Early identification of patients at risk for impaired tendon healing and corresponding novel therapeutic approaches are urgent medical needs. This study aimed to clarify the role of CD3+ T-cells during acute Achilles tendon (AT) healing. Blood and hematoma aspirate were taken from 26 patients during AT reconstruction, and additional blood samples were obtained during clinical follow-up at 6, 26 and 52 weeks after surgery. T-cell subsets were analyzed by flow cytometry using CD3, CD4, CD8, CD11a, CD57 and CD28 antibodies. Clinical follow-up included functional tests, MRI assessments, and subjective questionnaires. In vitro, the functional behavior of patient-derived tenocytes was investigated in co-cultures with autologous unpolarized CD4+ or CD8+ T-cells, or IFNy-polarized CD8+ or IL17-polarized CD4+ Tcells (n=5-6). This included alterations in gene expression (qPCR), MMP secretion (ELISA), migration rate (scratch wound healing assay) or contractility (collagen gels). Analysis revealed that elevated CD4+ T-cell levels and reduced CD8+ T-cell levels (increased CD4/CD8 ratio) in hematoma aspirate and pre-operative blood were associated with inferior clinical outcomes regarding pain and function at 26 and 52 weeks. Increased levels of CD8+ -memory T-cell subpopulations in blood 6 weeks after surgery were associated with less tendon elongation. In vitro, tenocytes showed increased MMP1/2/3 levels and collagen III/I ratio in co-culture with unpolarized and/or IL17-polarized CD4+ T-cells compared to unpolarized CD8+ T-cells. This coincided with increased IL17 receptor expression in tenocytes co-cultured with CD4+ T-cells. Exposure of tenocytes to IL17-polarized CD4+ T-cells decreased their migration rate and increased their matrix contractility, especially compared to IFNy-polarized CD8+ T-cells. The CD4+ /CD8+ T-cell ratio could serve as prognostic marker for early identification of patients with impaired AT healing potential. Local reduction of CD4+ T-cell levels or their IL17 secretion represent a potential therapeutic approach to improve AT healing and to prevent weakening of the tendon ECM.

The ability of the body to constantly maintain metabolism homeostasis while fulling the heightened energy and macromolecule demand is crucial to ensure successful tissue healing outcomes. Studies investigating the local metabolic environment during healing are scarce to date. Here, using Type 2 Diabetes (T2D) as a study model, we investigate the impact of metabolism dysregulation on scaffold-guided large-volume bone regeneration. Our study treated wild-type or T2D rats with 5 mm critical-sized femoral defects with 3D-printed polycaprolactone (PCL) scaffolds with 70% porosity. Metabolomics was leveraged for a holistic view of metabolism alteration as healing progress and correlated to regenerated bone tissue volume and quality assessed using micro-computed tomography (µ-CT), histology, and immunohistology. Semi-targeted metabolomics analysis indicated dysregulation in the glycolysis and TCA cycle – the main energy production pathways, in T2D compared to healthy animals. The abundance of metabolites substrates, i.e., amino acids – for protein/ extracellular matrix synthesis was also affected in T2D. Tissue-level metabolites observations aligned with morphological observation with less newly formed bone observed in T2D than wild-type rats. This study enlightens the metabolism landscape during scaffold-guided large-volume bone regeneration in wild-type vs. T2D to further guide the personalization of the scaffold to drive successful regeneration.

Introduction

Friction between head and cup is a primary factor for survival of total hip joint replacement (THR) and its gliding surfaces. In up to 40% of all revisions, the cup or inlay must be replaced as result of friction-induced wear [1]. Aim of the study was to measure the friction-induced temperature increase in vivo in THR and to identify possible individual parameters of influence.

Industrialized countries experience a population aging. Elderly patients, due to the experienced immunity, have a constant pro-inflammatory milieu. Little is known on how adaptive immunity impacts the tissue homeostasis and regeneration. The standardized housing of lab animals is specific pathogen free (SPF). However, this housing condition hinders antigen exposure and thus an aging of the adaptive immune system. We hypothesized that exposure to antigens and a developing adaptive immunity will impact tissue homeostasis and regeneration in mice. Mice kept under SPF housing or non-SPF were examined towards their immune status via flow cytometry, bone structure via microCT and bone competence via biomechanical torsional testing. MSCs from these mice were analyzed regarding their differentiation potential and ECM production under various immune cell signaling. Bone regeneration was analyzed in vivo in a mouse osteotomy model. The memory and effector compartment of the adaptive immunity was significantly increased in mice under non-SPF housing. This housing led to an increased femoral cortical thickness and torsional stiffness (p<0,05), whereas the tissue mineral density was not affected. The differentiation potential of stem cells under the influence of an aged immune milieu was significantly reduced. Bone formation was highly affected by the immune status and availed of a naïve immune cell milieu. Adaptive immunity directly impacts bone tissue formation, by exhibiting a constant stress, leading to structural differences in bone tissue organization as well as mechanical competence. For experimental settings, it appears highly relevant if mouse models have had the chance to develop an experienced immune system.

Recently, we could illustrate how tightly the bone and the immune system are interconnected during normal homeostasis but even stronger during bone regeneration. Specifically, the patient´s individual ratio of CD8+ effector T cells (TEFF, already identified as potential unfavorable cells for successful healing) to CD4+ regulatory T cells (TREG, one counterpart to CD8+ TEFF in controlling intratissue inflammation) prior to injury/ surgery appears to determine the healing outcome after fracture. We hypothesized that concentrating CD4+ TREG could serve as innovative therapeutic strategy to improve bone healing. We used an adoptive CD4+ TREG transfer in our well-established mouse osteotomy model. Before treatment, we identified the pre-surgery ratio of CD8+ TEFF/ CD4+ TREG by flow cytometry to characterize the healing potential of individual animals. Thereafter, we performed an adoptive CD4+ TREG transfer to reshape inflammation for supporting osteotomy healing. Across all groups, healing outcome was analyzed after 21 days post-surgery by µCT. Whereas TREG were highly supportive in SPF mice, we observed a heterogeneous clustered healing outcome in the non-SPF mice: TREG responder (improved healing outcome; p = 0.038) and TREG non-responder (impaired healing outcome; p = 0.024). Interestingly, the pre-/peri-surgery ratio of CD8+ TEFF/ CD4+ TREG was higher in the TREG non-responder (p=0.057). Thus, the amount of adoptively transferred CD4+ TREG was not sufficient to improve the healing outcome due to initial unfavorable high CD8+ TEFF/CD4+ TREG ratio. These results clearly show the importance of determining the individual immune status of each patient in the clinic before applying an immunotherapeutic approach.

Objectives

The objective of this study was to develop a test for the rapid (within 25 minutes) intraoperative detection of bacteria from synovial fluid to diagnose periprosthetic joint infection (PJI).

Aims

Tibiofemoral alignment is important to determine the rate of progression of osteoarthritis and implant survival after total knee arthroplasty (TKA). Normally, surgeons aim for neutral tibiofemoral alignment following TKA, but this has been questioned in recent years. The aim of this study was to evaluate whether varus or valgus alignment indeed leads to increased medial or lateral tibiofemoral forces during static and dynamic weight-bearing activities.

INTRODUCTION

The uncertainty of the biological effects of wear and corrosion from Metal-on-metal (MoM) implants has initiated a debate on their safety and use. Generally, the release of wear particles from MoM hip implants can clinically manifest in aseptic osteolysis. In our study, the effect of MoM-wear particles and particle originated Co and Cr ions on mesenchymal stromal cells (MSCs) was investigated [1]. The lead hypotheses were that (1) dissociated Co and Cr, originated from MoM-wear particles, accumulate in the bone marrow and (2) apparently impair the osteogenic function of local MSCs. This impairment could be one element contributing to the manifestation of periprosthetic osteolyses.

The accurate reconstruction of hip anatomy and biomechanics is thought to be important in achieveing good clinical outcomes following total hip arthroplasty (THA). To this end some newer hip designs have introduced further modularity into the design of the femoral component such that neck­shaft angle and anteversion, which can be adjusted intra-operatively. The clinical effect of this increased modularity is unknown. We have investigated the changes in these anatomical parameters following conventional THA with a prosthesis of predetermined neck–shaft angle and assessed the effect of changes in the hip anatomy on clinical outcomes.

In total, 44 patients (mean age 65.3 years (standard deviation (sd) 7); 17 male/27 female; mean body mass index 26.9 (kg/m²) (sd 3.1)) underwent a pre- and post-operative three-dimensional CT scanning of the hip. The pre- and post-operative neck–shaft angle, offset, hip centre of rotation, femoral anteversion, and stem alignment were measured. Additionally, a functional assessment and pain score were evaluated before surgery and at one year post-operatively and related to the post-operative anatomical changes.

The mean pre-operative neck–shaft angle was significantly increased by 2.8° from 128° (sd 6.2; 119° to 147°) to 131° (sd 2.1; 127° to 136°) (p = 0.009). The mean pre-operative anteversion was 24.9° (sd 8; 7.9 to 39.1) and reduced to 7.4° (sd 7.3; -11.6° to 25.9°) post-operatively (p < 0.001). The post-operative changes had no influence on function and pain. Using a standard uncemented femoral component, high pre- and post-operative variability of femoral anteversion and neck–shaft angles was found with a significant decrease of the post-operative anteversion and slight increase of the neck–shaft angles, but without any impact on clinical outcome.

Cite this article: Bone Joint J 2015;97-B:1615–22.

Objectives: Skeletal muscle trauma leads to severe functional deficits. Present therapeutic treatments are unsatisfying and insufficient posttraumatic regeneration is a problem in trauma and orthopaedic surgery. Mesenchymal stem cell (MSC) therapy is a promising tool in the regeneration of muscle function after severe trauma. Our group showed increased contraction forces compared to a non-treated control group 3 weeks after MSC transplantation (TX) into a skeletal muscle trauma. In addition we demonstrated a dose-response relationship of the amount of MSC and force enhancement. We furthermore investigated the fate of the transplanted MSC labelled with very small iron oxide particles using 7 Tesla-MRI. Histological analysis revealed fusion events between existing myofibers but only to a low amount. The increase of muscle force can not be explained by these events only. Before further steps are taken the impact of paracrine effects and the homing to the site of trauma of the MSC has to be evaluated. Experimental studies about the functional regeneration of traumatized skeletal muscule after systemic MSC-TX do not exist.

Methods: 36 female SD-rats received open crush trauma of the left soleus muscle. One week after trauma 2.5 x 106 autologous MSC, harvested from tibial biopsies, were transplanted intraarterially (i.a., femoral arte-ria, group 1) or intravenously (i.v., tail vein, group 2) (n=18). Control animals received saline (i.a.: group 3; i.v.: group 4) (n=18). Histological analysis and biomechanical evaluation by in vivo muscle force measurement was performed 3 weeks after TX.

Results: Twitch stimulation of the healthy right soleus muscles resulted in a contraction force of 0.52±0.14 N. Forces of tetanic contraction in the uninjured muscles reached 0.98±0.27 N. The i.a. MSC-TX improved the muscle force of the injured soleus significantly compared to control (twitch: 82,4%, p=0.02, tetany: 61.6%, p=0.02). Contraction forces of muscles treated i.v. (MSC vs. saline) showed no significant difference. The histological analysis showed no differences in the amount of fibrotic tissue.

Conclusions: The presented study demonstrates the effect of systemic MSC-TX in the treatment of severe skeletal muscle injuries. Interestingly, the functional regeneration could only be increased by i.a. application. The entrapment of MSC in the lungs and the dilution effect in the circulation, when injecting the MSC i.v. could be the reason. For possible future therapeutic approaches a systemic application is considered to be favourable compared to local injections due to the better distribution of the cells in the target muscle.

Skeletal muscle injuries often lead to severe functional deficits. Mesenchymal stem cell (MSC) therapy is a promising but still experimental tool in the regeneration of muscle function after severe trauma. One of the most important questions, which has to be answered prior to a possible future clinical application is the ideal time of transplantation. Due to the initial inflammatory environment we hypothesized that a local injection of the cells immediately after injury would result in an inferior functional outcome compared to a delayed transplantation.

Twenty-seven female Sprague Dawley rats were used for this study. Bone marrow was aspirated from both tibiae of each animal and autologous MSC cultures obtained from the material. The animals were separated into three groups (each n=9) and the left soleus muscles were bluntly crushed in a standardized manner. In group 1 2×106 MSCs were transplanted into the injured muscle immediately after trauma, whereas group 2 and 3 received an injection of saline. Another week later the left soleus muscles of the animals of group 2 were transplanted with the same number of MSCs. Group 1 and 3 received a sham treatment with the application of saline solution in an identical manner. In vivo functional muscle testing was performed four weeks after trauma to quantify muscle regeneration.

Maximum contraction forces after twitch stimulation decreased to 39 ± 18 % of the non injured right control side after crush trauma of the soleus muscles as measured in group 3. Tetanic stimulation showed a reduction of the maximum contraction capacity of 72 ± 12 % of the value obtained from intact internal control muscles. The transplantation of 2 x 106 MSCs one week after trauma improved the functional regeneration of the injured muscles as displayed by significantly higher contraction forces in group 2 (twitch: p = 0.014, tetany: p = 0.018). Local transplantation of the same number of MSCs immediately after crush injury was able to enhance the regeneration process to a similar extent with an increase of maximum twitch contraction forces by 73.3 % (p = 0.006) and of maximum tetanic contraction forces by 49.6 % (p = 0.037) compared to the control group.

The presented results underline the effectivity of MSC transplantation in the treatment of severe skeletal muscle injuries. The most surprising finding was that despite of the fundamental differences of the local environment into which MSCs had been transplanted, similar results could be obtained in respect to functional skeletal muscle regeneration. We assume that the effect of the MSC after immediate injection can partly be explained by their known immunomodulatory competences. The data of our study provide evidence for a large time window of MSC transplantation after muscle trauma.

The aim of this study was to examine the therapeutic potential of locally transplanted MSCs or osteoprogenitor cells (OPCs) in delayed unions. Autologous MSCs were cultured in DMEM or osteogenic medium. A femoral osteotomy was created in rats and stabilized with an external fixator. Except for the Control-group (C-group), a delayed union was induced by cauterization of the periosteum and bone marrow removal. After 2 days, these animals received an injection of DMEM in the gap containing MSCs (MSC-group), OPCs (OPC-group) or no cells (Sham-group). Histomorphometrical analysis showed significant differences in the fraction of mineralized bone, cartilage and connective tissue between the C- and the Sham-group after 2 (p=0.001) and 8 weeks (p≤0.009). After 2 weeks, the MSC- and OPC-groups developed a larger cartilage fraction (each p=0.019) compared to the Sham-group. Biomechanical testing after 8 weeks demonstrated a significantly lower torsional stiffness (p=0.001) in the Sham-group compared to the C-group. Both the MSC and OPC groups showed a higher torsional stiffness than the Sham-group with statistically significant differences (p< 0.002) in the OPC-group. Locally applied MSCs and OPCs slightly improved the healing in this model. The MSCs were less effective compared to the OPCs. The less than expected healing improvement of both cell treatments may be related to an unfavourable microenvironment at the application time. An explanation for the superior outcome of the OPCs might be that the OPCs may be protected by macroscopically visible matrix at the transplantation time point.

*Winner of ISFR Young Investigator Award

Purpose: The aim of this study was to compare the temporal expression pattern of factors related to cartilage and bone formation and endochondral ossification during standard and delayed bone healing for a more in-depth understanding of the molecular basis of disturbed bone healing and to elucidate suitable timing for substitution of factors to stimulate the healing process.

Methods: A tibial osteotomy was performed in two groups of sheep (n=30 each) and stabilized with either a rigid external fixator leading to standard healing or with a mechanically critical one leading to delayed healing. Hematoma/callus tissue was harvested 4, 7, 14, 21 and 42 days postop. qPCR was employed to determine the expression patterns of BMPs and other molecules.

Results: Gene expressions of BMP2, BMP4, BMP7, Noggin, MMP9 and MMP13 were distinctly lower in the delayed compared to the standard healing group at several time points from day 14, whilst no differential gene expression of Coll II and Coll X was found between both groups. Among the BMPs, BMP7 showed the most markedly differential expression. The first evident difference in BMP7 expression between both groups was found at day 14 suggesting that exogen substitution in the context of a therapeutic approach should be postponed. The differential expression pattern of both MMP9 and MMP13 suggests that there might be a failure or delay in endochondral ossification in delayed bone healing.

Conclusion: Downregulation in gene expression of osteogenic BMPs and cartilage matrix degrading MMPs may account for a considerable delay of bone healing.

Introduction: Currently used small animal models of a critical size defect do not sufficiently simulate the biologically unreactive situation in an atrophic non-union. Furthermore, models using intramedullary nails are of little, and poorly standardised, biomechanical stability. This is a characteristic known to promote callus formation though, rather leading to a hypertrophic non-union.

The aim of this study was to establish an atrophic non-union model in the rat femur under well defined biomechanical conditions and with minimised interactions between the processes in the healing zone and the implant by using external fixation.

MATERIALS AND METHODS: 80 male Sprague Dawley rats were randomly divided into two groups (non-union vs. control). All animals received an osteotomy (app. 0.5 mm gap) of the left femur, stabilised with a custom made external fixator. In the non-union group the periosteum was cauterised 2mm distal and proximal of the osteotomy, and the bone marrow was removed. X-rays were performed once weekly. Animals were sacrificed at 14 or 56 days post-operation. At both time points the femurs of 16 animals of each group underwent histological/histomorphometrical and immunhis-tochemical analyses (PMMA or paraffin embedding). Additionally at 56 days 8 animals of each group were tested biomechanically. The maximum torsional failure moment and the torsional stiffness were determined in relation to the intact femur. Post-mortem x-rays were evaluated in a descriptive manner.

RESULTS: At 14 days the histology and radiology showed considerable mineralised periosteal callus in the control group, while the non-union group only showed very little periosteal callus, distant to the osteotomy. At 56 days the control group was completely, or at least partially, bridged by mineralised callus. The non-union group did not show a bridging of the osteotomy gap in any of the animals, moreover the bone ends were resorbed and the gap widened. The relative mean torsional stiffness was significantly larger (p< 0.001) in the control group compared to the non-union group (136.2±34.5% vs. 2.3±1.2%). In the non-union group no maximal torsional failure moment could be detected for the osteotomised femurs. In the control group it was 134.2±79.1%, relative to the intact femur.

DISCUSSION: The cauterisation of the periosteum and the removal of the bone marrow, in combination with a high stiffness of the external fixator may create an atrophic non-union under well defined biomechanical conditions and with minimised interactions between the healing zone and the implant. This model will allow better standardised investigations on the subject of atrophic non-unions.

Introduction: Selective COX-2 (Cyclooxygenase-2) inhibitors have been found to impede fracture healing. The effect of selective COX-2 inhibitors on tendon healing in a bone tunnel, however, is unknown.

Methods: The authors performed bilateral anterior cruciate ligament reconstructions in 32 rabbits and used peripheral quantitative computed tomography (pQCT) to compare tendon-to-bone healing between tunnel aperture and midtunnel regarding bone mineral density (BMD) and ingrowth of new bone. Each animal was assigned to one of four groups. Two groups received selective COX-2 inhibitors orally for 3 weeks (Cele-coxib; 10 mg/kg/d), the two other groups received no COX-2 inhibitors (controls). The animals were sacrificed 3 and 6 weeks after surgery. In biomechanical testing maximum load to failure and stiffness of the tendon grafts were calculated from the load displacement curve and failure modes were recorded. To assess indirectly the effect on local COX-2 activity the synovial content of Prostaglandin E2 (PGE2), the major metabolite of arachnidonic acid metabolism and catalyzed by COX-2, was measured by Enzyme-linked Immunosorbent Assay (ELISA).

Results: Animals treated with selective COX-2 inhibitors had significantly lower BMD at the tunnel aperture (P=.02). In all groups the BMD at the tunnel aperture was significantly higher in comparison with the midtunnel (P< .05). In the controls ingrowth of new bone was greater at the tunnel aperture at 3 weeks (P=.028). After 3 weeks of COX-2 inhibitor administration synovial fluid concentrations of PGE2 were significantly lowered (P=.018) and increased more than threefold by 6 weeks after surgery and 3 weeks after last drug administration (P=.022), while in the controls there was a decrease in PGE2 between week 3 and 6. At 6 weeks the controls exhibited a twofold increase in maximum load to failure (3 weeeks: 28.2±20.9 N; 6 weeks: 59.6±53.6 N; P=.394), whereas the COX-2 inhibitor treated specimens decreased 1.9fold (3 weeks: 69.3±50.5 N; 6 weeks: 37.4±16.8 N; P=.24). Maximum load to failure values correlated with PGE2 changes, but not statistically significant (r²= −0,502; p=0,056). Failure modes at 3 and 6 weeks were rupture and degloving, respectively, of the tendon graft.

Discussion: This study revealed decreased bone mineral density at the tunnel aperture at 3 weeks, an increase of the inflammatory mediator PGE2 and decreased graft stability with time after treatment with selective COX-2 inhibitors. Untreated controls appeared to have a more physiological healing course with a continuous decrease in PGE2 and an increase in graft stability. Our results suggest, that selective COX-2 inhibitors may delay tendon healing in a bone tunnel.

Background: Autologous mesenchymal stem cells (MSC) have been shown to improve the functional outcome after severe skeletal muscle trauma. The reasons for this improvement have yet not been revealed. Up to now insufficient techniques of cell labelling, which could only be used for histologic analysis ex vivo, have been a problem.

The development of iron oxide nanoparticles, which are taken up and endosomally stored by stem cells, allows the evaluation of cellular behaviour in the muscle with the use of magnetic resonance imaging (MRI). Previous work has shown that labelling does not affect the proliferation and neurogenic differentiation capacity of embryonic stem cells. In the present study we are currently investigating the in vivo distribution and migration of locally transplanted MSC after blunt muscle trauma in a rat model.

Methods: MSC cultures are derived from tibial biopsies of Sprague Dawley rats via plastic adherence. A standardized open crush injury of the left soleus muscle is performed in each animal. 24 hours before transplantation cells are labelled with very small superparamagnetic iron oxid particles (VSOP-C200, Ferropharm, Teltow, Germany) and Green Fluorescent Protein (GFP). One week after trauma different amounts of stem cells (5×105, 1×106 and 5×106) are transplanted into the soleus muscle by local injection. Distribution and migration of the cells are evaluated over time by the repeated performance of high resolution-MRI at 7 Tesla (Bruker, Rheinstetten, Germany). At the endpoint of the study, three and six weeks after transplantation, the muscles are harvested and histologically and immunohistochemically analysed.

Results: Cells could be visualised inside the soleus muscle in the MRI 24 hours after transplantation showing characteristic signal extinctions in T2*-weighed images. The hypointense signal could be followed over the longest investigated time of six weeks and could be easily discriminated from the structures of the injured muscle. Preliminary results show that the cell pool changed its shape over time with the loss of an initially depicted injection canal and an increase in the surface/volume ratio. First histologic Prussian Blue stained sections showed co-localisation of the respective MRI signal and nanoparticle labelled cells. Fusion events of marked cells with regenerating myofibers could be observed.

Conclusion: Magnetic labelling of MSC is a powerful tool to analyse the in vivo behaviour of the cells after transplantation into a severly injured skeletal muscle. For the first time the observation of an intraindividual time course of the distribution of the transplanted cells is possible. Our preliminary results are promising and the ongoing work will further characterise migration processes and the correlation of the MRI results with muscle function evaluated by contraction force measurements.

Background: Scientific investigation of muscle trauma and regeneration is in need of well standardised models. These should mimic the clinical situation and be thoroughly described histologically and functionally. Existing models of blunt muscle injury are either based on segmental muscle damage or in case of whole muscle injury also affect the innervating structures. In this study we present a modified model of open crush injury to the whole soleus muscle of rats sparing the region of the neuromuscular junctions.

Methods: The left soleus muscles of male Sprague-Dawley rats were crushed with the use of a curved artery forceps. Functional regeneration was evaluated 1, 4 and 8 weeks after trauma (n = 6 per group) via in vivo measurement of muscle contraction force after fast twitch and tetanic stimulation of the sciatic nerve. The intact right soleus muscle served as an internal control. H & E staining was used for descriptive analysis of the trauma. The amount of fibrosis was determined histomorphologically on Picro-Sirius Red stained sections at each point of time.

Results: Across the evaluated regeneration period a continuous increase in contraction force after fast twitch as well as after tetanic stimulation could be observed – describing the functional regeneration of the traumatized soleus muscle over time. Tetanic force amounted to 0.34 ± 0.14 N, which are 23 ± 4% of the control side one week after trauma, and recovered to 55 ± 23% after eight weeks. Fast twitch contraction was reduced to 49 ± 7% of the control side at one week after injury and recovered to 68 ± 19% during the study period. Fibrotic tissue occupied 40 ± 4% of the traumatized muscles after the first week, decreased to approximately 25% after four weeks and remained at this value at eight weeks.

Conclusion: The trauma model characterised morphologically and functionally in the presented study allows the investigation of muscle regeneration caused by highly standardized injury exclusively to muscle fibers.

Introduction: The formation of new blood vessels is a prerequisite for bone healing. CYR61 (CCN1), an extracellular matrix-associated signaling protein, is a potent stimulator of angiogenesis and mesenchymal stem cell expansion and differentiation. A recent study showed that CYR61 is expressed during fracture healing and suggested that CYR61 plays a significant role in cartilage and bone formation. The hypothesis of the present study was that decreased fixation stability, which leads to a delay in healing, would lead to reduced CYR61 protein expression in fracture callus. The aim of the study was to quantitatively analyze CYR61 protein expression, vascularization, and tissue differentiation in the osteotomy gap and relate this to the mechanical fixation stability during the course of healing.

Materials and Methods: A mid-shaft osteotomy of the tibia was performed in two groups of sheep and stabilized with either a rigid or semirigid external fixator, each allowing different amounts of interfragmentary movement. The sheep were sacrificed at 2, 3, 6, and 9 weeks postoperatively. The tibiae were tested biomechanically and histological sections from the callus were analyzed immunohistochemically with regard to CYR61 protein expression and vascularization.

Results: Expression of CYR61 protein was upregulated at the early phase of fracture healing (2 weeks) and decreased over the healing time. Decreased fixation stability was associated with a reduced upregulation of the CYR61 protein expression and a reduced vascularization at 2 weeks leading to slower healing. The maximum cartilage callus fraction in both groups was reached at 3 weeks. However, the semirigid fixator group showed a significantly lower CYR61 immunoreactivity in cartilage than the rigid fixator group at this time point.

Discussion: The fraction of cartilage in the semirigid fixator group was not replaced by bone as quickly as in the rigid fixator group leading to an inferior histological and mechanical callus quality at 6 weeks and therefore to slower healing. The results supply further evidence that CYR61 may serve as an important regulator of bone healing.

The hypothesis of the current study was that the loading of the proximal femur is altered significantly by the surgical approach. The change in long-term periprosthetic bone mineral density in relation to the alteration of the musculature after the anterolateral (Group A) and transgluteal approaches (Group B) has been compared. Group A comprised 35 hip joints (30 patients) and Group B 47 hip joints (37 patients). No significant differences were seen between groups in respect to age, gender, or diaphyseal BMD distribution and in respect to average stem size in a Wilcoxon test. Measurement of BMD in femoral Gruen Zones I, II, VI, and VII revealed a significant bone loss in Group B compared with Group A; however the functional outcome showed no significant differences between the two groups postoperatively. Analysis of proximal femoral loading by means of a validated musculoskeletal model showed a considerable redistribution of the musculoskeletal loading across the hip during walking and stair climbing after a transgluteal compared with an anterolateral surgical approach. The muscular damage caused by the surgical approach seems to have a significant influence on the long-term bone loss and the initial postoperative loading of the proximal femur.