Real-time monitoring of amyloid fibrillation by electrical impedance spectroscopy.

Electrical impedance spectroscopy (EIS) appears a promising label-free methodology for the investigation of processes related to the aggregation of macromolecules in solution. Here, we explore the EIS technique as a convenient tool for studying the irreversible aggregation of human insulin and describing its corresponding fibrillation kinetics. The in situ measurement of the electrical response of pure insulin solutions at 60°C allows for the real-time monitoring of the protein fibrillation as a function of the incubation time. The fitting of the EIS data through an equivalent circuit based on a constant phase element provides a simple set of electric parameters whose abrupt changes can be associated to transitions occurring in the organization of the macromolecules. For establishing the reliability of the method proposed, we have compared the protein aggregation profile collected from the EIS data to that obtained from a conventional fluorescence methodology where Thioflavin T (ThT) is used as a dye probe. The description of the fibrillation process is quite similar in both cases, since characteristic times of the same order were found for the consecutive processes associated to the initial lag phase of insulin fibrillation, to the rapid growth of amyloidal aggregates and to the final saturation step. Our results suggest that in situ EIS can be considered as a promising approach for the real-time label-free monitoring of protein fibril formation.

Infliximab attenuates inflammatory osteolysis in a model of periodontitis in Wistar rats.

Periodontitis is a chronic inflammatory disease related to tooth loss in adults. Infliximab is a chimeric monoclonal antibody against TNF-α and is prescribed for the treatment of systemic inflammatory diseases. This study aimed to investigate the role of infliximab on experimental periodontal disease (EPD). EPD was induced by passing a 3.0 nylon thread around the upper left second molar in Wistar rats. Animals were either treated with intravenous infliximab (1, 5, 7, and 10 mg/kg) or saline solution 30 min before the periodontitis induction and were followed until they were sacrificed on the 11th day. A subset of rats was euthanized on the third day for analysis of gingival myeloperoxidase (MPO) and the blood MPO granulocyte index. In addition, we analyzed the bone loss index (BLI), the periodontal histopathological score, and the periodontal collagen network using confocal microscopy. We also analyzed metalloproteinase-1/-8, RANK, RANK-L, and osteoprotegerin in maxillary tissue by immunohistochemistry Gingival MPO, IL-1ß, TNF-α were measured by ELISA. EPD caused leukocytosis, significant increases in BLI and gingival pro-inflammatory cytokines and cell infiltrates, with worse histopathological scores and periodontal collagen derangement. Infliximab (5 mg/kg) reduced granulocyte blood counts, gingival IL-1ß, TNF-α, and MPO levels, diminished MMP-1/-8, RANK, and RANK-L bone immunolabeling with better periodontal histopathological scores and collagen network in comparison with the challenged saline group. We concluded that infliximab had significant anti-inflammatory and bone-protective effects in Wistar rats challenged by periodontitis.