Serotonin Modulates AhR Activation by Interfering with CYP1A1-Mediated Clearance of AhR Ligands.
Serotonin Modulates AhR Activation by Interfering with CYP1A1-Mediated Clearance of AhR Ligands.
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DOI：
10.33594/000000209
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发表时间：
2020-02-05
期刊：
影响因子：
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通讯作者：
Gill, Ravinder K
中科院分区：
文献类型：
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作者：
Manzella, Christopher R;Ackerman, Max;Gill, Ravinder K
背景/目标：5-羟色胺（5-HT）是一种神经递质和激素，在包括肠道在内的许多器官中具有重要的生理功能。我们以前已经表明，5-HT激活芳烃受体（AhR）在肠上皮细胞（IEC）通过5-羟色胺转运蛋白（SERT）依赖性机制。AhR是一种核受体，其结合多种分子，包括色氨酸（TRP）代谢物，以调节肠中的生理过程，包括异生物质解毒和免疫调节。我们假设5-HT通过干扰细胞色素P450 1A 1（CYP 1A 1）对AhR配体的代谢清除而间接激活AhR。使用荧光素酶和LC-MS/MS在人肠上皮细胞系Caco-2和重组CYP 1A 1微粒体中评估5-HT对CYP 1A 1活性的抑制。对于体外研究，通过RT-PCR测定CYP 1A 1和CYP 1B 1 mRNA表达水平，通过乙氧基试卤灵-O-脱乙基酶（EROD）测定测定CYP 1A 1活性。在体内研究中，AhR配体给予SERT KO小鼠和WT同窝仔和肠粘膜CYP 1A 1 mRNA的measured.RESULTS：我们发现，5-HT抑制代谢的两个前体在CYP 1A 1底物Luc-CEE以及高亲和力AhR配体6-甲酰吲哚[3，2-B]咔唑（FICZ）。重组CYP 1A 1试验表明，5-HT是由CYP 1A 1代谢的NADPH依赖性方式。在不含微量AhR配体的无TRP培养基中用5-HT处理表明，5-HT需要AhR配体的存在来激活AhR。用5-HT和FICZ共处理证实，5-HT增强AhR配体对AhR靶基因的诱导。然而，这仅适用于作为CYP 1A 1底物的配体，如FICZ。通过灌胃或吲哚-3-甲醇通过饮食SERT KO小鼠的管理表明，缺乏SERT损害肠道AhR activation.CONCLUSION：我们的研究提供了新的证据，串扰之间的α-羟色胺和AhR信号传导，5-HT可以影响AhR配体激活受体在肠道中的能力。
BACKGROUND/AIMS: Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter and hormone with important physiological functions in many organs, including the intestine. We have previously shown that 5-HT activates the aryl hydrocarbon receptor (AhR) in intestinal epithelial cells (IECs) via a serotonin transporter (SERT)-dependent mechanism. AhR is a nuclear receptor that binds a variety of molecules including tryptophan (TRP) metabolites to regulate physiological processes in the intestine including xenobiotic detoxification and immune modulation. We hypothesized that 5-HT activates AhR indirectly by interfering with metabolic clearance of AhR ligands by cytochrome P450 1A1 (CYP1A1).METHODS: Inhibition of CYP1A1 activity by 5-HT was assessed in the human intestinal epithelial cell line Caco-2 and recombinant CYP1A1 microsomes using both luciferase and LC-MS/MS. Degradation of 5-HT by recombinant CYP1A1 was measured by LC-MS/MS. For in vitro studies, CYP1A1 and CYP1B1 mRNA expression levels were measured by RT-PCR and CYP1A1 activity was measured by ethoxyresorufin-O-deethylase (EROD) assays. For in vivo studies, AhR ligands were administered to SERT KO mice and WT littermates and intestinal mucosa CYP1A1 mRNA was measured.RESULTS: We show that 5-HT inhibits metabolism of both the pro-luciferin CYP1A1 substrate Luc-CEE as well as the high affinity AhR ligand 6-formylindolo[3,2-b] carbazole (FICZ). Recombinant CYP1A1 assays revealed that 5-HT is metabolized by CYP1A1 in an NADPH dependent manner. Treatment with 5-HT in TRP-free medium, which is devoid of trace AhR ligands, showed that 5-HT requires the presence of AhR ligands to activate AhR. Cotreatment with 5-HT and FICZ confirmed that 5-HT potentiates induction of AhR target genes by AhR ligands. However, this was only true for ligands which are CYP1A1 substrates such as FICZ. Administration of beta-napthoflavone by gavage or indole-3-carbinol via diet to SERT KO mice revealed that lack of SERT impairs intestinal AhR activation.CONCLUSION: Our studies provide novel evidence of crosstalk between serotonergic and AhR signaling where 5-HT can influence the ability of AhR ligands to activate the receptor in the intestine.