Multicenter study on atypical femoral fractures in patients with bone metastases taking bone- modifying agents.

Bone-modifying agents (BMAs), with bone-resorptive inhibitory effects, such as zoledronic acid and denosumab, are widely used at higher doses for bone-related events caused by bone metastasis of malignant tumors. These drugs have been suggested to be associated with atypical femoral fractures (AFFs), and the relationship between BMAs and AFFs has attracted attention. To investigate the clinical features including bone union time of AFFs in patients administered BMA for bone metastasis, we conducted a retrospective multicenter study. Thirty AFFs from 19 patients were enrolled in this study. Thirteen patients had bilateral AFFs, and nineteen AFFs had prodromal symptoms. Eighteen AFFs underwent surgery after complete fracture, three failed to achieve bone union and required nonunion surgery, and 11 AFFs that achieved bone union had an average period until bone union of 16.2 months, which was much longer than that previously reported for ordinary AFFs. Seven patients discontinued the BMAs, but not due to AFFs. Stopping BMAs in patients with bone metastasis would make it difficult to secure their performance of activities of daily living, and AFF with BMA administration might require a longer time for union. Therefore, it would be important to prevent incomplete AFF from becoming complete AFF via prophylactic internal fixation.

Altered microRNA profile during fracture healing in rats with diabetes.

BACKGROUND: MicroRNAs (miRNAs) are a class of small non-coding RNA molecules that regulate gene expression. There is increasing evidence that some miRNAs are involved in the pathology of diabetes mellitus (DM) and its complications. We hypothesized that the functions of certain miRNAs and the changes in their patterns of expression may contribute to the pathogenesis of impaired fractures due to DM. METHODS: In this study, 108 male Sprague-Dawley rats were divided into DM and control groups. DM rats were created by a single intravenous injection of streptozotocin. Closed transverse femoral shaft fractures were created in both groups. On post-fracture days 5, 7, 11, 14, 21, and 28, miRNA was extracted from the newly generated tissue at the fracture site. Microarray analysis was conducted with miRNA samples from each group on post-fracture days 5 and 11. The microarray findings were validated by real-time polymerase chain reaction (PCR) analysis at each time point. RESULTS: Microarray analysis revealed that, on days 5 and 11, 368 and 207 miRNAs, respectively, were upregulated in the DM group, compared with the control group. The top four miRNAs on day 5 were miR-339-3p, miR451-5p, miR-532-5p, and miR-551b-3p. The top four miRNAs on day 11 were miR-221-3p, miR376a-3p, miR-379-3p, and miR-379-5p. Among these miRNAs, miR-221-3p, miR-339-3p, miR-376a-3p, miR-379-5p, and miR-451-5p were validated by real-time PCR analysis. Furthermore, PCR analysis revealed that these five miRNAs were differentially expressed with dynamic expression patterns during fracture healing in the DM group, compared with the control group. CONCLUSIONS: Our findings will aid in understanding the pathology of impaired fracture healing in DM and may support the development of molecular therapies using miRNAs for the treatment of impaired fracture healing in patients with DM.

Effects of the duration of transcutaneous CO2 application on the facilitatory effect in rat fracture repair.

BACKGROUND: Carbon dioxide therapy has been reported to be effective in treating certain cardiac diseases and skin problems. Although a previous study suggested that transcutaneous carbon dioxide application accelerated fracture repair in association with promotion of angiogenesis, blood flow, and endochondral ossification, the influence of the duration of carbon dioxide application on fracture repair is unknown. The aim of this study was to investigate the effect of the duration of transcutaneous carbon dioxide application on rat fracture repair. METHODS: A closed femoral shaft fracture was created in each rat. Animals were randomly divided into four groups: the control group; 1w-CO2 group, postoperative carbon dioxide treatment for 1 week; 2w-CO2 group, postoperative carbon dioxide treatment for 2 weeks; 3w-CO2 group, postoperative carbon dioxide treatment for 3 weeks. Transcutaneous carbon dioxide application was performed five times a week in the carbon dioxide groups. Sham treatment, where the carbon dioxide was replaced with air, was performed for the control group. Radiographic, histological, and biomechanical assessments were performed at 3 weeks after fracture. RESULTS: The fracture union rate was significantly higher in the 3w-CO2 group than in the control group (p < 0.05). Histological assessment revealed promotion of endochondral ossification in the 3w-CO2 group than in the control group. In the biomechanical assessment, all evaluation items related to bone strength were significantly higher in the 3w-CO2 group than in the control group (p < 0.05). CONCLUSIONS: The present study, conducted using an animal model, demonstrated that continuous carbon dioxide application throughout the process of fracture repair was effective in enhancing fracture healing.

Topical cutaneous application of carbon dioxide via a hydrogel for improved fracture repair: results of phase I clinical safety trial.

BACKGROUND: Clinicians have very limited options to improve fracture repair. Therefore, it is critical to develop a new clinically available therapeutic option to assist fracture repair biologically. We previously reported that the topical cutaneous application of carbon dioxide (CO2) via a CO2 absorption-enhancing hydrogel accelerates fracture repair in rats by increasing blood flow and angiogenesis and promoting endochondral ossification. The aim of this study was to assess the safety and efficacy of CO2 therapy in patients with fractures. METHODS: Patients with fractures of the femur and tibia were prospectively enrolled into this study with ethical approval and informed consent. The CO2 absorption-enhancing hydrogel was applied to the fractured lower limbs of patients, and then 100% CO2 was administered daily into a sealed space for 20 min over 4 weeks postoperatively. Safety was assessed based on vital signs, blood parameters, adverse events, and arterial and expired gas analyses. As the efficacy outcome, blood flow at the level of the fracture site and at a site 5 cm from the fracture in the affected limb was measured using a laser Doppler blood flow meter. RESULTS: Nineteen patients were subjected to complete analysis. No adverse events were observed. Arterial and expired gas analyses revealed no adverse systemic effects including hypercapnia. The mean ratio of blood flow 20 min after CO2 therapy compared with the pre-treatment level increased by approximately 2-fold in a time-dependent manner. CONCLUSIONS: The findings of the present study revealed that CO2 therapy is safe to apply to human patients and that it can enhance blood flow in the fractured limbs. TRIAL REGISTRATION: This study has been registered in the UMIN Clinical Trials Registry (Registration number: UMIN000013641, Date of registration: July 1, 2014).

Insufficiency and deficiency of vitamin D in elderly patients with fragility fractures of the hip in the Japanese population.

BACKGROUND: Data of vitamin D sufficiency in Asian patients with osteoporotic fragility hip fractures are limited. This study aimed to obtain data from the Japanese population. METHODS: Patients aged 60 years or older with hip fractures were prospectively enrolled. Serum 25-hydroxyvitamin D (25(OH)D) levels were measured. Levels were compared between patients receiving and not receiving treatment for osteoporosis, those with and without previous contralateral hip fractures, and those with femoral neck versus trochanteric fractures. Sex-based differences were also assessed. The serum levels in patients younger than 60 years with extremity fractures were assessed, and differences between elderly and younger patients were evaluated. The individual correlation between 25(OH)D levels and the ultraviolet (UV) index and age was analyzed in elderly patients with hip fractures. RESULTS: The data of 360 patients (aged 84.7 ± 8.2 years), comprising 80 men and 280 women, were analyzed. The mean 25(OH)D level was 16.5 ± 7.2 ng/mL. The prevalence of vitamin D insufficiency (25(OH)D <30 ng/mL) and deficiency (25(OH)D <20 ng/mL) was 93.9% and 71.7%, respectively. A significant difference was noted in the prevalence of vitamin D deficiency between patients with and without previous contralateral hip fractures. Age and 25(OH)D levels were found to be correlated, with no correlation between the UV index and the 25(OH)D levels. The 25(OH)D level in the younger population (n = 123) was 20.7 ± 8.6 ng/mL, which was significantly higher than that of the elderly. CONCLUSION: Perennial vitamin D insufficiency is prevalent in elderly Japanese patients with hip fractures.

Medial clavicle pseudarthrosis successfully treated with an inverted distal clavicle locking plate.

INTRODUCTION: Medial clavicle fractures are rare injuries. Symptomatic nonunion arises up to 8% of medial clavicle fractures when treated conservatively. PRESENTATION OF CASE: A 53-year-old man sustained a left medial clavicle fracture and was treated conservatively at another hospital. Nine months after his initial injury, he was referred to our institution. We diagnosed pseudarthrosis of the medial clavicle. We performed open reduction and internal fixation using an inverted distal clavicle locking plate. At the 1-year follow-up, radiographs showed bone union. DISCUSSION: This is the first reported case of medial clavicle pseudarthrosis treated with an inverted distal clavicle anatomical locking plate. There are several advantages in using this plate. CONCLUSION: This method is a good treatment option.

Escherichia coli-derived BMP-2-absorbed ß-TCP granules induce bone regeneration in rabbit critical-sized femoral segmental defects.

PURPOSE: This study investigated whether Escherichia coli-derived bone morphogenetic protein (BMP)-2 (E-BMP-2) adsorbed onto ß-tricalcium phosphate (ß-TCP) granules can induce bone regeneration in critical-size femoral segmental defects in rabbits. METHODS: Bone defects 20 mm in size and stabilized with an external fixator were created in the femur of New Zealand white rabbits, which were divided into BMP-2 and control groups. E-BMP-2-loaded ß-TCP granules were implanted into defects of the BMP-2 group, whereas defects in the controls were implanted with ß-TCP granules alone. At 12 and 24 weeks after surgery, radiographs were obtained of the femurs and histological and biomechanical assessments of the defect area were performed. Bone regeneration was quantified using micro-computed tomography at 24 weeks. RESULTS: Radiographic and histologic analyses revealed bone regeneration in the BMP-2 group but not the control group; no fracturing of newly formed bone occurred when the external fixator was removed at 12 weeks. At 24 weeks, tissue mineral density, the ratio of bone volume to total volume, and volumetric bone mineral density of the callus were higher in the BMP-2 group than in control animals. In the former, ultimate stress, extrinsic stiffness, and failure energy measurements for the femurs were higher at 24 weeks than at 12 weeks. CONCLUSION: E-BMP-2-loaded ß-TCP granules can effectively promote bone regeneration in long bone defects.

Locking plate osteosynthesis for a femoral fracture and subsequent nonunion in a patient with osteopetrosis.

INTRODUCTION: Osteopetrosis is a skeletal disorder characterized by increased osteodensity and a remodeling defect. The fragility of dense sclerotic bones may lead to an increased incidence of fractures. Although internal fixation can be performed, technical challenges may be experienced because of the increased bone density. Complications such as delayed union, nonunion, or implant failure may occur postoperatively. PRESENTATION OF CASE: We describe a patient with autosomal-dominant osteopetrosis type 2 who suffered a shaft fracture below a plate of his right femur. We performed osteosynthesis with a single locking plate. Union was delayed, and plate breakage occurred along with nonunion of the fracture. The nonunion was addressed using double locking plates, which secured fixation and allowed complete fracture healing. DISCUSSION: There were three reasons of nonunion in our case. First, we left gaps between the fragments. Second, we used mainly cerclage wires, rather than screws, for plate fixation, which led to inadequate stability. Third, the patient was large (height 167 cm, weight 93.1 kg), so the single plate provided insufficient fixing force. We then used double locking plates and attained stronger internal fixation with complete fracture healing. CONCLUSION: Double plating with locking plates may be an effective treatment option for femoral fractures in patients with osteopetrosis.

Triweekly administration of parathyroid hormone (1-34) accelerates bone healing in a rat refractory fracture model.

BACKGROUND: Some reports have shown that intermittent parathyroid hormone (PTH) (1-34) treatment for patients with delayed union or nonunion have led to successful healing. In this study, we investigated whether systemic intermittent administration of PTH (1-34) has a beneficial effect on bone healing in a rat refractory fracture model. METHODS: We created a refractory femoral fracture model in 32 rats with periosteal cauterization that leads to atrophic nonunion at 8 weeks after surgery. Half the rats received subcutaneous intermittent human PTH (1-34) injections at a dosage of 100 µg/kg, thrice a week for 8 weeks. The other half received the vehicle only. At 8 weeks after fracture, radiographic, histological and mechanical assessments were performed. RESULTS: Radiographic assessments showed that the union rate was significantly higher in the PTH group than in the control group (P < 0.05). The degree of fracture repair as scored using the Allen grading system in histological assessment was significantly greater in the PTH group than in the control group (P < 0.05). The ultimate stress and stiffness measurements were significantly greater in the PTH group than in the control group (p < 0.05). CONCLUSIONS: We demonstrated that triweekly administration of PTH (1-34) increased union rate and accelerated bone healing in a rat refractory fracture model, suggesting that systemic administration of PTH (1-34) could become a novel and useful therapy for accelerating fracture healing in patients at high risk of delayed union or nonunion.

Clinical Experience With the Use of Low-Intensity Pulsed Ultrasound (LIPUS) in the Treatment of Atypical Femoral Fractures.

OBJECTIVE: The aim of this study was to report the clinical results of atypical femoral fractures (AFFs) treated with low-intensity pulsed ultrasound (LIPUS). MATERIALS AND METHODS: The data on AFFs that were surgically treated in our hospital from 2010 to 2016 was retrospectively analyzed. AFF was diagnosed based on the criteria defined by the second report of an ASBMR task force. RESULTS: Seven fractures in 6 cases were included in this study. Two fractures were referred to us as being nonunion. Five fractures were subtrochanteric fractures and 2 fractures were shaft fractures. Five fresh AFFs were fixed with an intramedullary nail and 2 nonunion fractures were fixed with plates. LIPUS was used in 6 fractures. Bone union was achieved in 5 fractures with the average time to union being 17 months (5-29). In 4 out of the 6 fractures treated with LIPUS, bone union was achieved after 14 months on average. In the other 2 LIPUS-treated fractures, bone union was not achieved even at 1 year after surgery. DISCUSSION: It is known that AFF healing tends to be very slow. Some case reports indicate that AFF healing might be accelerated by LIPUS. In the current series, the subtrochanteric fracture that was not treated with LIPUS healed at 29 months after surgery, which was much longer than the average time to union in the 5 fractures that were treated with LIPUS. Although our number of cases is small, LIPUS may be a potentially useful tool for accelerating AFF repair.

Thrombin-antithrombin III complex tests.

PURPOSE: Patients with fractures of the pelvis and/or lower extremities are at a high risk of developing postoperative venous thromboembolism (VTE). The purpose of this study was to determine whether the thrombin-antithrombin III complex (TAT) tests could be used for postoperative screening of VTE in patients with lower limb or pelvic fractures. METHODS: We enrolled 133 patients who underwent surgical treatment for fracture of the pelvis or lower extremities. TAT and D-dimer levels were compared in patients with and without VTE. Receiver operating characteristic (ROC) curve analysis was done and the appropriate TAT and D-dimer cutoff levels were determined for VTE screening. RESULTS: VTE was diagnosed in 41 patients (30.8%). Patients with VTE had significantly higher levels of TAT and D-dimer on postoperative days 1, 3, and 7 than those without VTE, respectively. ROC curve analysis suggested that TAT test at postoperative day 7 had the highest accuracy for predicting postoperative VTE. With the optimal cutoff TAT level of 3.0 ng/mL, sensitivity and specificity were 93.3% and 70.1%, respectively. With the optimal cutoff D-dimer level of 7.4 µg/mL, sensitivity and specificity were 93.3% and 57.0%, respectively. CONCLUSION: TAT levels measured at postoperative day 7 could be the most useful parameter for screening postoperative VTE. TAT can be used as a screening tool for screening postoperative VTE in patients with lower limb and pelvic fractures.

Altered expression of SDF-1 and CXCR4 during fracture healing in diabetes mellitus.

PURPOSE: Diabetes mellitus (DM) is known to impair fracture healing. The purpose of this study was to elucidate and compare the gene expression patterns and localization of stromal cell-derived factor 1 (SDF-1) and CXC chemokine receptor 4 (CXCR4) during fracture healing of the femur in rats with and without DM. METHODS: Closed transverse fractures were created in the femurs of rats equally divided into a DM group and control group; DM was induced by streptozotocin. At post-fracture days five, seven, 11, 14, 21 and 28, total RNA was extracted from the fracture callus and mRNA expression levels of SDF-1 and CXCR4 were measured by real-time polymerase chain reaction. Localization of SDF-1 and CXCR4 proteins at the fracture site was determined by immunohistochemistry at days 21 and 28. RESULTS: SDF-1 expression was significantly lower in the DM group than in the healthy group on days 21 and 28, and showed a significant difference between days 14 and 21 in the healthy group. There was no significant difference in CXCR4 expression levels between the healthy and DM groups at any time point. On day 21 immunoreactivity of SDF-1 and CXCR4 was detected at the fracture site of the healthy group but no immunoreactivity was observed in the DM group. On day 28, immunoreactivity of SDF-1 and CXCR4 was detected at the fracture site in both groups. CONCLUSION: Gene expression and localization of SDF-1 and CXCR4 was altered during fracture healing, which may contribute to the impaired fracture healing in DM.

8. Low-Intensity Pulsed Ultrasound (LIPUS) Treatment for the Patient With Bone Reconstruction by the Masquelet Technique Using RIA.

BACKGROUND: The Masquelet technique is a relatively new option in the management of bone defects. It consists of 2-stage surgery. At first surgery, a cement spacer is placed to fill the bone defect. After a waiting period of 6-8 weeks, a pseudo-synovial membrane is formed around the cement. The induced membrane is rich in vascularity and osteogenic activity. At second surgery, the cement is replaced with an autologous bone graft and the graft is packed in the membrane. To harvest large quantities of bone graft, the reamer irrigator aspirator (RIA) is utilized. We report a case in which low-intensity pulsed ultrasound (LIPUS) was applied to bone reconstruction by the Masquelet technique using RIA. CASE: The patient was a 44-year-old man. He suffered a tibia fracture involving the proximal part and the shaft. Open reduction and internal fixation was performed at the previous hospital. However, MRSE infection was found at 6 months postoperatively and surgery to eradicate the infection was performed. Two months after surgery, pseudoarthrosis surgery was performed; however no progression in bone union was observed for 5 months. Therefore, the patient was referred to our hospital. At the first surgery in our hospital, plates and screws were removed, and the avascular bone was debrided. Vancomycin-impregnated cement was placed to fill the bone defect and an Ilizarov external fixator was applied. Eight weeks later, the cement was replaced with the autologous bone graft and the graft was packed in the induced membrane. Bone graft was harvested from the ipsilateral femur using RIA. We applied LIPUS to the affected site postoperatively. Now, 4 months after the second surgery, bone union is progressing and the patient can walk without pain. DISCUSSION: LIPUS may be used as an adjuvant therapy for a patient treated by the Masquelet technique using RIA.

Antibiotic-impregnated calcium phosphate cement as part of a comprehensive treatment for patients with established orthopaedic infection.

BACKGROUND: The treatment of established orthopaedic infection is challenging. While the main focus of treatment is wide surgical debridement, systemic and local antibiotic administration are important adjuvant therapies. Several reports have described the clinical use of antibiotic-impregnated calcium phosphate cement (CPC) to provide local antibiotic therapy for bone infections. However, these were all individual case reports, and no case series have been reported. We report a case series treated by a single surgeon using antibiotic-impregnated CPC as part of a comprehensive treatment plan in patients with established orthopaedic infection. METHODS: We enrolled 13 consecutive patients with osteomyelitis (n = 6) or infected non-union (n = 7). Implantation of antibiotic-impregnated CPC was performed to provide local antibiotic therapy as part of a comprehensive treatment plan that also included wide surgical debridement, systemic antibiotic therapy, and subsequent second-stage reconstruction surgery. We investigated the rate of successful infection eradication and systemic/local complications. The concentration of antibiotics in the surgical drainage fluids, blood, and recovered CPC (via elution into a phosphate-buffered saline bath) were measured. RESULTS: The mean follow-up period after surgery was 50.4 (range, 27-73) months. There were no cases of infection recurrence during follow-up. No systemic toxicity or local complications from the implantation of antibiotic-impregnated CPC were observed. The vancomycin concentration in the fluid from surgical drainage (n = 6) was 527.1 ± 363.9 µg/mL on postoperative day 1 and 224.5 ± 198.4 µg/mL on postoperative day 2. In patients who did not receive systemic vancomycin therapy (n = 3), the maximum serum vancomycin level was <0.8 µg/mL. In vitro vancomycin elution was observed from the CPC that was surgically retrieved (n = 2). CONCLUSIONS: Implantation of antibiotic-impregnated CPC is an option to provide local antibiotic therapy as part of a comprehensive treatment plan.

Treatment of ununited femoral neck fractures in young adults using low-intensity pulsed ultrasound: Report of 2 cases.

INTRODUCTION: Delayed union and non-union of displaced femoral neck fractures remains a difficult clinical problem for orthopaedic surgeons. In the physiologically young patient, every effort should be made to preserve the native hip joint. We present two cases of ununited femoral neck fractures in young adults who were successfully treated with low-intensity pulsed ultrasound (LIPUS) and showed satisfactory results. PRESENTATION OF CASE 1: A 41-year-old woman was involved in a motor vehicle crash and was diagnosed with displaced femoral neck fracture. Eleven months after internal fixation, a computed tomography (CT) scan revealed the presence of non-union of the femoral neck. LIPUS treatment was therefore initiated. After eight months, the fracture was completely consolidated. PRESENTATION OF CASE 2: A 39-year-old man was involved in a cycling accident and was diagnosed with displaced femoral neck fracture. Thirteen weeks after internal fixation, a CT scan revealed delayed union of the femoral neck. LIPUS treatment was therefore initiated. After six months, the fracture was completely consolidated. CONCLUSION: We suggest use of LIPUS as a possible treatment approach for delayed union and non-union of displaced femoral neck fractures in young patients before considering further surgical intervention.

Human pseudoarthrosis tissue contains cells with osteogenic potential.

INTRODUCTION: Nonunion is a challenging problem that may occur after certain bone fractures. The treatment of nonunion is closely related to its type. To develop an effective treatment strategy for each type of nonunion, biological analysis of nonunion tissue is essential. Pseudoarthrosis is a distinct pathologic entity of nonunion. To understand the pathology of pseudoarthrosis, we investigated the cellular properties of pseudoarthrosis tissue-derived cells (PCs) in vitro. PATIENTS AND METHODS: PCs were isolated from four patients with pseudoarthrosis and cultured. Cells were evaluated for cell-surface protein expression by using flow cytometry. Osteogenic differentiation capacity was assessed by using Alizarin Red S staining, alkaline phosphatase (ALP) activity assay, and reverse transcription polymerase chain reaction (RT-PCR) after osteogenic induction. Chondrogenic differentiation capacity was assessed via Safranin O staining and RT-PCR after chondrogenic induction. RESULTS: PCs were consistently positive for the mesenchymal stem cell-related markers CD29, CD44, CD105, and CD166, but were negative for the haematopoietic-lineage markers CD31, CD34, CD45, and CD133. Alizarin Red S staining revealed that PCs formed a mineralised matrix that was rich in calcium deposits after osteogenic induction. ALP activity under osteogenic conditions was significantly higher than that under control conditions. Gene expression of ALP, Runx2, osterix, osteocalcin, and bone sialoprotein was observed in PCs cultured under osteogenic conditions. Induced pellets were negatively stained by Safranin O staining. Gene expression of aggrecan, collagen II, collagen X, SOX5, and SOX9 was not observed. CONCLUSION: We have shown for the first time the properties of cells in patients with pseudoarthrosis. Our results indicated that osteogenic cells existed in the pseudoarthrosis tissue. This study might provide insights into understanding the pathology of pseudoarthrosis and improving the treatment for pseudoarthrosis.

Profiling microRNA expression during fracture healing.

BACKGROUND: The discovery of microRNA (miRNA) has revealed a novel type of regulatory control for gene expression. Increasing evidence suggests that miRNA regulates chondrocyte, osteoblast, and osteoclast differentiation and function, indicating miRNA as key regulators of bone formation, resorption, remodeling, and repair. We hypothesized that the functions of certain miRNAs and changes to their expression pattern may play crucial roles during the process of fracture healing. METHODS: Standard healing fractures and unhealing fractures produced by periosteal cauterization at the fracture site were created in femurs of seventy rats, with half assigned to the standard healing fracture group and half assigned to the nonunion group. At post-fracture days 3, 7, 10, 14, 21, and 28, total RNA including miRNA was extracted from the newly generated tissue at the fracture site. Microarray analysis was performed with miRNA samples from each group on post-fracture day 14. For further analysis, we selected highly up-regulated five miRNAs in the standard healing fracture group from the microarray data. Real-time PCR was performed with miRNA samples at each time point above mentioned to compare the expression levels of the selected miRNAs between standard healing fractures and unhealing fractures and investigate their time-course changes. RESULTS: Microarray and real-time polymerase chain reaction (PCR) analyses on day 14 revealed that five miRNAs, miR-140-3p, miR-140-5p, miR-181a-5p, miR-181d-5p, and miR-451a, were significantly highly expressed in standard healing fractures compared with unhealing fractures. Real-time PCR analysis further revealed that in standard healing fractures, the expression of all five of these miRNAs peaked on day 14 and declined thereafter. CONCLUSION: Our results suggest that the five miRNAs identified using microarray and real-time PCR analyses may play important roles during fracture healing. These findings provide valuable information to further understand the molecular mechanism of fracture healing and may lead to the development of miRNA-based tissue engineering strategies to promote fracture healing.

Regulation of lubricin/superficial zone protein by Wnt signalling in bovine synoviocytes.

Lubricin, homologous to superficial zone protein (SZP), functions as a boundary lubricant in articular cartilage and plays an essential role in the maintenance of joint function and homeostasis. Wnt signalling plays a key role in joint development, including synovial joint formation, and several Wnt proteins are expressed in the synovium and articular cartilage in arthritis. The aim of this study was to determine the role of Wnt signalling on SZP accumulation in synoviocytes. Isolated synoviocytes from bovine knee joints were cultured with Wnt proteins (Wnt-3a and Wnt-5a) and antagonists or agonists of the Wnt-ß-catenin pathway or Wnt-Ca(2+) pathway in serum-free chemically defined medium. SZP accumulation in the culture medium was determined by enzyme-linked immunosorbent assay. Wnt-3a suppressed SZP accumulation via a Wnt-ß-catenin-dependent pathway. In contrast, Wnt-5a stimulated SZP accumulation via a ß-catenin independent pathway. The present investigation provides novel insights into the role of the Wnt signalling pathways in SZP accumulation in synoviocytes and their roles in the homeostasis of normal joints.

Regeneration of Articular Cartilage Surface: Morphogens, Cells, and Extracellular Matrix Scaffolds.

The articular cartilage is a well-organized tissue for smooth and friction-free joint movement for locomotion in animals and humans. Adult articular cartilage has a very low self-regeneration capacity due to its avascular nature. The regeneration of articular cartilage surface is critical to prevent the progression to osteoarthritis (OA). Although various joint resurfacing procedures in experimental articular cartilage defects have been developed, no standardized clinical protocol has yet been established. The three critical ingredients for tissue regeneration are morphogens and growth factors, cells, and scaffolds. The concepts based on the regeneration triad have been extensively investigated in animal models. However, these studies in animal models have demonstrated variable results and outcomes. An optimal animal model must precisely mimic and model the sequence of events in articular cartilage regeneration in human. In this article, the progress and remaining challenges in articular cartilage regeneration in animal models are reviewed. The role of individual morphogens and growth factors in cartilage regeneration has been investigated. In normal articular cartilage homeostasis, morphogens and growth factors function sequentially in tissue regeneration. Mesenchymal stem cell-based repair of articular cartilage defects, performed with or without various growth factors and scaffolds, has been widely attempted in animal models. Stem cells, including embryonic and adult stem cells and induced pluripotent stem cells, have also been reported as attractive cell sources for articular cartilage surface regeneration. Several studies with regard to scaffolds have been advanced, including recent investigations based on nanomaterials, functional mechanocompatible scaffolds, multilayered scaffolds, and extracellular matrix scaffolds for articular cartilage surface regeneration. Continuous refinement of animal models in chondral and osteochondral defects provide opportunities that support further advances in tissue engineering for the optimal articular cartilage surface regeneration.

Rate of venous thromboembolism after complex lower-limb fracture surgery without pharmacological prophylaxis.

PURPOSE: To evaluate the rate of venous thromboembolism (VTE) after complex lower-limb fracture surgery without pharmacological prophylaxis. METHODS: Between July 2005 and April 2009, 11 men and 6 women (mean age, 45 years) underwent temporary external fixation followed by internal fixation for a total of 3 femoral and 20 tibial fractures. Patients were screened for VTE using contrast-enhanced computed tomography (CT) when the D-dimer value was beyond normal range. Between May 2009 and December 2012, 18 men and 4 women (mean age, 45 years) underwent temporary external fixation followed by internal fixation for a total of 8 femoral and 20 tibial fractures. Patients were routinely screened for deep vein thrombosis (DVT) using ultrasonography and for pulmonary embolism (PE) using contrast-enhanced CT (in high-risk patients with multiple injuries or pelvic and acetabular fractures). In both groups, pharmacological prophylaxis was not used; only mechanical prophylaxis was used. RESULTS: The 2 groups were comparable in terms of age, sex, and proportion of bilateral external fixation, open fracture, and associated injury, except that the duration of external fixation was longer in the early group (29.8±28.0 vs. 11.6±4.0 days, p=0.001). Of the 17 patients in the early group, 5 developed VTEs (3 DVTs and 2 PEs) at a mean of 13.6 (range, 6-20) days after injury; 2 of whom developed VTE during the external fixation period. Of the 22 patients in the later period, 10 developed VTEs (10 DVTs and 4 PEs) at a mean of 13 (range, 5-33) days after injury; 7 of whom developed VTE during the external fixation period. CONCLUSION: The rate of VTE was high in patients undergoing complex lower-limb fracture surgery without pharmacological prophylaxis.