Benzyl Butyl Phthalate Induced Early lncRNA H19 Regulation in C3H10T1/2 Stem Cell Line

Abstract

Exposure to endocrine-disrupting chemicals used in plastic manufacturing may contribute to the current obesity and diabetes epidemic. Our previous study demonstrated that benzyl butyl phthalate (BBP) induced adipogenesis in the C3H10T1/2 stem cell line. Here we investigated if BBP deregulated long noncoding RNA H19 and its downstream pathway and whether BBP plays a role in the insulin signaling pathway during adipocyte diiferentiation. Cells treated with an 8 day BBP regimen showed that H19 expression was decreased at day 2 with 50 μM BBP exposure (p < 0.05). However, no significant changes were observed from day 4 to day 8. Expression of miRNA-103/107, H19 regulated miRNAs, was upregulated at day 2 (p < 0.05) but not from day 4 to day 8. Similarly, expression of the let-7 family members (a, b, c, d, f, and g) was also significantly increased at day 2 (p < 0.05 or p < 0.01), except for let-7e. Both let-7 and miRNA-103/107 are targets of H19 and play roles in insulin signaling. Insulin receptor substrate (IRS)-1, one of the key insulin signal transduction regulators, was significantly downregulated from day 2 to day 8 (p < 0.05). Gene expression of insulin receptor (IR) and IRS-2 were not altered by BBP exposure. The ratio of IRS1/IRS2 was significantly decreased from day 2 to day 8. On day 4, phospho-Akt protein expression was significantly decreased (p < 0.05). In conclusion, BBP exposure may lead to metabolic dysregulation by altering vital epigenetic regulators such as lncRNA H19 and its target microRNAs at an earlier stage, which further regulates insulin signaling.

An Overview of Epigenetics in Obesity: The Role of Lifestyle and Therapeutic Interventions

“…Furthermore, phthalates were found to stimulate the expression of miR-34a-5p while decreasing the expression of its target genes, nicotinamide phosphoribosyltransferase (NAMPT) and sirtuin 1 (Sirt1), both of which are essential for energy homeostasis [53]. The lncRNA H19 and its downstream pathway related to insulin signaling were also altered in response to phthalates [54].…”

Application of In Vitro Models for Studying the Mechanisms Underlying the Obesogenic Action of Endocrine-Disrupting Chemicals (EDCs) as Food Contaminants—A Review

“…Results showed that exposure to butylparaben stimulated adipogenic differentiation via increased expression of PPARγ, C/EBPα and FABP4, as well as via decreased runt-related transcription factor 2 (RUNX2) mRNA levels, which plays an inhibitory role during adipocyte differentiation [ 130 ]. Other studies revealed that BBP induced adipocyte differentiation in C3H10T1/2 stem cells [ 137 , 138 ]. In turn, Zhang and Choudhury [ 138 ] showed that expression of the PPARγ and aP2 were significantly increased in C3H10T1/2 stem cells exposed to 50 µM BBP after 8 days of incubation.…”

“…In addition, decreased SIRT1 mRNA levels, as well as increased β-catenin and forkhead box protein O1 (FoxO1) acetylation under BBP exposure, were also associated with increased adipogenesis [ 138 ]. Moreover, the same authors [ 137 ] reported that 50 µM BBP significantly downregulated the expression of long non-coding H19 RNA and increased the expression of miR-103/107/let-7 (a, b, c, d, f and g) on day 2 of C3H10T1/2 cell differentiation, which probably stimulated adipogenesis. An adipogenic effect of DEHP, BPA, and TBT as a single compound was also observed in C3H10T1/2 cells by Biemann et al [ 139 ].…”

Epigenetic Mechanisms of Endocrine-Disrupting Chemicals in Obesity

“…In parallel, a decrease in the expression levels of Nampt and Sirt1 , two target genes of miR-34a-5p, is observed, along with another significant epigenetic regulator, Sirt3 [ 67 ]. Zhang and collaborators have demonstrated that BBP exposure may regulate insulin signaling by altering vital epigenetic regulators, such as long noncoding RNA H19, and their downstream pathways [ 70 ].…”