Bone marrow mesenchymal stem cells paracrine TGF-ß1 to mediate the biological activity of osteoblasts in bone repair.

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are an important source of seed cells for regenerative medicine and tissue engineering therapy. BMSCs have multiple differentiation potentials and can release paracrine factors to facilitate tissue repair. Although the role of the osteogenic differentiation of BMSCs has been fully confirmed, the function and mechanism of BMSC paracrine factors in bone repair are still largely unclear. This study aimed to determine the roles of transforming growth factor beta-1 (TGF-ß1) produced by BMSCs in bone tissue repair. METHODS: To confirm our hypothesis, we used a Transwell system to coculture hBMSCs and human osteoblast-like cells without contact, which could not only avoid the interference of the osteogenic differentiation of hBMSCs but also establish the cell-cell relationship between hBMSCs and human osteoblast-like cells and provide stable paracrine substances. In the transwell coculture system, alkaline phosphatase activity, mineralized nodule formation, cell migration and chemotaxis analysis assays were conducted. RESULTS: Osteogenesis, migration and chemotaxis of osteoblast-like cells were regulated by BMSCs in a paracrine manner via the upregulation of osteogenic and migration-associated genes. A TGF-ß receptor I inhibitor (LY3200882) significantly antagonized BMSC-induced biological activity and related gene expression in osteoblast-like cells. Interestingly, coculture with osteoblast-like cells significantly increased the production of TGF-ß1 by BMSCs, and there was potential intercellular communication between BMSCs and osteoblast-like cells. CONCLUSIONS: Our findings provide evidence that the biological mechanism of BMSC-produced TGF-ß1 promotes bone regeneration and repair, providing a theoretical basis and new directions for the application of BMSC transplantation in the treatment of osteonecrosis and bone injury.

Gallium ions promote osteoinduction of human and mouse osteoblasts via the TRPM7/Akt signaling pathway.

Gallium (Ga) ions have been widely utilized for biomedical applications; however, their role in osteoblast regulation is not completely understood. The aim of the present study was to investigate the potential effect of Ga ions on osteoinduction in two osteoblast cell lines and to explore the underlying mechanisms. Human hFOB1.19 and mouse MC3T3­E1 osteoblasts were treated with Ga nitride (GaN) at different concentrations, and the degree of osteoinduction was assessed. Ga ion treatment was found to increase alkaline phosphatase activity and accelerate calcium nodule formation, as assessed using ALP activity assay and Alizarin red S staining. Moreover, upregulated expression levels of osteogenic proteins in osteoblasts were identified using western blotting and reverse transcription­quantitative PCR. Western blotting was also performed to demonstrate that the biological action of Ga ions was closely associated with the transient receptor potential melastatin 7/Akt signaling pathway. Furthermore, it was found that Ga ions did not cause osteoblast apoptosis, as indicated via flow cytometry, but promoted osteoclast proliferation, migration or invasion. The present study investigated the potential role of Ga ions in regulating osteoinduction of osteoblasts, thereby providing a promising strategy for the treatment of osteoporosis.

Structural characterization of a safflower polysaccharide and its promotion effect on steroid-induced osteonecrosis in vivo.

A water-soluble polysaccharide (SPAW) was purified from Safflower and it was identified to be (1→3)-linked ß-d-Glucan. The therapeutic effect and underlying mechanism of SPAW on steroid-induced avascular necrosis of the femoral head (SANFH) in a rabbit model was performed here. The abnormal histopathologic changes and apoptosis of femoral head in model group were significantly reverted after SPAW (25, 100 and 200 mg/kg) administration for 60 days, as evidenced by the a decline of empty lacunae rate, the average bone marrow fat cell size and the proportion of apoptotic cells. Furthermore, administration of SPAW significantly decreased the Bax and caspase-3 protein expression, but increased the protein expression of Bcl-2 when compared these in model rabbits. Meanwhile, increased hydroxyproline (HOP) and decreased serum hexosamine (HOM) concentration in rabbit serum were turned to the opposite way. The present study suggested that SPAW may provide an alternative treatment for the treatment of SANFH.

Mesenchymal stem cell-loaded porous tantalum integrated with biomimetic 3D collagen-based scaffold to repair large osteochondral defects in goats.

BACKGROUND: The body is unable to repair and regenerate large area bone defects. Moreover, the repair capacity of articular cartilage is very limited. There has long been a lack of effective treatments for osteochondral lesions. The engineered tissue with biphase synthetic for osteochondral repair has become one of the hot research fields over the past few years. In this study, an integrated biomanufacturing platform was constructed with bone marrow mesenchymal stem cells (BMSCs)/porous tantalum (pTa) associated with chondrocytes/collagen membranes (CM) to repair large osteochondral defects in load-bearing areas of goats. METHODS: Twenty-four goats with a large osteochondral defect in the femoral heads of the left hind legs were randomly divided into three groups: eight were treated with chondrocytes/CM-BMSCs/pTa, eight were treated with pure CM-pTa composite, and the other eight goats were untreated. The repair effect was assessed by X-ray, gross observation, and histomorphology for 16 weeks after the operation. In addition, the biocompatibility of chondrocytes/CM-BMSCs/pTa was observed by flow cytometry, CCK8, immunocytochemistry, and Q-PCR. The characteristics of the chondrocytes/CM-BMSCs/pTa were evaluated using both scanning electron microscopy and mechanical testing machine. RESULTS: The integrated repair material consists of pTa, injectable fibrin sealant, and CM promoted adhesion and growth of BMSCs and chondrocytes. pTa played an important role in promoting the differentiation of BMSCs into osteoblasts. Three-dimensional CM maintained the phenotype of chondrocytes successfully and expressed chondrogenic genes highly. The in vivo study showed that after 16 weeks from implantation, osteochondral defects in almost half of the femoral heads had been successfully repaired by BMSC-loaded pTa associated with biomimetic 3D collagen-based scaffold. CONCLUSIONS: The chondrocytes/CM-BMSCs/pTa demonstrated significant therapeutic efficacy in goat models of large osteochondral defect. This provides a novel therapeutic strategy for large osteochondral lesions in load-bearing areas caused by severe injury, necrosis, infection, degeneration, and tumor resection with a high profile of safety, effectiveness, and simplicity.

Chemotherapy effectively suppresses interleukin-20, receptor activator of nuclear factor kappa-B ligand, and osteoprotegerin levels in patients with lung adenocarcinoma and bone metastasis.

BACKGROUND: Bone metastasis (BM) is common in patients with lung cancer. Osteolysis is caused by increased osteoclast activity. Interleukin-20 (IL-20) and receptor activator of nuclear factor kappa-B ligand (RANKL) are crucial for osteoclast formation. Osteoprotegerin (OPG) inhibits a receptor activator of RANKL/RANK signaling. The aims of this study were to analyze the serum levels of IL-20, OPG, and RANKL in patients with and without BM and to observe the effect of chemotherapy on these cytokines. PATIENTS AND METHODS: A total 54 cases of pathologically confirmed lung adenocarcinoma (ADC) and 18 healthy individuals (Control) were enrolled in this study. Eligible patients were divided into three groups (18 patients per group): ADC without BM (ADC), ADC with BM (ADC + BM), and ADC with BM treated with chemotherapy (ADC + BM + Chemo). Serum IL-20, RANKL, and OPG levels were analyzed by enzyme-linked immunosorbent assay. RESULTS: Serum IL-20, RANKL, and OPG levels in ADC + BM patients were significantly elevated compared with that in the Control or ADC groups (both P < 0.001). The serum cytokine levels were significantly lower following chemotherapy compared with that in patients who did not receive chemotherapy (P < 0.001). CONCLUSIONS: Serum IL-20, RANKL, and OPG levels increase in patients with lung cancer and BMs. Chemotherapy suppresses the elevation of these cytokines.

Vascular endothelial growth factor/bone morphogenetic protein-2 bone marrow combined modification of the mesenchymal stem cells to repair the avascular necrosis of the femoral head.

Vascular endothelial cell growth factor (VEGF) combined with bone morphogenetic protein (BMP) was used to repair avascular necrosis of the femoral head, which can maintain the osteogenic phenotype of seed cells, and effectively secrete VEGF and BMP-2, and effectively promote blood vessel regeneration and contribute to formation and revascularization of tissue engineered bone tissues. To observe the therapeutic effect on the treatment of avascular necrosis of the femoral head by using bone marrow mesenchymal stem cells (BMSCs) modified by VEGF-165 and BMP-2 in vitro. The models were avascular necrosis of femoral head of rabbits on right leg. There groups were single core decompression group, core decompression + BMSCs group, core decompression + VEGF-165/BMP-2 transfect BMSCs group. Necrotic bone was cleared out under arthroscope. Arthroscopic observation demonstrated that necrotic bone was cleared out in each group, and fresh blood flowed out. Histomorphology determination showed that blood vessel number and new bone area in the repair region were significantly greater at various time points following transplantation in the core decompression + VEGF-165/BMP-2 transfect BMSCs group compared with single core decompression group and core decompression + BMSCs group (P < 0.05). These suggested that VEGF-165/BMP-2 gene transfection strengthened osteogenic effects of BMSCs, elevated number and quality of new bones and accelerated the repair of osteonecrosis of the femoral head.

Combined vascularized iliac and greater trochanter graftings for reconstruction of the osteonecrosis femoral head with collapse: reports of three cases with 20 years follow-up.

In this report, we present the long-term results of using combined vascularized iliac and greater trochanter graftings for reconstruction of the osteonecrosis of the femoral head (ONFH) with collapse in three patients. Necrosis over two-thirds of the femoral head and collapse were observed in these patients, with Harris hip scores (HHS) of 46, 38, and 49 points, respectively. When the patients underwent the femoral head reconstruction procedures, the ages of the patients ranged from 20 to 28 years old. The patients were followed-up for 20-24 years. X-ray examinations showed no progress of necrosis or deformity in the femoral head of patients after surgery, with the exception of bone absorption in one patient with persistence of mild pain. The HHS in the three patients were 84, 65, and 86 points at the end of follow-up, respectively. These results show that the vascularized iliac and greater trochanter graftings may be a valuable option for reconstruction of the ONFH with collapse in younger patients.

Treatment of early stage osteonecrosis of the femoral head with autologous implantation of bone marrow-derived and cultured mesenchymal stem cells.

BACKGROUND: Treatment of early-stage osteonecrosis of the femoral head (ONFH) with autologous implantation of iliac crest bone marrow-derived mononuclear cells, which contain tens of thousands of bone marrow mesenchymal stem cells (BMMSCs), recently achieved a promising outcome. METHODS: One hundred patients with early-stage ONFH were recruited and randomly assigned to BMMSC treatment or core decompression (CD) treatment. Each BMMSC-treated hip received femoral head (FH) implantation of 2×10(6) autologous subtrochanteric bone marrow-derived and ex vivo expanded BMMSCs. The radiographic stage of ONFH according to the Association Research Circulation Osseous classification, Harris hip score (HHS), and the volume of the necrotic lesion or the low signal intensity zone (LowSIZ) in the FH were assessed before and 6, 12, 24, and 60 months after the initial operation. RESULTS: Sixty months after the operation, only 2 of the 53 BMMSC-treated hips progressed and underwent vascularized bone grafting. In CD group, 7 hips lost follow-up, and 10 of the rest 44 hips progressed and underwent vascularized bone grafting (5 hips) or total hip replacement (5 hips). Compared with the CD group, BMMSC treatment significantly improved the HHS as well as decreased the volume of femoral head LowSIZ of the hips preoperatively classified at stage IC, IIB, and IIC (P<0.05, respectively; stage IIA, P=0.06, respectively). No complication was observed in both treatment groups. CONCLUSIONS: Ex vivo expansion of autologous BMMSCs can reliably provide a greater number of BMMSCs for FH implantation. This intervention is safe and effective in delaying or avoiding FH collapse, which may necessitate total hip replacement.

[Treatment of avascular necrosis of femoral head after femoral neck fracture with pedicled iliac bone graft].

OBJECTIVE: To explore the effectiveness of pedicled iliac bone graft transposition for treatment of avascular necrosis of femoral head (ANFH) after femoral neck fracture. METHODS: Between June 2002 and December 2006, 22 cases (22 hips, 16 left hips and 6 right hips) of ANFH after femoral neck fracture were treated with iliac bone graft pedicled with ascending branch of the lateral femoral circumflex vessels. There were 18 males and 4 females with an age range from 28 to 48 years (mean, 37.5 years). The time from injury to internal fixation was 2-31 days, and all fractures healed within 12 months after internal fixation. The ANFH was diagnosed at 15-40 months (mean, 22 months) after internal fixation. The ANFH duration was 3-11 months (mean, 8 months). According to Association Research Circulation Osseous (ARCO) staging system, 2 hips were classified as stage IIa, 3 hips as stage IIb, 3 hips as stage IIc, 3 hips as stage IIIa, 7 hips as stage IIIb, and 4 hips as stage IIIc. The preoperative Harris hip score (HHS) was 64.10 +/- 5.95. RESULTS: All incisions healed by first intention and the patients had no complication of lung embolism, sciatic nerve injury, lower limb deep venous thrombosis, and numbness and pain of donor site. All patients were followed up 2.5 to 6.3 years (mean, 4.8 years). The fracture healing time was 8-12 months, and no femoral neck fracture recurred. The HHS was 90.20 +/- 5.35 at last follow-up, showing significant difference when compared with the preoperative value (t = -18.447, P = 0.000). The hip function were excellent in 11 hips, good in 10 hips, fair in 1 hip, and the excellent and good rate was 95.5%. Four hips were radiographically progressed in ARCO staging, 18 hips remained stable with a stable rate of 81.8%. CONCLUSION: Pedicled iliac bone graft transposition is an ideal option for treatment of ANFH after internal fixation of femoral neck fracture for the advantages of femoral head revascularization, sufficient cancellous bone supply, and relatively simple procedure.

[Three-dimensional gait analysis of patients with osteonecrosis of femoral head before and after treatments with vascularized greater trochanter bone flap].

OBJECTIVE: To provide the objective basis for the evaluation of the operative results of vascularized greater trochanter bone flap in treating osteonecrosis of the femoral head (ONFH) by three-dimensional gait analysis. METHODS: Between March 2006 and March 2007, 35 patients with ONFH were treated with vascularized greater trochanter bone flap, and gait analysis was made by using three-dimensional gait analysis system before operation and at 1, 2 years after operation. There were 23 males and 12 females, aged 21-52 years (mean, 35.2 years), including 8 cases of steroid-induced, 7 cases of traumatic, 6 cases of alcoholic, and 14 cases of idiopathic ONFH. The left side was involved in 15 cases, and right side in 20 cases. According to Association Research Circulation Osseous (ARCO) classification, all patients were diagnosed as having femoral-head necrosis at stage III. Preoperative Harris hip functional score (HHS) was 56.2 +/- 5.6. The disease duration was 1.5-18.6 years (mean, 5.2 years). RESULTS: All incisions healed at stage I without early postoperative complications of deep vein thrombosis and infections of incision. Thirty-five patients were followed up 2-3 years with an average of 2.5 years. At 2 years after operation, the HHS score was 85.8 +/- 4.1, showing significant difference when compared with the preoperative score (t = 23.200, P = 0.000). Before operation, patients showed a hip muscles gait, short gait, reduce pain gait, and the pathological gaits significantly improved at 1 year after operation. At 1 year and 2 years after operation, step frequency, pace, step length and hip flexion, hip extension, knee flexion, ankle flexion were significantly improved (P < 0.01). Acceleration-time curves showed that negative wave and spinous wave at acceleration-stance phase of front feet and hind feet in affected limb were obviously reduced at 1 year and 2 years after operation. Postoperative petronas wave appeared at swing phase; the preoperative situation was three normal phase waves. CONCLUSION: These results suggest that three-dimensional gait analysis before and after vascularized greater trochanter for ONFH can evaluate precisely hip vitodynamics variation.

[Gait analysis after anterior cruciate ligament reconstruction with different methods].

OBJECTIVE: To examine and compare the gait patterns of subjects with hamstring and patellar tendon graft reconstructions of anterior cruciate ligament (ACL). METHODS: Three groups were compared: 19 patellar tendon anterior cruciate ligament reconstruction patients (a mean of 11.8 weeks after injury), 19 hamstring tendon anterior cruciate ligament reconstruction patients (a mean of 10.4 weeks after injury) and 19 matched controls. Gait analysis was performed using the DorealSoft DVMC-8801 three-dimensional motion capture system. Kinematic data were recorded for the lower limb prior to and 3, 6, 9 and 12 months after ACL reconstructive surgery. Each subject was asked to walk on a motorized treadmill with different slopes and perform-after 6 minutes of familiarization time- at least 2 minutes of walking at a constant speed of 1.9 km/h. RESULTS: All the ACL-deficient patients exhibited a quadriceps avoidance pattern prior to and 3 months post-operation. In the operated individuals, the spatial-temporal parameters and the knee angle had already regained a normal pattern for the ACL-deficient limb during gait at 6 months post-operation. However, the angular acceleration showed no significant statistical difference with the values of healthy control group at just 9 months post-operation. The flexion angular acceleration at 15 degrees treadmill was significantly smaller than that in the hamstring tendon group and the extension angular acceleration was significantly smaller than that in the patellar tendon group at 6 months post-operation. CONCLUSION: ACL surgical repair significantly alters the lower-extremity gait patterns and the establishment of pre-injury gait patterns takes at least 9 months to occur. The graft-specific differences in knee biomechanics after anterior cruciate ligament reconstruction appear to be related with the donor site.

[Three-D gait patterns before and after total hip arthroplasty].

OBJECTIVE: To determine how selected gait parameters may change as a result of total hip arthroplasty (THA). METHODS: From February 2006 to February 2007 the study was performed on 53 osteonecrosis of femoral head subjects prior to and 6 weeks, 3, 6 and 12 months after total hip arthroplasty surgery. Gait analysis was performed using the three-dimensional Infrared-based system (Dalian Dong Fang Xin Rui Company, China). Kinematic data were recorded for the lower limb. The results obtained from the osteonecrosis of femoral head subjects were compared with those of 40 individuals without osteonecrosis. Using a three-dimensional motion analyzer, the following were measured: (1) gait speed, stride length, cadence; (2) step length, stride width; (3) standing phase time, walking phase time; (4) gait cycle time, Joint movement angular. RESULTS: Compared with persons before total hip arthroplasty, THA patients showed significantly increased gait speed, gait frequency, step length and affected limb stand phase (P < 0.01). Angle timetable: preoperative hip-maximum flexion was 15 degrees, post-extension was 8 degrees, knee-maximum flexion was 70 degrees, ankle-maximum flexion was 15 degrees. Postop hip-maximum flexion was 27, post-extension was 17 degrees, knee-maximum flexion was 50 degrees, ankle-maximum flexion was 14 degrees; parameters of hip, knee between preoperative and postoperative was significance (P < 0.01). CONCLUSION: These results suggest that 3-D gait analysis before and after total hip arthroplasty can evaluate precisely hip vitodynamics variation.

[Treatment of early osteonecrosis of femoral head by core decompression and autograft of mesenchymal stem cell cultured in vitro under arthroscopy: experiment with rabbits].

OBJECTIVE: To observe the therapeutic effect of core decompression and autograft of mesenchymal stem cells (MSCs) cultured in vitro under arthroscopy in treatment of early osteonecrosis of femoral head. METHODS: Thirty-six rabbits were made into models of early osteonecrosis of femoral head of the right leg, and then randomly divided into 3 equal groups: Group A group as control group, Group B group undergoing simple core decompression under arthroscopy by clearing out of necrotic bone, and Group C undergoing core decompression and implantation of MSCs. Four and 8 weeks later 6 rabbits from each group underwent roentgenography and then were killed with their femoral heads taken out to undergo histological examination. RESULTS: Four weeks later new bone was evident in the implantation field of Group C while fibrous tissues were evident in the femoral head of Group B. Pathological examination showed regular shape and normal density of femoral head in Group C and irregular shape and low density of femoral head in Group B. X-ray photograph 8 weeks after treatment showed that the femoral head articular surface of Group A collapsed, uneven density and defect of spongy bone were seen in Group B, and the shape of femoral head of Group C was regular without collapse and the original defect had been repaired. CONCLUSION: With strong osteogenic function, MSCs improve the repair of ischemic necrosis of femoral head and can be used in treatment of early osteonecrosis of femoral head via core decompression and autograft of MSCs cultured in vitro under arthroscopy.