Comparative Proteomics of Hepatocytes And Hepatic Cell Lines Using Swath-MS Reveals Significant Variations In...

Area of Science:

• Proteomics and Metabolomics
• Cellular Biology and Biochemistry

Background:

• Human hepatic carcinoma cell lines are vital in vitro models for studying liver metabolism and disease.
• Variability in protein expression between cell lines and primary human hepatocytes (cHH) challenges their translational validity.

Purpose of the Study:

• To quantitatively compare global protein abundance in HepaRG, HepG2, and Huh7 cell lines against cHH and human liver tissue homogenate (HLT).
• To assess the suitability of hepatic cell lines as in vitro models for drug metabolism and xenobiotic studies.

Main Methods:

• Utilized data-independent acquisition-based total protein approach (DIA-TPA) for global proteome quantification.
• Proteins from whole cell homogenates were digested using pressure-cycling technology (PCT).
• Sequential window acquisition of theoretical mass spectra (SWATH-MS) was performed, with quantification via Spectronaut™.

Main Results:

• Significant differences in global proteome were observed between cHH and hepatic cell lines.
• The HepaRG cell line demonstrated the highest correlation in global and ADME protein profiles with cHH.
• Quantifiable levels of clinically relevant drug-metabolizing enzymes (DMEs) like CYP450 and UGT families were found in HepaRG, cHH, and HLT.

Conclusions:

• Untargeted global proteomics effectively identifies protein expression differences among hepatic cell lines.
• The HepaRG cell line serves as a more representative model for studying hepatic lipid, xenobiotic metabolism, and glucose regulation compared to HepG2 and Huh7.
• This proteomic database aids in selecting appropriate cell lines for future research in liver biology and drug development.