Abstract 5573: Effects of endocrine disruptors on rat mammary transcriptome at critical developmental windows
Authors: Gopalakrishnan Kalpana , Li Qian , Ma Yula , Lambertini Luca , Chen Jia , Teitelbaum Susan , Manservisi Fabiana , Falcioni Laura , Bua Luciano , Belpoggi Fiorella 
Published: August 2015

Abstract
Background: Endocrine disruptors (EDs) are environmental chemicals that may disrupt endogenous hormones and have been linked to many adverse health outcomes including breast cancer. EDs have been shown to modulate the risk for mammary tumors in animal models, but their mechanisms at biologically relevant doses and during critical developmental windows remain poorly understood.
Methods: We examined the individual and combinatorial (MIX) effects of three EDs commonly found in personal care products - diethyl phthalate (DEP), methyl paraben (MPB) and triclosan (TCS). Sprague-Dawley rats were exposed at four developmental windows - prenatal, neonatal, prepubertal and pubertal - at doses comparable to human exposure based on urinary levels reported in NHANES. Whole-transcriptome profiling was done using Affymetrix rat gene 2.0 st arrays.
Results: Differential gene expression analysis by limma revealed that there were no substantial differences between the transcriptomes of prenatal and neonatal rats, so they were combined into a ‘perinatal’ group. Control rats: Expression levels of 702 transcripts significantly differed between the prepubertal and perinatal periods; gene ontology (GO) enrichment showed that GO terms related to development and response to estrogen stimuli were up-regulated and immune signaling pathways were down-regulated. Upstream analysis using Ingenuity Pathway Analysis showed that β-estradiol and estrogen receptor were among the top regulators up-regulated in the prepubertal window, and T-cell receptor and interleukins were among the top regulators down-regulated. The difference between later windows, i.e., between puberty and prepuberty was much less pronounced, with only 39 differentially expressed transcripts. ED-exposed rats: Large scale changes in mammary transcriptome were driven by developmental stage, while changes due to ED exposure appeared to be more subtle. One-way ANOVA among developmental windows demonstrated that only ∼3000 transcripts were significantly changed in DEP and MPB and ∼4000 transcripts changed in TCS and MIX compared to ∼6000 differentially expressed transcripts in control rats. Importantly, distinct signatures of genes were associated with each individual ED as well as MIX, suggesting distinctive mechanisms. Additionally, our data suggested that the pubertal window was the most susceptible; for example, 440 transcripts were differentially expressed in the MPB group relative to control only at the pubertal window and GO terms related to DNA damage and ion transport were up- and down-regulated, respectively.
Conclusions: Exposure to common EDs at levels comparable to human exposure resulted in measurable changes in mammary transcriptome and reveal novel mechanisms of ED action in mammary development. Findings from our study highlight the importance of taking into account the timing of exposure when studying the relationship between environment and disease.
Citation Format: Kalpana Gopalakrishnan, Qian Li, Yula Ma, Luca Lambertini, Jia Chen, Susan Teitelbaum, Fabiana Manservisi, Laura Falcioni, Luciano Bua, Fiorella Belpoggi. Effects of endocrine disruptors on rat mammary transcriptome at critical developmental windows. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5573. doi:10.1158/1538-7445.AM2015-5573