Despite moderate success in clinical applications, outcome of tendon grafts employed for anterior cruciate ligament (ACL) reconstruction remains unsatisfactory. This study investigated the effects of hyperbaric oxygen (HBO) on neovascularization at the tendon‐bone junction, collagen fibers of the tendon graft, and the tendon graft‐bony interface incorporated into the osseous tunnel in rabbits. Forty rabbits were assigned to two groups. The HBO group was exposed to 100% oxygen at 2.5 atmospheres pressure for 2 h daily, 5 consecutive days in a week. The control group was maintained in cages exposed to normal air. Histological studies of 12 rabbits were performed postoperatively at 6, 12, and 18 weeks. Biomechanical studies of 24 rabbits were conducted postoperatively at 12 and 18 weeks. Electron microscopy (EM) analyses of four rabbits were performed postoperatively at 18 weeks. Experimental results demonstrated that a higher number of Sharpey's fibers bridged the newly formed fibrocartilage and graft in the HBO group than in the control group. In addition, HBO treatment increased neovascularization and enhanced the incorporation of the progressive interface between tendon graft and bone. Biomechanical analysis showed that the HBO group achieved higher maximal pullout strength than the control group. Examination by EM showed that HBO treatment resulted in regenerated collagen fibers with increased compaction and regularity. Based on experimental results, HBO treatment is a treatment modality that potentially improves outcome following ACL reconstruction. © 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 25:636–645, 2007

This study presents a novel biodegradable alginate delivery system for antibiotics and bone cells to treat infected bone defects. About 2 × 10⁷ New Zealand rabbit mesenchymal stem cells (MSCs) and 5 mL vancomycin solution (50 mg/mL) were added to 5 mL of 2.5% (w/v) sodium alginate solution to form biodegradable antibiotic and MSCs alginate beads 3 mm in diameter. The alginate beads were then cultured in an osteogenic medium for 14 days. The profiles of antibiotics released from the alginate beads were evaluated using the method of high performance liquid chromatography (HPLC). The expression of osteogenic genes, including Cbfa1 and osteopontin, in the alginate beads was determined by reverse transcription‐polymerase chain reaction (RT‐PCR) analysis. The alkaline phosphatase activity, calcium level, and mineral deposition of the cultured cells within the alginate beads were assessed. Analytical results demonstrated that the concentrations of vancomycin eluted from the alginate beads were, for 14 days, well above the minimal inhibitory concentration of Staphylococcus aureus. Osteopontin and mRNA of Cbfa1 were detected and increased alkaline phosphatase activity and calcium levels were noted, as was a substantial mineral deposition in cultured MSCs. The PKH 26‐labeled MSCs and vancomycin alginate beads were implanted in rabbit bony cavities for in vivo analysis. Implanted PKH 26‐labeled MSCs were identified in the newly formed bony trabeculae in all specimens at 2 and 4 months after implantation and there was abundant mineral deposition. The results of in vitro study demonstrated sustained elution of vancomycin from the alginate carrier for 14 days and good osteogenic differentiation of cultured MSCs in the alginate carrier matrix. The results of in vivo study demonstrated the implanted MSCs participating in new bone formation. Based on experimental evidence, development of a biodegradable alginate carrier system for antibiotics and bone cells is possible, providing a potential treatment procedure for infected bone defects. © 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 25:62–72, 2007

We tested the hypothesis that the dose of recombinant human bone morphogenetic protein‐2 (rhBMP‐2) required to induce spine fusion can be reduced by combination with mesenchymal stem cells (MSCs). Twenty‐four adult rabbits underwent posterolateral intertransverse fusion at the L4–L5 level. The animals were divided into four groups based on the implant material: autologous iliac graft, Alginate‐MSCs composite, Alginate‐BMP‐2‐MSCs composite, and Alginate‐BMP‐2 composite. After 16 weeks, the rabbits were euthanized for radiographic examination, manual palpation, biomechanical testing, and histology. Radiographic union of 12 intertransverse fusion areas for the autogenous iliac graft, Alginate‐MSCs, Alginate‐BMP‐2‐MSCs, and Alginate‐BMP‐2 groups was 11, 8, 11, and 0, respectively. Moreover, manual palpation of six fusion segments in each subgroup found solid union to be 6, 1, 5, and 0, respectively. The average torques at failure of the first three groups were 2278 ± 135, 1943 ± 140, and 2334 ± 187 N‐mm, respectively. The failure torque did not differ significantly between the autograft and Alginate‐BMP‐2‐MSCs groups; both groups were significantly higher than the Alginate‐MSCs group. The results indicate that MSCs delivered with in vitro cellular doses of rhBMP‐2 are more osteoinductive than MSCs without rhBMP‐2. In combination with MSCs, a low dose (2.5 µg) of rh‐BMP‐2 could enhance bone formation and posterolateral spine fusion success in the rabbit model. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:380–384, 2009

The expression of both high-mobility group box 1 (HMGB1) and receptor for advanced glycation end-products (RAGE) is upregulated in degenerated discs. HMGB1 is known to function as a coupling factor between hypoxia and inflammation in arthritis, and this inflammatory response is modulated by microRNAs (miRNAs), with miR-107 expression downregulated during hypoxia. In this study, we investigated the regulation of the miR-107/HMGB1/RAGE pathway in degenerated nucleus pulposus cells (NPCs) after hyperbaric oxygen (HBO) treatment. NPCs were separated from human degenerated intervertebral disc tissues. The control cells were maintained in 5% CO2/95% air, and the hyperoxic cells were exposed to 100% O2 at 2.5 atmospheres absolute. MiRNA expression profiling was performed via microarray and confirmed by real-time PCR, and miRNA target genes were identified using bioinformatics and luciferase reporter assays. The cellular protein and mRNA levels of HMGB1, RAGE, and inducible nitric oxide synthase (iNOS) were assessed, and the phosphorylation of MAPK (p38MAPK, ERK, and JNK) was evaluated. Additionally, cytosolic and nuclear fractions of the IκBα and NF-κB p65 proteins were analyzed, and secreted HMGB1 and metalloprotease (MMP) levels in the conditioned media were quantified. Using microarray analyses, 96 miRNAs were identified as upregulated and 66 downregulated following HBO treatment. Based on these results, miR-107 was selected for further investigation. Bioinformatics analyses indicated that the 3′ untranslated region of the HMGB1 mRNA contained the “seed-matched-sequence” for hsa-miR-107, which was validated via dual-luciferase reporter assays. MiR-107 was markedly induced by HBO, and simultaneous suppression of HMGB1 was observed in NPCs. Knockdown of miR-107 resulted in upregulation of HMGB1 expression in HBO-treated cells, and HBO treatment downregulated the mRNA and protein levels of HMGB1, RAGE, and iNOS and the secretion of HMGB1. In addition, HBO treatment upregulated the protein levels of cytosolic IκBα and decreased the nuclear translocation of NF-κB in NPCs. Moreover, HBO treatment downregulated the phosphorylation of p38MAPK, ERK, and JNK and significantly decreased the secretion of MMP-3, MMP-9, and MMP-13. HBO inhibits pathways related to HMGB1/RAGE signaling via upregulation of miR-107 expression in degenerated human NPCs.

Atrophic nonunion of femoral shaft fracture after intramedullary (IM) nailing is uncommon. The treatment for femoral shaft aseptic atrophic non-union remained controversial. The aim of this study was to compare the surgical results between exchanging reamed nailing (ERN) and augmentative antirotational plating (AAP) for femoral shaft aseptic atrophic nonunion. We retrospectively reviewed the patients with femoral shaft nonunion between the year of 2014 and 2015. The patients with nonunion after plate osteosynthesis, septic nonunion, hypertrophic nonunion, additional surgery during revision surgery were excluded. All the patients were followed up at least 12 months. Overall, the union rate after revision surgery was 70.8%. The union rate was significantly higher in the AAP group than in the ERN group. Operating time was also significantly shorter in the AAP group. Regarding the location of nonunion, the union rate was comparable between groups for isthmic nonunions. However, for non-isthmic nonunions, the union rate was significantly higher and operating time was significantly shorter in the AAP group. AAP showed an overall higher union rate for management of femoral shaft aseptic atrophic nonunion compared with ERN. Especially for non-isthmic femoral shaft atrophic nonunions, AAP provided a significantly higher union rate and significantly shorter operating time.

Heat shock proteins (HSPs), inflammatory cytokines, nitric oxide (NO), and localized hypoxia‐induced apoptosis are thought to be correlated to the degree of cartilage injury. We investigated the effect of hyperbaric oxygen (HBO) on (1) interleukin‐1β (IL‐1β)‐induced NO production and apoptosis of rabbit chondrocytes and (2) healing of articular cartilage defects. For the in vitro study, RT‐PCR and Western blotting were performed to detect mRNA and protein expressions of HSP70, inducible NO synthase (iNOS), and caspase 3 in IL‐1β‐treated chondrocytes. To clarify that the HSP70 was necessary for anti‐iNOS and anti‐apoptotic activity by HBO, we treated the cells with an HSP70 inhibitor, KNK437. For the in vivo study, cartilage defects were created in rabbits. The HBO group was exposed to 100% oxygen at 2.5 ATA for 1.5 h a day for 10 weeks. The control group was exposed to normal air. After sacrifice, specimen sections were sent for examination using a scoring system. Immunohistochemical analyses were performed to detect the expressions of iNOS, HSP70, and caspase 3. Our results suggested that HBO upregulated the mRNA and protein expressions of HSP70 and suppressed those of iNOS and caspase 3 in chondrocytes. KNK437 inhibited the HBO‐induced downregulation of iNOS and casapase 3 activities. The histological scores showed that HBO markedly enhanced cartilage repair. Immunohistostaining showed that HBO enhanced HSP70 expression and suppressed iNOS and caspase 3 expressions in chondrocytes. Accordingly, HBO treatment prevents NO‐induced apoptosis in articular cartilage injury via enhancement of the expression of heat shock protein 70. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 376–384, 2013