Summary of Study ST003247

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 PR002016. The data can be accessed directly via it's Project DOI: 10.21228/M81C00 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	ST003247
Study Title	Effects of mitoregulin loss in aged female mice
Study Summary	Cardiac lipidome analysis in aged (21 to 23-months old) female wildtype and mtln knockout mice
Institute	University of Iowa
Last Name	Boudreau
First Name	Ryan
Address	4334 PBDB, 169 Newton Rd, Iowa City, IA 52242
Email	ryan-boudreau@uiowa.edu
Phone	3193535573
Submit Date	2024-06-04
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	LC-MS
Release Date	2024-12-31
Release Version	1

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Project:

Project ID:	PR002016
Project DOI:	doi: 10.21228/M81C00
Project Title:	Effects of mitoregulin loss on cardiac and mitochondrial lipids in mice
Project Summary:	We and others discovered a highly-conserved mitochondrial transmembrane microprotein, named Mitoregulin (Mtln), that supports lipid metabolism. We reported that Mtln strongly binds cardiolipin (CL), increases mitochondrial respiration and Ca2+ retention capacities, and reduces reactive oxygen species (ROS). Here we extend our observation of Mtln-CL binding and examine Mtln influence on cristae structure and mitochondrial membrane integrity during stress. We demonstrate that mitochondria from constitutive- and inducible Mtln-knockout (KO) mice are susceptible to membrane freeze-damage and that this can be rescued by acute Mtln re-expression. In mitochondrial-simulated lipid monolayers, we show that synthetic Mtln decreases lipid packing and monolayer elasticity. Lipidomics revealed that Mtln-KO heart tissues show broad decreases in 22:6-containing lipids and increased cardiolipin damage/remodeling. Lastly, we demonstrate that Mtln-KO mice suffer worse myocardial ischemia-reperfusion injury, hinting at a translationally-relevant role for Mtln in cardioprotection. Our work supports a model in which Mtln binds cardiolipin and stabilizes mitochondrial membranes to broadly influence diverse mitochondrial functions, including lipid metabolism, while also protecting against stress.
Institute:	University of Iowa
Last Name:	Boudreau
First Name:	Ryan
Address:	4334 PBDB, 169 Newton Rd, Iowa City, IA 52242
Email:	ryan-boudreau@uiowa.edu
Phone:	3193535573

Subject:

Subject ID:	SU003366
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090
Gender:	Female
Species Group:	Mammals

Factors:

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

mb_sample_id	local_sample_id	Genotype
SA353463	1133-1	KO
SA353464	1143-2	KO
SA353465	1143-3	KO
SA353466	1145-2	KO
SA353467	1152-1	KO
SA353468	1066-23	WT
SA353469	1066-4	WT
SA353470	1074-34	WT
SA353471	1103-12	WT
SA353472	WTF	WT

Showing results 1 to 10 of 10

Showing results 1 to 10 of 10

Collection:

Collection ID:	CO003359
Collection Summary:	Mtln-KO and WT mice were fasted for 4-6 h, and while under deep ketamine/xylazine anesthesia, the heart apex was snipped off and immediately snap-frozen by LN2 immersion.
Sample Type:	Heart

Treatment:

Treatment ID:	TR003375
Treatment Summary:	No treatment.

Sample Preparation:

Sampleprep ID:	SP003373
Sampleprep Summary:	Lipid extraction. Lipid extraction, based on Matyash et al.72, was performed as follows. All solutions were pre-chilled on ice. Tissues or mitochondrial pellets were transferred to labeled bead-mill tubes (1.4 mm, MoBio Cat# 13113-50) where lipids were extracted in a solution of 250 µL PBS, 225 µL MeOH containing internal standards, and 750 µL MTBE (methyl tert-butyl ether). Internal standards were Avanti SPLASH LipidoMix (Lot#12) at 10 µL per sample and Cambridge Isotope laboratories NSK-B and NSK-B-G1 (deuterated carnitines) at 10 µL per sample. The samples were homogenized in one 30 s cycle using the Omni Bead Ruptor followed by a rest on ice for 1 h. An addition of 188 µL PBS was made to induce phase separation. After centrifugation at 16,000 g for 5 minutes at 4 °C, the upper phases were collected and evaporated to dryness under a gentle nitrogen stream at room temperature. Lipid samples were reconstituted in 500 µL IPA (isopropyl alcohol) and transferred to an LC-MS vial with insert (Agilent 5182-0554 and 5183-2086) for analysis. Concurrently, a process blank sample and pooled quality control (QC) sample was prepared by taking equal volumes (~50 µL) from each sample after final resuspension.

Sampleprep ID:

SP003373

Sampleprep Summary:

Lipid extraction. Lipid extraction, based on Matyash et al.72, was performed as follows. All solutions were pre-chilled on ice. Tissues or mitochondrial pellets were transferred to labeled bead-mill tubes (1.4 mm, MoBio Cat# 13113-50) where lipids were extracted in a solution of 250 µL PBS, 225 µL MeOH containing internal standards, and 750 µL MTBE (methyl tert-butyl ether). Internal standards were Avanti SPLASH LipidoMix (Lot#12) at 10 µL per sample and Cambridge Isotope laboratories NSK-B and NSK-B-G1 (deuterated carnitines) at 10 µL per sample. The samples were homogenized in one 30 s cycle using the Omni Bead Ruptor followed by a rest on ice for 1 h. An addition of 188 µL PBS was made to induce phase separation. After centrifugation at 16,000 g for 5 minutes at 4 °C, the upper phases were collected and evaporated to dryness under a gentle nitrogen stream at room temperature. Lipid samples were reconstituted in 500 µL IPA (isopropyl alcohol) and transferred to an LC-MS vial with insert (Agilent 5182-0554 and 5183-2086) for analysis. Concurrently, a process blank sample and pooled quality control (QC) sample was prepared by taking equal volumes (~50 µL) from each sample after final resuspension.

Chromatography:

Chromatography ID:	CH004022
Chromatography Summary:	Positive Mode RP LCMS
Instrument Name:	Agilent 1290 Infinity
Column Name:	Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um)
Column Temperature:	65
Flow Gradient:	Started at 15% mobile phase B then increased to 30% B over 2.4 min. It sequentially increased to 48% B from 2.4 – 3.0 min, 82% B from 3 – 13.2 min, and 99% B from 13.2 – 13.8 min where it’s held until 16.7 min and returned to the initial conditions and equilibrated for 5 min.
Flow Rate:	0.4 mL min
Solvent A:	40% water/60% acetonitrile; 0.1% formic acid; 10 mM ammonium formate
Solvent B:	90% isopropanol/9% acetonitrile/1% water; 0.1% formic acid; 10 mM ammonium formate
Chromatography Type:	Reversed phase

Chromatography ID:	CH004023
Chromatography Summary:	Negative Mode RP LCMS
Instrument Name:	Agilent 1290 Infinity
Column Name:	Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um)
Column Temperature:	65
Flow Gradient:	Started at 15% mobile phase B then increased to 30% B over 2.4 min. It sequentially increased to 48% B from 2.4 – 3.0 min, 82% B from 3 – 13.2 min, and 99% B from 13.2 – 13.8 min where it’s held until 16.7 min and returned to the initial conditions and equilibrated for 5 min.
Flow Rate:	0.4 mL min
Solvent A:	40% water/60% acetonitrile; 10 mM ammonium formate
Solvent B:	90% isopropanol/9% acetonitrile/1% water; 10 mM ammonium acetate
Chromatography Type:	Reversed phase

Analysis:

Analysis ID:	AN005318
Analysis Type:	MS
Chromatography ID:	CH004022
Num Factors:	2
Num Metabolites:	192
Rt Units:	Minutes
Units:	pmol lipid per mg tissue

Analysis ID:	AN005319
Analysis Type:	MS
Chromatography ID:	CH004023
Num Factors:	2
Num Metabolites:	79
Rt Units:	Minutes
Units:	pmol lipid per mg tissue