Abstract.Various amino acids regulate cell growth and differentiation. In the present study, we examined the ability of HT-29 cells to differentiate into goblet cells in RPMI and DMEM which are largely different in the amounts of numerous amino acids. Most of the HT-29 cells differentiated into goblet cells downregulating the stem cell marker Lgr5 when cultured in DMEM, but remained undifferentiated in RPMI. The goblet cell differentiation in DMEM was inhibited by 1-methyl-tryptophan (1-MT), an inhibitor of indoleamine 2,3 dioxygenase-1 which is the initial enzyme in tryptophan metabolism along the kynurenine (KN) pathway, whereas tryptophan and KN induced goblet cell differentiation in RPMI. The levels of Notch1 and its activation product Notch intracytoplasmic domain in HT-29 cells were lower in DMEM than those in RPMI and were increased by 1-MT in both media. HT-29 cells grown in both media expressed β-catenin at the same level on day 2 when goblet cell differentiation was not observed. β-catenin expression, which was increased by 1-MT in both media, was decreased by KN. DMEM reduced Hes1 expression while enhancing Hath1 expression. Finally, aryl hydrocarbon receptor (AhR) activation moderately induced goblet cell differentiation. Our results suggest that KN promotes goblet cell differentiation by regulating Wnt, Notch, and AhR signals and expression of Hes1 and Hath1. IntroductionCell lines are usually maintained in defined media that commonly contain amino acids, vitamins, glucose and inorganic salts for normal cell metabolism. However, the composition of medium formulations vary widely in concentrations of glucose and other metabolic precursors, including amino acids. Thus, many types of cells showed different responses to culture media in regards to proliferation and differentiation. Th17 differentiation was induced more efficiently in Iscove's modified Dulbecco's medium (IMDM) than in RPMI (1). Bone marrow-derived dendritic cells in IMDM expressed higher levels of co-stimulatory and MHC II molecules, compared to the cells generated in RPMI (2). In addition, hepatocyte differentiation and propagation and phenotype of corneal stroma-derived stem cells were modulated by culture media (3,4). Subsequently, the underlying mechanisms of the medium-induced changes in cell proliferation and differentiation have been partially elucidated. Differentiation of Th17 cells was increased by endogenous aryl hydrocarbon receptor (AhR) agonists derived from aromatic amino acids such as tryptophan, which is underrepresented in RPMI compared with IMDM (1). Mesenchymal stem cells co-cultured with AML12 liver cells were able to differentiate into hepatocytelike cells expressing hepatocyte-specific markers including albumin, α-fetoprotein and cytokeratin 18 mRNAs, which were expressed at the initial 7-day culture in DMEM while being expressed during the 7-to 14-day culture in DMEM/ F12 (3). In a study using MDA-MB-231 breast cancer cells, ~25.6% of genes were expressed at significantly different levels in cells grown in MEM, DM...

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Aryl hydrocarbon receptor (AhR) regulates both innate and adaptive immune responses by sensing a variety of small synthetic and natural chemicals, which act as its ligands. AhR, which is expressed in dendritic cells (DCs), regulates the differentiation of DCs. However, effects of AhR on the differentiation of DCs are variable due to the heterogeneity of DCs in cell surface marker expression, anatomical location, and functional responses. The plasmacytoid DCs (pDCs), one of DC subsets, not only induce innate as well as adaptive immune responses by secreting type I interferons and pro-inflammatory cytokines, but also induce IL-10 producing regulatory T cell or anergy or deletion of antigen-specific T cells. We showed here that AhR ligands indoxyl 3-sulfate (I3S) and indole-3-carbinol (I3C) inhibited the development of pDCs derived from bone marrow (BM) precursors induced by FMS-like tyrosine kinase 3 ligand (Flt3L). I3S and I3C downregulated the expression of signal transducer and activator of transcription 3 (STAT3) and E2-2 (Tcf4). In mice orally treated with I3S and I3C, oral tolerance to dinitrofluorobenzene was impaired and the proportion of CD11c+B220+ cells in mesenteric lymph nodes was reduced. These data demonstrate that AhR negatively regulates the development of pDCs from BM precursors induced by Flt3L, probably via repressing the expression of STAT3.