Alteration of the N⁶-Methyladenosine Epitranscriptomic Profile in Synthetic Phthalate-Treated Human Induced Pluripotent Stem Cell-Derived Endothelial Cells

Abstract

Background: This study aimed to characterize the N⁶-methyladenosine epitranscriptomic profile induced by mono(2-ethylhexyl) phthalate (MEHP) exposure using a human-induced pluripotent stem cell-derived endothelial cell model. Methods: A multiomic approach was employed by performing RNA sequencing in parallel with an N⁶-methyladenosine-specific microarray to identify mRNAs, lncRNAs, and miRNAs affected by MEHP exposure. Results: An integrative multiomic analysis identified relevant biological features affected by MEHP, while functional assays provided a phenotypic characterization of these effects. Transcripts regulated by the epitranscriptome were validated with quantitative PCR and methylated RNA immunoprecipitation. Conclusion: The authors’ profiling of the epitranscriptome expands the scope of toxicological insights into known environmental toxins to under surveyed cellular contexts and emerging domains of regulation and is, therefore, a valuable resource to human health.

Plain language summary

Synthetic phthalates, such as mono(2-ethyhexyl) phthalate, have long been recognized as environmental toxins. What effect these compounds have on endothelial cells remains poorly understood. To address this, the authors utilized a human-induced pluripotent stem cell-derived endothelial cell model to screen for an environmental toxin. They then obtained a profile of the epitranscriptomic changes involving the N⁶-methyladensosine modification and performed biochemical and functional assays. Overall, this study demonstrated how stem cell-based approaches can be used for toxicological screening and provided a valuable resource that profiles the epitranscriptomic response, which was complemented with RNA sequencing and functional and biochemical assays. This study provides relevant toxicological insights into the context of human health.

Keywords::

• epitranscriptomic
• iPSCs
• mono(2-ethylhexyl) phthalate
• N⁶-methyladenosine
• synthetic phthalate
• toxicology screening

Supplementary data

To view the supplementary data that accompany this paper please visit the journal website at: www.tandfonline.com/doi/suppl/10.2217/epi-2022-0110

Author contributions

J Jousma carried out bioinformatic analysis, data visualization, cellular and molecular experimental methods as well as manuscript preparation and revisions. SB Nukala contributed to cellular experimental methods. Z Han contributed to cellular experimental methods. G Yan and HHT Le carried out validation of statistical analysis. Y Kwon and Y Li contributed cellular experimental methods. WH Lee and SG Ong carried out project conceptualization, design, supervision, resources and manuscript review. All authors read and approved the final manuscript. SB Ong made intellectual contributions during review and preparation.

Financial & competing interests disclosure

This paper was supported and funded by the National Heart, Lung, and Blood Institute (NHLBI) T32 HL007829 (JJ), (NHLBI) R00 HL130416 (S-GO) and R01 HL148756 (S-GO). The authors also received funding from the American Heart Association (AHA), Postdoctoral Fellowship 917176 (ZH), AHA Scientist Development Grant 16SDG27560003 (WHL). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Data availability

RNA sequencing and N⁶-methyladenosine microarray data will be made publicly accessible upon request to the corresponding authors and will be available in the Gene Expression Omnibus database at the accession number GSE214420.

Additional information

Funding

This paper was supported and funded by the National Heart, Lung, and Blood Institute (NHLBI) T32 HL007829 (JJ), (NHLBI) R00 HL130416 (S-GO) and R01 HL148756 (S-GO). The authors also received funding from the American Heart Association (AHA), Postdoctoral Fellowship 917176 (ZH), AHA Scientist Development Grant 16SDG27560003 (WHL). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.