Introduction. The hematoma occurring at a fracture site is known to play an important role in fracture healing. Previously, we demonstrated that fracture hematoma contained multilineage mesenchymal progenitor cells. On the other hand, the process of fracture healing is associated by two different mechanisms, intramembranous and endochondral. However, there are no reports proving the details about cellular analysis in the process of endochondoral ossification. Hypothesis. We hypothesized that one of the cell origins for endochondral ossification after fracture was hematoma. Materials & Methods. Fracture hematoma was obtained during osteosynthesis. Hematoma-derived cells were isolated and cultured for 5-weeks of chondrogenic induction followed by 2-weeks hypertrophic induction using pellet culture system. The pellets were analyzed histologically and immunohistochemically. The gene expression levels of chondrogenic, hypertrophic, osteogenic and angiogenic markers were measured by real-time PCR. Results. The histological and immunohistochemical analysis revealed that the Hematoma-derived cells differentiated into hypertrophic chondrocytes through chondrocytes, and finally differentiate into calcifying chondrocytes. The same trend was seen in the gene expression using real-time PCR analysis. Discussion & Conclusions. Our results suggest that fracture hematoma may be an origin of cells which play key roles in the process of endochondoral ossification during fracture healing

Background. An increased incidence of periprosthetic osteolysis, resulting in loss of biologic fixation, has been recently reported in contemporary THAs with low-carbide metal-on-metal compared to metal-on-polyethylene couple bearings. A hypersensitivity reaction due to Co and Cr debris is reported as a potential cause for failure of THAs with high-carbide bearings, but there are no evidence-based data for this reaction in low-carbide metal-on-metal bearings. Questions/purposes. We investigated whether there were differences in immunologic hypersensitivity reactions in retrievals from revised THAs with COP versus MOM bearing couples. Patients and Methods. We compared newly formed capsule and periprosthetic interface membranes retrieved from revision surgery due to aseptic failure in 20 patients with low-carbide bearings and 13 patients with ceramic-on-polyethylene bearings. For control tissue we obtained samples from the hip capsule during the primary THA implantation in 13 patients with low-carbide bearings and seven with ceramic-on-polyethylene. We examined the tissues with conventional histologic and immunohistochemical methods. Results. Compared to the controls and the tissue from patients with ceramic-on-polyethylene bearings, the tissues from patients with low-carbide metal-on-metal bearings were associated with (1) extensive necrosis and fibrin exudation in the newly formed hip capsule and (2) diffuse and perivascular lymphocytic infiltration of a higher degree than in the ceramic-on-polyethylene hips in conventional histologic examination and (3) more T than B cells. Conclusions. The conventional histologic and immunohistochemical findings in tissues retrieved from failed THAs with low-carbide metal-on-metal bearings are consistent with a link between hypersensitivity and osteolysis with low-carbide bearing couple THA

Introduction. Our previous study using microarray analysis showed that Rad (Ras associated with diabetes) was highly expressed in nonunion. The purpose of this study is to investigate the gene expression and immunolocalization of Rad, and other Ras-related G proteins: Rem1 and Rem2 in fracture/nonunion site using rat experimental models. Hypothesis. We hypothesized that Rad had a significant role in nonunion formation. Materials & Methods. For standard healing model, K-wire was inserted into the femur and a closed fracture was created. Nonunion model was produced by periosteal cauterization at the fracture site. At post-fracture days 3, 7, 10, 14, 21, and 28, RNA was extracted from callus or fibrous tissue for real-time PCR. At day 14, specimens were harvested for immunohistochemistry. Results. Significant difference of Rad gene expression was not observed between standard healing fracture and nonunion at the earlier time points. In contrast, significantly higher expression in nonunion was observed at the later time points. There were no significant differences between standard healing fracture and nonunion in gene expression of Rem1 and Rem2. In immunohistochemical analysis, Rad and Rem1 were detected in the fracture site, and Rem2 was not detected. On the other hand, Rad was only detected in fibrous tissue in nonunion. Discussion & Conclusion. Our results suggest a significant role of Rad in fracture healing and nonunion formation. Rad may become a target agent for treatment of nonunion

Femoroacetabular impingement (FAI) is a significant cause of osteoarthritis in young active individuals but the pathophysiology remains unclear. Increasing mechanistic studies point toward an inflammatory component in OA. This study aimed to characterise inflammatory cell subtypes in FAI by exploring the phenotype and quantification of inflammatory cells in FAI versus OA samples. Ten samples of labrum were obtained from patients with FAI (confirmed pathology) during open osteochondroplasty or hip arthroscopy. Control samples of labrum were collected from five patients with osteoarthritis undergoing total hip arthroplasty. Labral biopsies were evaluated immunohistochemically by quantifying the presence of macrophages (CD68 and CD202), T cells (CD3), mast cells (mast cell tryptase) and vascular endothelium (CD34). Labral biopsies obtained from patients with FAI exhibited significantly greater macrophage, mast cell and vascular endothelium expression compared to control samples. The most significant difference was noted in macrophage expression (p<0.01). Further sub typing of macrophages in FAI using CD202 tissue marker revealed and M2 phenotype suggesting that these cells are involved in a regenerate versus a degenerate process. There was a modest but significant correlation between mast cells and CD34 expression (r=0.4, p<0.05) in FAI samples. We provide evidence for an inflammatory cell infiltrate in femoroacetabular impingement. In particular, we demonstrate significant infiltration of mast cells and macrophages suggesting a role for innate immune pathways in the events that mediate hip impingement. Further mechanistic studies to evaluate the net contribution and hence therapeutic utility of these cellular lineages and their downstream processes may reveal novel therapeutic approaches to the management of early hip impingement

Injuries to growth plates may initiate the formation of reversible or irreversible bone-bridges, which may lead to partial or full closure of the growth plate resulting in bone length discrepancy, axis deviation or joint deformity. Blood vessels and vascular invasion are essential for the formation of new bone tissue. The aim of our study was to investigate the spatial and temporal expression VEGF and its receptors R1 and R2 as well as the ingrowth of vessels in the formation of bone bridges in a rat physeal injury model. Quantitative Real Time - Polymerase Chain Reaction (qRT-PCR) was performed for Vascular Endothelial Growth Factor (VEGF) and its R1 and R2 receptors. Samples from the proximal epiphysis, physis and metaphysis of the tibial bone were prepared for immunohistochemical analysis to demonstrate the spatial expression of VEGF and its R1 and R2 receptors as well as laminin. Kinetic expression of VEGF and VEGF-R1 mRNA documented a tendency towards an expression increase on day 7. Histological analysis showed a haematoma containing bone fragments on day 1which was replaced by a bony bridge by day 14. This remodelled and consolidated by day 82. These trabeculae were accompanied by vessel formation. Expression of VEGF was observed on the bone fragments and the haematoma from day 1 through to day 82. Although VEGF-R1 was expressed at all time points the expression of VEGF-R2 was noted until the 14th day. Physeal bone bridge formation is a combination of both enchondral and intramembranous ossification. This is in part triggered by the bony debris observed within the lesion in the first few days. By washing this debris out the likelihood of bone bridge formation may be reduced. We recommend this practice when operating on the physis in order to avoid iatrogenic physeal bar formation

INTRODUCTION. Fresh bipolar shell osteochondral allograft (FBOA) is a controversial treatment option for post-traumatic ankle arthritis. Immunological response to transplanted cartilage may play a role in failure. Aim of the study is to compare two groups of patients who received FBOA in association or not to immunosuppressive therapy. METHODS. 2 groups, of 20 patients each, underwent FBOA. Only one group (group-B) received immunosuppressive therapy. Pre-operative and follow-up evaluation were clinical (AOFAS) and radiographical (X-Rays, CT- scan, MRI). Bioptic samples harvested during II look were examined by histochemical, immunohistochemical (ICRS II score) and by genetic typing analyses. RESULTS. Group-A pre-operative AOFAS score improved from 28.2 ± 10.9, to 69.9 ± 18.2 at 24 months follow-up(p<0.005), while Group B improved from 26.2 ± 6.8 to 71.4 ± 7.3 (p<0.005). Comparison of clinical outcomes between the groups was non-significant. Group B showed better morphology of the grafts (ICRS II score mean of 68%) compared to Group A (mean of 40%) (p<0.05). Genetic typing showed a mixed recipient/donor DNA presence. Kendall ordinal correlation between groups and ICRS score was found. All the samples rated as 100 were in group B, while all the samples rated 0 were in group A (=0.506, p=0.008). CONCLUSIONS. Although clinical results were comparable in the two groups, better histological score in Group B evidentiated hyaline cartilage significatively better preserved. Genetic typing showed the presence of cells of the host into the transplanted cartilage suggesting a possible colonization of transplanted cartilage by host cells never described before

Objectives

The biomembrane (induced membrane) formed around polymethylmethacrylate (PMMA) spacers has value in clinical applications for bone defect reconstruction. Few studies have evaluated its cellular, molecular or stem cell features. Our objective was to characterise induced membrane morphology, molecular features and osteogenic stem cell characteristics.