Summary of Study ST003720
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 PR002309. The data can be accessed directly via it's Project DOI: 10.21228/M85J9C This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST003720 |
| Study Title | CD9-dependent extracellular vesicles mediate the propagation of ferroptosis |
| Study Summary | Ferroptosis is a form of regulated cell death that is mediated by accumulation of lipid peroxides. Emerging evidence suggests that ferroptosis as a cell-autonomous event can propagate to surrounding cells. Although a detailed mechanistic understanding of is still lacking, such a propagating feature of ferroptosis has far-reaching implications in developmental and pathological processes. Here, we show that the intercellular transmission of ferroptosis relies on an extracellular vesicle (EV)-dependent transport mechanism. Ferroptotic cells produce massive amounts of EVs carrying death signals. Proteomic analysis reveals that these EVs are CD9 positive and contain major components of the mitogen-activated protein kinase (MAPK) signaling. Further, untargeted lipidomics results show that EVs secreted from ferroptotic cells treated with RAS-selective lethal 3 (RSL3-EVs) and control EVs (DMSO-EVs) shared similar lipid composition. EV-transported MAP2K1/2 sensitizes target cells for ferroptosis through the action of peroxisome-proliferator activator receptor gamma (PPARG), a transcription factor that controls expression of various components of ferroptosis defense systems. Perturbation of EV-based transport protects adjacent tissue from ferroptosis in vivo. Our study thus uncovers an EV-dependent intercellular transport mechanism of ferroptosis that is mediated by the MAPK-responsive PPARG pathway. |
| Institute | Zhejiang University |
| Department | School of Medicine |
| Last Name | Li |
| First Name | Yue |
| Address | 866 Yuhangtang Road, Hangzhou, Zhejiang, 310058, China |
| liyue9104@zju.edu.cn | |
| Phone | (86)18869960769 |
| Submit Date | 2025-01-20 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2026-01-02 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002309 |
| Project DOI: | doi: 10.21228/M85J9C |
| Project Title: | CD9-dependent extracellular vesicles mediate the propagation of ferroptosis |
| Project Summary: | Ferroptosis is a form of regulated cell death that is mediated by accumulation of lipid peroxides. Emerging evidence suggests that ferroptosis as a cell-autonomous event can propagate to surrounding cells. Although a detailed mechanistic understanding of is still lacking, such a propagating feature of ferroptosis has far-reaching implications in developmental and pathological processes. Here, we show that the intercellular transmission of ferroptosis relies on an extracellular vesicle (EV)-dependent transport mechanism. Ferroptotic cells produce massive amounts of EVs carrying death signals. Proteomic analysis reveals that these EVs are CD9 positive and contain major components of the mitogen-activated protein kinase (MAPK) signaling. Further untargeted lipidomics results show that EVs secreted from ferroptotic cells (RSL3-EVs) and control EVs (DMSO-EVs) shared similar lipid composition. EV-transported MAP2K1/2 sensitizes target cells for ferroptosis through the action of peroxisome-proliferator activator receptor gamma (PPARG), a transcription factor that controls expression of various components of ferroptosis defense systems. Perturbation of EV-based transport protects adjacent tissue from ferroptosis in vivo. Our study thus uncovers an EV-dependent intercellular transport mechanism of ferroptosis that is mediated by the MAPK-responsive PPARG pathway. |
| Institute: | Zhejiang University |
| Department: | School of Medicine |
| Last Name: | Li |
| First Name: | Yue |
| Address: | 866 Yuhangtang Road, Hangzhou, Zhejiang, 310058, China |
| Email: | liyue9104@zju.edu.cn |
| Phone: | (86)18869960769 |
Subject:
| Subject ID: | SU003852 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Treatment |
|---|---|---|
| SA406882 | DMSO-EV-1 | DMSO |
| SA406883 | DMSO-EV-2 | DMSO |
| SA406884 | DMSO-EV-3 | DMSO |
| SA406885 | DMSO-EV-4 | DMSO |
| SA406886 | DMSO-EV-5 | DMSO |
| SA406887 | DMSO-EV-6 | DMSO |
| SA406888 | RSL3-EV-1 | RSL3 |
| SA406889 | RSL3-EV-2 | RSL3 |
| SA406890 | RSL3-EV-3 | RSL3 |
| SA406891 | RSL3-EV-4 | RSL3 |
| SA406892 | RSL3-EV-5 | RSL3 |
| SA406893 | RSL3-EV-6 | RSL3 |
| Showing results 1 to 12 of 12 |
Collection:
| Collection ID: | CO003845 |
| Collection Summary: | EVs secreted by DMSO- or RSL3-treated U-2 OS cells were harvested.The U-2 OS cell line is a human osteosarcoma cell line. RAS-selective lethal 3 (RSL3) is a drug that activates ferroptosis. Cells grew in DMEM medium till 60% confluency before being treated with 20 μM RSL3 or DMSO solvent for 18 hrs. Medium was removed and cells were washed before the incubation with plain DMEM medium for 24 hrs. Then conditioned culture media were collected and centrifuged at 4,500 × g for 30 min at 4°C. The supernatants were centrifuged at 10,000 g at 4°C for 1 hr. The pellets (EVs) were washed with PBS and centrifuged at 100,000 g at 4°C for 1 hr. The pellets (EVs) were resuspended in PBS. |
| Sample Type: | U-2 OS cell culture media |
Treatment:
| Treatment ID: | TR003861 |
| Treatment Summary: | U-2 OS cells were treated with 20 uM RSL3 or DMSO solvent for 18 hrs. Medium was removed and cells were incubated with fresh plain DMEM medium for 24 hrs before EV collection. |
Sample Preparation:
| Sampleprep ID: | SP003858 |
| Sampleprep Summary: | Lipid Extraction: A 200 μL sample was pipetted into an EP tube, mixed with 80 μL methanol and 400 μL MTBE, vortexed, and ultrasonicated at 5°C and 40 kHz for 30 min. After chilling at -20°C for 30 min, the mixture was centrifuged at 13,000 g at 4°C for 15 min. A 350 μL supernatant was collected, dried, and reconstituted in 100 μL of a 1:1 isopropanol:acetonitrile solution. The sample was vortexed for 30 s, then sonicated in an ice-water bath at 4°C and 40 kHz for 5 min. Following another centrifugation at 13,000 g at 4°C for 15 min, 2 μL of the clear supernatant was analyzed by UHPLC-MS/MS. |
Combined analysis:
| Analysis ID | AN006101 | AN006102 |
|---|---|---|
| Chromatography ID | CH004631 | CH004631 |
| MS ID | MS005807 | MS005808 |
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Thermo Vanquish | Thermo Vanquish |
| Column | Thermo Accucore C30 (100 x 2.1mm,2.6um) | Thermo Accucore C30 (100 x 2.1mm,2.6um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive HF-X Orbitrap | Thermo Q Exactive HF-X Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | log10(abundance) | log10(abundance) |
Chromatography:
| Chromatography ID: | CH004631 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Thermo Accucore C30 (100 x 2.1mm,2.6um) |
| Column Temperature: | 40 ℃ |
| Flow Gradient: | Linear gradient from 35% to 60%B, 0-4min; from 60% to 85% B, 4-12 min; from 85% to 100% B, 12-15 min; hold at 100% B, 15-17min; from 100% to 35% B, 17-18 min; hold at 35% B, until the end of separation. |
| Flow Rate: | 0.4 mL/min |
| Solvent A: | 50% acetonitrile/50% water; 10mM ammonium acetate; 0.1% formic acid |
| Solvent B: | 10% acetonitrile/88% isopropanol/2% water; 2mM ammonium acetate; 0.02% formic acid |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS005807 |
| Analysis ID: | AN006101 |
| Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometric data were acquired utilizing a Thermo UHPLC-Q-Exactive HF-X Benchtop Orbitrap Mass Spectrometer. The instrumental parameters were optimized as follows: sheath gas flow rate: 60 psi; auxiliary gas flow rate: 20 psi; auxiliary gas heater temperature: 370°C. The ion-spray voltage floating was adjusted to -3000 V for negative ion mode and +3000 V for positive ion mode. Rolling collision energies at 20-40-60 V. The Data Dependent Acquisition mode with a mass range of 200-2000 m/z were performed to collect data. Data Processing: The raw data were processed using LipidSearch (Thermo, CA) for peak detection, alignment and identification. The mass error was set to be smaller than 10 ppm. |
| Ion Mode: | POSITIVE |
| MS ID: | MS005808 |
| Analysis ID: | AN006102 |
| Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The mass spectrometric data were acquired utilizing a Thermo UHPLC-Q-Exactive HF-X Benchtop Orbitrap Mass Spectrometer. The instrumental parameters were optimized as follows: sheath gas flow rate: 60 psi; auxiliary gas flow rate: 20 psi; auxiliary gas heater temperature: 370°C. The ion-spray voltage floating was adjusted to -3000 V for negative ion mode and +3000 V for positive ion mode. Rolling collision energies at 20-40-60 V. The Data Dependent Acquisition mode with a mass range of 200-2000 m/z were performed to collect data. Data Processing: The raw data were processed using LipidSearch (Thermo, CA) for peak detection, alignment and identification. The mass error was set to be smaller than 10 ppm |
| Ion Mode: | NEGATIVE |
