Abstract:
Persistent environmental pollutants (POPs), such as polybrominated diphenyl ethers (PBDEs), and polychlorinated biphenyls (PCBs), are ubiquitously detected in the environment despite removal from commercial and industrial use. Due to their lipophilic and bioaccumulative nature, POPs have been found in close proximity to the human environment, including food (i.e., milk, eggs, meat, seafood), soil and water. In humans, such exposures have been linked to metabolic diseases and developmental disorders. The liver is as an essential organ for xenobiotic metabolism and nutrient homeostasis. Specifically, the liver serves as a central hub for digestion and homeostatic signaling, metabolism of carbohydrates, amino acids, nucleotides, fatty acids, cholesterol, hormones, and importantly, xenobiotics such as drugs, and chemicals present in our environment such as POPs. Interacting with the liver, the gut microbiome can modify xenobiotic biotransformation activities of the host and dysbiosis of the gut microbiome is appreciated as a critical regulator of disease susceptibility. However, little is known regarding how environmental toxicants and the gut microbiome interact to modulate the interface between xenobiotic and intermediary metabolism. Furthermore, it is increasingly recognized that there is a sensitive developmental time window for toxic exposures that may have life-long impacts on disease risk. Therefore, the goal of my thesis is to investigate, in an unbiased way, the acute effects of PCBs on the gut microbiome and liver (Aim 1), as well as the persistent effects from acute exposure to PBDEs, or PCBs, as compared to bisphenol A (BPA), a well-known epigenetic modifier (Aim 2). I hypothesized that the gut microbiome regulates PCB-mediated changes in the liver’s metabolic signatures and transcriptome, and that acute exposure to PBDEs and PCBs have persistent effects on the liver transcriptome. To achieve Aim 1, ninety-day-old female conventional (CV) and germ-free (GF) C57BL/6 mice were orally exposed to the PCB Fox River Mixture (an environmentally relevant synthetic PCB mixture, 6 or 30 mg/kg, respectively representing the low and high dose) or corn oil (vehicle control, 10 ml/kg), once daily for 3 consecutive days. Organs were collected 24 hours after the final dose. RNA-Seq was conducted on liver, and endogenous aqueous metabolites (i.e. amino acids, carbohydrates, and nucleotides) were measured in liver and serum by LC-MS. For Aim 2, two-day-old male and female C57BL/6 mice were supralingually exposed to corn oil, BPA (250 μg/kg), BDE-99 (an enriched PBDE congener found in the diet and in human tissues, 57 mg/kg), or the PCB Fox River Mixture (30 mg/kg), once daily for three days. RNA-Seq was conducted in mouse livers at postnatal (PND) day 5 (acute response) and PND 60 (persistent response). The prototypical target genes of the major xenobiotic-sensing transcription factors aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR), and constitutive androstane receptor (CAR) were more readily up-regulated by PCBs in CV than in GF conditions, indicating that the effect of PCBs on the hepatic transcriptome act partly through the gut microbiome. Xenobiotic and steroid metabolism pathways were up-regulated, whereas the response to incorrectly folded proteins pathway (unfolded protein response – UPR; also commonly called the endoplasmic reticulum (ER) stress response) was down-regulated by PCBs in a gut microbiome-dependent manner. At the high PCB dose, NADP and arginine appear to interact with genes that encode drug-metabolizing enzymes (Cyp1-3 family, DhcR7, and Nqo1), which are highly correlated with the presence of Anaerotruncus and Roseburia bacteria in CV mice, providing a novel explanation of gut-liver interaction from PCB exposure. In GF exposure groups, hepatic glucose was down-regulated, whereas fructose 6-phosphate and glucose 6-phosphate were up-regulated, indicating increased glucose utilization potentiated by the lack of gut microbiota. Through querying the LINCS L1000 chemical database, I predicted that therapeutic drugs targeting the anti-inflammatory and ER stress pathways are potential remedies to mitigate PCB toxicity. Interestingly, as opposed to Aim 1, the chemical effects on the hepatic transcriptomic changes were minimal in PND5 and were markedly amplified by PND60 (young adults), indicating that early life toxicant exposure shifted the developmental trajectory of the liver. The acute response showed signs of active metabolism of the exposed contaminants; these responses were then greatly amplified in young adults. In these young adults, all three chemicals persistently altered the expression of hepatic genes involved in the metabolism of drugs, carbohydrates, and lipids, oxidative stress and inflammation, as well as epigenetic modifications, with males being more susceptible than females. At the doses given, BPA and BDE-99 had more prominent effects compared to PCBs. The top most persistently altered predicted upstream transcriptional and cell signaling regulators include essential cell cycle genes (Trp53, Cdkn1a, E2f1 and E2f3), sterol biosynthesis (Scap), bile acid synthesis (Fgf15), as well as the nuclear receptors that regulate lipids (PPARs) and drug metabolism (RORs, CAR, PXR). In accordance with these observations, other notable trends included persistent down-regulation of genes that encode phase-I and phase-II drug-metabolizing enzymes, up-regulation of many membrane transporters, and up-regulation of oxidative stress and inflammation-related genes in young adults exposed neonatally to BPA and BDE-99. In addition, the expression of cancer-related lncRNAs and genes (i.e. H19, Snhg1 and Snhg4, Gsts) were persistently dysregulated following neonatal exposure to BPA or BDE-99. In conclusion, my findings demonstrate that the gut microbiota dramatically influence PCB-mediated hepatic responses, possibly due to crosstalk between the gut and liver. Furthermore, my results demonstrate that neonatal exposure to these environmental chemicals (especially BDE-99 and BPA) leads to persistent and amplified hepatic transcriptomic changes in young adulthood, with males being more sensitive than females. Persistent dysbiosis in the colon and potentially altered gut-liver axis were observed. Most notably, dysregulation of cell cycle, inflammation, oxidative stress, as well as lipid and drug metabolism pathways were observed, possibly due to cross-talk between epigenetic reprogramming and nuclear receptors/transcription factors.
URI
http://hdl.handle.net/1773/45988
