MgO-enhanced ß-TCP promotes osteogenesis in both in vitro and in vivo rat models.

Allogeneic bone grafts are used to treat bone defects in orthopedic surgery, but the osteogenic potential of artificial bones remains a challenge. In this study, we developed a ß-tricalcium phosphate (ß-TCP) formulation containing MgO, ZnO, SrO, and SiO2 and compared its bone-forming ability with that of ß-TCP without biological elements. We prepared ß-TCP discs with 60% porosity containing 1.0 wt% of these biological elements. ß-TCP scaffolds were loaded with bone marrow-derived mesenchymal stem cells (BMSC) from 7-week-old male rats and cultured for 2 weeks. ALP activity and mRNA expression of osteogenic markers were evaluated. In addition, scaffolds were implanted subcutaneously in rats and analyzed after 7 weeks. In vitro, the MgO group showed lower Ca concentrations and higher osteogenic marker expression compared to controls. In vivo, the MgO group showed higher ALP activity compared to controls, and RT-qPCR analysis showed significant expression of BMP2 and VEGF. Histopathology, fluorescent immunostaining, and micro-CT also showed relatively better bone formation in the MgO group. ß-TCP with MgO may enhance bone morphology in vitro and in vivo and improve the prognosis of patients with substantial and refractory bone defects.

Optimal retractor insertion point for nerve safety during total hip arthroplasty: an anatomical study on the femoral and sciatic nerves in relation to hip motion.

BACKGROUND: Nerve injury is one of the most serious complications of total hip arthroplasty (THA). It is suspected to be a result from nerve compression or direct injury caused by an acetabular retractor. The anatomical relationship between the acetabular rim and the femoral and sciatic nerves, including hip motion, has not been investigated. This study aimed to identify the optimal position for retractor insertion during THA to prevent nerve damage. METHODS: A total of 28 hip joints from 14 freshly frozen cadavers were used. Using an anterolateral approach, each cadaver was immobilised in the lateral decubitus position and deployed to measure the distance between the nerves and the acetabular rim, while the hip joint was changed to the extension, neutral, and flexion positions. RESULTS: Three femoral nerves were closest to the anterior margin of the acetabulum at 90° and 120° of extension and farthest away at 30° of flexion. The sciatic nerve was closest to the posterior margin of the acetabulum at 90° and 120° of flexion and farthest away at 30° and 150° of extension compared with the other points. CONCLUSIONS: To prevent nerve damage during THA, we suggest that the retractor be inserted at the points where the nerves are the farthest away, such as at 30° and 150°. The femoral and sciatic nerves vary in their movements depending on the hip position. Therefore, the safe insertion of a retractor is recommended for hip flexion of the femoral nerve and extension of the sciatic nerve. Additionally, it is important to carefully insert the retractor along the acetabular margin without penetrating the joint capsule. Overall, this study provides valuable insights into the anatomical location and movement of the femoral and sciatic nerves in relation to hip motion and can help inform surgical techniques for safer THA.

Silicate/zinc-substituted strontium apatite coating improves the osteoinductive properties of ß-tricalcium phosphate bone graft substitute.

BACKGROUND: ß-Tricalcium phosphate (ß-TCP) is a popular synthetic bone graft substitute with excellent osteoconductive properties and bioabsorbability. However, its osteoinductive properties are inferior to those of autologous or allogeneic bone. Trace elements such as strontium (Sr), silica (Si), and zinc (Zn) have been reported to promote osteogenesis in materials. In this study, we aimed to determine whether a Si/Zn-substituted Sr apatite coating of ß-TCP could enhance osteoinductive properties. METHODS: The apatite-coated ß-TCP disks were prepared using nanoparticle suspensions of silicate-substituted Sr apatite (SrSiP) or silicate- and Zn-co-substituted Sr apatite (SrZnSiP). Bone marrow mesenchymal cells (BMSCs) from rat femur were cultured and subsequently seeded at a density of 1.0 × 106/cm2 onto apatite-coated and non-coated ß-TCP disks. In vitro, the ß-TCP disks were then placed in osteogenic medium, and lactate dehydrogenase (LDH) activity was measured from supernatants after culture for 2 days. Additionally, after culture for 14 days, the mRNA expression of genes encoding osteocalcin (OC), alkaline phosphatase (ALP), bone morphogenetic protein-2 (BMP-2), and vascular endothelial growth factor (VEGF) was evaluated by qRT-PCR. In vivo, the ß-TCP disks were transplanted subcutaneously into rats that were sacrificed after 4 weeks. Then, the harvested disks were evaluated biochemically (ALP activity, OC content, mRNA expression of OC, ALP, BMP-2, and VEGF measured by qRT-PCR), radiologically, and histologically. RESULTS: Significantly higher mRNA expression of almost all evaluated osteogenic and angiogenic genes was observed in the SrZnSiP and SrSiP groups than in the non-coated group, with no significant cytotoxicity elicited by the apatite coating in vitro. Moreover, in vivo, the SrZnSiP and SrSiP groups showed significantly higher osteogenic and angiogenic gene expression and higher ALP activity and OC content than the non-coated group (P < 0.05). Radiological and histopathological findings revealed abundant bone formation in the apatite-coated group. CONCLUSIONS: Our findings indicate that apatite coating of ß-TCP improves osteoinductive properties without inducing significant cytotoxicity.

Histological Findings in Infected and Noninfected Second Stage Revision Knee Arthroplasties.

Tissues from a periprosthetic joint infection (PJI) of the knee contain a heavy neutrophil polymorph (NP) infiltrate (> 5 NPs per high-powered field [HPF] by Musculoskeletal Infection Society [MSIS] criteria). PJI of the knee can be treated by a two-stage procedure and our aim was to determine whether the MSIS histological criteria for PJI diagnosis are valid in a second-stage revision knee arthroplasty. Periprosthetic tissues from 45 second-stage revision knee cases were analyzed histologically by hematoxylin-eosin and chloroacetate esterase (CAE) staining for the identification of NPs. The number of NPs was determined semiquantitatively and results correlated with the microbiological and clinical findings. In 9 of the 45 cases, an organism was cultured in two or more samples, meeting MSIS microbiological criteria for a definite diagnosis of PJI; histologically, seven of these cases contained > 5 per NPs per HPF on average, with the remaining two cases containing 1 NP and 2 NPs per HPF. In noninfected second-stage revisions, NPs were not seen in 30 cases with 6 cases showing less than 1 NP per HPF on average. The sensitivity, specificity, accuracy, and positive and negative predictive values of MSIS histological criteria (> 5 NPs per HPF) to diagnose PJI were 78%, 100%, 96%, 100%, and 95%, respectively. MSIS histological criteria for the diagnosis of PJI are valid for most but not all infected second-stage revision knee arthroplasties. Correlation of histology with clinical, microbiology and other laboratory findings is required to establish a diagnosis of PJI in second-stage revision knee arthroplasties.

Supplying osteogenesis to dead bone using an osteogenic matrix cell sheet.

PURPOSE: To evaluate whether osteogenic matrix cell sheets can supply osteogenesis to dead bone. METHODS: Femur bone fragments (5 mm in length) were obtained from Fisher 344 rats and irradiated by a single exposure of 60 Gy to produce bones that were no longer viable. Osteogenic matrix cell sheets were created from rat bone marrow-derived stromal cells (BMSCs). After wrapping the dead bone with an osteogenic matrix cell sheet, it was subcutaneously transplanted into the back of a rat and harvested after 4 weeks. Bone formation around the dead bone was evaluated by X-ray imaging and histology. Alkaline phosphatase (ALP) and osteocalcin (OC) mRNA expression levels were measured to confirm osteogenesis of the transplanted bone. The contribution of donor cells to bone formation was assessed using the Sry gene and PKH26. RESULTS: After the cell sheet was transplanted together with dead bone, X-ray images showed abundant calcification around the dead bone. In contrast, no newly formed bone was seen in samples that were transplanted without the cell sheet. Histological sections also showed newly formed bone around dead bone in samples transplanted with the cell sheet, whereas many empty lacunae and no newly formed bone were observed in samples transplanted without the cell sheet. ALP and OC mRNA expression levels were significantly higher in dead bones transplanted with cell sheets than in those without a cell sheet (P < 0.01). Sry gene expression and cells derived from cell sheets labeled with PKH26 were detected in samples transplanted with a cell sheet, indicating survival of donor cells after transplantation. CONCLUSION: Our study indicates that osteogenic matrix cell sheet transplantation can supply osteogenesis to dead bone.

Role of LIGHT in the pathogenesis of joint destruction in rheumatoid arthritis.

AIM: To characterise the role of substitutes for receptor-activator nuclear factor kappa-B ligand (RANKL) in rheumatoid arthritis (RA) joint destruction. METHODS: Synovial fluid (SF) macrophages isolated from the knee joint of RA patients were incubated with 25 ng/mL macrophage-colony stimulating factor (M-CSF) and 50 ng/mL LIGHT (lymphotoxin-like, exhibits inducible expression and competes with herpes simplex virus glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes) in the presence and absence of 25 ng/mL RANKL and 100 ng/mL osteoprotegerin (OPG) on glass coverslips and dentine slices. Osteoclastogenesis was assessed by the formation of multinucleated cells (MNCs) expressing tartrate-resistant acid phosphatase (TRAP) on coverslips and the extent of lacunar resorption pit formation on dentine slices. The concentration of LIGHT in RA and osteoarthritis (OA) synovial fluid was measured by an enzyme-linked immunosorbent assay (ELISA) and the expression of LIGHT in RA and OA synovium was determined by immunohistochemistry using an indirect immunoperoxidase technique. RESULTS: In cultures of RA SF macrophages treated with LIGHT and M-CSF, there was significant formation of TRAP + MNCs on coverslips and extensive lacunar resorption pit formation on dentine slices. SF-macrophage-osteoclast differentiation was not inhibited by the addition of OPG, a decoy receptor for RANKL. Resorption pits were smaller and less confluent than in RANKL-treated cultures but the overall percentage area of the dentine slice resorbed was comparable in LIGHT- and RANKL-treated cultures. LIGHT significantly stimulated RANKL-induced lacunar resorption compared with RA SF macrophages treated with either RANKL or LIGHT alone. LIGHT was strongly expressed by synovial lining cells, subintimal macrophages and endothelial cells in RA synovium and the concentration of LIGHT was much higher in RA compared with OA SF. CONCLUSION: LIGHT is highly expressed in RA synovium and SF, stimulates RANKL-independent/dependent osteoclastogenesis from SF macrophages and may contribute to marginal erosion formation.

Talonavicular ligament: prevalence of injury in ankle sprains, histological analysis and hypothesis of its biomechanical function.

OBJECTIVE: To assess the prevalence of injury of the talonavicular ligament (TNL) in ankle sprains, its anatomy and the stability of the talonavicular joint (TNJ) before and after dividing the TNL in a cadaver. METHODS: During a prospective study of 100 patients to assess the outcome of ankle injuries, we noted high incidence of TNL injuries; we will discuss here the TNL findings. Each patient had undergone ultrasound and cone beam CT examination of the ankle. Six TNLs were dissected off fresh-frozen cadaveric feet for histological analysis. In further six cadaveric feet, the stability of the TNJ was assessed by mechanical stress before and after division of the TNL; movement at the joint was assessed by measuring the distance between the talus and navicular bone [talonavicular distance (TD)] using ultrasound. The TD was measured on ten randomly selected ultrasound images by three independent observers and repeated twice by a single observer to determine the inter- and intraobserver reliability. RESULTS: 21% of the patients had an injury to the TNL. Histological examination demonstrated a dense connective tissue composed of bundles of collagen in parallel arrangement along the ligament length. The interobserver and intraobserver reliability of the TD showed almost perfect agreement. Displacement at the TNJ after stress with the TNL intact measured 0.18 ± 0.08 cm and 0.29 ± 0.07 cm (p < 0.005) when divided. CONCLUSION: The TNL is surprisingly commonly injured in ankle sprains. Its anatomy and histology suggest a role in tensile force transmission during the windlass mechanism in gait. Advances in knowledge: Injury to the TNL is common and has not been described. Its anatomy suggests resistance to tensile forces and its injury allows excessive movement at the TNJ.

Utility of tricalcium phosphate and osteogenic matrix cell sheet constructs for bone defect reconstruction.

AIM: To determine the effects of transplanting osteogenic matrix cell sheets and beta-tricalcium phosphate (TCP) constructs on bone formation in bone defects. METHODS: Osteogenic matrix cell sheets were prepared from bone marrow stromal cells (BMSCs), and a porous TCP ceramic was used as a scaffold. Three experimental groups were prepared, comprised of TCP scaffolds (1) seeded with BMSCs; (2) wrapped with osteogenic matrix cell sheets; or (3) both. Constructs were implanted into a femoral defect model in rats and bone growth was evaluated by radiography, histology, biochemistry, and mechanical testing after 8 wk. RESULTS: In bone defects, constructs implanted with cell sheets showed callus formation with segmental or continuous bone formation at 8 wk, in contrast to TCP seeded with BMSCs, which resulted in bone non-union. Wrapping TCP constructs with osteogenic matrix cell sheets increased their osteogenic potential and resulting bone formation, compared with conventional bone tissue engineering TCP scaffolds seeded with BMSCs. The compressive stiffness (mean ± SD) values were 225.0 ± 95.7, 30.0 ± 11.5, and 26.3 ± 10.6 MPa for BMSC/TCP/Sheet constructs with continuous bone formation, BMSC/TCP/Sheet constructs with segmental bone formation, and BMSC/TCP constructs, respectively. The compressive stiffness of BMSC/TCP/Sheet constructs with continuous bone formation was significantly higher than those with segmental bone formation and BMSC/TCP constructs. CONCLUSION: This technique is an improvement over current methods, such as TCP substitution, and is useful for hard tissue reconstruction and inducing earlier bone union in defects.

Osteogenic Matrix Cell Sheets Facilitate Osteogenesis in Irradiated Rat Bone.

Reconstruction of large bone defects after resection of malignant musculoskeletal tumors is a significant challenge in orthopedic surgery. Extracorporeal autogenous irradiated bone grafting is a treatment option for bone reconstruction. However, nonunion often occurs because the osteogenic capacity is lost by irradiation. In the present study, we established an autogenous irradiated bone graft model in the rat femur to assess whether osteogenic matrix cell sheets improve osteogenesis of the irradiated bone. Osteogenic matrix cell sheets were prepared from bone marrow-derived stromal cells and co-transplanted with irradiated bone. X-ray images at 4 weeks after transplantation showed bridging callus formation around the irradiated bone. Micro-computed tomography images at 12 weeks postoperatively showed abundant callus formation in the whole circumference of the irradiated bone. Histology showed bone union between the irradiated bone and host femur. Mechanical testing showed that the failure force at the irradiated bone site was significantly higher than in the control group. Our study indicates that osteogenic matrix cell sheet transplantation might be a powerful method to facilitate osteogenesis in irradiated bones, which may become a treatment option for reconstruction of bone defects after resection of malignant musculoskeletal tumors.