Abstract
This study intended to evaluate the effects of a papain-gel with a red-light absorbing pigment (methylene blue – MB) to mediate photodynamic therapy (PDT) against Streptococcus mutans biofilms. The PapaMBlue was compared with free MB to generate reactive oxygen species using fluorescence probes (SOSG and HPF). PDT (660-nm light) was carried out against S. mutans biofilms grown on either plastic dishes or on collagen membrane and assayed by CFU, live-dead staining using confocal microscopy, transmission electron microscopy and H&E staining for collagen films. Cytotoxicity and subcellular localization was studied in human fibroblasts. Sponges of bioabsorbable type I collagen membrane were exposed to papain based gel, irradiated with laser and analyzed about their integrity by ATR-FTIR. The PapaMBlue produced higher amounts of singlet oxygen and hydroxyl radicals than free MB, possibly due to better disaggregation of the dye in solution. The PapaMBlue antimicrobial effects on biofilms proved to be capable of reducing the S. mutans. Both MTT and PrestoBlue assays showed higher cell viability and metabolism scores in fibroblasts treated with PapaMBlue and MB, possibly due to stimulation of mitochondrial activity and that collagen triple helix is unaffected. The PapaMBlue is equally effective as MB in destroying S. mutans biofilms growing on plastic or collagen without affecting fibroblasts.
Highlights
Currently considered the treatment of choice for deep carious lesions provided that certain treatment principles are observed[4]
This change was performed by adding a photosensitizer that could be activated by light in photodynamic therapy (PDT) and the interaction between the gel and the light resource, regarding that hypothesis to potentiate the antibacterial effect of Papain gel added the methylene blue (Fig. 1)
The present study is an important first step to launch a new product proving that the addition of methylene blue to Papacarie is reliable because: (1) it it does not change the absorption spectra of Papacárie (Fig. 2); (2) it preserves the biocompatibility of the gel (Fig. 4); (3) the PapaMBlue can be internalized by the cells and be identified in the mitochondria and lysosomes (Fig. 5); (4) the gel with Methylene blue (MB) is able to produce reactive oxygen species (ROS) (Fig. 3); (5) PapaMBlue is capable of reducing the S. mutans CFU of biofilm on plastic and (6) in caries lesion inducted by bacteria biofilm (Fig. 10) and (7) does not degrade collagen (Figs 8, 9 and 11)
Summary
Currently considered the treatment of choice for deep carious lesions provided that certain treatment principles are observed[4]. A controversy point of this gel is whether it reduces the bacteria colonies present in the dental tubules of affected dentin Those bacteria products are harmful to the pulp, mainly in deep cavities, where the remaining thin layer of dentin is less rigid, and if removed excessively there is a chance of a pulpal exposition. A change in Papacárie composition was suggested in order to improve its properties and applicability, so it might be used as an agent for removal the necrotic dentin tissue and as an antimicrobial agent This change was performed by adding a photosensitizer that could be activated by light in photodynamic therapy (PDT) and the interaction between the gel and the light resource, regarding that hypothesis to potentiate the antibacterial effect of Papain gel added the methylene blue (Fig. 1). Recent studies demonstrated that aPDT is an encouraging adjunctive therapy to the treatment of caries lesions, mainly in the minimally invasive approach to preserve dental tissue and favor its repair[16,17]
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