Valutazione in vitro del profilo tossicologico dei più rappresentativi composti dicarbonilici e relativi prodotti finali di glicazione avanzata correlati alla dieta

The Maillard reaction generate innumerable compounds that affect the organoleptic and toxicological properties of foods, in a non-enzymatic way. The products of this reaction include the dicarbonyl compounds and the advanced glycation end products (AGEs), derived from the first ones. This heterogeneous group of molecules may differently contribute to the diet-related exacerbation of oxidative stress, inflammation and protein modifications[1]. Previous in vitro toxicity studies have been carried out by treatment with a mixture of AGEs or by inducing their formation through the administration of reducing sugars at different temperatures. Due to the heterogenicity of these compounds, it is difficult to understand the contribution of each one[2]. Precisely for these reasons, in this study, through the use of the MTT test, ROS detection probes and Western Blot analysis, for the first time, AGEs and dicarbonyl compounds were tested individually on a cell line of human keratinocytes (HaCaT) evaluating the minimal toxic dose as well as the related molecular pathways involved. Among the compounds tested only dicarbonyl 3-deoxygalactosone (3-DGal), 3,4-dideoxyglucosone-3-ene (3,4-DGE), glyoxal (GO) and methylglyoxal (MGO) showed a significant decrease in cell viability. 3,4-DGE and 3-DGal, in particular, caused an increase in the intracellular ROS level. The antioxidant N-acetylcysteine (NAC) modified the effects of dicarbonyl compounds on cell viability. We discovered, thanks to the use of some cell death inhibitors, that the type of death triggered by 3-DGal and 3,4-DGE is not autophagy, ferroptosis or necroptosis, but still ROS dependent. We have found that the administration of 3-DGal and 3,4-DGE evoked the activation of the NF-κB inflammatory pathway in HaCaT cells, thus confirming the mechanisms of crosstalk between oxidative stress and inflammatory responses underlying their cellular toxicity. Furthermore, in the same treatment groups, the active form of GSDMD was significantly higher than in the control group as a consequence of the increased expression of NLRP3 inflammasome. The N-terminal residue of the GSDMD is responsible for the formation of pores in the membrane, causing the triggering of pyroptotic cell death therefore so, it is possible hypothesized that 3,4-DGE and 3-DGal can induce keratinocyte death with a pyroptotic mechanism. These results demonstrate the importance of testing compounds individually in fact, 3-DGal, but not its epimer 3-deoxyglucosone (3-DG), is toxic to human keratinocytes and exhibits the same toxicity mechanism as its dehydration product, 3,4-DGE (albeit at a concentration 10 times higher). In the future it will therefore be necessary to investigate which AGEs and their precursors are toxic to human health, in which concentrations and through which signaling pathways in order to develop precise and minimally invasive analytical techniques capable of predicting the onset and evolution of related pathologies.