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image of Comparative Proteomics Of Hepatocytes And Hepatic Cell Lines Using Swath-MS

Abstract

Introduction

Human hepatic carcinoma cell lines are widely used to study lipid and xenobiotic metabolism, as well as glucose regulation in both normal and diseased states. However, their validity is often questioned due to variability in protein expression compared to primary human hepatocytes (cHH). This study aimed to quantify protein abundance in various hepatic cell lines versus cHH and human liver tissue homogenate (HLT) using a data-independent acquisition-based total protein approach (DIA-TPA). We compared the global proteome from the whole cell homogenates of HepaRG, HepG2, and Huh7 cell lines with that of cHH and HLT.

Methods

Proteins in whole cell homogenates were digested in solution using pressure-cycling technology (PCT). DIA was performed via sequential window acquisition of theoretical mass spectra (SWATH-MS), and MS2 spectra were quantified using Spectronaut™, followed by analysis with TPA.

Results

We identified 2715, 2578, 2874, 2717, and 3083 proteins in HepaRG, HepG2, Huh7, cHH, and HLT, respectively, at a 1% FDR. The global proteome of cHH significantly differed from that of the cancer hepatic cell lines. Among the cell lines, the global and ADME protein profile of HepaRG most closely correlated with cHH, with 89 out of 101 ADME proteins identified. Clinically relevant DMEs from the CYP450 family (CYP2C9, CYP2C19, CYP2D6, and CYP3A4) and the UGT family (UGT1A1, UGT1A3, UGT1A6, UGT2B7, and UGT2B15) were quantifiable in human hepatocytes, human liver tissue, and the HepaRG cell line. The Huh7 cell line exhibited a higher abundance of proteins related to gluconeogenesis and glycolysis compared to other groups.

Conclusion

This study highlights the potential of untargeted global proteomics in detecting differences in protein expression among various hepatic cell lines and provides a comprehensive database to inform the choice of the cell line in future studies.

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2026-01-22
2026-01-31

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Comparative Proteomics Of Hepatocytes And Hepatic Cell Lines Using Swath-MS
Bentham Science Publishers ; https://doi.org/10.2174/0113892002403596251122091342
/content/journals/cdm/10.2174/0113892002403596251122091342
/content/journals/cdm/10.2174/0113892002403596251122091342
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  • Article Type:     Research Article
Keywords: proteomics ; drug metabolizing enzymes ; CYP450 ; HepaRG ; human hepatocytes ; SWATH-MS ; Huh7 ; HepG2

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