LncRNA HOTTIP enhances human osteogenic BMSCs differentiation via interaction with WDR5 and activation of Wnt/ß-catenin signalling pathway.

To uncover the underlying molecular mechanism of long non-coding RNA in the osteogenic differentiation process of bone marrow mesenchymal stem cells (BMSCs), HOXA transcript at the distal tip (HOTTIP) was selected by using a lncRNA microarray assay. Results showed that HOTTIP was significantly upregulated during osteogenic differentiation of human BMSCs. Downregulation of HOTTIP by shRNA inhibited the osteogenic differentiation of BMSCs. Overexpression of HOTTIP by lentiviral vector promoted human BMSCs osteogenic differentiation by increasing the transcription of ß-catenin. RIP assay and RNA pulldown assay confirmed the interaction between HOTTIP and WDR5, a transcription factor binding to the promoter of ß-catenin. The interaction promoted the translocation of WDR5 into the nucleus and increased the transcription of ß-catenin. Implanted HOTTIP-overexpressing BMSCs increased ectopic bone formation in nude mice. HOTTIP is a conservative long noncoding RNA that is essential for osteogenic differentiation of BMSC. HOTTIP enhances osteogenic differentiation via interaction with WDR5 and up-regulation of ß-catenin gene expression, therefore activating Wnt/ß-catenin signalling pathway.

Effects of tendon-bone healing of anterior cruciate ligament reconstruction by osteoprotegerin combined with deproteinized bovine bone.

BACKGROUND: The healing of a tendon graft in a bone tunnel depends on bone ingrowth into the interface between tendon and bone, or that can enhance tendon-bone healing, which is important to reduce the failure rate after ACL reconstruction. METHODS: Sixty skeletally mature, New Zealand white rabbits underwent left ACL reconstruction. OPG/DBB compound (concentration ratio of 30%, 60%, 100%) was delivered to the tendon-bone interface with use of a DBB carrier, and nothing as control group. Twenty animals were killed at 4, 8 and 12 weeks after surgery. I-IV levels of semi-quantitative and Sharpey fibers at the healing tendon-bone interface were evaluated, and the biomechanical properties were tested. RESULTS: A significantly greater amount of Sharpey fibers at the healing tendon-bone interface in the concentration ratio of 100% OPG/DBB-treated group was found compared with the others at all time-points (P<0.05), and it is the same to the Grade Scores at 12 weeks (P<0.05). The femur-ACL-tibia complex of the concentration ratio of 100% OPG/DBB-treated group has significantly increased stiffness compared with the others at 12 weeks (P<0.05). CONCLUSION: The concentration ratio of 100% OPG/DBB compound significantly improve bone formation around the grafted tendon and improve the stiffness at the healing tendon-bone junction in a rabbit model.

[PRELIMINARY STUDY ON EFFECT OF FEMORAL TUNNEL ANGLE ON FEMORAL TUNNEL AFTER ANTERIOR CRUCIATE LIGAMENT RECONSTRUCTION IN RABBITS].

OBJECTIVE: To investigate the effect of the femoral tunnel angle on the femoral tunnel after anterior cruciate ligament (ACL) reconstruction in rabbits. METHODS: Fifty-four healthy 4-5 months old rabbits (weighing, 1.8-2.3 kg, male or female) were randomly divided into 3 groups (n = 18). The ACL reconstruction models of the right knee were established in 3 experimental groups using its Achilles tendons, and the left knee served as the control group. On the coronal position, the angle between the femoral tunnel and the femoral shaft axis was 30 degrees, 45 degrees, and 60 degrees. The level of tumor necrosis factor α (TNF-α) in the synovial fluid at 1, 2, and 4 weeks, the maximum load of the ligament and the rate of bone tunnel enlargement at 4, 8, and 12 weeks were detected. RESULTS: The level of TNF-α significantly increased, and the maximum load of the ligament significantly decreased in the 3 experimental groups when compared with ones in the control group (P < 0.05), but no significant difference was found among 3 experimental groups (P > 0.05). The bone tunnel enlargement was observed in 3 experimental groups at each time point and reached the peak at 4 weeks, but no significant difference was shown among 3 groups (P > 0.05). CONCLUSION: The 30-60 degrees angle between the femoral tunnel and the femoral shaft axis in the coronal position has no significant effect on the femoral tunnel enlargement after ACL reconstruction in rabbits.

[EXPERIMENTAL STUDY ON DIFFERENT CONCENTRATION RATIOS OF OSTEOPROTEGERIN COMBINED WITH DEPROTEINIZED BONE ON BONE TUNNEL AFTER ANTERIOR CRUCIATE LIGAMENT RECONSTRUCTION].

OBJECTIVE: To investigate the effects of different concentrations of osteoprotegerin (OPG) combined with deproteinized bone (DPB) on the bone tunnel after the anterior cruciate ligament (ACL) reconstruction. METHODS: The femoral epiphyseal side was harvested from newborn calf, and allogenic DPB were prepared by hydrogen peroxide-chloroform/methanol method. Then, DPB were immersed in 3 concentrations levels of OPG (30, 60, 100 µg/mL) and 3 concentration ratios (30%, 60%, 100%) of the gel complex were prepared. Sixty healthy New Zealand white rabbits, male or female, weighing (2.7 ± 0.4) kg, were divided randomly into 4 groups (n = 15): control group (group A), 30% (group B), 60% (group C), and 100% (group D) OPG/DPB gel complex. The ACL reconstruction models were established by autologous Achilles tendon. Different ratios of OPG/DPB gel complex were implanted in the femoral and tibial bone tunnel of groups B, C, and D, but group A was not treated. The pathology observation (including the percentage of the femoral bone tunnel enlargement) and histological observation were performed and the biomechanical properties were measured at 4, 8, and 12 weeks after operation. RESULTS: One rabbit died of infection in groups A and D, 2 rabbits in groups B and C respectively, and were added. General pathology observation showed that the internal orifices of the femoral and tibia tunnels were covered by a little of scar tissue at 4 weeks in all groups. At 8 weeks, white chondroid tissues were observed around the internal orifices of the femoral and tibia tunnels, especially in groups C and D. At 12 weeks, the internal orifices of the femoral and tibia tunnels enlarged in groups A, B, and C, but it was completely closed in group D. At each time point, the rates of the femoral bone tunnel enlargement in groups B, C, and D were significantly lower than that in group A, and group D was significantly lower than groups B and C (P < 0.05); group C was significantly lower than group B at 8 weeks, but no significant difference was found at 4 and 12 weeks (P < 0.05). Hisological observation showed that fresh fibrous connective tissue was observed in 4 groups at 4 weeks; there was various arrangements of Sharpey fiber in all groups at 8 weeks and the atypical 4-layer structure of bone was seen in group D; at 12 weeks, Sharpey fiber arranged regularly in all groups, with typical 4-layer structure of bone in groups B, C, and D, and an irregular "tidal line" formed, especially in group D. Biomechanics measurement showed that the maximum tensile load in group D was significantly higher than that in groups A and B at 4 weeks (P < 0.05), but no significant difference was shown among groups A, B, and C, and between groups C and D (P > 0.05); at 8 weeks, it was significantly higher in groups C and group D than group A, and in group D than group B (P < 0.05), but there was no significant difference between groups A, C and group B (P > 0.05); at 12 weeks, it was significantly higher in groups C and D than groups A and B, and in group D than group C (P < 0.05), but difference was not significant between groups A and B (P > 0.05). CONCLUSION: Different concentrations ratios of OPG/DPB gel complexes have different effects on the bone tunnel after ACL reconstruction. 100% OPG/DPB gel complex has significant effects to prevent the enlargement of bone tunnel and to enhance tendon bone healing.