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The increasing incidence of physiologic/pathologic conditions that impair the otherwise routine healing of endochondral bone fractures and the occurrence of severe bone injuries necessitate novel approaches to enhance clinically challenging bone fracture repair. To promote the healing of nonunion fractures, we tested an approach that used two small molecules to sequentially enhance cartilage development and conversion to the bone in the callus of a murine femoral segmental defect nonunion model of bone injury. Systemic injections of smoothened agonist 21k (SAG21k) were used to stimulate chondrogenesis through the activation of the sonic hedgehog (SHH) pathway early in bone repair, while injections of the prolyl hydroxylase domain (PHD)2 inhibitor, IOX2, were used to stimulate hypoxia signaling‐mediated endochondral bone formation. The expression of SHH pathway genes and Phd2 target genes was increased in chondrocyte cell lines in response to SAG21k and IOX2 treatment, respectively. The segmental defect responded to sequential systemic administration of these small molecules with increased chondrocyte expression of PTCH1, GLI1, and SOX9 in response to SAG and increased expression of hypoxia‐induced factor‐1α and vascular endothelial growth factor‐A in the defect tissues in response to IOX2. At 6 weeks postsurgery, the combined SAG–IOX2 therapy produced increased bone formation in the defect with the bony union over the injury. Clinical significance: This therapeutic approach was successful in promoting cartilage and bone formation within a critical‐size segmental defect and established the utility of a sequential small molecule therapy for the enhancement of fracture callus development in clinically challenging bone injuries. [ABSTRACT FROM AUTHOR]

Some of the earliest success in de novo tissue generation was in bone tissue, and advances, facilitated by the use of endogenous and exogenous progenitor cells, continue unabated. The concept of one health promotes shared discoveries among medical disciplines to overcome health challenges that afflict numerous species. Carefully selected animal models are vital to development and translation of targeted therapies that improve the health and well‐being of humans and animals alike. While inherent differences among species limit direct translation of scientific knowledge between them, rapid progress in ex vivo and in vivo de novo tissue generation is propelling revolutionary innovation to reality among all musculoskeletal specialties. This review contains a comparison of bone deposition among species and descriptions of animal models of bone restoration designed to replicate a multitude of bone injuries and pathology, including impaired osteogenic capacity. [ABSTRACT FROM AUTHOR]

Bone stress injuries are commonly due to repetitive loading, as often described in competitive athletes or military recruits. The underlying pathophysiology of bone stress injuries is multifactorial. The present cross‐sectional study investigated (i) cortical and trabecular bone microstructure as well as volumetric bone mineral density in subjects with bone stress injuries at the tibial diaphysis, measured at the distal tibia and the distal radius by means of high‐resolution peripheral quantitative computed tomography (CT), (ii) areal bone mineral density using dual‐energy X‐ray absorptiometry as well as calcaneal dual X‐ray absorptiometry and laser, and (iii) the influence on bone turnover markers of formation and resorption at the early phase after injury. A total of 26 Caucasian male professional soldiers with post‐training bone stress injury at the tibial diaphysis were included (case group). A total of 50 male, Caucasian professional soldiers from the same military institution served as controls (control group). High‐resolution peripheral quantitative CT revealed a higher total area at the radius within the case group. Cortical bone mineral density was reduced at the radius and tibia within the case group. The trabecular number and trabecular thickness were reduced at the tibia in the case group. The trabecular network was more inhomogeneous at the radius and tibia within the case group. Calcaneal dual X‐ray absorptiometry and laser was significantly reduced in the case group. This study quantified differences in bone microstructure among otherwise healthy individuals. Differences in bone microarchitecture may impair the biomechanical properties by increasing the susceptibility to sustain bone stress injuries. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2516–2523, 2019 [ABSTRACT FROM AUTHOR]

Traumatic composite bone‐muscle injuries, such as open fractures, often require multiple surgical interventions and still typically result in long‐term disability. Clinically, a critical indicator of composite injury severity is vascular integrity; vascular damage alone is sufficient to assign an open fracture to the most severe category. Challenging bone injuries are often treated with bone morphogenetic protein 2 (BMP‐2), an osteoinductive growth factor, delivered on collagen sponge. Previous studies in a composite defect model found that a minimally bridging dose in the segmental defect model was unable to overcome concomitant muscle damage, but the effect of BMP dose on composite injuries has not yet been studied. Here, we test the hypotheses that BMP‐2‐mediated functional regeneration of composite extremity injuries is dose dependent and can be further enhanced via co‐delivery of adipose‐derived microvascular fragments (MVF), which have been previously shown to increase tissue vascular volume. Although MVF did not improve healing outcomes, we observed a significant BMP‐2 dose‐dependent increase in regenerated bone volume and biomechanical properties. This is the first known report of an increased BMP‐2 dose improving bone healing with concomitant muscle damage. While high dose BMP‐2 delivery can induce heterotopic ossification (HO) and increased inflammation, the maximum 10 μg dose used in this study did not result in HO and was associated with a lower circulating inflammatory cytokine profile than the low dose (2.5 μg) group. These data support the potential benefits of an increased, though still moderate, BMP‐2 dose for treatment of bone defects with concomitant muscle damage. Published 2019. This article is a U.S. Government work and is in the public domain in the USA. J Orthop Res [ABSTRACT FROM AUTHOR]

ABSTRACT Insufficient primary stability of screws in bone leads to screw loosening and failure. Unlike conventional continuum finite-element models, micro-CT based finite-element analysis (micro-FE) is capable of capturing the patient-specific bone micro-architecture, providing accurate estimates of bone stiffness. However, such in silico models for screws in bone highly overestimate the apparent stiffness. We hypothesized that a more accurate prediction of primary implant stability of screws in bone is possible by considering insertion-related bone damage. We assessed two different screw types and loading scenarios in 20 trabecular bone specimens extracted from 12 cadaveric human femoral heads ( N = 5 for each case). In the micro-FE model, we predicted specimen-specific Young's moduli of the peri-implant bone damage region based on morphometric parameters such that the apparent stiffness of each in silico model matched the experimentally measured stiffness of the corresponding in vitro specimen as closely as possible. The standard micro-FE models assuming perfectly intact peri-implant bone overestimated the stiffness by over 330%. The consideration of insertion related damaged peri-implant bone corrected the mean absolute percentage error down to 11.4% for both loading scenarios and screw types. Cross-validation revealed a mean absolute percentage error of 14.2%. We present the validation of a novel micro-FE modeling technique to quantify the apparent stiffness of screws in trabecular bone. While the standard micro-FE model overestimated the bone-implant stiffness, the consideration of insertion-related bone damage was crucial for an accurate stiffness prediction. This approach provides an important step toward more accurate specimen-specific micro-FE models. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2415-2424, 2017. [ABSTRACT FROM AUTHOR]

ABSTRACT Healing of injured cancellous bone is characterized by a transient stage of rapid bone formation throughout the traumatized bone volume, often followed by similarly rapid resorption. This is different from the slower diaphyseal healing via an external callus. We, therefore, hypothesized that antiresorptive treatment might have an earlier positive effect in cancellous bone healing than in diaphyseal fractures. One hundred and twenty-three male C57bl6 mice received either an internally stabilized diaphyseal osteotomy of the femur or a screw inserted into the tibial metaphysis. The mice were randomized to daily alendronate injections (200 μg/kg/day), or control injections, and killed for mechanical testing after 14, 21, or 28 days. The hypothesis was tested by a three-way Anova (time, site, and drug). The ultimate force was increased by bisphosphonate treatment in both models. There was a significant interaction between time, site, and drug ( p < 0.001) so that the full positive effect of alendronate was evident in the metaphysis at 14 days, but first after 28 days in the diaphysis. While the early effect in the metaphysis might be translated into earlier healing, the late effect in the diaphysis was due to delayed remodeling of the callus, which might have less clinical importance. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:793-799, 2017. [ABSTRACT FROM AUTHOR]

ABSTRACT Patients with tibial pilon fractures have a higher incidence of post-traumatic osteoarthritis than those with fractures of the tibial plateau. This may indicate that pilon fractures present a greater mechanical insult to the joint than do plateau fractures. We tested the hypothesis that fracture energy and articular fracture edge length, two independent indicators of severity, are higher in pilon than plateau fractures. We also evaluated whether clinical fracture classification systems accurately reflect severity. Seventy-five tibial plateau fractures and 52 tibial pilon fractures from a multi-institutional study were selected to span the spectrum of severity. Fracture severity measures were calculated using objective CT-based image analysis methods. The ranges of fracture energies measured for tibial plateau and pilon fractures were 3.2-33.2 Joules (J) and 3.6-32.2 J, respectively, and articular fracture edge lengths were 68.0-493.0 mm and 56.1-288.6 mm, respectively. There were no differences in the fracture energies between the two fracture types, but plateau fractures had greater articular fracture edge lengths ( p < 0.001). The clinical fracture classifications generally reflected severity, but there was substantial overlap of fracture severity measures between different classes. Similar fracture energies with different degrees of articular surface involvement suggest a possible explanation for dissimilar rates of post-traumatic osteoarthritis for fractures of the tibial plateau compared to the tibial pilon. The substantial overlap of severity measures between different fracture classes may well have confounded prior clinical studies relying on fracture classification as a surrogate for severity. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:618-624, 2017. [ABSTRACT FROM AUTHOR]

ABSTRACT Tissue engineering promises therapies ideal for treating conventional large bone injuries and defects. In the present study, we developed a novel Si-HA scaffold loaded with a synthetic BMP-2-related peptide, P28, and tested its ability to repair a critical-sized calvarial defect. We created a calvarial defect (5 mm in diameter) in the parietal bone of 32 rats and implanted one of the following biomaterials: No implant (control), Si-HA, P28/Si-HA, or rhBMP-2/Si-HA. As assessed by micro CT imaging and histological evaluations, the P28/Si-HA scaffold promoted bone recovery to a similar degree as the rhBMP-2/Si-HA scaffold. In addition, both P28/Si-HA and rhBMP-2/Si-HA promoted recovery better than Si-HA alone. The novel P28/Si-HA scaffold might represent a promising biomaterial for future bone tissue engineering applications. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1874-1882, 2016. [ABSTRACT FROM AUTHOR]

ABSTRACT The development of software applications that assist the radiographic evaluation of fracture healing could advance clinical diagnosis and expedite the identification of effective treatment strategies. A radiographic feature regularly used as an outcome measure for basic and clinical fracture healing research is new bone growth, or fracture callus. In this study, we developed OrthoRead, a portable software application that uses image-processing algorithms to detect and measure fracture callus in plain radiographs. OrthoRead utilizes an optimal boundary tracking algorithm to semi-automatically segment the cortical surface, and a novel iterative thresholding selection algorithm to then automatically segment the fracture callus. The software was validated in three steps. First, algorithm accuracy and sensitivity were analyzed using surrogate models with known callus size. Second, the callus area of distal femur fractures measured using OrthoRead was compared to callus area manually outlined by orthopaedic surgeons. Third, the callus area of ovine tibial fractures was measured using OrthoRead and compared to callus volume measured from micro-CT. The software had less than a 5% error in measuring surrogate callus, and was insensitive to changes in image resolution, image rotation, and the size of the analyzed region of interest. Strong positive correlations existed between OrthoRead and clinicians ( R2 = 0.98), and between 2D callus area and 3D callus volume ( R2 = 0.70). The average run time for OrthoRead was 3 s when using a 2.7 GHz processor. By being accurate, fast, and robust, OrthoRead can support prospective and retrospective clinical studies investigating implant efficacy, and can assist research on fracture healing mechanobiology. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1224-1233, 2016. [ABSTRACT FROM AUTHOR]

ABSTRACT Chronic opioid therapy is associated with bone loss. This led us to hypothesize that the opioid antagonists, that include naloxone, would stimulate bone formation by regulating MSC differentiation. The opioid growth factor receptor (OGFR) is a non-canonical opioid receptor that binds naloxone with high affinity whereas the native opioid growth factor, met5-enkephalin (met5), binds both the OGFR and the canonical delta opioid receptor (OPRD). Naloxone and an shRNA OGFR lentivirus were employed to disrupt the OGFR-signaling axis in cultured MSC. In parallel, naloxone was administered to bone marrow using a mouse unicortical defect model. OPRD, OGFR, and the met5-ligand were highly expressed in MSC and osteoblasts. A pulse-dose of naloxone increased mineral formation in MSC cultures in contrast to MSC treated with continuous naloxone or OGFR deficient MSC. Importantly, SMAD1 and SMAD8/9 expression increased after a pulse dose of naloxone whereas SMAD1, SMAD7, and ID1 were increased in the OGFR deficient MSC. Inhibited OGFR signaling decreased proliferation and increased p21 expression. The addition of naloxone to the unicortical defect resulted in increased bone formation within the defect. Our data suggest that novel mechanism through which signaling through the OGFR regulates osteogenesis via negative regulation of SMAD1 and p21. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1195-1205, 2016. [ABSTRACT FROM AUTHOR]

ABSTRACT Significant research exists regarding heat production during single-hole bone drilling. No published data exist regarding repetitive sequential drilling. This study elucidates the phenomenon of heat accumulation for sequential drilling with both Kirschner wires (K wires) and standard two-flute twist drills. It was hypothesized that cumulative heat would result in a higher temperature with each subsequent drill pass. Nine holes in a 3 × 3 array were drilled sequentially on moistened cadaveric tibia bone kept at body temperature (about 37°C). Four thermocouples were placed at the center of four adjacent holes and 2 mm below the surface. A battery-driven hand drill guided by a servo-controlled motion system was used. Six samples were drilled with each tool (2.0 mm K wire and 2.0 and 2.5 mm standard drills). K wire drilling increased temperature from 5°C at the first hole to 20°C at holes 6 through 9. A similar trend was found in standard drills with less significant increments. The maximum temperatures of both tools increased from <0.5°C to nearly 13°C. The difference between drill sizes was found to be insignificant ( P > 0.05). In conclusion, heat accumulated during sequential drilling, with size difference being insignificant. K wire produced more heat than its twist-drill counterparts. This study has demonstrated the heat accumulation phenomenon and its significant effect on temperature. Maximizing the drilling field and reducing the number of drill passes may decrease bone injury. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:463-470, 2016. [ABSTRACT FROM AUTHOR]

ABSTRACT The relationship between modern clinical diagnostic data, such as from radiographs or computed tomography, and the temporal biomechanical integrity of bone fracture healing has not been well-established. A diagnostic tool that could quantitatively describe the biomechanical stability of the fracture site in order to predict the course of healing would represent a paradigm shift in the way fracture healing is evaluated. This paper describes the development and evaluation of a wireless, biocompatible, implantable, microelectromechanical system (bioMEMS) sensor, and its implementation in a large animal (ovine) model, that utilized both normal and delayed healing variants. The in vivo data indicated that the bioMEMS sensor was capable of detecting statistically significant differences ( p-value <0.04) between the two fracture healing groups as early as 21 days post-fracture. In addition, post-sacrifice micro-computed tomography, and histology data demonstrated that the two model variants represented significantly different fracture healing outcomes, providing explicit supporting evidence that the sensor has the ability to predict differential healing cascades. These data verify that the bioMEMS sensor can be used as a diagnostic tool for detecting the in vivo course of fracture healing in the acute post-treatment period. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 33:1439-1446, 2015. [ABSTRACT FROM AUTHOR]

ABSTRACT Impaired bone healing can occur with numerous pathologic conditions like trauma, osteoporosis, and infection. Therefore tissue-engineering strategies that aim to enhance osteogenic differentiation of stem cells in order to accelerate bone healing are a major goal of contemporary regenerative research. In this study we cultivated mesenchymal stem cells (MSC) together with the recently patented programmable cells of monocytic origin (PCMO) to test whether co-cultures promote an osteogenic differentiation process. PCMO have recently been shown to have pluripotent characteristics and do support the regeneration processes of liver and heart diseases. Quantitative real time PCR expression profiles of osteogenic marker genes such as alkaline phosphatase in co-cultures of PCMO and MSC showed that MSC differentiated into osteoblast-like cells more rapidly as compared to mono-cultures. Alkaline phosphatase expression and enzyme activity levels were highly increased in co-cultures compared to mono-cultures of MSC. Tests for mineralized matrix formation also indicated that PCMO have a positive effect on co-cultured MSC under osteogenic culture conditions. However, analysis of collagen 1A did not show enhanced expression. In summary, PCMO obviously have the ability to promote osteogenic differentiation of MSC in vitro while their own pluripotent potential is not sufficient to develop osteoblast-like characteristics themselves. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1264-1270, 2014. [ABSTRACT FROM AUTHOR]

ABSTRACT We compared the effect of a sclerostin antibody to that of a clinically relevant dose of parathyroid hormone (PTH) in a rat model for metaphyseal bone healing. Screws of steel or poly methyl methacrylate (PMMA) were inserted bilaterally into the proximal tibia of young male rats. During 4 weeks the animals then received injections of either phosphate buffered saline (control), sclerostin antibody (25 mg/kg, twice weekly) or PTH (5 µg/kg, daily). The healing response around the screws was then assessed by mechanical testing and X-ray microtomography (µCT). To distinguish between effects on healing and general effects on the skeleton, other untraumatized bone sites and serum biomarkers were also assessed. After 4 weeks of treatment, PTH yielded a 48% increase in screw pull-out force compared to control ( p = 0.03), while the antibody had no significant effect. In contrast, the antibody increased femoral cortical and vertebral strength where PTH had no significant effect. µCT showed only slight changes that were statistically significant for the antibody mainly at cortical sites. The results suggest that a relatively low dose of PTH stimulates metaphyseal repair (screw fixation) specifically, whereas the sclerostin antibody has wide-spread effects, mainly on cortical bone, with less influence on metaphyseal healing. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:471-476, 2014. [ABSTRACT FROM AUTHOR]

ABSTRACT Biological adaptation following placement of a total knee replacements (TKRs) affects peri-implant bone mineral density (BMD) and implant fixation. We quantified the proximal tibial bone strain and implant-bone micro-motion for functioning postmortem retrieved TKRs and assessed the strain/micro-motion relationships with chronological (donor age and time in service) and patient (body weight and BMD) factors. Twenty-two tibial constructs were functionally loaded to one body weight (60% medial/40% lateral), and the bone strains and tray/bone micro-motions were measured using a digital image correlation system. Donors with more time in service had higher bone strains ( p = 0.044), but there was not a significant ( p = 0.333) contribution from donor age. Donors with lower peri-implant BMD ( p = 0.0039) and higher body weight ( p = 0.0286) had higher bone strains. Long term implants (>11 years) had proximal bone strains 900 µϵ that were almost twice as high as short term (<5 years) implants 570 µϵ. Micro-motion was greater for younger donors ( p = 0.0161) and longer time in service ( p = 0.0008). Increased bone strain with long term in vivo service could contribute to loosening of TKRs by failure of the tibial peri-implant bone. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:355-361, 2014. [ABSTRACT FROM AUTHOR]

ABSTRACT The objective of this study was to understand the effect of pre-repair rotator cuff chronicity on post-repair healing outcomes using a chronic and acute multi-tendon rat rotator cuff injury model. Full-thickness dual tendon injuries (supra- and infraspinatus) were created unilaterally in adult male Sprague Dawley rats, and left chronically detached for 8 or 16 weeks. After chronic detachment, tears were repaired and acute dual tendon injuries were created and immediately repaired on contralateral shoulders. Tissue level outcomes for bone, tendon, and muscle were assessed 4 or 8 weeks after repair using histology, microcomputed tomography, biomechanical testing, and biochemical assays. Substantial gap formation was seen in 35% of acute repairs and 44% of chronic repairs. Gap formation negatively correlated with mechanical and structural outcomes for both healing time points regardless of injury duration. Bone and histomorphometry, as well as biomechanics, were similar between acute and chronic injury and repair regardless of chronicity and duration of healing. This study was the first to implement a multi-tendon rotator cuff injury with surgical repair following both chronic and acute injuries. Massive tear in a rodent model resulted in gap formation regardless of injury duration which had detrimental effects on repair outcomes. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:439-447, 2014. [ABSTRACT FROM AUTHOR]

ABSTRACT We compared the mechanical force of tendon-to-bone repair techniques for flexor tendon reconstruction. Thirty-six flexor digitorum profundus (FDP) tendons were divided into three groups based upon the repair technique: (1) suture/button repair using FDP tendon (Pullout button group), (2) suture bony anchor using FDP tendon (Suture anchor group), and (3) suture/button repair using FDP tendon with its bony attachment preserved (Bony attachment group). The repair failure force and stiffness were measured. The mean load to failure and stiffness in the bony attachment group were significantly higher than that in the pullout button and suture anchor groups. No significant difference was found in failure force and stiffness between the pullout button and suture anchor groups. An intrasynovial flexor tendon graft with its bony attachment has significantly improved tensile properties at the distal repair site when compared with a typical tendon-to-bone attachment with a button or suture anchor. The improvement in the tensile properties at the repair site may facilitate postoperative rehabilitation and reduce the risk of graft rupture. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31:1720-1724, 2013 [ABSTRACT FROM AUTHOR]

Local cooling is very common after bone and joint surgery. Therefore the knowledge of bone blood flow during local cooling is of substantial interest. Previous studies revealed that hypothermia leads to vasoconstriction followed by decreased blood flow levels. The aim of this study was to characterize if local cooling is capable of inducing reduced blood flow in bone tissue using a stepwise-reduced temperature protocol in experimental rabbits. To examine bone blood flow we utilized the fluorescent microsphere (FM) method. In New Zealand white rabbits one randomly chosen hind limb was cooled stepwise from 32 to 2°C, whereas the contra lateral hind limb served as control. Injection of microspheres was performed after stabilization of bone and muscle temperature at each temperature level. Bones were removed, dissected and fluorescence intensity was determined to calculate blood flow values. We found that blood flow of all cooled regions decreased relative to the applied external temperature. At maximum cooling blood flow was almost completely disrupted, indicating local cooling as powerful regulatory mechanism for regional bone blood flow (RBBF). Postoperative cooling therefore may lead to strongly decreased bone blood flow values. As a result external cooling has capacity to both diminish bone healing and reduce bleeding complications. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31:1820-1827, 2013 [ABSTRACT FROM AUTHOR]

Although mechanical loading is known to influence trabecular bone adaptation, the role of specific loading parameters requires further investigation. Previous studies demonstrated that the number of loading cycles and loading duration modulate the adaptive response of trabecular bone in a rabbit model of applied loading. In the current study, we investigated the influence of load magnitude on the adaptive response of trabecular bone using the rabbit model. Cyclic compressive loads, producing peak pressures of either 0.5 or 1.0 MPa, were applied daily (5 days/week) at 1 Hz and 50 cycles/day for 4 weeks post-operatively to the trabecular bone on the lateral side of the distal right femur, while the left side served as an nonloaded control. The adaptive response was characterized by microcomputed tomography and histomorphometry. Bone volume fraction, bone mineral content, tissue mineral density, and mineral apposition rate (MAR) increased in loaded limbs compared to the contralateral control limbs. No load magnitude dependent difference was observed, which may reflect the critical role of loading compared to the operated, nonloaded contralateral limb. The increased MAR suggests that loading stimulated new bone formation rather than just maintaining bone volume. The absence of a dose-dependent response of trabecular bone observed in this study suggests that a range of load magnitudes should be examined for biophysical therapies aimed at augmenting current treatments to enhance long-term fixation of orthopedic devices. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 930-934, 2013 [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a leading cause of disability worldwide. We hypothesized that inflammation following isolated intra-articular bone injury can stimulate post-traumatic OA and developed a rabbit model to test that concept. Sixty female New Zealand White Rabbits were used. Twenty-six experimental animals had two holes drilled into their right femoral-notch, 18 rabbits had sham surgery, and 16 were un-operated controls. Rabbits were euthanized in subgroups at 72 h, 3, 6, 9, and 52 weeks. Knees were assessed grossly and tissues collected. Cartilage and synovium were analyzed with histology and qPCR and subgroups compared statistically. All surgical joints showed gross and histological (modified Mankin score) cartilage damage after surgery, with experimentals worsening with time ( p < 0.05). Cartilage qPCR showed fivefold increases in TGFβ ( p < 0.05) expression at 72 h and 3 weeks with sixfold increases in MMP13 ( p < 0.025) expression at 72 h. By 6 weeks, expression of these markers was similar to baseline levels. Synovial membrane thickening with increased cellularity was seen at both 9 and 52 weeks ( p < 0.05). Short-term synovial inflammatory marker (IL-1β, IL-Ra, IL-6, and IL-8) expression was three- to fourfold increase in experimentals at 72 h ( p < 0.01) returning to baseline levels by 3 weeks. Intra-articular bone injury creates early joint inflammation with some chronic synovial changes and progressive cartilage damage consistent with OA in adult rabbits. This model provides an exciting new avenue to potentially explore some relevant inflammatory drivers of OA without major mechanical variables. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 914-920, 2013 [ABSTRACT FROM AUTHOR]

Stem cells, such as mesenchymal stem cells (MSCs), contribute to bone fracture repair if they are delivered to the injury site. However, it is difficult to assess the retention and differentiation of these cells after implantation. Current options for non-invasively tracking the transplanted stem cells are limited. Cell-based therapies using MSCs would benefit greatly through the use of an imaging methodology that allows cells to be tracked in vivo and in a timely fashion. In this study, we implemented an in vivo imaging methodology to specifically track early events such as differentiation of implanted human MSCs (hMSCs). This system uses the collagen type 1 (Col1α1) promoter to drive expression of firefly luciferase (luc) in addition to a constitutively active promoter to drive the expression of green fluorescent protein (GFP). The resulting dual-promoter reporter gene system provides the opportunity for osteogenic differentiation-specific luc expression for in vivo imaging and constitutive expression of GFP for cell sorting. The function of this dual-promoter reporter gene was validated both in vitro and in vivo. In addition, the ability of this dual-promoter reporter system to image an early event of osteogenic differentiation of hMSCs was demonstrated in a murine segmental bone defect model in which reporter-labeled hMSCs were seeded into an alginate hydrogel scaffold and implanted directly into the defect. Bioluminescence imaging (BLI) was performed to visualize the turn-on of Col1α1 upon osteogenic differentiation and followed by X-ray imaging to assess the healing process for correlation with histological analyses. © 2013 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res XX:XXX-XXX, 2013 © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 871-879, 2013 [ABSTRACT FROM AUTHOR]

Persistent pain is an important cause of patient dissatisfaction after unicompartmental knee replacement (UKR) and has been correlated with localized tibial strain. However, the factors that influence these strains are not well understood. To address this issue, we created finite element models to examine the effect on tibial strain of: (1) muscle forces (estimated using instrumented knee data) acting on attachment sites on the proximal tibia, (2) UKR implantation, (3) loading position, and (4) changes in gait pattern. Muscle forces acting on the tibia had no significant influence on strains within the periprosthetic region, but UKR implantation increased strain by 20%. Strain also significantly increased if the region of load application was moved >3 mm medially. The strain within the periprosthetic region was found to be dependent on gait pattern and was influenced by both medial and lateral loads, with the medial load having a greater effect (regression coefficients: medial = 0.74, lateral = 0.30). These findings suggest that tibial strain is increased after UKR and may be a cause of pain. It may be possible to reduce pain through modification of surgical factors or through altered gait patterns. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 821-828, 2013 [ABSTRACT FROM AUTHOR]

Treatment of delayed bone healing and non-unions after fractures, osteotomies or arthrodesis still is a relevant clinical challenge. Artificially applied growth factors can increase bone healing and progressively gain importance in clinical routine. The aim of this study was to determine the effects of rhPDGF-BB, rhVEGF-165, and rhBMP-2 in fibrin matrix on bone healing in a delayed-union rat model. Thirty-seven rats underwent a first operation where a standardized femoral critical size defect was created. A silicone spacer was implanted to impair vascularization within the defect. At 4 weeks the spacer was removed in a second operation and rhPDGF-BB, rhVEGF-165, or rhBMP-2 were applied in a fibrin clot. Animals in a fourth group received a fibrin clot without growth factors. At 8 weeks fibrin bound rhBMP-2 treated animals showed a significantly increased union rate and bone volume within the defect compared to the other groups. Single application of fibrin bound rhPDGF-BB and rhVEGF-165 failed to increase bone healing in our atrophic non-union model. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1563-1569, 2012 [ABSTRACT FROM AUTHOR]

Fracture healing is highly sensitive to mechanical conditions; however, the effects of mechanical loading on large bone defect regeneration have not been evaluated. In this study, we investigated the effects of functional loading on repair of critically sized segmental bone defects. About 6-mm defects were created in rat femora, and each defect received 5 µg recombinant human bone morphogenetic protein-2 (rhBMP-2), delivered in alginate hydrogel. Limbs were stabilized by either stiff fixation plates for the duration of the study or compliant plates that allowed transfer of compressive ambulatory loads beginning at week 4. Healing was assessed by digital radiography, microcomputed tomography, mechanical testing, histology, and finite element modeling. Loading significantly increased regenerate bone volume and average polar moment of inertia. The response to loading was location-dependent with the polar moment of inertia increased at the proximal end of the defect but not the distal end. As a result, torsional stiffness was 58% higher in the compliant plate group, but failure torque was not altered. In single samples assessed for histology from each group, a qualitatively greater amount of cartilage and a lesser degree of remodeling to lamellar bone occurred in the loaded group compared to the stiff plate group. Finally, principal strain histograms, calculated by FE modeling, revealed that the compliant plate samples had adapted to more efficiently distribute loads in the defects. Together, these data demonstrate that functional transfer of axial loads alters BMP-induced large bone defect repair by increasing the amount and distribution of bone formed within the defect. © 2011 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 30:1067-1075, 2012 [ABSTRACT FROM AUTHOR]

Bone exhibits rate-dependent failure behavior, suggesting that viscoelasticity is a factor in the damage and fracture of bone. Microdamage initiates at scales below the macroscopic porosity in bone, and, as such, is affected by the intrinsic viscoelasticity of the bone tissue. The viscoelasticity of the bone tissue can be measured by nanoindentation and recording the creep behavior at constant load. The viscoelastic properties have been used to assess differences in tissue behavior with respect to fracture healing, aging, and mouse strains. In this study, we compared the viscoelastic behavior of human cortical bone between genders by using nanoindentation at a fixed load of 10 mN to measure the creep time constant. Bones from females had a significantly greater time constant, indicating slower creep and relaxation, than bones from males. The creep time constants decreased with increasing tissue modulus. The mineralization, collagen content, and collagen cross-link density, which were bulk measurements, were analyzed to determine if the differences in viscoelastic behavior were explained by compositional differences in the bone. However, none of the parameters differed between genders, nor were they correlated to the viscoelastic time constant. As such, the difference must depend on other matrix proteins that we did not assess or differences in the microstructural organization. This is one of the only intrinsic bone material properties that has been found to differ between males and females, and it may be important for assessing differences in fracture risk, since crack propagation is generally sensitive to viscoelastic properties. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:693-699, 2012 [ABSTRACT FROM AUTHOR]

The potential for peri-implant bone yielding and subsequent loosening of Ilizarov ring-wire external fixation systems was investigated using non-linear finite element (FE) analyses. A strain-based plasticity model was employed to simulate bone yielding. FE models also incorporated contact behavior at the wire-bone interface, orthotropic elasticity, and periosteal-endosteal variation of bone properties. These simulations were used to determine the extent and location of yielding with change in age-related bone structure and properties for the bone-Ilizarov construct at the tibial midshaft. At critical wire-bone interfaces, the predicted volume of yielded bone with four wires (on either side of the fracture) was ∼40% of that with two wires. Old-aged cases showed considerably greater bone yielding at the wire-bone interface than young cases (1.7-2.2 times greater volumes of yielded bone). The volume of yielded bone at all wire-bone interfaces decreased with an increase in wire pre-tension. The absence of continuous through-thickness yielding offers an explanation for the clinical observation that Ilizarov ring-wire fixation can provide stable fracture fixation even in bone with high porosity. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:726-732, 2012 [ABSTRACT FROM AUTHOR]

Appropriate and controlled chondrogenesis and endochondral ossification play fundamental roles in the fracture healing cascade, a regenerative process involved in highly coordinated biological events, including the Wnt/β-catenin signaling pathway. To examine the role and importance of this pathway in chondrocytes, we studied bone repair of closed tibias fractures in Col2a1-ICAT transgenic mice, in which the Wnt/β-catenin signaling pathway is specially inhibited in chondrocytes. Radiological, histological, and histomorphometric analyses at 7, 9, 12, 14, 21, and 28 days after fracture demonstrated the bone repairs were retarded in Col2a1-ICAT transgenic mice, due to reduced and delayed cartilage formation, chondrocyte hypertrophy, and bone generation. In addition, at 5 weeks, Col2a1-ICAT transgenic mice exhibited a weak mechanical tolerance to four-point bending. Furthermore, quantitative-PCR analysis revealed that the expression of genes associated specifically with cartilage extracellular matrix formation (collagen II, collagen X, and mmp13), bone remodeling (alp, collagen I, and osteocalcin), and vascular extravagation (vegf), and transcriptional activators involved in cartilage generation and ossification (sox9 and runx2) was decreased and delayed in the fracture sites of Col2a1-ICAT transgenic mice during healing. Collectively, these results suggest that Wnt/β-catenin signaling is critical for fracture healing, especially with respect to chondrogenesis and endochondral ossification. Thus, our study provides insight into the possible mechanisms of and therapeutic targets for improving normal facture repair and the healing of non-union fractures. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:304-310, 2012 [ABSTRACT FROM AUTHOR]

Low bone quality, such as induced by osteoporosis, is considered a main factor leading to failure of fracture fixations. Peri-implant bone augmentation has been proposed as a means of reducing failure rates in osteoporotic bone by improving implant stability. The beneficial effects of pharmacological augmentation of bone in the immediate vicinity of the implant have been demonstrated. Yet, a quantitative understanding of the role of peri-implant bone in implant stability is lacking. Therefore, the aim of our study was to quantify the effects of bone loss and peri-implant bone augmentation on implant stability using image-based finite element analyses. Using a validated model, we simulated how osteoporotic bone loss would affect implant stability in human humeral heads. We also quantified how augmentation of peri-implant bone can enhance implant stability. Our simulations revealed that a 30% reduction in bone mass led to a 50% decrease in implant stability. We also found that peri-implant bone augmentation increased implant stability and that the efficiency of bone augmentation decreased with increasing peri-implant distance. These findings highlight the strong effect that bone loss has on implant fixation and the potential of peri-implant bone augmentation for improving implant anchorage in low quality bone. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:178-184, 2012 [ABSTRACT FROM AUTHOR]

Individuals with nerofibromatosis Type 1 (NF1) frequently suffer a spectrum of bone pathologies, such as abnormal skeletal development (scoliosis, congenital bowing, and congenital pseudoarthroses, etc), lower bone mineral density with increased fracture risk. These skeletal problems may result, in part, from abnormal osteoclastogenesis. Enhanced RAS/PI3K activity has been reported to contribute to abnormal osteoclastogenesis in Nf1 heterozygous ( Nf1+/−) mice. However, the specific downstream pathways linked to NF1 abnormal osteoclastogenesis have not been defined. Our aim was to determine whether mammalian target of rapamycin (mTOR) was a key effector responsible for abnormal osteoclastogenesis in NF1. Primary osteoclast-like cells (OCLs) were cultured from Nf1 wild-type ( Nf1+/+) and Nf1+/− mice. Compared to Nf1+/+ controls, there were 20% more OCLs induced from Nf1+/− mice. Nf1+/− OCLs were larger and contained more nuclei. Hyperactive mTOR signaling was detected in Nf1+/− OCLs. Inhibition of mTOR signaling by rapamycin in Nf1+/− OCLs abrogated abnormalities in cellular size and number. Moreover, we found that hyperactive mTOR signaling induced abnormal osteoclastogenesis major through hyper-proliferation. Our research suggests that neurofibromin directly regulates osteoclastogenesis through mTOR signaling pathway. Inhibiting mTOR may represent a viable strategy to treat NF1 bone diseases. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:144-152, 2012 [ABSTRACT FROM AUTHOR]

Our objective was to examine the load-bearing capacity of the transverse processes of human cadaveric thoracic vertebrae to vertical loads and axial rotation moments (i.e., moment applied in the transverse plane). A secondary objective was to examine the effect of the attached rib stumps. We wanted to demonstrate that the transverse process is durable enough to support the CAB hook-a complementary hook to the CD system-and can handle the vertical load or axial rotation moment during correction of scoliosis. We used 107 thoracic vertebrae removed from 10 cadavers. They were prepared in vertebral pairs, and were fixed into a material testing apparatus. Superoinferior vertical loads and axial rotation moments were applied to the transverse process using the CAB hooks at a rate of 30 mm/min and 8.5°/s respectively until it fractured. We recorded 142 measurements, 99 were for vertical load and 43 for axial rotation moment. The average ultimate vertical load was 338 (SD = 128) N and the average ultimate axial rotation moment was 14.4 (SD = 4.52) Nm. The ultimate axial rotation moment for specimens with rib stumps attached was significantly greater than for specimens without rib stumps 15.9 (SD = 4.1) Nm versus 12.5 (SD = 4.4) Nm. Our results showed that both the vertical and axial rotation loading capability of the transverse process are large enough to withstand significant correctional forces, without fracture, through the CAB hooks. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1904-1909, 2011 [ABSTRACT FROM AUTHOR]

Alterations in microdamage morphology and accumulation are typically attributed to impaired remodeling, but may also result from changes in microdamage initiation and propagation. Such alterations are relevant for cancellous bone with high metabolic activity and numerous bone quality changes. This study investigates the role of trabecular microarchitecture on morphology and accumulation of microdamage in human cancellous bone. Trabecular bone cores from donors of varying ages and bone volume fraction (BV/TV) were separated into high and low BV/TV groups. Samples were subjected to no load or uniaxial compression to 0.6% (pre-yield) or 1.1% (post-yield) strain. Microdamage was stained with lead uranyl acetate and specimens were imaged via microcomputed tomography to quantify microdamage and determine its morphology in three-dimensions (3D). Donors with high BV/TV had greater post-yield strain and were tougher than low BV/TV donors. High BV/TV bone had less microdamage than low BV/TV bone under post- but not pre-yield loading. Microdamage under both loading conditions showed significant correlations with microarchitecture and BV/TV, but the key predictor was structure model index (SMI). As SMI increased (more trabecular rods), microdamage morphology became crack-like. Thus, low BV/TV and increased SMI have strong influences on microdamage accumulation in bone through altered initiation. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1739-1744, 2011 [ABSTRACT FROM AUTHOR]

Rupture of the insertion of the pectoralis major muscle to the proximal humerus is becoming a common injury. Repair of these ruptures increases patient satisfaction, strength, and cosmesis, and shortens return to competitive sports. Several repair techniques have been described, but recently many surgeons are using suture anchors. The traditional repair technique uses transosseous sutures, but no study has biomechanically compared the strength of these two repair techniques in human cadavers. Twelve fresh-frozen human shoulder specimens were dissected. The pectoralis major tendon insertion was cut from the bone and repaired using one of the two repair techniques: specimens were randomly assigned to transosseous trough with suture tied over bone versus four suture anchors. The fixation constructs were pulled to failure at 4 mm/s on a materials testing system. The mean ultimate failure load of the transosseous repairs was 611 N and the mean ultimate failure load of the suture anchor repair was 620 N. The mean stiffness of the transosseous repair was 32 and 28 N/mm for the suture anchor group. We found no statistically significant difference between these two repair techniques. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1783-1787, 2011 [ABSTRACT FROM AUTHOR]

Fracture healing is a well-organized process between several molecules and mediators. As known from other diseases, genetic polymorphisms may exhibit different expression patterns in these mediators. Concerning fracture healing, this may lead to an extended healing process or non-union. We investigated the incidence of polymorphisms in patients with aseptic non-unions after femoral and tibial shaft fractures as compared to patients with uneventful healing. Exclusion criteria were smoking, diabetes, bilateral fractures, systemic corticoid therapy, and septic non-unions. Analysis of allele frequencies and genotype distribution of various mediators were carried out following PCR. Clinical parameters such as injury severity and in-hospital were analyzed. Fifty patients following non-union (group NU) were enrolled, the control group consisted of 44 patients (group H). A significant association of a PDGF haplotype and non-unions following fracture could be observed. There was a significantly increased in-hospital time and amount of surgical procedures in group NU. Polymorphisms within the PDGF gene seem to be a genetic risk factor for the development of non-unions of the lower extremity following fracture. The early identification of high risk patients could result in an adapted therapeutical strategy and might contribute to a significant decrease of posttraumatic non-unions. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1724-1731, 2011 [ABSTRACT FROM AUTHOR]

The aim of the present study was to investigate the effect of exercise on angiogenesis during bone defect healing in mice. We evaluated angiogenesis during cranial bone defect healing by intravital fluorescence microscopy (IVM) at days 0-21. To characterize the type of bone repair, we performed additional histomorphometric analyses at days 3-15. IVM was conducted in mice, which were housed in cages supplied with running wheels (exercise group; n = 7) and compared to IVM results of mice, which were housed in cages without running wheels (controls; n = 7). In the exercise group, we additionally performed correlation analyses between results of the IVM and the running distance. IVM showed an accelerated decrease of bone defect area in the exercise group compared to the control group. This was associated with a significantly higher blood vessel diameter in animals undergoing exercise at days 9 and 12 and a significant correlation between running distance and blood vessel density at day 9 ( r = 0.74). Histomorphometry showed osseous bridging of the defect at day 9. The newly woven bone was covered by a neo-periosteum containing those blood vessels, which were visible by IVM. We conclude that exercise accelerates bone defect healing and stimulates angiogenesis during bore repair. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29: 1086-1092, 2011 [ABSTRACT FROM AUTHOR]

Even with current treatments of acute joint injuries, more than 40% of people who suffer significant ligament or meniscus tears, or articular surface injuries, will develop osteoarthritis (OA). Correspondingly, 12% or more of all patients with lower extremity OA have a history of joint injury. Recent research suggests that acute joint damage that occurs at the time of an injury initiates a sequence of events that can lead to progressive articular surface damage. New molecular interventions, combined with evolving surgical methods, aim to minimize or prevent progressive tissue damage triggered by joint injury. Seizing the potential for progress in the treatment of joint injuries to forestall OA will depend on advances in (1) quantitative methods of assessing the injury severity, including both structural damage and biologic responses, (2) understanding of the pathogenesis of post-traumatic OA, taking into account potential interactions among the different tissues and the role of post-traumatic incongruity and instability, and (3) application of engineering and molecular research to develop new methods of treating injured joints. This paper highlights recent advances in understanding of the structural damage and the acute biological response following joint injury, and it identifies important directions for future research. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:802-809 [ABSTRACT FROM AUTHOR]

We evaluated the gait characteristics of patients who had short or long-segment fusion after thoracolumbar burst fractures. The study included 12 patients (5 male, 7 female) who had vertebral fusion after traumatic thoracolumbar region (T12-L2) burst fractures. Patients were divided into two groups based on fixation type (short or long segment). Twelve healthy age and sex-matched subjects (seven male, five female) served as controls. Quantitative gait data, including all time-distance (walking velocity, cadence, step time, step length, double support time), kinematic (joint rotation angles of pelvis, hip, knee, and ankle), and kinetic data (moments of hip in sagittal and coronal plane) were collected. Three patients had a T12, six patients an L1, and three patients had an L2 lesion. Patients in the two groups were similar in terms of age, sex, and time since operation. No significant differences were found in the time-distance, kinematic, and kinetic gait characteristics between the two groups. This preliminary study reveals that the gait characteristics of patients with short and long-segment fusion after thoracolumbar burst fractures were similar to those of healthy subjects and did not differ from each other. However, the present results should be interpreted with caution due to the small sample size. Future studies with larger groups are needed to provide additional data to validate these findings. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:915-918 [ABSTRACT FROM AUTHOR]

In poly-traumatic patients a blunt chest trauma is an important trigger of the posttraumatic systemic inflammatory response. There is clinical evidence that fracture healing is delayed in such patients, however, experimental data are lacking. Therefore, we investigated the influence of a thoracic trauma on fracture healing in a rat model. Male Wistar rats received either a blunt chest trauma combined with a femur osteotomy or an isolated osteotomy. A more rigid or a more flexible external fixator was used for fracture stabilization to analyze whether the thoracic trauma influences regular healing and mechanically induced delayed bone healing differently. The blunt chest trauma induced a significant increase of IL-6 serum levels after 6 and 24 h, suggesting the induction of a systemic inflammation, whereas the isolated fracture had no effect. Under a more rigid fixation the thoracic trauma considerably impaired fracture healing after 35 days, reflected by a significantly reduced flexural rigidity (three-point-bending test), as well as a significantly diminished callus volume, moment of inertia, and relative bone surface (µCT analysis). In confirming the clinical evidence, this study reports for the first time that a blunt chest trauma considerably impaired bone healing, possibly via the interaction of the induced systemic inflammation with local inflammatory processes. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:734-739, 2011 [ABSTRACT FROM AUTHOR]

Low-magnitude high-frequency vibration (LMHFV) (35 Hz, 0.3 g) accelerates fracture healing by enhancing callus formation and mineralization for both normal and osteoporotic rats in our previous studies. We hypothesized that LMHFV enhances fracture healing through bone remodeling. Ibandronate was used to suppress LMHFV-stimulated bone remodeling and changes in remodeling were investigated to verify our hypothesis. Closed femoral fractures were created in 80 osteoporotic female Sprague-Dawley rats. The rats were randomly assigned into control (CG), LMHFV (VG) (20 min/day, 5 days/week), ibandronate (BG) (7 µg/kg/week), or LMHFV + ibandronate (VBG) for a treatment duration of 2, 4, 6, or 8 weeks. Blood was taken and the femora were harvested for histological and radiological analyses. VG had the fastest drop in callus area (CA) and width (CW), and bone volume to tissue volume ratio (BV/TV); whereas, a plateaued trend in BG and VBG was observed. The fastest callus reduction, highest mineral apposition rate at week 6, and increased serum concentration of osteocalcin and TRAP5b in VG suggested enhanced remodeling. LMHFV partially reversed the inhibition of bone remodeling by ibandronate suggested LMHFV had an opposite effect on bone remodeling to ibandronate. In conclusion, LMHFV accelerated fracture healing by enhancing bone remodeling and the administration of ibandronate can impair this enhancement. LMHFV has great potential in improving fracture outcome clinically. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:746-752, 2011 [ABSTRACT FROM AUTHOR]

SIRT1 has been implicated as a key factor in aging-related diseases. Nevertheless, the role of SIRT1 in the pathogenesis of osteoarthritis (OA) is still unknown. We examined the expression of SIRT1 in cartilage samples and the effect of SIRT1 inhibition on chondrocyte gene expression changes to elucidate the role of SIRT1 in chondrocytes. SIRT1 expression was examined using cartilage samples from patients undergoing total knee arthroplasty and femoral head replacement by immunohistochemistry. The effect of SIRT1 inhibition by siRNA on chondrocyte gene expression was examined by real-time PCR and Western blotting. SIRT1 expression was barely detectable in the severely degenerated cartilage while SIRT1 was clearly expressed in the less damaged cartilage. The inhibition of SIRT1 by siRNA induced OA-like gene expression changes, namely the significant down-regulation of aggrecan and up-regulation of COL10A1 and ADAMTS-5. Our observations suggest that SIRT1 expression decreases with development of OA and the reduction of SIRT1 in chondrocytes may cause chondrocyte hypertrophy and cartilage matrix loss. SIRT1 might play important roles in the pathogenesis of OA. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:511-515, 2011 [ABSTRACT FROM AUTHOR]

Progression to osteoarthritis (OA) is a frequent sequela of severe articular fracture, particularly when weight-bearing joints are involved. Prevention from post-traumatic OA remains a challenge. Hyaluronan (HA) therapy is reported to represent a safe and effective treatment for patients with OA and rheumatoid arthritis. However, the capacity of HA to prevent the occurrence of osteoarthritic changes in fractured joints has not been demonstrated. The present study was undertaken to examine the effects of HA on expression of six OA-related proteins in fibroblast-like synoviocytes (FLS) from 10 patients with tibia plateau fracture. OA-related factors were quantified using a sandwich enzyme-linked immunosorbent assay. Regardless of induction of the FLS with interleukin (IL)-1β, HA was found to down-regulate expression of catabolic factors (IL-1β, matrix metalloproteinase-3, and tumor necrosis factor-α) and to up-regulate production of anti-catabolic factors (tissue inhibitors of metalloproteinase-1 and metalloproteinase-2). HA also enhanced expression of IL-10, an anti-inflammatory cytokine, in FLS. Our results indicated that HA can promote the expression of both antiinflammatory and structure-protective factors in FLS of patients with tibia plateau fracture. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:495-500, 2011 [ABSTRACT FROM AUTHOR]

The primary function of the disc is mechanical; therefore, degenerative changes in disc mechanics and the interactions between the annulus fibrosus (AF) and nucleus pulposus (NP) in nondegenerate and degenerate discs are important to functional evaluation. The disc experiences complex loading conditions, including mechanical interactions between the pressurized NP and the surrounding fiber-reinforced AF. Our objective was to noninvasively evaluate the internal deformations of nondegenerate and degenerate human discs under axial compression with flexion, neutral, and extension positions using magnetic resonance imaging and image correlation. The side of applied bending (e.g., anterior AF in flexion) had higher tensile radial and compressive axial strains, and the opposite side of bending exhibited tensile axial strains even though the disc was loaded under axial compression. Degenerated discs exhibited higher compressive axial and tensile radial strains, which suggest that load distribution through the disc subcomponents are altered with degeneration, likely due to the depressurized NP placing more of the applied load directly on the AF. The posterior AF exhibited higher compressive axial and higher tensile radial strains than the other AF regions, and the strains were not correlated with degeneration, suggesting this region undergoes high strains throughout life, which may predispose it to failure and tears. In addition to understanding internal disc mechanics, this study provides important new data into the changes in internal strain with degeneration, data for validation of finite element models, and provides a technique and baseline data for evaluating surgical treatments. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29: 547-555, 2011 [ABSTRACT FROM AUTHOR]

Beyond its classical role in regulation of erythropoiesis, erythropoietin (EPO) has been shown to exert protective and regenerative actions in a variety of non-hematopoietic tissues. However, little is known about potential actions in bone regeneration. To analyze fracture healing in mice, a femoral 0.25 mm osteotomy gap was stabilized with a pin-clip technique. Animals were treated with 500 U EPO/kg bw per day or with vehicle only. After 2 and 5 weeks, fracture healing was analyzed biomechanically, radiologically and histologically. Expression of PCNA and NFκB was examined by Western blot analysis. Vascularization was analyzed by immunohistochemical staining of PECAM-1. Circulating endothelial progenitor cells were measured by flow-cytometry. Herein, we demonstrate that EPO-treatment significantly accelerates bone healing in mice. This is indicated by a significantly greater biomechanical stiffness and a higher radiological density of the periosteal callus at 2 and 5 weeks after fracture and stabilization. Histological analysis demonstrated significantly more bone and less cartilage and fibrous tissue in the periosteal callus. Endosteal vascularization was significantly increased in EPO-treated animals when compared to controls. The number of circulating endothelial progenitor cells was significantly greater in EPO-treated animals. The herein shown acceleration of healing by EPO may represent a promising novel treatment strategy for fractures with delayed healing and non-union formation. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:165-172, 2011 [ABSTRACT FROM AUTHOR]

Fracture healing is a complex process, which is further complicated if the bone is osteoporotic. Calcium is one of the important minerals in bone and has been found to prevent osteoporosis but its role in fracture healing of osteoporotic bone is still unclear. We carried out a study on the effects of calcium supplementation on the late phase healing of fractured osteoporotic bone using an ovariectomized rat model. Twenty-four female Sprague-Dawley rats were divided into three groups: sham-operated (SO), ovariectomized-control (OVXC), and ovariectomized + calcium supplements (Ca). The right femurs of all the rats were fractured at mid-epiphysis and a K-wire was inserted for internal fixation. After 2 months of treatment, the rats were sacrificed and the femora were dissected out for radiological and biomechanical assessment. As expected, osteoporosis resulted in impaired healing as shown by the poor radiological and biomechanical properties of the OVXC group. CT scans showed significantly lower callus volumes in the SO and Ca groups compared to the OVXC group. Radiological scoring of fracture healing and callus staging of the SO and Ca groups were better than the OVXC group. However, the biomechanical parameters of the Ca group were significantly lower than the SO group and similar to the OVXC group. Therefore, calcium supplements may appear to improve fracture healing of osteoporotic bone but failed to improve strength. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1651-1656, 2010 [ABSTRACT FROM AUTHOR]

Substance P (SP) has been shown in vitro to stimulate both formation and resorption of bone. This seemingly contradictory observation could be explained by in vivo variations in skeletal loading and rate of bone turnover, features which may be explored during different phases of fracture healing. In 50 SD rats, the right tibia was fractured and fixed with an intramedullary pin in straight alignment and in anterior angulation resulting in a convex and concave side under different load. Fracture repair was assessed by radiography, histology, and semi-quantitative immunohistochemistry of SP nerve fiber occurrence at days 7, 21, 35, 56, and 84 post-fracture. During regeneration, days 7-35, abundant SP-nerve ingrowth was observed in the fracture callus reaching a side-symmetrical peak at day 21 in straight fractures. In angulated fractures, the SP peak was also observed at day 21 on the concave loaded side, but not until day 35 on the convex unloaded side. Each SP-peak coincided with cortical bridging. During remodeling, days 35-84, a side-symmetrical disappearance of SP-positive fibers was seen in straight fractures. The same pattern was seen on the concave loaded side of angulated fractures. However, on the convex unloaded side, where resorption now took place, SP-fibers remained until the end of the experiment. Our study suggests that neuronal SP during bone regeneration has a stimulatory role on bone formation, while during remodeling increased SP fiber density in unloaded areas may be related to bone resorption. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1643-1650, 2010 [ABSTRACT FROM AUTHOR]

Secondary fracture healing in long bones leads to the successive formation of intricate patterns of tissues in the newly formed callus. The main aim of this work was to quantitatively describe the topology of these tissue patterns at different stages of the healing process and to generate averaged images of tissue distribution. This averaging procedure was based on stained histological sections (2, 3, 6, and 9 weeks post-operatively) of 64 sheep with a 3 mm tibial mid-shaft osteotomy, stabilized either with a rigid or a semi-rigid external fixator. Before averaging, histological images were sorted for topology according to six identified tissue patterns. The averaged images were obtained for both fixation types and the lateral and medial side separately. For each case, the result of the averaging procedure was a collection of six images characterizing quantitatively the progression of the healing process. In addition, quantified descriptions of the newly formed cartilage and the bone area fractions (BA/TA) of the bony callus are presented. For all cases, a linear increase in the BA/TA of the bony callus was observed. The slope was greatest in the case of the most rigid stabilization and lowest in the case of the least stiff. This topological description of the progression of bone healing will allow quantitative validation (or falsification) of current mechano-biological theories. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1440-1447, 2010 [ABSTRACT FROM AUTHOR]

The role of platelet-rich plasma (PRP) as a promoter of bone healing remains controversial. The aim of this study was to investigate the effect of PRP in combination with calcium phosphate granules (CPG) on bone defect healing in a metaphyseal long bone defect. A metaphyseal bone defect at the proximal tibia of 16 mini-pigs was filled with CPG combined with autologous PRP or CPG solely (control group). The PRP showed 4.4-fold more platelets compared to peripheral blood. Six weeks after surgery the radiological and histomorphometrical evaluations showed significantly more bone formation in the PRP group in the central area of the defect zone ( p < 0.01) as well as the cortical defect zone ( p < 0.04). Furthermore, the resorption rate of CPG was increased in animals who received PRP. Nevertheless there were only isolated instances of complete osseous bridging of the bone defects even in the PRP group. This study demonstrates that a PRP-CPG composit promotes bone regeneration but does not lead to a solid fusion of a tibial defect in mini-pigs. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1448-1455, 2010 [ABSTRACT FROM AUTHOR]

GFP reporter mice previously developed to assess levels of osteoblast differentiation were employed in a tibial long bone fracture model using a histological method that preserves fluorescent signals in non-decalcified sections of bone. Two reporters, based on Col1A1 (Col3.6GFPcyan) and osteocalcin (OcGFPtpz) promoter fragments, were bred into the same mice to reflect an early and late stage of osteoblast differentiation. Three observations were apparent from this examination. First, the osteoprogenitor cells that arise from the flanking periosteum proliferate and progress to fill the fracture zone. These cells differentiate to osteoblasts, chondrocytes, to from the outer cortical shell. Second, the hypertrophic chondrocytes are dispersed and the cartilage matrix mineralized by the advancing Col3.6+ osteoblasts. The endochondral matrix is removed by the following osteoclasts. Third, a new cortical shell develops over the cartilage core and undergoes a remodeling process of bone formation on the inner surface and resorption on the outer surface. The original fractured cortex undergoes resorption as the outer cortical shell remodels inward to become the new diaphyseal bone. The fluorescent microscopy and GFP reporter mice used in this study provide a powerful tool for appreciating the molecular and cellular processes that control these fundamental steps in fracture repair, and may provide a basis for understanding fracture nonunion. Published by Wiley Periodicals, Inc. J Orthop Res 28:1338-1347, 2010 [ABSTRACT FROM AUTHOR]

Age significantly reduces the regenerative capacity of the skeleton, but the underlying causes are unknown. Here, we tested whether the functional status of inflammatory cells contributes to delayed healing in aged animals. We created chimeric mice by bone marrow transplantation after lethal irradiation. In this model, chondrocytes and osteoblasts in the regenerate are derived exclusively from host cells while inflammatory cells are derived from the donor. Using this model, the inflammatory system of middle-aged mice (12 month old) was replaced by transplanted bone marrow from juvenile mice (4 weeks old), or age-matched controls. We found that the middle-aged mice receiving juvenile bone marrow had larger calluses and more bone formation during early stages and faster callus remodeling at late stages of fracture healing, indicating that inflammatory cells derived from the juvenile bone marrow accelerated bone repair in the middle-aged animals. In contrast, transplanting bone marrow from middle-aged mice to juvenile mice did not alter the process of fracture healing in juvenile mice. Thus, the roles of inflammatory cells in fracture healing may be age-related, suggesting the possibility of enhancing fracture healing in aged animals by manipulating the inflammatory system. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1000-1006, 2010 [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the effects of local endothelial progenitor cell (EPC) therapy on bone regeneration in a rat model. A segmental bone defect (5 mm) was created in the femur and fixed with a mini-plate. There were two groups: EPC-treated ( N = 28) and control ( N = 28). Seven animals were sacrificed from each group at 1, 2, 3, and 10 weeks postoperatively. Healing of the defect was evaluated with radiographic, histological, and quantitative micro-computed tomography (micro-CT) scans. Radiographically, mean scores of the EPC and control groups were, respectively, 1.16-0.61 ( p < 0.05) at 1 week, 2.53-1.54 ( p < 0.05) at 2 weeks, and 4.58-2.35 at 3 weeks ( p < 0.05). At 10 weeks, all the animals in the EPC-treated group had complete union (7/7), but in the control group none achieved union (0/7). Histological evaluation revealed that specimens from EPC-treated animals had abundant new bone and vessel formation compared to that in controls. Micro-CT assessment of the samples from the animals sacrificed at 10 weeks ( N = 14) showed significantly improved parameters of bone volume (36.58-10.57, p = 0.000), bone volume density (0.26-0.17, p = 0.000), model index −2.22-2.79, p = 0.000), trabecular number (1.28-0.91, p = 0.063), trabecular thickness (0.21-0.15, p = 0.001), trabecular spacing (0.63-1.07, p = 0.022), bone surface (353.75-152.08, p = 0.000), and bone surface to bone volume ratio (9.54-14.24, p = 0.004) for the EPC group compared to control, respectively. In conclusion, local EPC therapy significantly enhanced bone regeneration in a segmental defect model in rat femur diaphysis. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1007-1014, 2010 [ABSTRACT FROM AUTHOR]

While the importance of Wnt signaling in skeletal development and homeostasis is well documented, little is known regarding its function in fracture repair. We hypothesized that activation and inactivation of Wnt signaling would enhance and impair fracture repair, respectively. Femoral fractures were generated in Lrp5 knockout mice (Lrp5−/−) and wild-type littermates (Lrp5+/+), as well as C57BL/6 mice. Lrp5−/− and Lrp5+/+ mice were untreated, while C57BL/6 mice were treated 2×/week with vehicle or anti-Dkk1 antibodies (Dkk1 Ab) initiated immediately postoperatively (Day 0) or 4 days postoperatively (Day 4). Fractures were radiographed weekly until sacrifice at day 28, followed by DXA, pQCT, and biomechanical analyses. Lrp5−/− mice showed impaired repair compared to Lrp5+/+ mice, as evidenced by reduced callus area, BMC, BMD, and biomechanical properties. The effects of Dkk1 Ab treatment depended on the timing of initiation. Day 0 initiation enhanced repair, with significant gains seen for callus area, BMC, BMD, and biomechanical properties, whereas Day 4 initiation had no effect. These results validated our hypothesis that Wnt signaling influences fracture repair, with prompt activation enhancing repair and inactivation impairing it. Furthermore, these data suggest that activation of Wnt signaling during fracture repair may have clinical utility in facilitating fracture repair. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:928-936, 2010 [ABSTRACT FROM AUTHOR]