The 'triple contrast' method in experimental wound ballistics and backspatter analysis.

In practical forensic casework, backspatter recovered from shooters' hands can be an indicator of self-inflicted gunshot wounds to the head. In such cases, backspatter retrieved from inside the barrel indicates that the weapon found at the death scene was involved in causing the injury to the head. However, systematic research on the aspects conditioning presence, amount and specific patterns of backspatter is lacking so far. Herein, a new concept of backspatter investigation is presented, comprising staining technique, weapon and target medium: the 'triple contrast method' was developed, tested and is introduced for experimental backspatter analysis. First, mixtures of various proportions of acrylic paint for optical detection, barium sulphate for radiocontrast imaging in computed tomography and fresh human blood for PCR-based DNA profiling were generated (triple mixture) and tested for DNA quantification and short tandem repeat (STR) typing success. All tested mixtures yielded sufficient DNA that produced full STR profiles suitable for forensic identification. Then, for backspatter analysis, sealed foil bags containing the triple mixture were attached to plastic bottles filled with 10 % ballistic gelatine and covered by a 2-3-mm layer of silicone. To simulate backspatter, close contact shots were fired at these models. Endoscopy of the barrel inside revealed coloured backspatter containing typable DNA and radiographic imaging showed a contrasted bullet path in the gelatine. Cross sections of the gelatine core exhibited cracks and fissures stained by the acrylic paint facilitating wound ballistic analysis.

HLA class II antigen expression in colorectal carcinoma tumors as a favorable prognostic marker.

The goal of this study was to determine the frequency of HLA class II antigen expression in colorectal carcinoma (CRC) tumors, its association with the clinical course of the disease, and the underlying mechanism(s). Two tissue microarrays constructed with 220 and 778 CRC tumors were stained with HLA-DR, DQ, and DP antigen-specific monoclonal antibody LGII-612.14, using the immunoperoxidase staining technique. The immunohistochemical staining results were correlated with the clinical course of the disease. The functional role of HLA class II antigens expressed on CRC cells was analyzed by investigating their in vitro interactions with immune cells. HLA class II antigens were expressed in about 25% of the 220 and 21% of the 778 tumors analyzed with an overall frequency of 23%. HLA class II antigens were detected in 19% of colorectal adenomas. Importantly, the percentage of stained cells and the staining intensity were significantly lower than those detected in CRC tumors. However, HLA class II antigen staining was weakly detected only in 5.4% of 37 normal mucosa tissues. HLA class II antigen expression was associated with a favorable clinical course of the disease. In vitro stimulation with interferon gamma (IFNγ) induced HLA class II antigen expression on two of the four CRC cell lines tested. HLA class II antigen expression on CRC cells triggered interleukin-1β (IL-1β) production by resting monocytes. HLA class II antigen expression in CRC tumors is a favorable prognostic marker. This association may reflect stimulation of IL-1β production by monocytes.

Intra-arterial injection of neural stem cells using a microneedle technique does not cause microembolic strokes

Intra-arterial (IA) injection represents an experimental avenue for minimally invasive delivery of stem cells to the injured brain. It has however been reported that IA injection of stem cells carries the risk of reduction in cerebral blood flow (CBF) and microstrokes. Here we evaluate the safety of IA neural progenitor cell (NPC) delivery to the brain. Cerebral blood flow of rats was monitored during IA injection of single cell suspensions of NPCs after stroke. Animals received 1 × 10(6) NPCs either injected via a microneedle (microneedle group) into the patent common carotid artery (CCA) or via a catheter into the proximally ligated CCA (catheter group). Controls included saline-only injections and cell injections into non-stroked sham animals. Cerebral blood flow in the microneedle group remained at baseline, whereas in the catheter group a persistent (15 minutes) decrease to 78% of baseline occurred (P<0.001). In non-stroked controls, NPCs injected via the catheter method resulted in higher levels of Iba-1-positive inflammatory cells (P=0.003), higher numbers of degenerating neurons as seen in Fluoro-Jade C staining (P<0.0001) and ischemic changes on diffusion weighted imaging. With an appropriate technique, reduction in CBF and microstrokes do not occur with IA transplantation of NPCs.

Confocal imaging protocols for live/dead staining in three-dimensional carriers

In tissue engineering, a variety of methods are commonly used to evaluate survival of cells inside tissues or three-dimensional (3D) carriers. Among these methods confocal laser scanning microscopy opened accessibility of 3D tissue using live cell imaging into the tissue or 3D scaffolds. However, although this technique is ideally applied to 3D tissue or scaffolds with thickness up to several millimetres, this application is surprisingly rare and scans are often done on slices with thickness <20 μm. Here, we present novel protocols for the staining of 3D tissue (e.g. intervertebral disc tissue) and scaffolds, such as fibrin gels or alginate beads.

A Minimally Invasive Technique for the Detection and Analysis of Pulmonary Fat Embolism: A Feasibility Study*

We investigated the feasibility of postmortem percutaneous needle biopsy (PNB) for obtaining pulmonary samples adequate for the study of pulmonary fat embolism (PFE). Samples of both lungs were obtained from 26 cadavers via two different methods: (i) PNB and (ii) the double-edged knife technique, the gold standard at our institute. After water storage and Sudan III staining, six forensic pathologists independently examined all samples for the presence and severity of PFE. The results were compared and analyzed in each case regarding the vitality of the PFE and its relationship to the cause of death. The results showed that PFE was almost identically diagnosed and graded on the samples obtained via both methods. The discrepancies between the two techniques did not affect the diagnoses of vitality or cause of death related to PFE. This study demonstrates the feasibility of the PNB sampling method for the diagnosis and interpretation of PFE in the postmortem setting.

Tight contact technique during side-to-side laser tissue soldering of rabbit aortas improves tensile strength

BACKGROUND: Cerebral revascularization may be indicated either for blood flow preservation or flow augmentation, often in clinical situations where neither endovascular nor standard surgical intervention can be performed. Cerebral revascularization can be performed by using a temporary occlusive or a non-occlusive technique. Both of these possibilities have their specific range of feasibility. Therefore non-occlusive revascularization techniques have been developed. To further reduce the risks for patients, less time consuming, sutureless techniques such as laser tissue soldering are currently being investigated. METHOD: In the present study, a new technique for side-to-side anastomosis was developed. Using a "sandwich technique", two vessels are kept in close contact during the laser soldering. Thoraco-abdominal aortas from 24 different rabbits were analyzed for laser irradiation induced tensile strength. Two different irradiation modes (continuous and pulsed) were used. The results were compared to conventional, noncontact laser soldering. Histology was performed using HE, Mason's Trichrome staining. FINDINGS: The achieved tensile strengths were significantly higher using the close contact "sandwich technique" as compared to the conventional adaptation technique. Furthermore, tensile strength was higher in the continuously irradiated specimen as compared to the specimen undergoing pulsed laser irradiation. The histology showed similar denaturation areas in both groups. The addition of a collagen membrane between vessel components reduced the tensile strength. CONCLUSION: These first results proved the importance of close and tight contact during the laser soldering procedure thus enabling the development of a "sandwich laser irradiation device" for in vivo application in the rabbit.