Hydrogen peroxide is an oxidative agent commonly used for dental bleaching procedures. The structural and biochemical responses of enamel, dentin, and pulp tissues to the in vivo bleaching of human (n = 20) premolars were investigated in this study. Atomic force microscopy (AFM) was used to observe enamel nanostructure. The chemical composition of enamel and dentin was analyzed by infrared spectroscopy (FTIR). The enzymatic activities of dental cathepsin B and matrix metalloproteinases (MMPs) were monitored with fluorogenic substrates. The amount of collagen in dentin was measured by emission of collagen autofluorescence with confocal fluorescence microscopy. The presence of Reactive Oxygen Species (ROS) in the pulp was evaluated with a fluorogenic 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) probe. Vital bleaching of teeth significantly altered all tested parameters: AFM images revealed a corrosion of surface enamel nanostructure; FTIR analysis showed a loss of carbonate and proteins from enamel and dentin, along with an increase in the proteolytic activity of cathepsin-B and MMPs; and there was a reduction in the autofluorescence of collagen and an increase in both cathepsin-B activity and ROS in pulp tissues. Together, these results indicate that 35% hydrogen peroxide used in clinical bleaching protocols dramatically alters the structural and biochemical properties of dental hard and soft pulp tissue.

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c Paracoccidioidomycosis (PCM), caused by Paracoccidioides species, is the main cause of death due to systemic mycoses in Brazil and other Latin American countries. Therapeutic options for PCM and other systemic mycoses are limited and time-consuming, and there are high rates of noncompliance, relapses, toxic side effects, and sequelae. Previous work has shown that the cyclopalladated 7a compound is effective in treating several kinds of cancer and parasitic Chagas disease without significant toxicity in animals. Here we show that cyclopalladated 7a inhibited the in vitro growth of Paracoccidioides lutzii Pb01 and P. brasiliensis isolates Pb18 (highly virulent), Pb2, Pb3, and Pb4 (less virulent) in a dose-response manner. Pb18 was the most resistant. Opportunistic Candida albicans and Cryptococcus neoformans were also sensitive. BALB/c mice showed significantly lighter lung fungal burdens when treated twice a day for 20 days with a low cyclopalladated 7a dose of 30 g/ml/day for 30 days after intratracheal infection with Pb18. Electron microscopy images suggested that apoptosis-and autophagy-like mechanisms are involved in the fungal killing mechanism of cyclopalladated 7a. Pb18 yeast cells incubated with the 7a compound showed remarkable chromatin condensation, DNA degradation, superoxide anion production, and increased metacaspase activity suggestive of apoptosis. Autophagy-related killing mechanisms were suggested by increased autophagic vacuole numbers and acidification, as indicated by an increase in LysoTracker and monodansylcadaverine (MDC) staining in cyclopalladated 7a-treated Pb18 yeast cells. Considering that cyclopalladated 7a is highly tolerated in vivo and affects yeast fungal growth through general apoptosisand autophagy-like mechanisms, it is a novel promising drug for the treatment of PCM and other mycoses.