Summary of Study ST002186

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001393. The data can be accessed directly via it's Project DOI: 10.21228/M8NT4K This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study ID	ST002186
Study Title	An early-life microbiota metabolite protects against obesity via intestinal PPAR-gamma
Study Type	untargeted metabolomics analysis
Study Summary	The mechanisms by which the early-life microbiota protects against environmental factors that promote childhood obesity remain largely unknown. Using a mouse model in which young mice are simultaneously exposed to antibiotics and a high-fat diet, we show that Lactobacillus species, predominant members of the small intestine microbiota, regulate intestinal epithelial cells (IECs) to limit diet-induced obesity during early-life. A Lactobacillus-derived metabolite, phenyllactic acid (PLA), protected against metabolic dysfunction caused by early-life exposure to antibiotics and a high-fat diet by increasing the abundance of peroxisome proliferator activated receptor gamma (PPAR-gamma) in the small intestine IECs. Therefore, PLA is a microbiota-derived metabolite that activates protective pathways in the small intestine epithelium to regulate fat absorption and prevent obesity during early life.
Institute	Vanderbilt University
Department	Chemistry
Laboratory	Center for Innovative Technology
Last Name	Codreanu
First Name	Simona Gabriella
Address	1234 Stevenson Center Lane
Email	SIMONA.CODREANU@VANDERBILT.EDU
Phone	6158758422
Submit Date	2022-06-03
Num Groups	4
Total Subjects	20
Num Males	20
Raw Data Available	Yes
Raw Data File Type(s)	raw(Thermo)
Analysis Type Detail	LC-MS
Release Date	2023-06-06
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR001393
Project DOI:	doi: 10.21228/M8NT4K
Project Title:	An early-life microbiota metabolite protects against obesity via intestinal PPAR-Gamma
Project Type:	Untargeted Metabolomics analysis
Project Summary:	The mechanisms by which the early-life microbiota protects against environmental factors that promote childhood obesity remain largely unknown. Using a mouse model in which young mice are simultaneously exposed to antibiotics and a high-fat diet, we show that Lactobacillus species, predominant members of the small intestine microbiota, regulate intestinal epithelial cells (IECs) to limit diet-induced obesity during early-life. A Lactobacillus-derived metabolite, phenyllactic acid (PLA), protected against metabolic dysfunction caused by early-life exposure to antibiotics and a high-fat diet by increasing the abundance of peroxisome proliferator activated receptor gamma (PPAR-gamma) in the small intestine IECs. Therefore, PLA is a microbiota-derived metabolite that activates protective pathways in the small intestine epithelium to regulate fat absorption and prevent obesity during early life.
Institute:	Vanderbilt University
Department:	Chemistry
Laboratory:	Center for Innovative Technology
Last Name:	Codreanu
First Name:	Simona Gabriella
Address:	1234 Stevenson Center Lane
Email:	SIMONA.CODREANU@VANDERBILT.EDU
Phone:	16158758422

Subject:

Subject ID:	SU002272
Subject Type:	Other organism
Subject Species:	Mus musculus
Taxonomy ID:	10090
Genotype Strain:	C57BL/6N mice
Age Or Age Range:	5 weeks
Gender:	Male
Species Group:	Mammals

Factors:

Subject type: Other organism; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id	local_sample_id	Treatment
SA210048	HFP_381_S16	HF diet + LDP (HFP)
SA210049	HFP_382_S17	HF diet + LDP (HFP)
SA210050	HFP_383_S18	HF diet + LDP (HFP)
SA210051	HFP_384_S19	HF diet + LDP (HFP)
SA210052	HFP_386_S20	HF diet + LDP (HFP)
SA210053	HF_374_S13	High fat (HF) diet
SA210054	HF_375_S14	High fat (HF) diet
SA210055	HF_376_S15	High fat (HF) diet
SA210056	HF_372_S12	High fat (HF) diet
SA210057	HF_371_S11	High fat (HF) diet
SA210058	LF_356_S05	LF diet
SA210059	LF_354_S04	LF diet
SA210060	LF_353_S03	LF diet
SA210061	LF_352_S02	LF diet
SA210062	LFP_361_S06	LF diet + low dose penicillin (LFP)
SA210063	LFP_362_S07	LF diet + low dose penicillin (LFP)
SA210064	LFP_365_S10	LF diet + low dose penicillin (LFP)
SA210065	LFP_364_S09	LF diet + low dose penicillin (LFP)
SA210066	LFP_363_S08	LF diet + low dose penicillin (LFP)
SA210067	LF_351_S01	Low Fat (LF) diet

Showing results 1 to 20 of 20

Showing results 1 to 20 of 20

Collection:

Collection ID:	CO002265
Collection Summary:	At the end of the experiment (5-weeks)after starting diet and antibiotic treatments, mice were humanely euthanized by CO2 administration. Afterwards, ileum (distal small intestine) content was collected.
Sample Type:	Feces
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR002284
Treatment Summary:	Animals were fed either a 60% fat diet (HF) (OpenSource Diets, #D12492) or a 10% fat control diet (LF) (OpenSource Diets, #D12450J) for 5 weeks. Groups of LF or HF diet mice were also given low doses of penicillin (Sigma Aldrich #P1382)(LDP) (6.67 mg/L, (10)) in their drinking water throughout the experiment or clinical doses of penicillin (0.167 g/L, (35)) in their drinking water from days 0 – 5, 15 – 20, and 30 – 35. For long-term experiments, mice were fed a HF diet or given a HF diet and LDP for 5 weeks before being switched to either a LF or HF diet alone for an additional 5 weeks.

Treatment ID:

TR002284

Treatment Summary:

Animals were fed either a 60% fat diet (HF) (OpenSource Diets, #D12492) or a 10% fat control diet (LF) (OpenSource Diets, #D12450J) for 5 weeks. Groups of LF or HF diet mice were also given low doses of penicillin (Sigma Aldrich #P1382)(LDP) (6.67 mg/L, (10)) in their drinking water throughout the experiment or clinical doses of penicillin (0.167 g/L, (35)) in their drinking water from days 0 – 5, 15 – 20, and 30 – 35. For long-term experiments, mice were fed a HF diet or given a HF diet and LDP for 5 weeks before being switched to either a LF or HF diet alone for an additional 5 weeks.

Sample Preparation:

Sampleprep ID:	SP002278
Sampleprep Summary:	Frozen mouse intestinal content samples (n=20, 5 biological replicates for each sample group) were lysed in 500 µl ice-cold lysis buffer (1:1:2, v:v:v, acetonitrile: methanol: ammonium bicarbonate 0.1M - pH 8.0) and sonicated individually using a probe tip sonicator at 50% power (10 pulses). The lysis buffer contained isotopically labeled standards (n=2) to determine sample process variability. Homogenized samples were normalized by weight to the smallest amount of tissue sample such that each sample contained an equal amount of tissue. Proteins were precipitated from individual samples by addition of 800 µL of ice-cold methanol followed by overnight incubation at -80°C. Precipitated proteins were pelleted by centrifugation (15,000 rpm, 15 min) and metabolite extracts were dried down in vacuo and stored at -80°C. Individual samples were reconstituted in 100 μL of reverse phase liquid chromatography reconstitution buffer (acetonitrile/water with 0.1% formic acid, 3:97, v/v) containing isotopically labeled standards (n=2) to assess instrument variability. A pooled quality control (QC) sample was prepared by pooling equal volumes (10 μL) from each individual sample following reconstitution.
Processing Storage Conditions:	-80℃
Extraction Method:	Following lysis and standard addition, protein precipitation was performed by adding 800µL of ice-cold methanol (4x by volume). Samples were incubated at -80°C overnight. Following incubation, samples were centrifuged at 10,000 rpm for 10 min to eliminate proteins. The supernatants containing metabolites were dried via speed-vacuum.
Extract Storage:	-80℃

Combined analysis:

Analysis ID	AN003580
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Thermo Vanquish
Column	Thermo Hypersil Gold (100 x 2. mm,1.9um)
MS Type	ESI
MS instrument type	QTRAP
MS instrument name	Thermo Q Exactive HF hybrid Orbitrap
Ion Mode	NEGATIVE
Units	time_m/z

Chromatography:

Chromatography ID:	CH002647
Chromatography Summary:	Prepared samples were analyzed by RPLC-HRMS/MS in the Vanderbilt Center for Innovative Technology (CIT) using a modified version of a reversed phase chromatography negative ionization method. Metabolites were separated on a Thermo Fisher Scientific (Waltham, MA) Hypersil Gold C18 column (100 x 2.1 mm, 1.9 μm particle size) using water/acetonitrile gradient with formic acid (0.1%) added to both mobile phases.
Instrument Name:	Thermo Vanquish
Column Name:	Thermo Hypersil Gold (100 x 2. mm,1.9um)
Column Temperature:	40
Flow Rate:	0.25 mL/min
Solvent A:	100% water; 0.1% formic acid
Solvent B:	80% acetonitrile/20% water; 0.1% formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS003337
Analysis ID:	AN003580
Instrument Name:	Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:	QTRAP
MS Type:	ESI
MS Comments:	Progenesis QI software
Ion Mode:	NEGATIVE