{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002837","ANALYSIS_ID":"AN004638","VERSION":"1","CREATED_ON":"August 22, 2023, 12:07 pm"},

"PROJECT":{"PROJECT_TITLE":"Bacterial tryptophan metabolites increased by prebiotic galactooligosaccharide reduce microglial reactivity and are associated with lower anxiety-like behavior","PROJECT_SUMMARY":"Prebiotic galactooligosaccharides (GOS) reduce anxiety-like behaviors in mice and humans. However, the biological pathways behind these behavioral changes are not well understood. To begin to study these pathways, C57BL/6 mice were fed a standard diet with or without GOS supplementation for 3 weeks prior to testing on the open field. After behavioral testing, colonic contents and serum were collected for bacteriome (16S rRNA gene sequencing, colonic contents only) and metabolome (UPLC-MS, colonic contents and serum data) analyses. As expected, GOS significantly reduced anxiety-like behavior (i.e., increased time in the center (p < 0.05)). Time in the center was significantly correlated with serum methyl-indole-3-acetate (mI3A). This metabolite, which is a methylated form of indole-3-acetate, is derived from bacterial metabolism of tryptophan. Sequencing analyses showed that GOS significantly increased Akkermansia, which is known to metabolize both GOS and tryptophan. To test the hypothesis that mI3A can reduce anxiety-like behavior and affect microglial activity, we first tested mI3A effects on LPS-stimulated BV2 microglia. Cells treated with mI3A produced significantly less CCL2 and TNF-α than vehicle-treated cells (p<.05). We then treated mice with an intraperitoneal injection of mI3A or vehicle control, and found that mice given mI3A had lower CCL2 in the prefrontal cortex and hippocampus, as well as a reduction in a composite behavioral score in the open field. Together, these data support a novel pathway through which GOS reduces anxiety-like behaviors in mice, and suggests that the bacterial metabolite mI3A, which is elevated by GOS, reduces microglial CCL2, which in turn reduces anxiety-like behavior.","INSTITUTE":"National Center for Advancing Translational Sciences","LAST_NAME":"Spencer","FIRST_NAME":"Kyle","ADDRESS":"9800 Medical Center Dr, Rockville, MD 20850","EMAIL":"kyle.spencer@nih.gov","PHONE":"9897084858"},

"STUDY":{"STUDY_TITLE":"Bacterial tryptophan metabolites increased by prebiotic galactooligosaccharide reduce microglial reactivity and are associated with lower anxiety-like behavior (Blood)","STUDY_SUMMARY":"Prebiotic galactooligosaccharides (GOS) reduce anxiety-like behaviors in mice and humans. However, the biological pathways behind these behavioral changes are not well understood. To begin to study these pathways, C57BL/6 mice were fed a standard diet with or without GOS supplementation for 3 weeks prior to testing on the open field. After behavioral testing, colonic contents and serum were collected for bacteriome (16S rRNA gene sequencing, colonic contents only) and metabolome (UPLC-MS, colonic contents and serum data) analyses. As expected, GOS significantly reduced anxiety-like behavior (i.e., increased time in the center (p < 0.05)). Time in the center was significantly correlated with serum methyl-indole-3-acetate (mI3A). This metabolite, which is a methylated form of indole-3-acetate, is derived from bacterial metabolism of tryptophan. Sequencing analyses showed that GOS significantly increased Akkermansia, which is known to metabolize both GOS and tryptophan. To test the hypothesis that mI3A can reduce anxiety-like behavior and affect microglial activity, we first tested mI3A effects on LPS-stimulated BV2 microglia. Cells treated with mI3A produced significantly less CCL2 and TNF-α than vehicle-treated cells (p<.05). We then treated mice with an intraperitoneal injection of mI3A or vehicle control, and found that mice given mI3A had lower CCL2 in the prefrontal cortex and hippocampus, as well as a reduction in a composite behavioral score in the open field. Together, these data support a novel pathway through which GOS reduces anxiety-like behaviors in mice, and suggests that the bacterial metabolite mI3A, which is elevated by GOS, reduces microglial CCL2, which in turn reduces anxiety-like behavior.","INSTITUTE":"National Center for Advancing Translational Sciences","LAST_NAME":"Spencer","FIRST_NAME":"Kyle","ADDRESS":"9800 Medical Center Dr, Rockville, MD 20850","EMAIL":"kyle.spencer@nih.gov","PHONE":"9897084858"},

"SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"NACH-00271",
"Factors":{"GROUP_ID":"GOS"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00272",
"Factors":{"GROUP_ID":"GOS"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00273",
"Factors":{"GROUP_ID":"GOS"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00274",
"Factors":{"GROUP_ID":"GOS"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00275",
"Factors":{"GROUP_ID":"GOS"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00276",
"Factors":{"GROUP_ID":"CON"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00277",
"Factors":{"GROUP_ID":"CON"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00278",
"Factors":{"GROUP_ID":"CON"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00279",
"Factors":{"GROUP_ID":"CON"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00280",
"Factors":{"GROUP_ID":"CON"}
},
{
"Subject ID":"-",
"Sample ID":"NACH-00281",
"Factors":{"GROUP_ID":"CON"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"After euthanasia the serum of the mice were collected and sent to Metabolon Inc for UPLC-MS Analysis.","SAMPLE_TYPE":"Blood (serum)"},

"TREATMENT":{"TREATMENT_SUMMARY":"The mice were fed either a control diet or a diet supplemented with 10% galactooligosaccharide for 3 weeks. After the 3 weeks had elapsed the mice were euthanized and the colonic contents were collected."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"The colonic contents were sent to Metabolon who then performed the sample preparation prior to UPLC-MS analysis."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"Low pH Lipophilic (LC/MS Pos late)","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Waters Acquity","COLUMN_NAME":"Waters Acquity BEH C18 (100 x 2mm, 1.7um)","SOLVENT_A":"-","SOLVENT_B":"-","FLOW_GRADIENT":"-","FLOW_RATE":"-","COLUMN_TEMPERATURE":"-"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"Other","ION_MODE":"POSITIVE","MS_COMMENTS":"Metabolon (LC/MS Pos late)"},

"MS_METABOLITE_DATA":{
"Units":"Relative Abundance",

"Data":[{"Metabolite":"1,2-dilinoleoyl-GPC (18:2/18:2)","NACH-00271":"0.663","NACH-00272":"1.155","NACH-00273":"0.8284","NACH-00274":"1.098","NACH-00275":"0.9648","NACH-00276":"1.2468","NACH-00277":"1","NACH-00278":"0.8796","NACH-00279":"1.2247","NACH-00280":"0.8985","NACH-00281":"1.0437"},{"Metabolite":"1,2-dilinoleoyl-GPE (18:2/18:2)*","NACH-00271":"1","NACH-00272":"1.0235","NACH-00273":"0.5449","NACH-00274":"0.7097","NACH-00275":"1.2621","NACH-00276":"1.1141","NACH-00277":"0.7605","NACH-00278":"0.6237","NACH-00279":"1.0862","NACH-00280":"0.9509","NACH-00281":"1.2539"},{"Metabolite":"1,2-dipalmitoyl-GPC (16:0/16:0)","NACH-00271":"1.4013","NACH-00272":"0.989","NACH-00273":"1.0651","NACH-00274":"1","NACH-00275":"0.9099","NACH-00276":"0.897","NACH-00277":"0.9033","NACH-00278":"0.8021","NACH-00279":"1.2142","NACH-00280":"1.1536","NACH-00281":"2.5302"},{"Metabolite":"1,2-dipalmitoyl-GPE (16:0/16:0)*","NACH-00271":"0.873","NACH-00272":"1.0817","NACH-00273":"2.5205","NACH-00274":"0.7618","NACH-00275":"0.7618","NACH-00276":"0.9183","NACH-00277":"0.7831","NACH-00278":"0.7618","NACH-00279":"0.7618","NACH-00280":"2.7221","NACH-00281":"2.6626"},{"Metabolite":"1-(1-enyl-oleoyl)-GPE (P-18:1)*","NACH-00271":"0.9768","NACH-00272":"1.3809","NACH-00273":"0.6873","NACH-00274":"0.8961","NACH-00275":"0.7528","NACH-00276":"1.1759","NACH-00277":"1.3222","NACH-00278":"0.5009","NACH-00279":"1.5637","NACH-00280":"1.3511","NACH-00281":"1"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-arachidonoyl-GPC (P-16:0/20:4)*","NACH-00271":"1.3136","NACH-00272":"0.8361","NACH-00273":"0.9232","NACH-00274":"1.1772","NACH-00275":"0.9922","NACH-00276":"0.995","NACH-00277":"1.1258","NACH-00278":"0.7966","NACH-00279":"1.4155","NACH-00280":"0.7966","NACH-00281":"1.005"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-arachidonoyl-GPE (P-16:0/20:4)*","NACH-00271":"0.7689","NACH-00272":"0.8822","NACH-00273":"1.0067","NACH-00274":"1","NACH-00275":"1.0954","NACH-00276":"0.9906","NACH-00277":"0.9172","NACH-00278":"0.8604","NACH-00279":"1.4096","NACH-00280":"1.2208","NACH-00281":"1.8166"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-linoleoyl-GPC (P-16:0/18:2)*","NACH-00271":"0.7278","NACH-00272":"1.0172","NACH-00273":"0.6541","NACH-00274":"1.1889","NACH-00275":"1.1679","NACH-00276":"1.3934","NACH-00277":"1","NACH-00278":"0.7527","NACH-00279":"1.4322","NACH-00280":"0.9347","NACH-00281":"0.9754"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-linoleoyl-GPE (P-16:0/18:2)*","NACH-00271":"0.4748","NACH-00272":"0.4748","NACH-00273":"0.6361","NACH-00274":"0.4748","NACH-00275":"1.9355","NACH-00276":"1.0854","NACH-00277":"0.4748","NACH-00278":"1.0347","NACH-00279":"0.7388","NACH-00280":"0.4748","NACH-00281":"1"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-oleoyl-GPC (P-16:0/18:1)*","NACH-00271":"0.9446","NACH-00272":"1","NACH-00273":"0.5469","NACH-00274":"1.1415","NACH-00275":"0.9599","NACH-00276":"1.0751","NACH-00277":"1.0336","NACH-00278":"0.9434","NACH-00279":"1.4352","NACH-00280":"0.9604","NACH-00281":"1.0946"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-oleoyl-GPE (P-16:0/18:1)*","NACH-00271":"0.7505","NACH-00272":"0.8316","NACH-00273":"1.2825","NACH-00274":"0.8075","NACH-00275":"1.1322","NACH-00276":"1","NACH-00277":"0.994","NACH-00278":"0.6286","NACH-00279":"1.3486","NACH-00280":"1.2918","NACH-00281":"2.0513"},{"Metabolite":"1-(1-enyl-palmitoyl)-2-palmitoyl-GPC (P-16:0/16:0)*","NACH-00271":"1","NACH-00272":"0.9073","NACH-00273":"0.799","NACH-00274":"0.9385","NACH-00275":"1.062","NACH-00276":"0.9298","NACH-00277":"1.0884","NACH-00278":"0.9753","NACH-00279":"1.0949","NACH-00280":"1.1134","NACH-00281":"1.2318"},{"Metabolite":"1-(1-enyl-palmitoyl)-GPC (P-16:0)*","NACH-00271":"0.7936","NACH-00272":"1","NACH-00273":"0.7923","NACH-00274":"1.046","NACH-00275":"1.0028","NACH-00276":"1.1023","NACH-00277":"1.0864","NACH-00278":"0.7726","NACH-00279":"1.0231","NACH-00280":"0.9544","NACH-00281":"0.7618"},{"Metabolite":"1-(1-enyl-palmitoyl)-GPE (P-16:0)*","NACH-00271":"0.889","NACH-00272":"1.2514","NACH-00273":"0.8428","NACH-00274":"1","NACH-00275":"0.9527","NACH-00276":"1.1479","NACH-00277":"1.262","NACH-00278":"0.7319","NACH-00279":"1.2538","NACH-00280":"1.301","NACH-00281":"0.9806"},{"Metabolite":"1-(1-enyl-stearoyl)-2-arachidonoyl-GPE (P-18:0/20:4)*","NACH-00271":"0.5976","NACH-00272":"0.8836","NACH-00273":"0.8582","NACH-00274":"0.7956","NACH-00275":"1.0381","NACH-00276":"1.1264","NACH-00277":"1","NACH-00278":"0.9076","NACH-00279":"1.4478","NACH-00280":"1.1524","NACH-00281":"1.0615"},{"Metabolite":"1-(1-enyl-stearoyl)-2-linoleoyl-GPE (P-18:0/18:2)*","NACH-00271":"0.5919","NACH-00272":"0.9144","NACH-00273":"0.8406","NACH-00274":"0.922","NACH-00275":"1.0955","NACH-00276":"1.3725","NACH-00277":"1.071","NACH-00278":"1","NACH-00279":"1.4067","NACH-00280":"1.0336","NACH-00281":"0.9411"},{"Metabolite":"1-(1-enyl-stearoyl)-2-oleoyl-GPE (P-18:0/18:1)","NACH-00271":"0.5005","NACH-00272":"0.5005","NACH-00273":"0.8854","NACH-00274":"0.5005","NACH-00275":"1.1373","NACH-00276":"1.8592","NACH-00277":"0.5005","NACH-00278":"0.5526","NACH-00279":"0.5005","NACH-00280":"1.1146","NACH-00281":"0.5005"},{"Metabolite":"1-(1-enyl-stearoyl)-GPE (P-18:0)*","NACH-00271":"1","NACH-00272":"1.2365","NACH-00273":"0.8618","NACH-00274":"0.9675","NACH-00275":"0.9658","NACH-00276":"1.4171","NACH-00277":"1.3041","NACH-00278":"0.8357","NACH-00279":"1.372","NACH-00280":"1.1709","NACH-00281":"0.8264"},{"Metabolite":"1-arachidonoyl-GPC (20:4n6)*","NACH-00271":"0.7146","NACH-00272":"1.0302","NACH-00273":"0.844","NACH-00274":"1","NACH-00275":"0.78","NACH-00276":"1.1839","NACH-00277":"1.0617","NACH-00278":"0.8453","NACH-00279":"1.2952","NACH-00280":"1.034","NACH-00281":"0.8499"},{"Metabolite":"1-arachidonoyl-GPE (20:4n6)*","NACH-00271":"0.526","NACH-00272":"0.9702","NACH-00273":"1.0665","NACH-00274":"1.5907","NACH-00275":"1","NACH-00276":"0.9523","NACH-00277":"0.9273","NACH-00278":"0.8919","NACH-00279":"1.4859","NACH-00280":"1.3399","NACH-00281":"1.3632"},{"Metabolite":"1-lignoceroyl-GPC (24:0)","NACH-00271":"0.557","NACH-00272":"1.073","NACH-00273":"0.7184","NACH-00274":"1","NACH-00275":"0.9998","NACH-00276":"1.1155","NACH-00277":"1.1662","NACH-00278":"0.8221","NACH-00279":"1.3528","NACH-00280":"1.1885","NACH-00281":"0.6856"},{"Metabolite":"1-linolenoyl-GPC (18:3)*","NACH-00271":"0.8287","NACH-00272":"1.1519","NACH-00273":"0.9117","NACH-00274":"1.3426","NACH-00275":"1.2167","NACH-00276":"1.2349","NACH-00277":"1.091","NACH-00278":"0.9814","NACH-00279":"1","NACH-00280":"0.9592","NACH-00281":"0.8841"},{"Metabolite":"1-linoleoyl-2-arachidonoyl-GPC (18:2/20:4n6)*","NACH-00271":"0.2941","NACH-00272":"1.1965","NACH-00273":"0.7869","NACH-00274":"1","NACH-00275":"0.8826","NACH-00276":"1.4216","NACH-00277":"1.2412","NACH-00278":"0.8044","NACH-00279":"1.3017","NACH-00280":"1.1415","NACH-00281":"0.9295"},{"Metabolite":"1-linoleoyl-2-arachidonoyl-GPE (18:2/20:4)*","NACH-00271":"1.0032","NACH-00272":"0.9649","NACH-00273":"1.1131","NACH-00274":"0.8733","NACH-00275":"0.8527","NACH-00276":"1","NACH-00277":"0.978","NACH-00278":"0.6324","NACH-00279":"1.0784","NACH-00280":"1.7563","NACH-00281":"2.1535"},{"Metabolite":"1-linoleoyl-2-linolenoyl-GPC (18:2/18:3)*","NACH-00271":"0.524","NACH-00272":"1.1361","NACH-00273":"0.7543","NACH-00274":"1.0661","NACH-00275":"1.2745","NACH-00276":"1.3161","NACH-00277":"1.162","NACH-00278":"0.9449","NACH-00279":"1","NACH-00280":"0.8792","NACH-00281":"0.8791"},{"Metabolite":"1-linoleoyl-GPC (18:2)","NACH-00271":"0.7357","NACH-00272":"1.0578","NACH-00273":"0.8648","NACH-00274":"1.1712","NACH-00275":"0.9825","NACH-00276":"1.1851","NACH-00277":"1.0253","NACH-00278":"0.9436","NACH-00279":"1.1703","NACH-00280":"1","NACH-00281":"0.9074"},{"Metabolite":"1-linoleoyl-GPE (18:2)*","NACH-00271":"0.9655","NACH-00272":"1.0211","NACH-00273":"0.8257","NACH-00274":"1.2875","NACH-00275":"1.1642","NACH-00276":"1","NACH-00277":"0.789","NACH-00278":"0.7859","NACH-00279":"1.2686","NACH-00280":"0.9204","NACH-00281":"1.0574"},{"Metabolite":"1-myristoyl-2-arachidonoyl-GPC (14:0/20:4)*","NACH-00271":"0.2877","NACH-00272":"1.1028","NACH-00273":"0.5389","NACH-00274":"0.9226","NACH-00275":"1","NACH-00276":"1.0115","NACH-00277":"1.0634","NACH-00278":"0.723","NACH-00279":"1.0692","NACH-00280":"1.0364","NACH-00281":"0.7262"},{"Metabolite":"1-myristoyl-2-palmitoyl-GPC (14:0/16:0)","NACH-00271":"1.1183","NACH-00272":"0.9772","NACH-00273":"1.1263","NACH-00274":"1.1119","NACH-00275":"0.9598","NACH-00276":"0.8905","NACH-00277":"0.9805","NACH-00278":"0.6665","NACH-00279":"1.2592","NACH-00280":"1","NACH-00281":"4.9047"},{"Metabolite":"1-oleoyl-2-arachidonoyl-GPE (18:1/20:4)*","NACH-00271":"0.5622","NACH-00272":"1.0733","NACH-00273":"0.8719","NACH-00274":"0.9795","NACH-00275":"0.9805","NACH-00276":"1.022","NACH-00277":"1","NACH-00278":"0.6658","NACH-00279":"1.3728","NACH-00280":"1.7142","NACH-00281":"1.8425"},{"Metabolite":"1-oleoyl-2-arachidonoyl-GPI (18:1/20:4)*","NACH-00271":"0.5147","NACH-00272":"1.5667","NACH-00273":"0.9392","NACH-00274":"0.9725","NACH-00275":"0.6626","NACH-00276":"1.4917","NACH-00277":"1","NACH-00278":"0.7433","NACH-00279":"1.4527","NACH-00280":"1.5587","NACH-00281":"1.3782"},{"Metabolite":"1-oleoyl-2-docosahexaenoyl-GPC (18:1/22:6)*","NACH-00271":"0.2434","NACH-00272":"1","NACH-00273":"0.7976","NACH-00274":"1.0839","NACH-00275":"0.7007","NACH-00276":"1.0725","NACH-00277":"0.9406","NACH-00278":"0.6421","NACH-00279":"1.3997","NACH-00280":"1.1434","NACH-00281":"1.2243"},{"Metabolite":"1-oleoyl-2-docosahexaenoyl-GPE (18:1/22:6)*","NACH-00271":"0.5799","NACH-00272":"0.9438","NACH-00273":"1.0026","NACH-00274":"1","NACH-00275":"0.9625","NACH-00276":"0.9878","NACH-00277":"1.0699","NACH-00278":"0.7205","NACH-00279":"1.108","NACH-00280":"1.5831","NACH-00281":"1.7299"},{"Metabolite":"1-oleoyl-2-linoleoyl-GPC (18:1/18:2)*","NACH-00271":"0.7419","NACH-00272":"0.9989","NACH-00273":"0.8138","NACH-00274":"1.2721","NACH-00275":"0.632","NACH-00276":"1.0207","NACH-00277":"1","NACH-00278":"0.6575","NACH-00279":"1.2391","NACH-00280":"1.0702","NACH-00281":"1.4621"},{"Metabolite":"1-oleoyl-2-linoleoyl-GPE (18:1/18:2)*","NACH-00271":"0.8472","NACH-00272":"1.0664","NACH-00273":"0.6536","NACH-00274":"0.9096","NACH-00275":"1.2547","NACH-00276":"1","NACH-00277":"0.8266","NACH-00278":"0.6554","NACH-00279":"1.1177","NACH-00280":"1.2102","NACH-00281":"1.2846"},{"Metabolite":"1-oleoyl-GPC (18:1)","NACH-00271":"0.4571","NACH-00272":"1","NACH-00273":"0.7861","NACH-00274":"1.1561","NACH-00275":"0.7376","NACH-00276":"1.2478","NACH-00277":"0.9462","NACH-00278":"0.7387","NACH-00279":"1.3529","NACH-00280":"1.1697","NACH-00281":"1.0059"},{"Metabolite":"1-oleoyl-GPE 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}