(PTFS22-03-24) Dubosiella newyorkensis Modulates Tryptophan and Serotonin Production and Related Signaling Pathways in the Enteroendocrine Cell Line STC1

Objectives: The newly identified gut microbial species D. newyorkensis has been shown to exhibit probiotic functions, possibly through anti-inflammatory pathways. We observed gut dysbiosis in murine models of colitis and colon inflammation, including the heterologous nuclear ribonucleoprotein (hnRNPI) knockout (KO) mouse model. Single-cell sequencing data revealed an upregulation of the tryptophan hydroxylase1 (Tph1) gene, a limiting key enzyme in the serotonin production from tryptophan. The increased Tph1 correlates with the elevated serotonin levels in the KO mice colons. Fecal gut microbiota analysis through amplicon sequencing of the 16S rRNA gene uncovered a significant decrease in the abundance of D. newyorkensis in the KO mice. Therefore, we aimed to investigate the role of this bacteria and its metabolites in modulating Tph1 gene expression using an in vitro model.

Methods: D. newyorkensis was cultured in the mTGE media for 24h and supernatant was collected. STC1 cells were seeded onto 60-mm cell culture dishes. The cells were exposed to D. newyorkensis metabolites for 24h. RNA from the cells was isolated, and gene expression of the key factors in the serotonin pathway, including Tph1, was quantified using quantitative PCR (qPCR) analysis, western blot analysis, and immunohistochemical analysis.

Results: Various concentrations of bacterial metabolites were used to treat STC1 cells. STC1 cells treated with D. newyorkensis metabolites at 10% exhibited a decrease in Tph1 gene expression levels when compared to cells treated with mTGE (bacterial media). Consistent with these gene expression findings, a concurrent reduction in serotonin protein expression was noted in cells treated with bacterial metabolites as opposed to those treated with mTGE.

Conclusions: Our experimental findings indicate that metabolites derived from gut microbiota can engage in crosstalk with EECs. Our data shows promise for establishing the potential of D. newyorkensis as a probiotic capable of regulating EECs and potentially reducing serotonin levels in gastrointestinal disorders such as inflammatory bowel diseases and colorectal cancer.

Funding Sources: USDA Cooperative State Research, Education and Extension Service (Hatch project numbers # ILLU-971351 and ILLU-698923), and the Office of the Vice Chancellor for Research in the University of Illinois at Urbana-Champaign.