Summary of Study ST002508

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 PR001619. The data can be accessed directly via it's Project DOI: 10.21228/M8F41P 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.

Study ID	ST002508
Study Title	13C-isotopic labeling of mouse eye organoids under three conditions.
Study Summary	To examine the potential involvement of metabolites, we performed 13C-glucose labeling under three conditions: 13C-glucose = control, 13C-glucose + GNE-140 = LHDi, 13C-lactate in glucose free media mimicking the lack of glucose. Cell Name AES0145 : Rx-GFP K/I EB5 (RIKEN Cell Bank): Organoid method (PMID: 21475194).
Institute	Northwestern University
Last Name	TAKATA
First Name	NOZOMU
Address	303 East Superior Street, 10-220, Chicago, Illinois, 60611, USA
Email	nozomu.takata@northwestern.edu
Phone	13125036066
Submit Date	2023-03-15
Raw Data Available	Yes
Raw Data File Type(s)	raw(Thermo)
Analysis Type Detail	LC-MS
Release Date	2023-06-01
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR001619
Project DOI:	doi: 10.21228/M8F41P
Project Title:	Lactate-dependent Transcriptional Regulation Controls Mammalian Eye Morphogenesis
Project Summary:	Mammalian retinal metabolism favors aerobic glycolysis. However, the role of glycolytic metabolism in retinal morphogenesis remains unknown. Here we report that aerobic glycolysis is necessary for the early stages of retinal development. Taking advantage of an unbiased approach that combines the use of eye organoids and single-cell RNA sequencing we identified specific glucose transporters and glycolytic genes in retinal progenitors. Next, we determined that the optic vesicle territory of mouse embryos displays elevated levels of glycolytic activity. At the functional level, we found that removal of Glut1 (also known as Slc2a1) and Lactate dehydrogenase A (Ldha) gene activity from developing retinal progenitors arrested eye morphogenesis. Surprisingly, we uncovered that lactate-mediated upregulation of key eye-field transcription factors was controlled by the epigenetic modification of histone H3 acetylation through histone deacetylase (Hdac) activity. Our results identify a novel bioenergetic independent role of lactate as a signaling molecule necessary for mammalian eye morphogenesis.
Institute:	Northwestern University
Last Name:	TAKATA
First Name:	NOZOMU
Address:	303 East Superior Street, 10-220, Chicago, Illinois, 60611, USA
Email:	nozomu.takata@northwestern.edu
Phone:	13125036066

Subject:

Subject ID:	SU002608
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

Factors:

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

mb_sample_id	local_sample_id	Genotype	Treatment
SA252376	Cont1	Wild-type	Control
SA252377	Cont3	Wild-type	Control
SA252378	Cont2	Wild-type	Control
SA252382	Lactate3	Wild-type	Lactate
SA252383	Lactate2	Wild-type	Lactate
SA252384	Lactate1	Wild-type	Lactate
SA252379	LDHi3	Wild-type	LDHi
SA252380	LDHi2	Wild-type	LDHi
SA252381	LDHi1	Wild-type	LDHi

Showing results 1 to 9 of 9

Showing results 1 to 9 of 9

Collection:

Collection ID:	CO002601
Collection Summary:	The eye organoids were collected and washed twice with PBS before pellets were flash-frozen and stored at −80°C until metabolite extraction. Three biologically independent repeated samples were collected.
Sample Type:	Retina

Treatment:

Treatment ID:	TR002620
Treatment Summary:	13C-glucose labeling under three conditions: 13C-glucose = control, 13C-glucose + GNE-140 = LHDi, 13C-lactate in glucose free media mimicking the lack of glucose. All the organoids (50 organoids per condition) were washed with blank SILAC before being reconstituted in 13C medium at a concentration of 10 eye organoids/ml.

Sample Preparation:

Sampleprep ID:	SP002614
Sampleprep Summary:	The eye organoids were collected and washed twice with PBS before pellets were flash-frozen and stored at −80°C until metabolite extraction. Metabolite analysis was performed as described previously (PMID: 33931446). Briefly, for metabolite extraction, 80% methanol was used followed by the rapid freeze-thaw method to break the tissues. The supernatant underwent speedvac drying. The samples were prepared in 80% acetonitrile and were analyzed by High-Resolution Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS).

Sampleprep ID:

SP002614

Sampleprep Summary:

The eye organoids were collected and washed twice with PBS before pellets were flash-frozen and stored at −80°C until metabolite extraction. Metabolite analysis was performed as described previously (PMID: 33931446). Briefly, for metabolite extraction, 80% methanol was used followed by the rapid freeze-thaw method to break the tissues. The supernatant underwent speedvac drying. The samples were prepared in 80% acetonitrile and were analyzed by High-Resolution Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS).

Combined analysis:

Analysis ID	AN004130
Analysis type	MS
Chromatography type	HILIC
Chromatography system	Q-exactive
Column	Waters XBridge BEH Amide (100 x 3.0mm, 3.5um)
MS Type	ESI
MS instrument type	Orbitrap
MS instrument name	Thermo Q Exactive Orbitrap
Ion Mode	UNSPECIFIED
Units	Peak area

Chromatography:

Chromatography ID:	CH003060
Instrument Name:	Q-exactive
Column Name:	Waters XBridge BEH Amide (100 x 3.0mm, 3.5um)
Column Temperature:	40
Flow Gradient:	0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1-18 min, 75% A; 18-25 min, 15% A with a flow rate of 150 μL/min
Flow Rate:	150 μL/mi
Solvent A:	95% water/5% acetonitrile; 10 mM ammonium hydroxide; 10 mM ammonium acetate, pH 9.0
Solvent B:	100% acetonitrile
Chromatography Type:	HILIC

MS:

MS ID:	MS003877
Analysis ID:	AN004130
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	The capillary of the ESI source was set to 275 °C, with sheath gas at 35 arbitrary units, auxiliary gas at 5 arbitrary units and the spray voltage at 4.0 kV. In positive/negative polarity switching mode, an m/z scan range from 60 to 900 was chosen and MS1 data was collected at a resolution of 70,000. The automatic gain control (AGC) target was set at 1 × 106 and the maximum injection time was 200 ms. The top 5 precursor ions were subsequently fragmented, in a data-dependent manner, using the higher energy collisional dissociation (HCD) cell set to 30% normalized collision energy in MS2 at a resolution power of 17,500. Besides matching m/z, metabolites are identified by matching either retention time with analytical standards and/or MS2 fragmentation pattern. Data acquisition and analysis were carried out by Xcalibur 4.1 software and Tracefinder 4.1 software, respectively (both from Thermo Fisher Scientific)
Ion Mode:	UNSPECIFIED