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Proteomic Analysis of Extracellular Vesicles Derived from Human Pluripotent Stem Cells

ABSTRACT: Extracellular vesicles (EVs) and secretory factors are key to intercellular communication, yet their molecular dynamics in human pluripotent stem cells (hPSCs) are not well understood. We identified a key EV surface protein that regulates hPSC stemness, enhances growth, and promotes EV endocytosis. EV internalization helps reduce oxidative stress and cell death by transporting redox and stress response proteins, highlighting the importance of EV surface proteins in hPSC self-renewal and survival.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Embryo

SUBMITTER: Youngseok Lee

LAB HEAD: Jong-Hoon Kim

PROVIDER: PXD056373 | Pride | 2025-02-14

REPOSITORIES: pride

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Project description:Multi-omics integrates diverse types of biological information from genomic, proteomic, and metabolomics experiments to achieve a comprehensive understanding of complex cellular mechanisms. However, this approach is also challenging due to technical issues such as limited sample quantities, complexity of data pre-processing, and reproducibility concerns. Although conventional pre-processing methods in multi-omics research are standardized to ensure consistency; their simultaneous application can obscure specific details. Here, findings obtained from various omics approaches were profiled using various extraction methods (methanol extraction, Folch method, and Matyash methods for metabolites and lipids) and two digestion methods (Filter-aided sample preparation (FASP) and suspension traps (S-Trap)) for resuspended proteins. FASP was found to be more effective for separation of membrane-related proteins, whereas S-Trap excelled in isolating nuclear-related and RNA processing proteins. Thus, ASP may be suitable for investigating the immune response and bacterial infection pathways, whereas S-Trap may be more effective for studies focused on the mechanisms of neurodegenerative diseases. Moreover, the choice of extraction method, either single-phase MeOH or two-phase using Folch and Matyash methods, significantly influences the types of compounds identified, reflecting distinct profiles in different omics data sets. Among metabolites, the single-phase methd identified organic compounds and compounds related to fatty acids, whereas the two-phase extraction identified more hydrophilic compounds such as nucleotides. Lipids with strong hydrophobicity, such as ChE and TG, were identified in the two-phase extraction results. These findings highlight that significant differences between small molecules identified are primarily due to varying polarities of extraction solvents. To address human error and batch effects, a strategy that optimizes the balance between efficiency and the quality of the results is also proposed here. Our study reaffirms the impact of choice of pre-processing method in multi-omics, and also provides specific profiles of several protein and metabolite clusters as well as lipid classes.

			Action	DRS
	L22JSP2857.msf	Msf
	L22JSP2857.pdStudy	Other
	L22JSP2857_ANOVA.pdResult	Other
	L22JSP2857_EV1.raw	Raw
	L22JSP2857_EV2.raw	Raw

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Proteomic Analysis of Extracellular Vesicles Derived from Human Pluripotent Stem Cells

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