Safety Assessment of the Modified Lactoperoxidase System—In Vitro Studies on Human Gingival Fibroblasts

Abstract

One strategy in caries prevention is to inhibit the formation of cariogenic biofilms. Attempts are being made to develop oral hygiene products enriched with various antimicrobial agents. One of them is lactoperoxidase—an enzyme that can oxidise (pseudo)halide ions to reactive products with antimicrobial activity. Currently, commercially available products utilise thiocyanate as a substrate; however, several alternatives that are oxidised to products with greater antimicrobial potential have been found. In this study, toxicity against human gingival fibroblasts of the lactoperoxidase system was evaluated using four different (pseudo)halide substrate systems—thiocyanate, iodide, selenocyanate, and a mixture of thiocyanate and iodide. For this purpose, cells were treated with the systems and then apoptosis, cell cycle, intracellular glutathione concentration, and mitochondrial superoxide production were assessed. The results showed that each system, after generating 250 µM of the product, inhibited cell divisions, increased apoptosis, and increased the percentage of dead cells. It was concluded that the mechanism of the observed phenomena was not related to increased superoxide production or the depletion of glutathione concentration. These findings emphasised the need for the further in vitro and in vivo toxicity investigation of the modified lactoperoxidase system to assess its safety and the possibility of use in oral hygiene products.

Modified Lactoperoxidase System as a Promising Anticaries Agent: In Vitro Studies on Streptococcus mutans Biofilms

“…Finally, a necessary step will be to determine the lowest concentration of LPO system components that exhibit significant in vitro and in vivo antibiofilm effects while remaining nontoxic to host cells. In our previous studies on the toxicity of the described modifications in an LPO system against human gingival fibroblasts, we found that the LPO-I − system was characterised by the highest toxicity and the physiological system (LPO-SCN − ) by the lowest toxicity [29]. The results showed that modifying the LPO system enhances both its antibiofilm activity and its negative effects on host cells.…”

“…In the presented research, bovine milk LPO was used due to its high level of similarity (structural and functional) to the human LPO molecule [13]. A buffered solution of 50 nmol/L LPO was used throughout the study that reflected the physiological enzyme concentration found in saliva [29]. All LPO reactions were performed in phosphate buffered solution (PBS) at pH 7.4, which reflected the environmental conditions of the oral cavity of a healthy person between meals.…”

“…In the available studies, two approaches to exposing microorganisms to the LPO system can be found [29,30]. The first consists of creating a mixture of the substrates and the enzyme and triggering the reaction; then, after it reaches completion, the resulting mixture containing the products is added to the tested microorganisms.…”

“…The first consists of creating a mixture of the substrates and the enzyme and triggering the reaction; then, after it reaches completion, the resulting mixture containing the products is added to the tested microorganisms. The second approach is based on running the reaction in situ, already in the presence of tested microorganisms [29].…”