymal stem cells into cartilage injured region of rabbits,which is covered by periosteum.DESIGN,TIME AND SETTING:The randomized,controlled animal experiment was performed at the Laboratory of Department of Orthopaedics,Second Hospital,Shanxi Medical University from July 2005 to July 2007.MATERIALS:A total of 24 New Zealand rabbits were selected for creating models of articular cartilage injury.METHODS:Rabbit bone marrow mesenchymal stem cells were collected and purified using density gradient centrifugation,and amplified by in vitro adherence method.Twenty-four rabbits were randomly assigned into three groups(n=8).In the non-induced bone marrow mesenchymal stem cells + periosteal flap group,bone marrow mesenchymal stem cells were implanted into articular cartilage defects of rabbit knees and covered by autologous periosteal flap.In the periosteal flap group,periosteal flap was used to cover the injured region.Rabbits in the blank control group were left intact.MAIN OUTCOME MEASURES:Repair tissues of rabbits were examined at 6 and 12 weeks after surgery.RESULTS:In the non-induced bone marrow mesenchymal stem cells + periosteal flap group,the defects were filled with hyaline-like cartilage at 6 weeks.Cartilage and the subchodral bone were remodeled at 12 weeks after surgery.The expression of type Ⅱ collagen in the repair tissues was verified by immunohistochemistry.In the periosteal flap group,the defects were partly filled with chondrocytes-like in the surface and basement.In the blank control group,fibrous tissue repair was presented,with a few cartilage-like cells in the basement.CONCLUSION:Following comparison of results from three groups,in vitro cultured rabbit bone marrow mesenchymal stem cells and periosteal transplantation can enhance repair of articular cartilage injury.This method can be an effective way for repairing injured articular cartilage,presently.

BACKGROUND:Such methods as transplanting autologous periosteum or autologous bone marrow mesenchymal stem cells (BMSCs) can promote the repair of articular cartilage defects for sure. But they all have their own limits in chondrogenic abilities,which results in an unsatisfactory curative effect.OBJECTIVE:To study the effect of transplanting BMSCs (which were induced into chondrocytes) combined with autogenous periosteum on repairing articular cartilage defects in rabbits.MATERIALS:A total of 18 New Zealand rabbits,aged 6-8 months,were divided with random digits table method into 3 groups,namely,periosteum+BMSCs group,periosteum group and blank control group,with 6 ones (12 knee joint samples) in each group. METHODS:In periosteum+BMSCs group,BMSCs were harvested and adherently cultured with trypsin digestion method. Then they were induced by transforming growth factor 81 into chondrocytes. At the same time,immunofluorescence labeling was performed to BMSCs membranes with PKH-26. Full-thickness articular cartilage defects (diameter:3mm,depth:3mm) were made to bilateral condylus medialis femoris of all rabbits. In periosteum+BMSCs group and periosteum group,defects were covered by homolateral autogenous proximal tibia periosteums,with germinal layer facing to cavitas medullaris. After that,the periosteum+BMSCs group received 3 sutures,followed by injection of 20 μL BMSCs suspension (1×109/L) into the defects,after which the last suture was taken. The periosteum group underwent coverage with periosteum on defects only. The blank control group underwent perforate only.MAIN OUTCOME MEASURES:General observation,histological observation,Wakitani's score,immunohistochemical and in situ hybridization detection of collagen type Ⅱ were performed to defects at 6 and 12 weeks following operation.RESULTS:No sutured periosteums were found desquamate. In periosteum+BMSCs group,defects were filled with hyaline cartilage-like repairing tissues at week 6 following operation;Week 12 following operation saw remodeled tissues whose cells were mainly the implanted cells labeled with PKH-26. In periosteum group,repairing tissues in defect areas were ivory white,smooth with light introcession and distinctively different from the surrounding normal cartilage tissue. In the blank control group,clearer introcession or irregular appearance,even broken surrounding cartilage tissues could be seen in the defect area. Both Wakitani's score and histological score were highest in periosteum+BMSCs group at week 6 and 12 following operation (P＜0.05),with higher ones in periosteum group than in the control group (P＜0.05). What'more,matrix around cells in the repairing tissues showed positive results to both immunohistochemical and in situ hybridization staining of collagen typeⅡ,which proved that cells in repairing tissues were the implanted ones.CONCLUSION:Transplanted BMSCs (which were induced into chondrocytes) combined with autogenous periosteum can form hyaline cartilage-like repairing tissues through which articular cartilage defects are repaired.

BACKGROUND:Chondrocytes may dedifferentiate when they are cultured in vitro,and the capacity of synthetizing glycosaminoglycan (GAG)is also reduced,how to delay the dedifferentiation of chondrocytes is a crucial topic in the field of tissue engineering.OBJECTIVE:To observe the performance of chondrocytes synthetizing GAG at different inoculum densities.DESIGN,TIME AND SETTING:A controlled cellular experiment was performed at the laboratory of Department of Orthopaedics in the Second Hospital of Shanxi Medical University between January 2007 and May 2007.MATERIALS:Five New Zealand rabbits of one month old were used in this study.METHODS:Articular chondrocytes were isolated from both knees and digested using 0.4% pronase enzyme and 0.025% Ⅱ type collagenase.The chondrocytes harvested from the same rabbit were divided into two sets,one was seeded at a constant density of 2×104/cm2 in primary and subculture,the other was cultured at a reduced density of 2×103/cm2 following cellular adhesion.Cellular morphology and proliferation were observed under inverted microscope.The culture media were renewed after the primary cells and passage 1 cells were confluent.MAIN OUTCOME MEASURES:GAG concentration was determined using the modified precipitation method with Alcian blue at 12,24,36,48 and 60 hours following the renewal of culture media.RESULTS:Articular chondrocytes in the primary high-density culture group were polygonal with clear boundaries,they have shown to form colony at 3-4 days.Cells around colonies were more slender than those in the center of colonies,shaping as long polygon.There was no obvious change observed in the morphology of passage 1 cells.In the low-density culture group,cells scattered at early stage and formed colonies at 7 days,cellular morphology showed no significant differences in comparison with high-density culture group.The time of primary cells becoming confluent in the low-density culture group was prolonged compared with high-density culture group.The GAG concentration in supernatants in the primary cells of low-density culture group was significantly lower than that in primary cells and passage 1 cells of high-density culture group (P＜0.001,P＜0.05).The GAG concentration showed a greater difference along with the prolonging of culture time.CONCLUSION:High-density culture is better then low-density culture to enhance the performance of chondrocytes synthetizing GAG and to retard the velocity of chondrocytes dedifferentiation,which suggests high-density culture contributes to maintain the chondrocytes phenotype and can be considered as a good way of plate culture.

Objective To observe the change of collagen in osteoarthritis and study the pathogenesis of osteoarthritis.Methods The right hind limb of twenty-four adult New Zealand White rabbits were immobilized with plaster cast in extension position for 10,20,30 and 40 days,respectively.Six animals without immobilization served as control.The articular cartilage of the medial femur condyle was harvested for transmission electron microscopy,in-situ hybridization of collagenⅡ,and immunohistochemistry of collagenⅠ,Ⅱ,Ⅲ.Results Transmission electron microscopy showed articular cartilage was destructed from the fine collagen fiber network of tangential zone.The fine collagen fiber network did not contain chondrocyte.Immunohistochemistry and in-situ hybridization showed that in earlier period of osteoarthritis,the collagen typeⅡand its gene expression firstly increased,then decreased with destruction of ultrastructure,and chondrocytes enhanced type Ⅱ collagen expressing and synthesizing mainly in transition zone and upper deep zone.In articular cartilage of osteoarthritis there was type Ⅲ collagen,instead of typeⅠcollagen.Conclusion In osteoarthritis,articular cartilage degenerated from tangential layer,in which collagen can not be repaired after destruction,this may contribute to the chondral degeneration.

Objective:To explore the effects of Alpha-2-macroglobulin-rich serum (A2MRS) on knee post-traumatic osteoarthritis (PTOA).Methods:The knee PTOA models were constructed by transection of the anterior cruciate ligament (ACL) in 80 SD male rats, aged 2 months and weighing from 250 to 300 g, which were randomized into 4 groups ( n=20): a high dose group (A2MRS containing 20 μg/μL A2M administered), a low dose group (A2MRS containing 10 μg/μL A2M administered), a positive control group (normal saline administered), and a blank control group (the knee joint cut pseudooperatively and normal saline administered). HE, toluidine blue staining, safranine O staining, modified Mankin scoring and Osteoarthritis Research Society International (OARSI) scoring were conducted to evaluate and compare the therapeutic effects of A2MRS on the knee PTOA among the 4 groups. Results:The rat cartilage was thinner with patchy and cracked surface, and the chondrocytes were reduced and distributed unevenly in the positive control group, compared with the blank control group. The modified Mankin score (3.89±0.93) and OARSI score (10.05±0.72) in the positive control group were significantly higher than those in the blank control group (0.67±0.07 and 3.10±0.29) ( P<0.05). The rat cartilage was thicker with basically complete and crack-free surface, and the chondrocytes were increased and distributed more evenly in the high dose group and the low dose group, compared with the positive control group. The modified Mankin scores (1.33±0.50 and 1.56±0.53) and OARSI scores (6.30±0.64 and 4.75±0.66) in the high dose group and the low dose group were significantly lower than those in the positive control group ( P<0.05). However, there were no such differences between the high dose group and the low dose group ( P>0.05). Conclusion:A2MRS effectively delays the pathological process of knee PTOA.

Objective To identify the risk factors for the fractures secondary to percutaneous vertebroplasty for osteoporotic vertebral compression fractures.Methods A comprehensive search was conducted for the studies from January 2006 to September 2016 on the risk factors for secondary fractures after percutaneous vertebroplasty in the Cochrane Library,PubMed Data,CNKI,Chinese Biomedical Database,Wanfang Data and manually as well.After retrieval of the eligible data,software Revman5.0 was used to perform the heterogeneity test and calculate the pooled odds ratio (OR),weighted mean difference(WMD) value and 95％ confidence interval (CI).Results Twenty studies involving 3,602 patients,627 of whom had fracture secondary to the surgery,were included in this meta-analysis.Meta-analyses showed the secondary fracture after percutaneous vertebroplasty for osteoporotic vertebral compression fracture was related to bone mineral density [WMD =-0.66,95％ CI (-0.97,-0.36),P ＜ 0.05] and kyphosis after primary operation [WMD =4.51,95％ CI (3.02,6.00),P ＜ 0.05],but not to gender [OR =0.98,95％ CI (0.77,1.25),P＞ 0.05],age [WMD=1.48,95％CI (-0.13,3.09),P＞ 0.05],body mass index [WMD=-0.76,95％ CI(-1.61,0.08),P＞ 0.05],cement volume [WMD=-0.15,95％CI (-0.60,0.30),P＞0.05],intradiscal cement [0R=1.11,95％CI (0.56,2.22),P＞0.05],number of vertebrae primarily treated [OR=0.74,95％ CI (0.09,6.45),P ＞ 0.05],thoracolumbar spine [OR =0.86,95％ CI (0.63,1.18),-P ＞ 0.05],or cement injection approach [OR =1.58,95％ CI (0.74,3.37),P ＞ 0.05].Conclusions Bone mineral density and kyphosis after primary operation may be the risk factors closely correlative to the secondary fracture after percutaneous vertebroplasty.There has not been enough evidence to support the associations between the secondary fracture and gender,age,body mass index,cement volume,intradiscal cement,number of vertebrae primarily treated,thoracolumbar spine,or cement injection approach.

Objective:To compare the efficacy of augmentation suture and direct suture in treatment of acute Achilles tendon ruptures so as to provide evidences for the selection of surgical methods.Methods:A systematic search was conducted in PubMed, Web of Science, EMBASE, CNKI and Wanfang database for studies of augmentation suture versus direct suture of acute Achilles tendon ruptures. Literatures were retrieved from January 1990 to January 2019. Literatures were screened according to the pre-set inclusion and exclusion criteria, and the rating of included literature was strictly evaluated by the Cochrane System Evaluation Manual and Newcastle-Ottawa scale (NOS). The relevant data were analyzed using RevMan 5.3 to compare patient satisfaction, rerupture rate, infection rate, ankle's range of motion and other complications (postoperative calf atrophy, plantar flexion, plantar flexion strength).Results:Eight articles involving 558 patients were identified including 273 patients in augmentation suture group and 285 patients in direct suture group. There were no significant differences between two groups in aspects of patient satisfaction ( OR=1.00, 95% CI 0.54-1.84, P>0.05), rerupture rate ( OR=0.75, 95% CI 0.34-1.66, P>0.05), infection rate ( OR=1.73, 95% CI 0.76-3.94, P>0.05), ankle's range of motion ( OR=0.97, 95% CI 0.33-2.87, P>0.05) and other complications rate ( OR=1.93, 95% CI 0.91-4.09, P>0.05). Meantime, the two groups showed similar effect on postoperative calf atrophy and average isokinetic calf muscle strength ( P>0.05). There was a significant difference in plantar flexion of injured limbs after augmentation suture( P<0.05). Conclusion:Compared with direct suture, augmentation suture cannot improve patient satisfaction or reduce complication rate in repair of acute Achilles tendon ruptures.

Objective To compare the efficacy and safety between minimally invasive plate osteosynthesis (MIPO) and open reduction and internal fixation (ORIF) in the treatment of humeral shaft fracture.Methods Case-control studies and randomized clinical trials comparing MIPO with ORIF in the treatment of humeral shaft fracture from January 2010 to June 2018 were retrieved from PubMed Library,Cochrane Library,Embase Library,China National Knowledge Internet and Wanfang Data.Methodological quality of the studies and trials was critically assessed.REVMANS.3 was used for data analysis.The 2 groups of patients were compared in terms of University of California,Los Angeles shoulder rating scale (UCLA),Mayo Elbow Performance Score (MEPS),union time,nonunion rate and complications.Results A total of 452 patients from 9 articles were included.There were 216 cases in the MIPO group and 236 ones in the ORIF group.The Meta analyses showed that the MIPO group had a significantly higher UCLA score (WMD =0.36,P=0.03),significantly lower incidences of complications (OR =-0.15,P ＜ 0.05) and iatrogenie radial nerve palsy (OR =0.24,P ＜ 0.05),and significantly shorter union time (SMD =-0.36,P =0.02) than the ORIF group.There were no significant differences between the 2 groups in MEPS (WMD =-0.48,P =0.43) or nonunion rate (OR =0.45,P =0.11).Conclusion MIPO may be a better choice for humeral shaft fracture than ORIF in regards to postoperative shoulder functions,union time,and incidences of complications and iatrogenic radial nerve palsy.

Objective We performed comprehensive proteomic analyses of synovial fluid by using the isobaric tags for relative and absolute quantitation (iTRAQ) method and LC-MS/MS, and searched for candidate biomarkers for osteoarthritis. Methods Synovial fluid was collected from patients with late-stage OA, earlystage OA and the control group. Molecular variations were detected by the iTRAQ method. T test was used for statistical analysis. Results Using the iTRAQ method, we identified 1 283 proteins from synovial fluid of patients with osteoarthritis, of which 268 proteins were not reported previously. There were 72 proteins upregulated and 249 proteins down-regulated between moderate OA group and controls. One hundred and twentyeight proteins were up-regulated and 141 proteins were down-regulated between severe OA group and controls.One hundred and ninety-two proteins were up-regulated and 76 proteins were down-regulated between severe OA group and moderate OA group. Eight proteins were found to be up-regulated in the three groups.Conclusion This is an in-depth analysis of the synovial fluid proteome from patients with osteoarthritis by iTRAQ method. The catalog of proteins generated in this study will further expand our knowledge regarding the pathophysiology of osteoarthritis and help us in identifying good biomarkers for early diagnosis.

Objective:To investigate the differences in gene expression levels in knee chondrocytes and chondrons in vitro.Methods:The chondrocytes and chondrons were isolated from full thickness of the 8-months ( n=5) rabbit knees cartilage. Chondrons from right knee were enzymatically isolated using 0.3% dispase and 0.2% collagenase-2 with shaking for 3 hours. Chondrocytes were isolated by 0.4% Pronase and 0.025% collagenase-2 from left knee. The mRNA levels in chondrocytes and chondrons were analyzed by quantitative real-time PCR, including matrix proteins [aggrecan(Agg), collagen(Col-2), Col-6A6, Col-10, Col-11], MMPs and inhibitors (MMP-1, MMP-3, MMP-9, MMP-13, TIMP-1, TIMP-2, TIMP-3), cytoskeletal proteins (Sox-9, vinculin, tubulin, actin), cytokines (IL-β, TNF-α). Analysis was performed using SPSS 16.0 statistical software, and the two-group comparisons were considered as significant by t-test at P<0.05. Results:Compared to the chondrocytes, the Agg [(5.78±0.90) vs (1.89±0.27), t=9.26, P<0.001], Col-2 [(6.29±0.76) vs (3.06±0.60), t=7.46, P<0.001], Col-6A6 [(0.89±0.18) vs (0.22±0.06), t=7.90, P<0.001], Col-10 [(3.83±0.76) vs (1.00±0.26), t=7.88, P<0.001] and TIMP-1 [(1.98±0.85) vs (1.03±0.34), t=2.32, P=0.049], TIMP-2[(3.46±1.50) vs (1.52±1.06), t=2.36, P=0.046], TIMP-3 [(3.96±0.50) vs (1.36±0.18), t=10.94, P<0.001], Sox-9 [(7.09±2.93) vs (3.24±0.77), t=2.84, P=0.022], vinculin [(3.42±1.69) vs (1.46±0.68), t=2.41, P=0.043], tubulin[(9.34±0.71) vs (2.35±0.80), t=14.61, P<0.001] showed higher expression in the chondrons. Compared to the chondrocytes, the MMP-1 [(1.02±0.30) vs (2.67±0.45), t=6.91, P<0.001], MMP-3 [(1.21±0.32) vs (2.52±0.79), t=3.44, P=0.009], MMP-13 [(1.23±0.34) vs (3.42±0.86), t=5.30, P=0.007], IL-1β [(1.02±0.14) vs (2.70±0.49), t=7.37, P<0.001], TNF-α [(0.99±0.08) vs (3.15±0.54), t=8.85, P<0.001] showed lower expression in the chondrons. There were no difference between chondrons and chondrocytes for Col-11, MMP-9, actin ( P>0.05). Conclusion:The gene expression of extracellular matrix components are higher and the gene expression levels of inflammatory factors and MMPs are decreased in chondrons compared with the chondrocytes, suggesting the chondrons have more multiplication potential as seeding cells for tissue-engineered cartilage.