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Objective:This study aimed to investigate the effect of differentiation osteogenic bone marrow mesenchymal stem cells (De-BMSCs) transplantation on the promotion of bone formation at the tendon-bone interface after anterior cruciate ligament reconstruction (ACLR), and further explored the molecular mechanism of the enhanced osteogenic effect of De-BMSCs.Methods:BMSCs from femur and tibia of New Zealand White rabbit were subjected to osteogenic induction and then cultured in no osteogenic factor medium; the obtained cell population was termed De-BMSCs. De-BMSCs were induced into osteo-, chondro-and adipo-differentiation in vitro to examine the characteristics of primitive stem cells. ACLR model with a semitendinosus tendon were performed in 48 adult rabbits, three groups were established: control group with alginate gel injectionat the tendon-bone interface, BMSCs group with the injection of alginate gel containing BMSCs, De-BMSCs group with the injection of alginate gel containing De-BMSCs. At 4 and 12 weeks after surgery, rabbits in each group were sacrificed to evaluate tendon-bone healing by histologic staining, micro-CT examination, and biomechanical test. During osteogenic differentiation of De-BMSCs, si-RNA of nuclear factor of activated T cells 2 (NFATc2) si-RNA of nuclear factor of activated T cells 1 (NFATc1) were used to verify the molecular mechanism of enhanced osteogenic effect of De-BMSCs.Results:De-BMSCs exhibited some properties similar to BMSCs including multiple differentiation potential and cell surface marker. At 4 weeks after surgery, the BV/TV value of the De-BMSCs group 0.36±0.01 was significantly higher than that of the control group 0.24±0.03 and BMSCs group 0.30±0.02 (all P<0.05), and the maximum load 40.34±1.19 N and stiffness 20.67±2.14 N/mm were significantly higher than those in the control group 14.88±2.74N, 8.67±2.19 N/mm and the BMSCs group 26.31±1.76 N, 13.81±2.14 N/mm (all P<0.05). At 12 weeks after surgery, the BV/TV value of the De-BMSCs transplantation group 0.47±0.02 was significantly higher than that of the control group 0.30±0.02 and the BMSCs group 0.35±0.03 (all P<0.05), and the maximum load 64.46±6.69 N and stiffness 25.18±3.11 N/mm were significantly higher than those in the control group 41.01±6.12 N, 11.59±2.54 N/mm and the BMSCs group 48.21±4.12 N, 15.89±2.94 N/mm (all P<0.05). During the osteogenic differentiation of De-BMSCs, the expressions of Nanog and NFATc1 were synergistically increased which promoted interaction of NFATc1 and Osterix ( P< 0.05), resulting in the increased expression of osteoblast marker genessuch as COL1A, OCN, OPN (all P< 0.05). Conclusion:De-BMSCs transplantation could promote bone formation at the tendon-bone interface after ACLR,Nanog/NFATc1/Osterix signaling pathway mediated the enhancement of the osteogenic differentiation effect of De-BMSCs.
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Objective To compare the bone mineral density of the regions of femoral and tibial tunnels for anterior cruciate ligament(ACL)reconstruction in order to provide reference for the choice of optimal screw diameter for interference fixation.Methods Thirty healthy volunteers aged from 18 to 35 years were enrolled in our study,and the dual-energy X-ray absorptiometry(DEXA)was used to measure the bone mineral density of femoral and tibial tunnel regions of the right knee.All the right knees of the volunteers were also scanned by spiral CT and three-dimensional reconstruction technique was utilized to determine the circular sections that pass through the longitudinal axis of the femoral and tibial tunnels separately.The CT gray scale values of the Sections were measured.From August to October 2010,9 patients who had been diagnosed as ACL rupture underwent the operation of ACL reconstruction,and cylindrical cancellous bone peg was removed from the femoral and tibial tunnel respectively during the operation.Volumetric bone mineral density of the bone pegs were measured by using a standardized immersion technique according to Archimedes’ principle.Results Measured by DEXA,bone density of the femoral tunnel region arid tibial tunnel region were(1.162±0.034)g/cm2 and(0.814±0.038)g/cm2 respectively.The difference was significant between the femoral and tibial tunnel region(t=9.11,P=0.000).The CT gray scale value of the section for femoral tunnel region was(211.7±11.5)Hu,while that of the tibial tunnel region was(104.9±7.4)Hu.There was statistically significant difference between them(t=10.15,P=0.000).The volumetric bone mineral density of the bone peg from femoral tunnel and tibial tunnel were(2.80±0.88)g/cm3 and(1.88±0.59)g/cm3 respectively.The difference was statistically significant(t=4.32,P=0.002).Conclusion For ACL reconstruction,bone mineral density of femoral tunnel region is higher than that of the tibial tunnel.