High-Content Imaging and Transcriptomic Analyses of the Effects of Bisphenol S and Alternative Color Developers on KGN Granulosa Cells

Concerns about the adverse effects of bisphenol A (BPA), a chemical used for the production of polycarbonate plastics and epoxy resins as well as a color developer in thermal papers, have led to an increase in the use of 4,4-sulfonyldiphenol (bisphenol S; BPS), bis(3-allyl-4-hydroxyphenyl) sulfone (TGSA), 4-hydroxyphenyl 4-isoprooxyphenylsulfone (D-8), and [3-[(4-methylphenyl)sulfonylcarbamoylamino]phenyl] 4-methylbenzenesulfonate (Pergafast-201; PF-201), and 2,4-bis(phenylsulfonyl)phenol (DBSP) as alternative color developers. Data on these chemicals is scarce and little is known about their potential toxicity. We therefore determined the effects of BPS, TGSA, D-8, PF-201, and DBSP on the phenotype, function, and transcriptome of KGN human granulosa cells. Using high-content imaging, we observed that TGSA was the most cytotoxic compound tested, followed by D-8, DBSP, PF-201, and BPS. While the effects of these compounds on lysosomes, mitochondria, and oxidative stress were minimal, TGSA, D-8, and PF-201 drastically increased the number and total area of lipid droplets compared to control. RNA sequencing analyses revealed that TGSA and D-8 exposure differentially regulated 2,414 and 2,563 genes, respectively. PF-201 was the least transcriptionally active chemical, significantly affecting only six genes. Activation of pathways related to the extracellular matrix was the predominant effect of TGSA, while together, TGSA and D-8 commonly inhibited pathways involved in cell cycle regulation, DNA replication, and DNA repair. Such mechanisms may be underlying the cytotoxicity and lipid droplet accumulation observed in KGN cells. These data suggest that alternative color developers such as TGSA, D-8, and PF-201 may not be responsible replacements for BPA and BPS in thermal papers. Overall design: Total RNA was extracted from KGN human granulosa cells exposed for 48 h to the vehicle control (DMSO) or 30 µM of TGSA, D-8, or PF-201; n = 5.