The role of endothelin B receptor in bone modelling during orthodontic tooth movement: a study on ETB knockout rats.

The endothelin system has an important role in bone modelling during orthodontic tooth movement (OTM); however, little is known about the involvement of endothelin B receptors (ETB) in this process. The aim of this study was to evaluate the role of ETB in bone modelling during OTM using ETB knockout rats (ETB-KO). Thirty-two male rats were divided into 4 groups (n = 8 per group): the ETB-KO appliance group, ETB-KO control group, wild type (ETB-WT) appliance group, and ETB-WT control group. The appliance consisted of a super-elastic closed-coil spring placed between the first and second left maxillary molar and the incisors. Tooth movement was measured on days 0 and 35, and maxillary alveolar bone volume, osteoblast, and osteoclast volume were determined histomorphometrically on day 35 of OTM. Next, we determined the serum endothelin 1 (ET-1) level and gene expression levels of the osteoclast activity marker cathepsin K and osteoblast activity markers osteocalcin and dentin matrix acidic phosphoprotein 1 (DMP1) on day 35. The ETB-KO appliance group showed significantly lower osteoblast activity, diminished alveolar bone volume and less OTM than the ETB-WT appliance group. Our results showed that ETB is involved in bone modelling in the late stage of OTM.

Magnetic interactions and in vitro study of biocompatible hydrocaffeic acid-stabilized Fe-Pt clusters as MRI contrast agents.

A detailed magnetic study of separated Fe-Pt NPs and Fe-Pt clusters was performed to predict their optimal size and morphology for the maximum saturation magnetization, a factor that is known to influence the performance of a magnetic-resonance-imaging (MRI) contrast agent. Excellent stability and biocompatibility of the nanoparticle suspension was achieved using a novel coating based on hydrocaffeic acid (HCA), which was confirmed with a detailed Fourier-transform infrared spectroscopy (FTIR) study. An in vitro study on a human-bladder papillary urothelial neoplasm RT4 cell line confirmed that HCA-Fe-Pt nanoparticles showed no cytotoxicity, even at a very high concentration (550 µg Fe-Pt per mL), with no delayed cytotoxic effect being detected. This indicates that the HCA coating provides excellent biocompatibility of the nanoparticles, which is a prerequisite for the material to be used as a safe contrast agent for MRI. The cellular uptake and internalization mechanism were studied using ICP-MS and TEM analyses. Furthermore, it was shown that even a very low concentration of Fe-Pt nanoparticles (<10 µg mL-1) in the cells is enough to decrease the T 2 relaxation times by 70%. In terms of the MRI imaging, this means a large improvement in the contrast, even at a low nanoparticle concentration and an easier visualization of the tissues containing nanoparticles, proving that HCA-coated Fe-Pt nanoparticles have the potential to be used as an efficient and safe MRI contrast agent.

Hybrid FePt/SiO2/Au nanoparticles as a theranostic tool: in vitro photo-thermal treatment and MRI imaging.

We have produced an innovative, theranostic material based on FePt/SiO2/Au hybrid nanoparticles (NPs) for both, photo-thermal therapy and magnetic resonance imaging (MRI). Furthermore, a new synthesis approach, i.e., Au double seeding, for the preparation of Au nanoshells around the FePt/SiO2 cores, is proposed. The photo-thermal and the MRI response were first demonstrated on an aqueous suspension of hybrid FePt/SiO2/Au NPs. The cytotoxicity together with the internalization mechanism and the intracellular fate of the hybrid NPs were evaluated in vitro on a normal (NPU) and a half-differentiated cancerous cell line (RT4). The control samples as well as the normal cell line incubated with the NPs showed no significant temperature increase during the in vitro photo-thermal treatment (ΔT < 0.8 °C) and thus the cell viability remained high (∼90%). In contrast, due to the high NP uptake by the cancerous RT4 cell line, significant heating of the sample was observed (ΔT = 4 °C) and, consequently, after laser irradiation the cell viability dropped significantly to ∼60%. These results further confirm that the hybrid FePt/SiO2/Au NPs developed in the scope of this work were not only efficient but also highly selective photo-thermal agents. Furthermore, the improvement in the contrast and the easier distinction between the healthy and the cancerous tissues were clearly demonstrated with in vitro MRI experiments, proving that hybrid NPs have an excellent potential to be used as contrast agents.

Establishment and characterization of primary and subsequent subcultures of normal mouse urothelial cells.

In this study, we report a reliable technique for the harvest, cultivation and expansion of monoculture of NMU. The NMU were harvested by two methods, directly from the urothelium in vivo and indirectly from the urothelial outgrowths of bladder explant cultures. Primary cultures and subsequent subcultures were propagated in the mixture of media MCDB 153 and Advanced-DMEM, and conditioned medium. Primary urothelial cells required an initial plating density of 1 x 10(5) viable cells/cm2 for survival, while passaged cells needed lower plating densities (1 x 10(4) viable cells per cm2). The cultured cells were identified as urothelial by their epithelioid morphology and by the positive immunofluorescence labelling of tight junctional proteins, occludin and ZO-1, adherens protein E-cadherin and cytoskeletal protein cytokeratin 7. Markers of highly differentiated urothelial cells, cytokeratin 20 and uroplakins, were not expressed. Furthermore, the immunofluorescence labelling of occludin and cytokeratin 7 was not detected in later passages when urothelial cells replicated at a high rate. In spite of the use of conditioned medium derived from V79 fibroblast cell culture supernatant, the NMU in the primary cultures and subsequent subcultures expressed a basal/intermediate cell phenotype. In conclusion, we demonstrate that homogeneous long-term culture of NMU can be developed. Since powerful transgenic tools exist to manipulate the mouse genome, our findings should help design the mouse in vitro systems for studying the control mechanisms of urothelial cell proliferation, stratification and differentiation in health and disease.