No abstract

Polychrome sequential labeling with fluorochromes is a standard technique for the investigation of bone formation and regeneration processes in vivo. However, for human application, only tetracycline and its derivates are approved as fluorochromes. Therefore, the aim of this study was to determine the fluorescence characteristics of the different tetracycline derivates to assess the feasibility of sequential in vivo bone labeling using distinguishable fluorochromes. Eight different tetracycline derivates were injected subcutaneously into growing rats as a single dose or sequentially in different combinations. After preparation of resin-embedded undecalcified bone sections, the fluorescence properties of the tetracycline derivates in bone were analyzed using conventional fluorescence microscopy, spectral image analysis and confocal laser scanning microscopy. Each tetracycline derivate exhibited a characteristic fluorescence spectrum, but the differences between them were small. Chlortetracycline could be discriminated reliably from all other derivates and could therefore be combined with any other tetracycline derivate for reliably distinguishable double labeling. Tetracycline itself exhibited the brightest fluorescence of all the investigated derivates. Interestingly, in conventional microscopy the same tetracycline derivative can appear in different colours to the human eye, even if spectral analysis confirmed identical emission peaks. In conclusion, the data suggest that fluorescence double labeling of bone is feasible using appropriate tetracycline derivates in combination with spectral imaging modalities.

Fluorochrome sequential labelling of mineralizing tissues is commonly used in different fields of clinical and basic research. Recently we improved polychrome fluorescent sequential labelling of bone by applying spectral image analysis to discriminate seven different fluorochromes. Although basic mineralization processes of bone and teeth follow comparable principles, the respective tissues differ in terms of matrix composition and mineral assembly.The aim of this study therefore was to investigate the feasibility of this new technique for polychrome sequential labelling of teeth and to demonstrate the advantages in the field of dentistry. Furthermore, the exact labelled area of each fluorochrome could be measured, even in regions of overlapping fluorochromes. The technique presented may provide a basis for further investigations of mineralization processes of different anatomical dental structures.

The human Y-box binding protein 1 (YB-1) is known to be a promising target for cancer therapy. We have demonstrated that YB-1 plays an important role in the adenoviral life cycle by regulating the adenoviral E2-gene expression. Thus, we studied the oncolytic effect of the recombinant adenovirus Ad-Delo3-RGD, in which the transactivation domain CR3 of the E1A protein is ablated to enable viral replication only in YB-1 positive cancer cells. In vitro Southern Blot analysis and cytopathic effect assays demonstrate high anti-glioma potency, which was significantly increased in combination with temozolomide (TMZ), daunorubicin and cisplatin. Since vascular endothelial growth factor (VEGF) is thought to promote the hypervascular phenotype of primary, malignant brain tumors, we also tested Ad-Delo3-RGD in regard to the inhibition of VEGF expression. Indeed, we found that Ad-Delo3-RGD induced VEGF down regulation, which was even amplified under hypoxic conditions. Tumor-bearing nudemice treated with the YB-1 dependent oncolytic adenovirus showed significantly smaller tumors than untreated controls. Furthermore, combination therapy with TMZ led to a regression in all treated animals with complete tumor regression in 33 % of analyzed mice, which was verified by bioluminescence imaging and histological studies. In addition, histopathological evaluation revealed enhanced apoptosis and a reduction in tumor vessel formation, indicating that AdDelo3-RGD has an anti-angiogenic effect in addition to its oncolytic capacity in vivo. Hence, our results demonstrate that the combination therapy of YB-1 dependent virotherapy and TMZ is effective in a xenograft glioma mouse model and might be useful in a YB-1 based clinical setting.

Concerning dental implant systems, a main problem is the adhesion of peri-implant mucosa in the cervical region. The aim of the present study was to use a laser for modifying titanium implants to promote mucosal adhesion, which is indispensable as a biological barrier against bacterial infection. By the use of a KrF excimer laser, it was possible to induce a holey structure on the polished area of the implant surface, which was analysed by a scanning electron microscope. In addition, the attachment of fibroblast cells to the created structures was investigated with the aid of an environmental scanning electron microscope. It turned out that the cells preferentially attach to the holey structure. Thereby, the cells form bridges inside, leading to a complete covering of the hole. In this way, a more effective biological barrier against bacteria can be created.