CIRS-LAS - a novel approach to increase transparency in laboratory animal science for improving animal welfare by reducing laboratory animal distress.

The 3Rs principle is highly topical in animal-based research. These include, above all, new scientific methods for conducting experiments without an animal model, by using non-animal models (Replace), reducing the number of laboratory animals (Reduction) or taking measures to keep the stress on the laboratory animal as low as possible (Refinement). Despite numerous modern alternative approaches, the complete replacement of animal experiments is not yet possible. The exchange in the team about the daily work with laboratory animals, about open questions and problems, contributes to a reflection of one's own work and to a better understanding of the work of the others. CIRS-LAS (Critical Incident Reporting System in Laboratory Animal Science) represents a reporting system for incidents in laboratory animal science. It is urgently needed because the lack of transparency about incidents leads to the repetition of failed experiments. Negative experiences from animal-based experiments are often not mentioned in publications, and the fear of hostility is still very high. Therefore, a constructive approach to errors is not a matter of course. To overcome this barrier, CIRS-LAS was created as a web-based database. It addresses the areas of reduction and refinement of the 3Rs principle by providing a platform to collect and analyze incidents. CIRS-LAS is open to all individuals working with laboratory animals worldwide and currently exists with 303 registered members, 52 reports, and an average of 71 visitors per month. The development of CIRS-LAS shows, that an open and constructive error culture is difficult to establish. Nevertheless, the upload of a case report or the search in the database leads to an active reflection of critical occurrences. Thus, it is an important step towards more transparency in laboratory animal science. As expected, the collected events in the database concern different categories and animal species and are primarily reported by persons involved in an experiment. However, reliable conclusions about observed effects require further analysis and continuous collection of case reports. Looking at the development of CIRS-LAS, its high potential is shown in considering the 3Rs principle in daily scientific work.

Iodine doping of a photocuring, one-component embedding medium for µCT and histology.

OBJECTIVE: Evaluation of µCT scans of bone implant complexes often shows a specific problem: if an implant material has a very similar radiopacity as the embedding medium (e.g. methacrylate resin), the implant is not visible in the µCT image. Segmentation is not possible, and especially osseointegration as one of the most important parameter for biocompatibility is not evaluable. METHODS: To ensure µCT visualisation and contrast enhancement of the evaluated materials, the embedding medium Technovit® VLC7200 was doped with an iodine monomer for higher radiopacity in different concentrations and tested regarding to handling, polymerisation, and histological preparation, and visualisation in µCT. Six different µCT devices were used and compared with regard to scan conditions, contrast, artefacts, image noise, and spatial resolution for the evaluation of the bone-implant blocks. RESULTS: Visualisation and evaluation of all target structures showed very good results in all µCT scans as well as in histology and histological staining, without negative effects caused by iodine doping. Subsequent evaluation of explants of in vivo experiments without losing important information was possible with iodine doped embedding medium. CONCLUSION: Visualisation of implants with a similar radiopacity as the embedding medium could be considerably improved. µCT scan settings should be selected with the highest possible resolution, and different implant materials should be scanned individually for optimal segmentation. µCT devices with higher resolutions should be preferred. ADVANCES IN KNOWLEDGE: Iodine doped embedding medium is a useful option to increase radiopacity for better visualisation and evaluation of special target structures in µCT.

Electrospun mucosal wound dressings containing styptics for bleeding control.

Two major aspects need to be focused to accelerate wound healing of mucosal damages especially in the field of otorhinolaryngology. (i) The problem of application due to the small access during surgery, (ii) the fixation of the wound dressing to reveal a stable healing process. In the present work the high request to a mucosal wound dressing which additionally support hemostasis was addressed. We developed an electrospun fabric made of poly(l-lactide-co-d/l-lactide) (PLA) which can be loaded with the hemostatic agents adrenaline and tranexamic acid to cover mucosal lesions analogues to common skin patches. These loaded electrospun fabrics were demonstrated to be biocompatible, thin and flexible, and thus could be adapted individually to the mucosal defect with respect to localization and size of the lesion. The treatment of mucosal defects with these loaded PLA wound dressings induced a faster and time controlled hemostatic reaction, which significantly improved the healing process.

Malignancy of bladder cancer cells is enhanced by tumor-associated fibroblasts through a multifaceted cytokine-chemokine loop.

The microenvironment of tumor cells is critically involved in tumor development and progression. Tumor-associated fibroblasts (TAFs) represent a major constituent of the tumor stroma. Tumor cells are operative in the activation of TAFs, whereas TAFs in turn contribute to tumor cell malignancy. This report describes mechanisms of communication between fibroblasts and urinary bladder cancer (UBC) cells. Migration of bladder cancer cell lines RT112 and Cal-29, representing two different grades of dedifferentiation, was enhanced by cocultivation with TAFs. Conditioned medium from tumor cells induced the release of interleukin (IL)-8, hepatocyte growth factor (HGF), matrix metalloproteinase-2, granulocyte macrophage colony-stimulating factor, and monocyte chemotactic protein (MCP)-1 by TAFs. Tumor cell-derived IL-1α was identified as a major mediator of these stimulatory effects. Fibroblasts, on the other hand, exerted a migration and invasion stimulating influence on UBC cells. MCP-1 and HGF were shown to promote cell migration of both bladder cancer cell lines.

FISH analysis of washing urine from the upper urinary tract for the detection of urothelial cancers.

INTRODUCTION: To evaluate FISH analysis of washing urine from the upper urinary tract (UUT) in comparison with cytology (Cyt) for the detection of urothelial cancers. PATIENTS AND METHODS: In 82 patients with symptoms or abnormalities of the UUT sampling of washing urine for FISH and Cyt and a stepwise diagnostic work-up (e.g. retrograde ureteropyelography, ureterorenoscopy and endoscopic biopsy) were performed. In case of endoscopically and/or histologically proven malignancy patients either underwent nephroureterectomy, partial ureterectomy or local treatment. Sensitivity and specificity for FISH and Cyt as well as its combination were determined. RESULTS: Urothelial cancer of the UUT was detected in 20 patients. Eleven patients underwent nephroureterectomy, six partial ureterectomy and three endoscopic tumour treatment. This revealed nine pTa, three pT1 and seven muscle-invasive tumours. Twelve tumours were classified as low and seven as high-grade tumours. In one patient with a macroscopic unequivocal finding of tumour, endoscopic laser ablation without histologic confirmation was performed. FISH was evaluable in 76 patients and detected 16 tumours with a sensitivity and specificity of 84.2 and 91.1 %, respectively. Cyt was performed in 79 and was evaluable in 78 patients. It detected ten tumours with a sensitivity and specificity of 52.6 and 91.4 %, respectively. Cyt and FISH together detected 19 tumours with (sensitivity 100 % and specificity 83.6 %). CONCLUSION: FISH was more sensitive than and equally specific to Cyt in the detection of urothelial cancers of the UUT. Both markers in combination revealed the best sensitivity, making it a possible approach in future settings.

Specific protein and miRNA patterns characterise tumour-associated fibroblasts in bladder cancer.

PURPOSE: Tumour development and progression are strongly affected by interaction of tumour cells and tumour stroma. Different tumour models demonstrate a supportive effect of tumour-associated fibroblasts (TAF) on the tumour genesis. Aims of the present study are the isolation of TAF from primary urinary bladder tumour specimens and the proteomic and epigenetic characterisation. METHODS: TAF were isolated from cultured urinary bladder tumour specimens. Therefore, primary tumour material was treated with EDTA followed by two separated detachment steps. Non-tumour fibroblasts were isolated from foreskin and normal bladder tissues. Proteins and total RNA were isolated from cultured fibroblasts. Protein pattern analyses were carried out by SELDI-TOF-MS. The miRNA expression profile was analysed by miRNA microarray. RESULTS: By optimising cell culture routines, we achieved to isolate and subsequently cultivate TAF from primary tumour material of the urinary bladder. SELDI-TOF-MS measurements reveal distinct differences in the proteomic patterns of TAF and non-tumour fibroblasts. Microarray analyses indicate specific expression of several miRNAs in TAF and non-tumour fibroblasts. CONCLUSION: In summary, we determined proteomic and epigenetic differences between non-tumour fibroblasts and TAF of urinary bladder carcinoma and identified specific protein expression patterns as well as miRNA profiles of TAF in comparison with non-tumour fibroblasts. These findings provide more insights into the complex tumour network and a good starting point for the identification of markers for the prediction of tumour development and progression based on specific TAF expression patterns.