Detection of gelatinolytic activity in developing basement membranes of the mouse embryo head by combining sensitive in situ zymography with immunolabeling

Genetic evidence indicates that the major gelatinases MMP-2 and MMP-9 are involved in mammalian craniofacial development. Since these matrix metalloproteinases are secreted as proenzymes that require activation, their tissue distribution does not necessarily reflect the sites of enzymatic activity. Information regarding the spatial and temporal expression of gelatinolytic activity in the head of the mammalian embryo is sparse. Sensitive in situ zymography with dye-quenched gelatin (DQ-gelatin) has been introduced recently; gelatinolytic activity results in a local increase in fluorescence. Using frontal sections of wild-type mouse embryo heads from embryonic day 14.5-15.5, we optimized and validated a simple double-labeling in situ technique for combining DQ-gelatin zymography with immunofluorescence staining. MMP inhibitors were tested to confirm the specificity of the reaction in situ, and results were compared to standard SDS-gel zymography of tissue extracts. Double-labeling was used to show the spatial relationship in situ between gelatinolytic activity and immunostaining for gelatinases MMP-2 and MMP-9, collagenase 3 (MMP-13) and MT1-MMP (MMP-14), a major activator of pro-gelatinases. Strong gelatinolytic activity, which partially overlapped with MMP proteins, was confirmed for Meckel's cartilage and developing mandibular bone. In addition, we combined in situ zymography with immunostaining for extracellular matrix proteins that are potential gelatinase substrates. Interestingly, gelatinolytic activity colocalized precisely with laminin-positive basement membranes at specific sites around growing epithelia in the developing mouse head, such as the ducts of salivary glands or the epithelial fold between tongue and lower jaw region. Thus, this sensitive method allows to associate, with high spatial resolution, gelatinolytic activity with epithelial morphogenesis in the embryo.

Sciatic-femoral nerve block with bupivacaine in goats undergoing elective stifle arthrotomy

The aim of this study was to describe the sciatic-femoral nerve block (SFNB) in goats and to evaluate the peri-operative analgesia when the goats underwent stifle arthrotomy. The animals were randomly assigned to one of four treatment groups: groups 0.25, 0.5 and 0.75 received 0.25%, 0.5% and 0.75% of bupivacaine, respectively, while group C (control group) received 0.9% NaCl. In all groups, the volume administered was 0.2 mL/kg. Intra-operatively, the proportion of animals receiving rescue propofol was significantly lower in groups 0.5 and 0.75, compared to group C. Post-operatively, the visual analogue scale (VAS) and total pain score were significantly higher in group C than in the other groups. Group 0.75 had the highest percentage of animals showing motor blockade. SFNB performed with bupivacaine resulted in better intra- and post-operative analgesia than SFNB performed with saline. Compared to the other concentrations, 0.5% bupivacaine resulted in satisfactory analgesia with acceptable side effects.

The clinical impact of p16 status in fine-needle aspirates of cervical lymph node metastasis of head and neck squamous cell carcinomas

Lymph node involvement is prognostically the most determinant clinical factor for patients with head and neck squamous cell carcinomas (HNSCCs). Ultrasound of the neck and fine-needle aspiration (FNA) cytology is one of the first diagnostic procedures and the most accurate diagnostic staging tool for the neck. Patients with HPV-positive oropharyngeal carcinomas (OPSCC) show a significantly better prognosis when compared with HPV-negative OPSCC. P16 overexpression is accepted as surrogate marker for HPV-positive in OPSCC. These HPV/p16-positive OPSCC are localized either in the palatal tonsils or the base of tongue and frequently present with lymph node metastases. We analyzed the correlation and reliability of p16 expression of the FNA of the lymph node metastasis with the immunohistochemical expression of p16 of the same lymph node metastasis and its corresponding primary tumor, as it could be of importance for determining the localization and different prognosis of the primary tumor. 54 HNSCC patients were evaluated, p16 expression of the primary tumors and their lymph node metastases correlated precisely. In 25 of the 54 HNSCC patients, a FNA of the lymph node metastases was taken before the treatment. The positive cytological and immunohistochemical p16 staining correlated exactly. Of the 17 histologically p16-negative lymph node metastases 15 FNA were p16-negative, whereas two samples were p16-positive. In our view, a cytological p16 analysis of cervical lymph node metastasis can facilitate the correct localization of the primary tumor and discriminate reliably HPV-positive OPSCC from HPV-negative HNSCC with their significantly diverse prognosis.

Reasons for ordering computed tomography scans of the head in patients with minor brain injury

Minor brain injury is a frequent condition. Validated clinical decision rules can help in deciding whether a computed tomogram (CT) of the head is required. We hypothesized that institutional guidelines are not frequently used, and that psychological factors are a common reason for ordering an unnecessary CT.

Inhibition of endogenous antagonists with an engineered BMP-2 variant increases BMP-2 efficacy in rat femoral defect healing

Bone morphogenetic proteins (BMP) have been used successfully by orthopedic clinicians to augment bone healing. However, these osteoinductive proteins must be applied at high concentrations to induce bone formation. The limited therapeutic efficacy may be due to the local expression of BMP antagonists such as Noggin that neutralize exogenous and endogenous BMPs. If so, inhibiting BMP antagonists may provide an attractive option to augment BMP induced bone formation. The engineered BMP-2 variant L51P is deficient in BMP receptor type I binding, but maintains its affinity for BMP receptor type II and BMP antagonists including Noggin, Chordin and Gremlin. This modification makes L51P a BMP receptor-inactive inhibitor of BMP antagonists. We implanted β-tricalcium phosphate ceramics loaded with BMP-2 and/or L51P into a critical size defect model in the rat femur to investigate whether the inhibition of BMP antagonist with L51P enhances the therapeutic efficacy of exogenous BMP-2. Our study reveals that L51P reduces the demand of exogenous BMP-2 to induce bone healing markedly, without promoting bone formation directly when applied alone.

Predictors of severe late radiotherapy-related toxicity after hyperfractionated radiotherapy with or without concomitant cisplatin in locally advanced head and neck cancer. Secondary retrospective analysis of a randomized phase III trial (SAKK 10/94)

This secondary analysis was performed to identify predictive factors for severe late radiotherapy (RT)-related toxicity after treatment with hyperfractionated RT +/- concomitant cisplatin in locally advanced head and neck cancer.

What is the evidence supporting the prevention of osteoarthritis and improved femoral coverage after shelf procedure for Legg-Calvé-Perthes disease?

The evidence supporting continued use of shelf acetabuloplasty in Legg-Calvé-Perthes disease (LCPD) is not well-defined, and there is controversy regarding the long-term benefits related to clinical and functional improvement.

Clinical stability of slipped capital femoral epiphysis does not correlate with intraoperative stability

The most important objective of clinical classifications of slipped capital femoral epiphysis (SCFE) is to identify hips associated with a high risk of avascular necrosis (AVN)--so-called unstable or acute slips; however, closed surgery makes confirmation of physeal stability difficult. Performing the capital realignment procedure in SCFE treatment we observed that clinical estimation of physeal stability did not always correlate with intraoperative findings at open surgery. This motivated us to perform a systematic comparison of the clinical classification systems with the intraoperative observations.

Small-bowel MRI in children and young adults with Crohn disease: retrospective head-to-head comparison of contrast-enhanced and diffusion-weighted MRI

Small-bowel MRI based on contrast-enhanced T1-weighted sequences has been challenged by diffusion-weighted imaging (DWI) for detection of inflammatory bowel lesions and complications in patients with Crohn disease.

High-accuracy patient-to-image registration for the facilitation of image-guided robotic microsurgery on the head

Image-guided microsurgery requires accuracies an order of magnitude higher than today's navigation systems provide. A critical step toward the achievement of such low-error requirements is a highly accurate and verified patient-to-image registration. With the aim of reducing target registration error to a level that would facilitate the use of image-guided robotic microsurgery on the rigid anatomy of the head, we have developed a semiautomatic fiducial detection technique. Automatic force-controlled localization of fiducials on the patient is achieved through the implementation of a robotic-controlled tactile search within the head of a standard surgical screw. Precise detection of the corresponding fiducials in the image data is realized using an automated model-based matching algorithm on high-resolution, isometric cone beam CT images. Verification of the registration technique on phantoms demonstrated that through the elimination of user variability, clinically relevant target registration errors of approximately 0.1 mm could be achieved.