Summary of Study ST004072

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 PR002556. The data can be accessed directly via it's Project DOI: 10.21228/M8824K This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php

Study ID	ST004072
Study Title	Metabolic perturbation in ependymal cells leads to local and distant neurodegeneration and cognitive decline - Study 3 of 3 (1-month Glut1KO against Ctrl)
Study Summary	Ependymal cells (ECs) are specialized multi-ciliated glial cells that line the ventricular system of the brain, regulating cerebrospinal fluid flow (CSF) and the neighbouring neural stem cell (NSC) niche. However, their role in maintaining brain homeostasis or in disease pathogenesis remains unclear. To elucidate their function, we disrupted ependymal glucose metabolism by genetically deleting glucose-transporter-1 (GLUT1/Slc2a1) in postnatal ECs. Analyses were carried out across three separate studies (batches), with one study at 1 month (6 mice) and two studies at 12 months (3 mice, 5 mice). Results from this project confirm CSF flow changes and disrupted NSC differentiation and neuroblast migration. These mice also exhibited periventricular lipid droplet accumulation similar to Alzheimer’s disease brains. Aged cKO mice exhibited progressive cognitive and motor dysfunction, and onset of seizure activity. These behavioral deficits were coincident with various neurodegenerative pathologies, including dysmyelination, microglia-associated inflammation, and lipid imbalance. When combined with metabolic perturbation in ECs, 5xFAD mice exhibited accelerated disease onset. These findings suggest that ECs are important regulators of brain homeostasis, and their dysfunction may contribute to the pathogenesis of neurodegenerative diseases. In this part, 6x mice were analyzed at 1-months under Glut1ko (3 mice) or Control (3 mice) conditions.
Institute	University of Calgary
Department	Veterinary Medicine
Laboratory	Biernaskie Lab
Last Name	Colter
First Name	James
Address	2500 University Drive NW, Calgary AB Canada, T2N1N4
Email	jdcolter@ucalgary.ca
Phone	+1 (403) 210-7306
Submit Date	2025-06-14
Num Groups	4
Total Subjects	13
Raw Data Available	Yes
Raw Data File Type(s)	mcf
Analysis Type Detail	MALDI-MS
Release Date	2025-08-15
Release Version	1

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Project:

Project ID:	PR002556
Project DOI:	doi: 10.21228/M8824K
Project Title:	Metabolic perturbation in ependymal cells leads to local and distant neurodegeneration and cognitive decline
Project Type:	MS Imaging Analysis
Project Summary:	Ependymal cells (ECs) are specialized multi-ciliated glial cells that line the ventricular system of the brain, regulating cerebrospinal fluid flow (CSF) and the neighbouring neural stem cell (NSC) niche. However, their role in maintaining brain homeostasis or in disease pathogenesis remains unclear. To elucidate their function, we disrupted ependymal glucose metabolism by genetically deleting glucose-transporter-1 (GLUT1/Slc2a1) in postnatal ECs. Analyses were carried out across three separate studies (batches), with one study at 1 month (6 mice) and two studies at 12 months (3 mice, 5 mice). Results from this project confirm CSF flow changes and disrupted NSC differentiation and neuroblast migration. These mice also exhibited periventricular lipid droplet accumulation similar to Alzheimer’s disease brains. Aged cKO mice exhibited progressive cognitive and motor dysfunction, and onset of seizure activity. These behavioral deficits were coincident with various neurodegenerative pathologies, including dysmyelination, microglia-associated inflammation, and lipid imbalance. When combined with metabolic perturbation in ECs, 5xFAD mice exhibited accelerated disease onset. These findings suggest that ECs are important regulators of brain homeostasis, and their dysfunction may contribute to the pathogenesis of neurodegenerative diseases.
Institute:	University of Calgary
Department:	Veterinary Medicine
Laboratory:	Biernaskie Lab
Last Name:	Colter
First Name:	James
Address:	2500 University Drive NW
Email:	jdcolter@ucalgary.ca
Phone:	+1 (403) 210-7306
Funding Source:	CIHR
Project Comments:	Part 1 of 3
Publications:	(Under Review)
Contributors:	Nilesh Sharma, Alexander Pun, James Colter, Leslie Cao, Nicole Rosin, Dominic Gerding, Isabel Rea, Apolline Pistek, Qandeel Shafqat, Sarthak Sinha, Elodie Labit, Eren Kutluberk, Caleb Small, Reese Landes, Tak Ho Chu, Kartikeya Murari, E. Dale Abel, Jeffrey T. Joseph, Rehana Leak, Jeff Dunn, and Jeff Biernaskie

Subject:

Subject ID:	SU004218
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090
Age Or Age Range:	1 month

Factors:

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

mb_sample_id	local_sample_id	Sample source	Condition	Sample source
SA472710	286	240301_youngmice	Control	Brain
SA472711	287	240301_youngmice	Control	Brain
SA472712	288	240301_youngmice	Control	Brain
SA472713	509	240301_youngmice	Knockout	Brain
SA472714	521	240301_youngmice	Knockout	Brain
SA472715	522	240301_youngmice	Knockout	Brain

Showing results 1 to 6 of 6

Showing results 1 to 6 of 6

Collection:

Collection ID:	CO004211
Collection Summary:	The brain was collected as mentioned below. After the mice were euthanized, it was transcardially perfused with PBS. The brain is dissected out and then flash frozen in liquid nitrogen. The flash frozen brain tissue was then sectioned at 12 μm thickness on a cryostat (Leica Biosystems).
Sample Type:	Brain

Treatment:

Treatment ID:	TR004227
Treatment Summary:	The brain was collected as mentioned with minor modifications. After the mice were euthanized, it was transcardially perfused with PBS. The brain is dissected out and then flash frozen in liquid nitrogen. The flash frozen brain tissue was then sectioned at 12 μm thickness on a cryostat (Leica Biosystems). MALDI matrix (9-aminoacridine, 9AA) (Sigma-Aldrich) was spray-coated onto the target slides in an automated fashion using a TM Sprayer (HTX Imaging). 9-AA was made up as a 5 mg/ml solution in 90% methanol. Four passes were used with a nozzle temperature of 85°, a flowrate of 0.15 ml/min, 2-mm track spacing, and a stage velocity of 700 mm/min. Nitrogen was used as the nebulization gas and was set to 10 psi. Images were acquired on a 15T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS, Solarix, Bruker Daltonics) equipped with an Apollo II dual ion source and Smartbeam II 2kHz Nd:YAG laser that was frequency tripled to 355 nm. Data were collected in the negative ion mode with the laser operating at 2 kHz at 50 μm resolution. Tentative metabolite identifications were made by accurate mass, typically better than 1 ppm. Images were analyzed with flexImaging software (Bruker), while average spectra were exported to mMass for visualization of differences.

Treatment ID:

TR004227

Treatment Summary:

The brain was collected as mentioned with minor modifications. After the mice were euthanized, it was transcardially perfused with PBS. The brain is dissected out and then flash frozen in liquid nitrogen. The flash frozen brain tissue was then sectioned at 12 μm thickness on a cryostat (Leica Biosystems). MALDI matrix (9-aminoacridine, 9AA) (Sigma-Aldrich) was spray-coated onto the target slides in an automated fashion using a TM Sprayer (HTX Imaging). 9-AA was made up as a 5 mg/ml solution in 90% methanol. Four passes were used with a nozzle temperature of 85°, a flowrate of 0.15 ml/min, 2-mm track spacing, and a stage velocity of 700 mm/min. Nitrogen was used as the nebulization gas and was set to 10 psi. Images were acquired on a 15T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS, Solarix, Bruker Daltonics) equipped with an Apollo II dual ion source and Smartbeam II 2kHz Nd:YAG laser that was frequency tripled to 355 nm. Data were collected in the negative ion mode with the laser operating at 2 kHz at 50 μm resolution. Tentative metabolite identifications were made by accurate mass, typically better than 1 ppm. Images were analyzed with flexImaging software (Bruker), while average spectra were exported to mMass for visualization of differences.

Sample Preparation:

Sampleprep ID:	SP004224
Sampleprep Summary:	MALDI matrix (9-aminoacridine, 9AA) (Sigma-Aldrich) was spray-coated onto the target slides in an automated fashion using a TM Sprayer (HTX Imaging). 9-AA was made up as a 5 mg/ml solution in 90% methanol. Four passes were used with a nozzle temperature of 85°, a flowrate of 0.15 ml/min, 2-mm track spacing, and a stage velocity of 700 mm/min. Nitrogen was used as the nebulization gas and was set to 10 psi. Images were acquired on a 15T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS, Solarix, Bruker Daltonics) equipped with an Apollo II dual ion source and Smartbeam II 2kHz Nd:YAG laser that was frequency tripled to 355 nm. Data were collected in the negative ion mode with the laser operating at 2 kHz at 50 μm resolution. Tentative metabolite identifications were made by accurate mass, typically better than 1 ppm. Images were analyzed with flexImaging software (Bruker), while average spectra were exported to mMass for visualization of differences.

Sampleprep ID:

SP004224

Sampleprep Summary:

MALDI matrix (9-aminoacridine, 9AA) (Sigma-Aldrich) was spray-coated onto the target slides in an automated fashion using a TM Sprayer (HTX Imaging). 9-AA was made up as a 5 mg/ml solution in 90% methanol. Four passes were used with a nozzle temperature of 85°, a flowrate of 0.15 ml/min, 2-mm track spacing, and a stage velocity of 700 mm/min. Nitrogen was used as the nebulization gas and was set to 10 psi. Images were acquired on a 15T Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS, Solarix, Bruker Daltonics) equipped with an Apollo II dual ion source and Smartbeam II 2kHz Nd:YAG laser that was frequency tripled to 355 nm. Data were collected in the negative ion mode with the laser operating at 2 kHz at 50 μm resolution. Tentative metabolite identifications were made by accurate mass, typically better than 1 ppm. Images were analyzed with flexImaging software (Bruker), while average spectra were exported to mMass for visualization of differences.

Combined analysis:

Analysis ID	AN006738
Chromatography ID	CH005120
MS ID	MS006437
Analysis type	MS
Chromatography type	None (Direct infusion)
Chromatography system	none
Column	none
MS Type	MALDI
MS instrument type	MALDI-TOF
MS instrument name	Bruker Solarix FT-ICR-MS
Ion Mode	NEGATIVE
Units	n/a

Chromatography:

Chromatography ID:	CH005120
Instrument Name:	none
Column Name:	none
Column Temperature:	none
Flow Gradient:	none
Flow Rate:	none
Solvent A:	none
Solvent B:	none
Chromatography Type:	None (Direct infusion)

MS:

MS ID:	MS006437
Analysis ID:	AN006738
Instrument Name:	Bruker Solarix FT-ICR-MS
Instrument Type:	MALDI-TOF
MS Type:	MALDI
MS Comments:	Methods below. Please note that the processed data files included in this submission only provide the identified metabolites in the set, since the data format for workbench does not allow a 3-dimensional approach (each pixel contains a spectrum, and sample brains within an experiment are sectioned by brain region and analyzed by metabolite across spatial coordinates). Please refer to github.com/BiernaskieLab for .csv files containing spectral intensities by metabolite across spatial coordinates for specific brain regions by sample. MALDI-TOF spectral data was acquired using Bruker Compass FlexImaging software. The raw dataset was exported to Bruker SciLS Lab for peak-finding, isolation of spectral data by region. Reorganized data was exported as .csv files for cross-dataset comparisons utilizing Python. Total ion count (TIC) normalization was applied prior to exporting numerical intensity data from SciLS Lab or plotting and exporting images from FlexImaging. Automated peak-finding was applied within Bruker SciLS Lab to TIC-normalized datasets to generate an ion list for further processing. This ion list was cross-referenced between datasets to harmonize the ion list across brain sections imaged in all experiments. The COMP_DB database at https://www.lipidmaps.org was applied to annotate ions with potential lipid hits. A delta m/z of ±0.01 was used. If no matches were returned, this delta was increased to ±0.02, and repeated at ±0.05 if nothing was returned. Ions that had no matches in the database beyond ±0.05 were excluded from the set. Given the limitations of assessing fragmentation profiles of the mass spectral data, ions with potential matches from multiple lipid groups were classified as ‘unknown’. Those ions with multiple possible matches from the same group were included in their respective lipid group and annotated with all possible matches. This resulted in a list of 48 ions with known lipid groups.
Ion Mode:	NEGATIVE