Summary of Study ST003019

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 PR001759. The data can be accessed directly via it's Project DOI: 10.21228/M8BH9K This work is supported by NIH grant, U2C- DK119886.

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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 IDST003019
Study TitleMetabolomic profiling of PMM2-CDG brain organoids by GC/MS
Study SummaryPMM2-CDG is a rare inborn error of metabolism caused by deficiency of the PMM enzyme, which leads to impaired protein glycosylation. While the disorder has primarily neurological presentation, there is limited knowledge about the specific brain-related changes that result from PMM deficiency. Utilizing 3D brain organoids derived from individuals with PMM2-CDG, we identified abnormal glucose metabolism in PMM2-CDG organoids, indicating disturbances in metabolic utilization. In this experiment, day 160 organoids were used.
Institute
Mayo Clinic
Last NameRadenkovic
First NameSilvia
Address200 2nd Ave SW Rochester MN, USA
Emailradenkovic.silvia@mayo.edu
Phone507(77) 6-6107
Submit Date2023-12-19
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailGC-MS
Release Date2024-12-20
Release Version1
Silvia Radenkovic Silvia Radenkovic
https://dx.doi.org/10.21228/M8BH9K
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001759
Project DOI:doi: 10.21228/M8BH9K
Project Title:Metabolomic profiling of PMM2-CDG brain organoids
Project Summary:PMM2-CDG is a rare inborn error of metabolism caused by deficiency of the PMM enzyme, which leads to impaired protein glycosylation. While the disorder has primarily neurological presentation, there is limited knowledge about the specific brain-related changes that result from PMM deficiency. Utilizing 3D brain organoids derived from individuals with PMM2-CDG, we identified abnormal glucose metabolism in PMM2-CDG organoids, indicating disturbances in metabolic utilization.
Institute:Mayo Clinic
Last Name:Radenkovic
First Name:Silvia
Address:200 2nd Ave SW Rochester MN
Email:radenkovic.silvia@mayo.edu
Phone:507(77) 6-6107
Funding Source:NIH, KU Leuven

Subject:

Subject ID:SU003133
Subject Type:Mammal
Subject Species:Homo sapiens
Taxonomy ID:9606
Genotype Strain:WT/PMM2-CDG
Age Or Age Range:5-25
Gender:Male and female
Species Group:Mammals

Factors:

Subject type: Mammal; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Genotype
SA327663S5CTR
SA327664S1CTR
SA327665S3CTR
SA327666S6PMM2
SA327667S4PMM2
SA327668S2PMM2
Showing results 1 to 6 of 6

Collection:

Collection ID:CO003126
Collection Summary:Brain were collected at day in vitro 160. Briefly, 5 organoids per cell line were collected and washed three times in DPBS (Gibco). The DPBS was then removed, the organoids flash frozen and kept in -80 °C prior to metabolomics experiments. The metabolites were extracted using two phase extraction protocol. First, the organoids were transferred to lysing matrix tube and 350 µL of ice-cold extraction buffer (80 % MeOH, IS) was added to the sample. Next, the organoids were lyzed with ribolyzer, the lysate transferred to 1.5 mL Eppendorf tube and placed overnight at -80 °C. Further, the samples were centrifuged at 15,000 rpm, 4 °C, 20 min. 100 µL of supernatant was transferred to a fresh Eppendorf tube and 35 µL of ddH20 was added, followed by 800 µL 100% chloroform. The samples were then vortexed and stored at 4 °C overnight. Next, the polar phase was the used for Gas Chromatography/Mass Spectrometry (GC/MS)
Sample Type:Brain organoids
Storage Conditions:-80℃

Treatment:

Treatment ID:TR003142
Treatment Summary:Samples were not treated

Sample Preparation:

Sampleprep ID:SP003139
Sampleprep Summary:The metabolites were extracted from the brain organoids using double phase metabolite extration. Standards containing metabolites of interest were prepared simultaneously and dried overnight together with cellular extracts by vacuum centrifugation at 4 °C. The following day, 20 µL of methoxyamine (MOX) (Sigma Aldrich) was added to the samples and they were incubated for 90 min at 37 °C. Next, 60 µL of N, O-bis(trimethylsilyl)trifluoroacetamide (TMS) (Sigma Aldrich) was added to the samples, which were then incubated at 60 °C for 30 min. Samples were kept overnight in a dry and cool place to allow further derivatization with TMS
Processing Storage Conditions:-80℃
Extract Storage:-80℃
Sample Derivatization:MOX, TMS

Combined analysis:

Analysis ID AN004953
Analysis type MS
Chromatography type GC
Chromatography system Agilent 7890A
Column Agilent HP-5ms (30m x 0.25mm, 0.25um)
MS Type EI
MS instrument type Triple quadrupole
MS instrument name Agilent 7000B
Ion Mode POSITIVE
Units AUC

Chromatography:

Chromatography ID:CH003737
Chromatography Summary:Agilent 7890A GC (Agilent Technologies) coupled with an HP-5 ms 5 % phenyl methyl silox capillary column (30 m, 0.25 mm, 0.25 um, Agilent Technologies). The temperature gradient applied was as follows: 2 min 100°C, next 175 °C at Δ20 °C/min, followed by an increase to 230 °C at Δ4 °C/min, kept at 230 °C for 3 min, increased to 300 °C at Δ 40°C/min and lastly kept at 300 °C for 5 min. The column was further baked for another 3 min at 325 °C after the gradient.
Instrument Name:Agilent 7890A
Column Name:Agilent HP-5ms (30m x 0.25mm, 0.25um)
Column Temperature:Gradient
Flow Gradient:N/A
Flow Rate:NA
Solvent A:N/A
Solvent B:N/A
Chromatography Type:GC

MS:

MS ID:MS004693
Analysis ID:AN004953
Instrument Name:Agilent 7000B
Instrument Type:Triple quadrupole
MS Type:EI
MS Comments:Mass Hunter Workstation software with the Quantitative Analysis Version B.06.00/Build 6.0.388.0 and el-maven polly software. Specific metabolites were identified based on their fragment formula/elution time
Ion Mode:POSITIVE
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