PXD056319-1

PXD056319 is an original dataset announced via ProteomeXchange.

Title	Comparison of serum-free cultivated HepG2 cells with cells cultivated under FBS containing conditions using proteomic analysis
Description	With an increasing focus in science, politics and society on animal welfare, human in vitro systems are gaining particular interest as potential replacement tools for animal experimentation. However, animal components are still commonly used in cell culture models, thereby compromising the true potential of in vitro systems as “pure” animal replacement tools. Especially fetal bovine serum (FBS) is heavily used in cell culture models, sparking debates about the ethical and scientific defensibleness of the usage of FBS in in vitro cell culture systems. In this study, we performed a comparative proteomic characterization of HepG2 cells adapted to serum-free conditions with cells cultivated under FBS-containing conditions. Four different serum-free media were used, of which two were previously shown to support long-term serum-free cultivation of HepG2 cells. Notably, while two media can be characterized as xeno-free, two media still contain animal components (bovine transferrin and/or bovine serum albumin). As we have previously shown increase sensitivity of serum-free cultivated HepG2 cells in cytotoxicity assays, as well as increased oxidative stress protection, we set a particular focus on drug-metabolizing enzymes and NRF2 regulated proteins. Using principal component analysis and pathways analysis, we identified nutrient related stress and energy generation as key factors for survival under serum-free conditions, either due to a decreased/altered availability of substrates for utilization or missing micronutrients. Furthermore, we observed predicted up-regulation of multiple pathways associated with drug metabolism and oxidative stress protection, including; “phase I – functionalization of compounds”, “phase II – conjugation of compounds” and “NRF2 mediated oxidative stress response”. We observed significant up-regulation of multiple members of the glutathione-s-transferase and glutathione peroxidase families and confirmed that increased antioxidant enzyme expression correlates with higher enzyme activity in vitro. These findings support our previously generated results derived from different toxicity assays and provide insights into the changes inflicted upon HepG2 cells by altered culturing conditions.
HostingRepository	PRIDE
AnnounceDate	2025-08-25
AnnouncementXML	Submission_2025-08-25_02:44:19.456.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Luisa-Marie Pfeifer
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	timsTOF Pro

Revision	Datetime	Status	ChangeLog Entry
0	2024-09-27 02:26:19	ID requested
⏵ 1	2025-08-25 02:44:20	announced

10.1021/acs.jproteome.5c00100;

Pfeifer LM, Singh J, Zimmer A, Krenkel M, Fischer F, Sensbach J, Pipp F, Werkmann D, Hewitt P, Comparative Proteomic Characterization of Serum-Free Cultivated HepG2 Cells Reveals Upregulated Drug Metabolism and Increased Oxidative Stress Protection. J Proteome Res, 24(8):3961-3978(2025) [pubmed]

submitter keyword: Human, HepG2 cells, FBS, serum-free

Dr. Philip Hewitt
contact affiliation	Merck Healthcare KGaA
contact email	Philip.hewitt@merckgroup.com
lab head
Luisa-Marie Pfeifer
contact affiliation	Merck Healthcare KGaA
contact email	luisa-marie.a.pfeifer@external.merckgroup.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2025/08/PXD056319

PRIDE project URI

• PRIDE
  1. PXD056319
     1. Label: PRIDE project
     2. Name: Comparison of serum-free cultivated HepG2 cells with cells cultivated under FBS containing conditions using proteomic analysis