In vivo anti-melanoma activities of the Melan-A/MART-1101–115 T CD4+ cell peptide

Malignant melanoma causes significant health problems. The identification of tumour-associated antigens has led to novel approaches to increase T cell mediated anti-tumour immune response. Melan-A/MART-1 has been use as target antigen for several T cell based immunotherapeutic treatments. More recently, the critical role of CD4+ T cells in inducing and maintaining anti-tumour immunity has been increasingly recognized. In order to optimize tumour immunotherapy, greater efforts have been concentrated on the identification of tumour antigens presented by MHC class II molecules to CD4+ T cells. In a publication, Tiwari et al. (2004) [1] have identified by a computational approach the 15-mer amino-acid sequence 101–115 (PPAYEKLSAEQSPPP) of the Melan-A/MART-1 as a good target for a vigorous and safe immunotherapy. Therefore, we have investigated the in vivo anti-tumour activity of this peptide in a murine melanoma model. For the prophylactic treatment, 20 μg or 50 μg peptide was subcutaneously injected in mice once a week during 3 weeks before tumour induction. Treatment with 50 μg peptide significantly affected tumour development. Thus, our preliminary data demonstrate potential in vivo prophylactic activity of the 101–115 peptide-based vaccine to control melanoma growth.     

Early Results of the Mau Osteotomy for Correction of Moderate to Severe Hallux Valgus: A Review of 24 Cases

In our retrospective study, we report the objective results of the Mau osteotomy in the treatment of hallux valgus. We reviewed the results of 24 cases of moderate to severe hallux valgus deformities corrected with the Mau osteotomy of the first metatarsal combined with a distal soft-tissue procedure. Follow-up was possible in 24 cases. Preoperatively the mean hallux valgus and first intermetatarsal angles were 31.3 degrees and 16.6 degrees respectively, and were corrected postoperatively to an average of 13.00 degrees+/-7.15 degrees and 9.80 degrees+/-2.43 degrees respectively (P< .001). In the sagittal plane, the first metatarsal was shortened by an average of 2.00 mm. Two (8.3%) cases had dorsal elevation of the osteotomy fragment. Complications included 3 recurrences of the deformity, 1 frank nonunion, 8 dorsal cortical nonunions, 5 cases of undercorrection, and 1 case of broken hardware that was present in the nonunion that went on to revision. There were no superficial or deep infections, and no cases of transfer metatarsalgia were noted. In this series, the use of an oblique first metatarsal osteotomy with a dorsal shelf resulted in reliable and powerful correction of the first intermetatarsal angle in patients with moderate to severe hallux valgus. Particular attention should be paid to severe IM angles and the possibility of undercorrections. Despite ambulation postoperatively, the Mau osteotomy minimized dorsal malunion and the incidence of transfer metatarsalgia. LEVEL OF CLINICAL EVIDENCE: 4.     

CNS 7056: A Novel Ultra-short-acting Benzodiazepine

Background: A new benzodiazepine derivative, CNS 7056, has been developed to permit a superior sedative profile to current agents, i.e., more predictable fast onset, short duration of sedative action, and rapid recovery profile. This goal has been achieved by rendering the compound susceptible to metabolism via esterases. The authors now report on the profile of CNS 7056 in vitro and in vivo.

Methods: The affinity of CNS 7056 and its carboxylic acid metabolite, CNS 7054, for benzodiazepine receptors and their selectivity profiles were evaluated using radioligand binding. The activity of CNS 7056 and midazolam at subtypes ([alpha]1[beta]2[gamma]2, [alpha]2[beta]2[gamma]2, [alpha]3[beta]2[gamma]2, [alpha]5[beta]2[gamma]2) of the [gamma]-aminobutyric acid type A (GABAA) receptor was evaluated using the whole cell patch clamp technique. The activity of CNS 7056 at brain benzodiazepine receptors in vivo was measured in rats using extracellular electrophysiology in the substantia nigra pars reticulata. The sedative profile was measured in rodents using the loss of righting reflex test.

Results: CNS 7056 bound to brain benzodiazepine sites with high affinity. The carboxylic acid metabolite, CNS 7054, showed around 300 times lower affinity. CNS 7056 and CNS 7054 (10 [mu]m) showed no affinity for a range of other receptors. CNS 7056 enhanced GABA currents in cells stably transfected with subtypes of the GABAA receptor. CNS 7056, like midazolam and other classic benzodiazepines, did not show clear selectivity between subtypes of the GABAA receptor. CNS 7056 (intravenous) caused a dose-dependent inhibition of substantia nigra pars reticulata neuronal firing and recovery to baseline firing rates was reached rapidly. CNS 7056 (intravenous) induced loss of the righting reflex in rodents. The duration of loss of righting reflex was short (< 10 min) and was inhibited by pretreatment with flumazenil.     

Comparison between traditional 2-dimensional cephalometry and a 3-dimensional approach on human dry skulls.

The cephalogram is the standard used by orthodontists to assess skeletal, dental, and soft tissue relationships. This approach, however, is based on 2-dimensional (2D) views used to analyze 3-dimensional (3D) objects. The purpose of this project was to evaluate and compare a 3D imaging system and traditional 2D cephalometry for accuracy in recording the anatomical truth as defined by physical measurements with a calibrated caliper. Thirteen skeletal landmarks were located by both radiographic methods on 9 dry human skulls. Intraclass correlation (0.995), variance (0.054 mm(2)), and standard deviation (SD) (0.237 mm) were averaged over 76 measurements and derived from precision calipers to establish these physical measurements as a reliable gold standard to make comparisons of the 2D and 3D radiographic methods. The results showed great variability of the 2D from the gold standard, with the range varying from -17.68 mm (underestimation of Gn-Zyg R) to +15.52 mm (overestimation of Zyg L-Zyg R). In contrast, the 3D method (Sculptor, Glendora, Calif) indicated a range of the SD from -3.99 (underestimation) mm to +2.96 mm (overestimation). The 3D evaluation was much more precise, within approximately 1 mm of the gold standard. These results indicate that, when the actual distance is measured on a human skull in its true dimensions of 3D space, the Sculptor program, by using a 3D method, is more precise and 4 to 5 times more accurate than the 2D approach. Evaluating distances in 3D space with a 2D image grossly exaggerates the true measure and offers a distorted view of craniofacial growth. There is an inherent problem of representing a linear measure occupying a 3D space with a 2D image.     

Hybrid Anterior Cruciate Ligament Reconstruction: Introduction of a New Technique for Anatomic Anterior Cruciate Ligament Reconstruction

Recently, anatomic or double-bundle reconstruction of the anterior cruciate ligament (ACL) has been presented in an effort to more accurately restore the native anatomy. These techniques create 2 tunnels in both the femur and tibia to reproduce the bundles of the ACL. However, the increased number of tunnels, particularly on the femoral side, has raised some concerns among authors and surgeons. We describe a technique to reconstruct the 2 distinct bundles of the ACL by using a single femoral tunnel and 2 tibial tunnels, the “hybrid” ACL reconstruction. The femoral tunnel is drilled through an anteromedial arthroscopy portal, which allows placement in a more anatomic position. Fixation in the femur is achieved with a novel device that separates a soft-tissue graft into 2 independently functioning bundles. Once fixed in the femur, the anteromedial and posterolateral bundles of the graft are passed through respective tunnels at the anatomic footprint on the tibia. These bundles are independently tensioned, which creates a reconconstruction that is similar to the native ACL. The technique presented provides surgeons with an alternative to other double-bundle techniques involving 4 tunnels.     

Differential gene expression in cultured osteoblasts and bone marrow stromal cells from patients with Paget's disease of bone.

Paget's disease is a focal condition of bone. To study changes in cells within pagetic lesions, we cultured osteoblasts and stromal cells from 22 patients and compared gene expression in these cells to cells from healthy bone. We identified several differentially regulated genes, and we suggest that these changes could lead to the formation of the lesions. INTRODUCTION: Paget's disease is a focal condition of bone of unknown cause. Although it is regarded as primarily an osteoclast disorder, the tight coupling of the activity of osteoclasts and osteoblasts suggests that the osteoblast could play a key role in its pathogenesis. The aim of the study was to identify possible changes in pagetic osteoblasts and stromal cells that might contribute to the development of pagetic lesions. MATERIALS AND METHODS: Candidate genes were identified based on known bone cell regulators, supplemented with microarray analysis. Gene expression was determined by real-time PCR in primary cultures of osteoblasts and bone marrow stromal cells from pagetic patients and control subjects. Concentrations of secreted proteins were determined by ELISA. RESULTS: Dickkopf1 mRNA and protein levels were increased in both pagetic osteoblast and stromal cell cultures, and interleukin (IL)-1 and IL-6 were overexpressed in pagetic osteoblasts. These changes parallel recent findings in myeloma bone disease, which shares some clinical similarities with Paget's disease. Alkaline phosphatase was overexpressed, and bone sialoprotein and osteocalcin were underexpressed in pagetic osteoblasts, consistent with their circulating levels in pagetic patients. It is hypothesized that overexpression of Dickkopf1, IL-1, and IL-6 would result in stimulation of osteoclast proliferation and inhibition of osteoblast growth, leading to the development of the characteristic lytic bone lesions. By stimulating osteoblast differentiation, Dickkopf1 and IL-6 may also promote mineralization, leading to the conversion of lytic lesions to sclerotic. CONCLUSIONS: These findings suggest that dysregulated gene expression in pagetic osteoblasts could cause the changes in bone cell number and function characteristic of Paget's disease.     

Enhanced chondrogenesis and Wnt signaling in PTH-treated fractures.

Studies have shown that systemic PTH treatment enhanced the rate of bone repair in rodent models. However, the mechanisms through which PTH affects bone repair have not been elucidated. In these studies we show that PTH primarily enhanced the earliest stages of endochondral bone repair by increasing chondrocyte recruitment and rate of differentiation. In coordination with these cellular events, we observed an increased level of canonical Wnt-signaling in PTH-treated bones at multiple time-points across the time-course of fracture repair, supporting the conclusion that PTH responses are at least in part mediated through Wnt signaling. INTRODUCTION: Since FDA approval of PTH [PTH(1-34); Forteo] as a treatment for osteoporosis, there has been interest in its use in other musculoskeletal conditions. Fracture repair is one area in which PTH may have a significant clinical impact. Multiple animal studies have shown that systemic PTH treatment of healing fractures increased both callus volume and return of mechanical competence in models of fracture healing. Whereas the potential for PTH has been established, the mechanism(s) by which PTH produces these effects remain elusive. MATERIALS AND METHODS: Closed femoral fractures were generated in 8-wk-old male C57Bl/6 mice followed by daily systemic injections of either saline (control) or 30 microg/kg PTH(1-34) for 14 days after fracture. Bones were harvested at days 2, 3, 5, 7, 10, 14, 21, and 28 after fracture and analyzed at the tissue level by radiography and histomorphometry and at the molecular and biochemical levels level by RNase protection assay (RPA), real-time PCR, and Western blot analysis. RESULTS: Quantitative muCT analysis showed that PTH treatment induced a larger callus cross-sectional area, length, and total volume compared with controls. Molecular analysis of the expression of extracellular matrix genes associated with chondrogenesis and osteogenesis showed that PTH treated fractures displayed a 3-fold greater increase in chondrogenesis relative to osteogenesis over the course of the repair process. In addition, chondrocyte hypertrophy occurred earlier in the PTH-treated callus tissues. Analysis of the expression of potential mediators of PTH actions showed that PTH treatment significantly induced the expression of Wnts 4, 5a, 5b, and 10b and increased levels of unphosphorylated, nuclear localized beta-catenin protein, a central feature of canonical Wnt signaling. CONCLUSIONS: These results showed that the PTH-mediated enhancement of fracture repair is primarily associated with an amplification of chondrocyte recruitment and maturation in the early fracture callus. Associated with these cellular effects, we observed an increase in canonical Wnt signaling supporting the conclusion that PTH effects on bone repair are mediated at least in part through the activation of Wnt-signaling pathways.     

The anterolateral neck dissection with special reference to the digastric and hypoglossal tunnels

The elective surgical treatment of the neck for advanced T-stage laryngeal presentations of squamous cell carcinoma is typically the anterolateral neck dissection. Modifications of the anterior-based technique that include the use of digastric and hypoglossal tunnels to establish reliable landmarks result in a reproducible technique. The complex anatomic relationships of the anterior and superior part of the neck dissection are exposed and protected early in the technique, making this approach ideal for use in an academic setting.