## 4 Discussion
### AhR Transcriptional Activation Representing Host-Microbe Interaction  
Host-microbe interaction in the human gastrointestinal tract is a complex dynamic involving multiple pathways. Among these is the activation of xenobiotic sensors such as AhR. This receptor processes signals from endogenous and exogenous chemicals that trigger important molecular pathways through the induction of its transcriptional activity [125]. AhR signaling has been associated with the regulation at different levels of the immune response of the host organism, and it is considered a key element in the maintenance of homeostasis in the gut microbial ecosystem of mammalians [126; 127]. The human colorectal adenocarcinoma HT29 is a cell model successfully used in the study of host-microbiome interactions, as it can express characteristics of mature intestinal cells [128]. There are several advantages of HT29 compared to the Caco-2 cell line (also derived from human colorectal adenocarcinoma) when studying the effects on the gut mucus layer [128; 129]. HT29 culture models have been used, for example, to evaluate the impact of colonic fermentation of prebiotic flours (e.g., rye, lentils, wheat) [130], and the protective effects of probiotics (e.g., _Streptococcus thermophilus_, _Lactobacillus acidophilus_) [131]. In this study, the HT29-Lucia cell line was utilized to identify the potential activation of AhR transactivity caused by microbial-derived metabolites in the colonic environment. However, several gut microbial-derived metabolites could reach the liver or be incorporated into the systemic circulation, modulating health and disease through AhR [10; 132]. Therefore, the potential activation of AhR transcriptional activity was also evaluated outside the intestinal context through HepG2 and T47D human cell models (Figure 5). The results obtained revealed that the metabolic output of the healthy human microbiota fermentation is able to induce AhR transactivity in intestinal cells (HT29) two-fold times higher than the well-known AhR agonist TCDD (Figure 5). Meanwhile, T47D cells were even more sensitive to the fermentation supernatant, with an AhR activation above 20-fold response by the second and third day of fermentation. The induced activation of AhR transcriptional machinery in the intestine is probably caused by a synergistic effect of several metabolites. For example, indole derivatives (e.g., Indolo[3,2-b]carbazole (ICZ), 3,3\({}^{\prime}\)-dimidolymet methane), arachidonic acid metabolites (e.g., Lipoxin 4A, Prostaglandin PGG2), heme-derived metabolites (e.g., bilirubin, biliverdin), and tryptophan metabolites (e.g., kynure, tryptamine, 6-formylindol[3,2-b]carbazole (FICZ)) are known endogenous ligands of the AhR derived from diet, photodoxidation, host or microbiota metabolisms, indistinctly [30]. Mechanistic studies have shown how the AhR transcriptional activity induced by tryptophan metabolites is crucial to maintaining the fragile equilibrium between the microbiota and the host cells [133]. Thus, tryptophan catabolites derived from gut microbiota are dysbiosis biomarkers, and their production is a commonly suggested pharmacological target [14,15,82,134]. Tryptamine, as previously mentioned, was stably produced during the 72-h experiment, and it is a long-known AhR agonist [135]. The first studies of tryptamine-induced activation of AhR suggested that effective tryptamine concentrations were between 80-100 \(\upmu\)M [123]. However, it has been later revealed in cell models that tryptamine can induce CYP1A1 gene expression even at very low concentrations (\(\sim\)5 \(\upmu\)M) [136]. Therefore, the concentrations of tryptamine detected in the human microbiota herein modeled (10-30 \(\upmu\)M) probably contribute to the AhR activation displayed by the metabolic output (Figure 5). Additionally, SCFAs are probably major contributors to the AhR-induced activity, since they have all proven to be AhR modulators.