Mouse Mammary Gland Whole Mount Density Assessment across Different Morphologies Using a Bifurcated Program for Image Processing.

Mammographic density is associated with increased breast cancer risk. Conventional visual assessment of murine mouse models does not include quantified total density analysis. A bifurcated method was sufficient to obtain relative density scores on a broad range of two-dimensional whole mount images that contained both normal and abnormal findings. Image processing techniques, including a ridge operator and a gaussian denoising method, were used to isolate background away from mammary epithelium and use mean pixel intensity to represent mammary density on genetically engineered mouse models for breast cancer in mice 4 to 29 months of age. The bifurcated method allowed for application of an optimal image processing approach for the structural elements present in the whole mount images. Gaussian denoising was the optimal approach when more dense lobular growth and tertiary branching dominate and a ridge operator when epithelial growth was more sparse and secondary branching was the more dominant structural feature. The two processing approaches were combined in a single experimental flow program using an initial image density measurement as the decision point between the two approaches. Higher density was associated with lobular growth, tertiary branching, fibrotic stroma, and presence of cancer. The significance of the study is development of a readily accessible program for digital assessment of mammary gland whole mount density across a range of mammary gland morphologies.

Improving outcomes in acute and chronic periprosthetic hip and knee joint infection with a multidisciplinary approach.

AIMS: Periprosthetic hip and knee infection remains one of the most severe complications following arthroplasty, with an incidence between 0.5% to 1%. This study compares the outcomes of revision surgery for periprosthetic joint infection (PJI) following hip and knee arthroplasty prior to and after implementation of a specialist PJI multidisciplinary team (MDT). METHODS: Data was retrospectively analyzed from a single centre. In all, 29 consecutive joints prior to the implementation of an infection MDT in November 2016 were compared with 29 consecutive joints subsequent to the MDT conception. All individuals who underwent a debridement antibiotics and implant retention (DAIR) procedure, a one-stage revision, or a two-stage revision for an acute or chronic PJI in this time period were included. The definition of successfully treated PJI was based on the Delphi international multidisciplinary consensus. RESULTS: There were no statistically significant differences in patient demographics or comorbidities between the groups. There was also no significant difference in length of overall hospital stay (p = 0.530). The time taken for formal microbiology advice was significantly shorter in the post MDT group (p = 0.0001). There was a significant difference in failure rates between the two groups (p = 0.001), with 12 individuals (41.38%) pre-MDT requiring further revision surgery compared with one individual (6.67%) post-MDT inception. CONCLUSION: Our standardized multidisciplinary approach for periprosthetic knee and hip joint infection shows a significant reduction in failure rates following revision surgery. Following implementation of our MDT, our success rate in treating PJI is 96.55%, higher than what current literature suggests. We advocate the role of a specialist infection MDT in the management of patients with a PJI to allow an individualized patient-centred approach and care plan, thereby reducing postoperative complications and failure rates. Cite this article: Bone Jt Open 2021;2(7):509-514.

Vancomycin Wrap for Anterior Cruciate Ligament Surgery: Molecular Insights.

BACKGROUND: The use of the vancomycin wrap to pretreat the hamstring graft in anterior cruciate ligament reconstruction (ACLR) has grown in popularity since it was first described in 2012 and has significantly reduced rates of postoperative infection. However, it remains unknown if this antibiotic treatment affects the molecular composition of the graft. PURPOSE: To establish whether treatment with vancomycin at 5 mg/mL, the most commonly used concentration, alters the molecular function of the hamstring graft in ACLR. STUDY DESIGN: Controlled laboratory study. METHODS: Surplus hamstring tendon collected after routine ACLR surgery was used for in vitro cell culture and ex vivo tissue experiments. Vancomycin was used at 5 mg/mL in RPMI or saline diluent to treat cells and tendon tissue, respectively, with diluent control conditions. Cell viability at 30, 60, and 120 minutes was assessed via colorimetric viability assay. Tendon cells treated with control and experimental conditions for 1 hour was evaluated using semiquantitative reverse transcription analysis, immunohistochemistry staining, and protein quantitation via enzyme-linked immunosorbent assay for changes in apoptotic, matrix, and inflammatory gene and protein expression. RESULTS: Vancomycin treatment at 5 mg/mL significantly reduced tenocyte viability in vitro after 60 minutes of treatment (P < .05); however, this was not sustained at 120 minutes. Vancomycin-treated tendon tissue showed no significant increase in apoptotic gene expression, or apoptotic protein levels in tissue or supernatant, ex vivo. Vancomycin was associated with a reduction in inflammatory proteins from treated tendon supernatants (IL-6; P < .05). CONCLUSION: Vancomycin did not significantly alter the molecular structure of the hamstring graft. Reductions in matrix protein and inflammatory cytokine release point to a potential beneficial effect of vancomycin in generating a homeostatic environment. CLINICAL RELEVANCE: Vancomycin ACL wrap does not alter the molecular structure of the ACL hamstring graft and may improve graft integrity.

IL-17A mediates inflammatory and tissue remodelling events in early human tendinopathy.

Increasingly, inflammatory mediators are considered crucial to the onset and perpetuation of tendinopathy. We sought evidence of interleukin 17A (IL-17A) expression in early human tendinopathy and thereafter, explored mechanisms whereby IL-17A mediated inflammation and tissue remodeling in human tenocytes. Torn supraspinatus tendon (established pathology) and matched intact subscapularis tendon (representing 'early pathology') along with control biopsies were collected from patients undergoing shoulder surgery. Markers of inflammation and IL-17A were quantified by RT-PCR and immunohistochemistry. Human tendon cells were derived from hamstring tendon obtained during ACL reconstruction. In vitro effects of IL-17A upon tenocytes were measured using RT-PCR, multiplex cytokine assays, apoptotic proteomic profiling, immunohistochemistry and annexin V FACS staining. Increased expression of IL-17A was detected in 'early tendinopathy' compared to both matched samples and non-matched control samples (p < 0.01) by RT-PCR and immunostaining. Double immunofluoresence staining revealed IL-17A expression in leukocyte subsets including mast cells, macrophages and T cells. IL-17A treated tenocytes exhibited increased production of proinflammatory cytokines (p < 0.001), altered matrix regulation (p < 0.01) with increased Collagen type III and increased expression of several apoptosis related factors. We propose IL-17A as an inflammatory mediator within the early tendinopathy processes thus providing novel therapeutic approaches in the management of tendon disorders.

IL-21 receptor expression in human tendinopathy.

The pathogenetic mechanisms underlying tendinopathy remain unclear, with much debate as to whether inflammation or degradation has the prominent role. Increasing evidence points toward an early inflammatory infiltrate and associated inflammatory cytokine production in human and animal models of tendon disease. The IL-21/IL-21R axis is a proinflammatory cytokine complex that has been associated with chronic inflammatory diseases including rheumatoid arthritis and inflammatory bowel disease. This project aimed to investigate the role and expression of the cytokine/receptor pair IL-21/IL-21R in human tendinopathy. We found significantly elevated expression of IL-21 receptor message and protein in human tendon samples but found no convincing evidence of the presence of IL-21 at message or protein level. The level of expression of IL-21R message/protein in human tenocytes was significantly upregulated by proinflammatory cytokines (TNFα/IL-1ß) in vitro. These findings demonstrate that IL-21R is present in early human tendinopathy mainly expressed by tenocytes and macrophages. Despite a lack of IL-21 expression, these data again suggest that early tendinopathy has an inflammatory/cytokine phenotype, which may provide novel translational targets in the treatment of tendinopathy.

Does degree of trochlear dysplasia and position of femoral tunnel influence outcome after medial patellofemoral ligament reconstruction?

BACKGROUND: The medial patellofemoral ligament (MPFL) is the main restraining force against lateral patellar displacement. It is disrupted after patellar subluxation or dislocation. Reconstruction of the MPFL is frequently performed when nonoperative management fails and the patient experiences recurrent patellar dislocation. PURPOSE: To determine the relationship between the degree of trochlear dysplasia and femoral tunnel position and outcome after MPFL reconstruction. STUDY DESIGN: Case series; Level of evidence, 4. METHODS: A total of 68 patients (72 knees) with recurrent dislocation of the patella underwent MPFL reconstruction. The mean follow-up was 31.3 months (range, 13-72 months). Clinical and functional outcomes were recorded using the Kujala, Lysholm, and Tegner scores. Postoperative complications, participation in sporting activity, and overall patient satisfaction were determined. Radiographs were analyzed to evaluate congruence angle, lateral patellofemoral angle, patellar height, trochlear dysplasia, trochlear boss height, and position of the femoral tunnel. RESULTS: The mean Kujala, Lysholm, and Tegner scores postoperatively were 76.2, 73.8, and 3.6, respectively (n = 61). The mean congruence angle (n = 30) improved from 22.5° to 1.0° postoperatively (P = .000038), the lateral patellofemoral angle (n = 30) improved from 7.4° to 7.8° postoperatively (P = .048), and the patellar height (n = 46) using the Caton-Deschamps method improved from 1.1 to 1.0 postoperatively (P = .000016). Mild trochlear dysplasia grade A/B was found in 89% of patients (n = 54), and 11% of patients (n = 7) had severe grade C/D dysplasia. The mean distance from the anatomic insertion of the MPFL to the center of the tunnel was 9.3 mm (range, 0.5-28.2 mm), with 71.7% thought to be within 10 mm of the anatomic position defined by Schottle (n = 46). When patients with high-grade trochlear dysplasia were excluded, anatomically placed femoral tunnels demonstrated significantly better clinical scores than did tunnels not placed anatomically (Kujala score, P = .028; Lysholm score, P = .012). A multivariate logistic regression analysis also demonstrated that the distance of the femoral tunnel from the anatomic position predicted clinical outcome (Kujala score, P = .043; Lysholm score, P = .028). All of the patients with severe trochlear dysplasia (n = 7) suffered from recurrent dislocations postoperatively, compared with only 9.3% of patients (n = 5) with mild trochlear dysplasia (P = .0001). Four patients had patellar fractures postoperatively. Of patients with mild dysplasia, 83% were either very satisfied or satisfied with the outcome of their surgery compared with only 57% with severe dysplasia (P = .05). Of patients with mild trochlear dysplasia, 56% returned to sport postoperatively compared with only 43% of patients with severe trochlear dysplasia (P = .526). CONCLUSION: This study demonstrates the importance of restoration of the anatomic insertion point of the MPFL when performing MPFL reconstruction and proposes that this procedure should not be performed in isolation in patients with high-grade trochlear dysplasia.

Hypoxia: a critical regulator of early human tendinopathy.

OBJECTIVES: To seek evidence for the role of hypoxia in early human tendinopathy, and thereafter to explore mechanisms whereby tissue hypoxia may regulate apoptosis, inflammatory mediator expression and matrix regulation in human tenocytes. METHODS: Fifteen torn supraspinatus tendon (established pathology) and matched intact subscapularis tendon (representing 'early pathology') biopsies were collected from patients undergoing arthroscopic shoulder surgery. Control samples of the subscapularis tendon were collected from 10 patients undergoing arthroscopic stabilisation surgery. Markers of hypoxia were quantified by immunohistochemical methods. Human tendon-derived primary cells were derived from hamstring tendon tissue obtained during hamstring tendon anterior cruciate ligament reconstruction. The impact of hypoxia upon tenocyte biology ex vivo was measured using quantitative real-time PCR, multiplex cytokine assays, apoptotic proteomic profiling, immunohistochemistry and annexin V fluorescence-activated cell sorter staining. RESULTS: Increased expression of hypoxia-inducible factor 1α, Bcl-2 and clusterin was detected in subscapularis tendon samples compared with both matched torn samples and non-matched control samples (p<0.01). Hypoxic tenocytes exhibited increased production of proinflammatory cytokines (p<0.001), altered matrix regulation (p<0.01) with increased production of collagen type III operating through a mitogen-activated protein kinase-dependent pathway. Finally, hypoxia increased the expression of several mediators of apoptosis and thereby promoted tenocyte apoptosis. CONCLUSION: Hypoxia promotes the expression of proinflammatory cytokines, key apoptotic mediators and drives matrix component synthesis towards a collagen type III profile by human tenocytes. The authors propose hypoxic cell injury as a critical pathophysiological mechanism in early tendinopathy offering novel therapeutic opportunities in the management of tendon disorders.

A prospective comparison of two treatment options for tuberosity fractures of the proximal fifth metatarsal.

BACKGROUND: Tuberosity 'avulsion' fractures of the proximal fifth metatarsal are common injuries, but with few comparisons of conservative treatment options. OBJECTIVE: This study prospectively compared two commonly used conservative treatment methods. METHODS: 37 patients were allocated to treatment in either a plaster slipper (n=20) or tubi-grip support (n=17) for a period of 6 weeks. Patients were assessed at 2, 6 and 12 weeks after injury using a modified foot score which measured the level of pain and dysfunction. A radiograph was taken at 12 weeks. A Mann-Whitney U test compared median foot scores between the treatment groups at each review (p-value of 0.05 or less was deemed significant). RESULTS: A significantly (p=0.02) better foot score was measured in the plaster group at 2 weeks after injury (median score 68 (plaster group) compared to 57 (tubi-grip group)). However by the 6- and 12-week stages, the scores were comparable. Radiographs indicated two patients in each group with no clear evidence of union after 12 weeks. One patient remained symptomatic and required surgical treatment. CONCLUSIONS: The eventual clinical and radiological outcomes were comparable. During the initial 2 weeks, patients treated in plaster had a better-combined level of pain and function.