Differential proteomics analysis of JEG-3 and JAR placental cell models and the effect of androgen treatment

Characterization of Xenobiotic and Steroid Disposition Potential of Human Placental Tissue and Cell Lines (BeWo, JEG-3, JAR, and HTR-8/SVneo) by Quantitative Proteomics

“…Consistent with our previous study (Kruger et al, 2022), JEG-3 cells also expressed BCRP albeit at lower levels than BeWo (1.4-fold higher than placenta). However, unlike our previous finding, the level of BCRP was below the limit of quantification in JAR cells which likely indicates inconsistent expression in this cell line.…”

“…For example, in this study we observed that BeWo, JEG-3, and JAR cells all expressed aromatase while HTR-8/SVneo cells did not. Even though both JAR and JEG-3 cells have been used to study the effects of xenobiotics on aromatase (Letcher et al, 1999;Mor et al, 2001;Zhang et al, 2011), our previous study showed that JEG-3 cells express CYP19A1 more consistently than JAR cells (Kruger et al, 2022). This means that results utilizing JAR cells to study CYP19A1 may fluctuate due to the inconsistent levels expressed within the cells.…”

“…Placenta tissue and cell lines were digested following a previously optimized procedure (Kruger et al, 2022). Briefly, 80 µg of protein sample suspended in 180 µL of 100 mM ABC was mixed with a solution containing 20 µL BSA (0.02 mg/mL), 10 µL DTT (250 mM), and 30 µL of ABC (100 mM).…”

“…A previously described LC-MS/MS method was used for proteomics data acquisition (Kruger et al, 2022). Briefly, the digested proteins were loaded onto an Easy Spray 1200 series nanoLC coupled Q-Exactive HF MS (Thermo Fisher Scientific, Waltham, MA) using a Thermo Scientific TM Acclaim TM PepMap TM 100 C18 HPLC column (75 µm x 250 mm) at 1 µg/µL with the column temperature of 35 °C and the mobile phase consisting of 0.1% formic acid in water (A) and 0.1% formic acid in 80% acetonitrile (B).…”

“…The suspension was centrifuged at 16000 xg for 10 min at 4 °C and 900 µL of the supernatant was collected. The Data visualization and statistical analysis: DDA data were analyzed using MaxQuant as previously described (Kruger et al, 2022), except the false discovery rate was set to 1% for both peptides and proteins. MaxQuant used the Andromeda search engine to search the human proteome database curated from Uniprot against the high-resolution MS/MS spectra.…”