Dataset Information

Proteomics reveals PFOS-induced alteration of lipid utilization and xenobiotic metabolism in a model of PFOS-augmented diet-induced fatty liver

ABSTRACT: Perfluorooctanesulfonic acid (PFOS) is a persistent, bio-accumulative pollutant that has been used for the last 60+ years in numerous industrial and commercial applications. In mice, PFOS administration is known to induce hepatomegaly and hepatic steatosis. The aim of the present study was to evaluate potential PFOS and diet interactions and explore the mechanism of PFOS induced liver lipid accumulation. Prior to PFOS administration, mice were fed either a standard chow diet (SD) or 60% kCal high fat diet (HFD) for 4 weeks to establish significant body weight increase. After 4 weeks of diet acclimation, the treatment groups received 0.0003% PFOS in diet for an additional 10 weeks. In addition, a subset of the mice fed HFD were switched to a SD (H-SD) to mimic weight-loss induced improvement of hepatic steatosis. A total of six treatment groups: i) SD, ii) HSD, iii) HFD (H), iv) SD +PFOS(SDP), v) H-SD +PFOS (HSDP), and vi) HFD +PFOS (HP) were included. PFOS and lipid concentrations were measured in both serum and liver. Relative liver mRNA expression was determined by targeted bead array and proteins were quantified using untargeted mass spectrometry. PFOS exposure increased liver weight, and in the HFD increased liver triglycerides and liver cholesterol content. Gene and protein expression in the liver demonstrated that PFOS exposure induced lipid utilization and xenobiotic metabolism pathways, and in a HFD, induced lipid synthesis. The data suggests that PFOS exposure acts on lipid utilization genes and exacerbates hepatic steatosis in mice fed a HFD.

INSTRUMENT(S):

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Liver

DISEASE(S): Metabolic Dysfunction-associated Steatotic Liver Disease

SUBMITTER: Emily Marques

LAB HEAD: Angela L. Slitt

PROVIDER: PXD016161 | Pride | 2020-09-30

REPOSITORIES: Pride

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Perfluorooctanesulfonic acid (PFOS) administration shifts the hepatic proteome and augments dietary outcomes related to hepatic steatosis in mice.

Marques Emily E Pfohl Marisa M Auclair Adam A Jamwal Rohitash R Barlock Benjamin J BJ Sammoura Ferass M FM Goedken Michael M Akhlaghi Fatemeh F Slitt Angela L AL

Toxicology and applied pharmacology 20200923

Hepatic steatosis increases risk of fatty liver and cardiovascular disease. Perfluorooctanesulfonic acid (PFOS) is a persistent, bio-accumulative pollutant that has been used in industrial and commercial applications. PFOS administration induces hepatic steatosis in rodents and increases lipogenic gene expression signatures in cultured hepatocytes. We hypothesized that PFOS treatment interferes with lipid loss when switching from a high fat diet (HFD) to a standard diet (SD), and augments HFD-in ...[more]

PMID: 32979393

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Project description:Background: Exposure to pollutants including the ubiquitous ‘forever chemical’, Perfluorooctane sulfonate (PFOS) has increasingly been associated with metabolic dysfunction-associated steatotic liver disease (MASLD). Recent epidemiological evidence has identified associations between Per- and polyfluoroalkyl substances (PFAS) exposure and increased liver injury in alcohol consumers, suggesting potential interactions between these exposures. However, the intersection of pollutant exposures and alcohol-associated liver disease (ALD) is not well studied. We hypothesize that pollutants may disrupt hepatic metabolism to modify ALD severity. Recently, we developed a two-hit (ethanol plus pollutant) mouse model, enabling testing of this hypothesis. Here, we elucidate the metabolic and disease-modifying effects of PFOS in this model. Methods: Male C57BL/6J mice were fed isocaloric control or 5% Ethanol (EtOH) Lieber-DeCarli diet for 15 days. From day 6 of feeding, mice were concurrently gavaged with 1 mg/kg PFOS or 2% tween-80 vehicle for 10 days, followed by a 5 g/kg EtOH binge dose and euthanized 5-6 hours later. Results: Approximately 60% of the administered PFOS dose accumulated in liver. PFOS exacerbated EtOH-induced hepatic steatosis and was associated by higher levels of plasma very low-density lipoprotein (vLDL) and alanine aminotransferase (ALT). PFOS upregulated hepatic ethanol-metabolizing enzymes and lowered blood alcohol levels. Ingenuity Pathway Analysis (IPA) Top Toxicity Functions/Lists associated with hepatic gene expression following PFOS co-exposure in EtOH-fed mice included: fatty acid metabolism and liver steatosis; nuclear receptor activation, cytochrome P450, and reactive oxygen species (ROS); apoptosis; liver fibrosis; and hepatocellular carcinoma (HCC). GO/KEGG analyses similarly revealed enrichment in fatty acid, xenobiotic, alcohol, or glutathione metabolic processes; and Peroxisome proliferator-activated receptor (PPAR) signaling. PFOS upregulated hepatic expression of several nuclear receptors (e.g., Pparα, Car, and Pxr) and their P450 target genes (e.g., Cyp4a10, Cyp2b10, and Cyp3a11) by RT-PCR or Western blot, confirming key IPA predictions. Conclusions: PFOS is a metabolism disrupting chemical that worsened ALD severity. PFOS activated hepatic nuclear receptors and enriched hepatic transcriptional pathways associated with steatosis, xenobiotic metabolism, oxidative stress, cell death, fibrosis, and HCC. These data demonstrate a novel mechanism whereby PFOS exacerbates ALD through coordinated dysregulation of lipid homeostasis and liver injury, potentially mediated by nuclear receptor activation. The identification of PFOS as an ALD risk modifier highlight the critical need to evaluate environmental pollutants as potential contributors to liver disease progression in vulnerable populations. More data are required on environmental pollution as a disease modifying factor in ALD.

Project description:Perfluorooctanesulfonate (PFOS) is a widespread environmental pollutant with a long half-life and clearly negative outcomes on metabolic diseases such as fatty liver and diabetes. Male and female Cyp2b-null and humanized CYP2B6-transgenic (hCYP2B6-Tg) mice were treated with 0, 1, or 10 mg/kg/day PFOS for 21 days, and surprisingly it was found that PFOS was retained at greater concentrations in the serum and liver of hCYP2B6-Tg mice than Cyp2b-null mice with greater differences in the females. Thus, Cyp2b-null and hCYP2B6-Tg mice provide new models for investigating individual mechanisms for PFOS bioaccumulation and toxicity. Overt toxicity was greater in hCYP2B6-Tg mice (especially females) as measured by mortality; however, steato-sis occurred more readily in Cyp2b-null mice despite the lower PFOS liver concentrations. Tar-geted lipidomics and transcriptomics from PFOS treated Cyp2b-null and hCYP2B6-Tg mouse livers were performed and compared to PFOS retention and serum markers of toxicity by PCA. Several oxylipins, including prostaglandins, thromboxanes, and docosahexaenoic acid metabo-lites are associated or inversely associated with PFOS toxicity. Both lipidomics and tran-scriptomics indicate PFOS toxicity is associated with PPAR activity in all models. GO terms as-sociated with reduced steatosis were sexually dimorphic with lipid metabolism and transport increased in females and circadian rhythm associated genes increased in males. However, we can rule out that steatosis was initially protective from PFOS toxicity. Moreover, several transporters are associated with increased retention probably due to increased uptake. The strongest associa-tions are the organic anion transport proteins (Oatp1a4-6) genes and a long-chain fatty acid transport protein (fatp1), enriched in female hCYP2B6-Tg mice. PFOS uptake was also reduced in cultured murine hepatocytes by OATP inhibitors. The role of OATP1A6 and FATP1 in PFOS transport has not been tested. In summary, Cyp2b-null and hCYP2B6-Tg mice provided unique models for estimating the importance of novel mechanisms in PFOS retention and toxicity.

Project description:This series was used for two studies: Study 1: Recently it was discovered that the perfluorooctane sulfonate (PFOS) detected in wildlife, such as fish-eating birds, had a greater proportion of linear PFOS (L-PFOS) than the manufactured technical product (T-PFOS), which contains linear and branched isomers. This suggests toxicological studies based on T-PFOS data may inaccurately assess exposure risk to wildlife. To determine if PFOS effects were influenced by isomer content we compared the transcriptional profiles of cultured chicken embryonic hepatocytes (CEH) exposed to either L-PFOS or T-PFOS using Agilent microarrays. At equal concentrations (10 ?M), T-PFOS altered the expression of more transcripts (340, >1.5 fold change, p<0.05) compared to L-PFOS (130 transcripts). Higher concentrations of L-PFOS (40 ?M) were also less transcriptionally disruptive (217 transcripts) than T-PFOS at 10 ?M. Functional analysis showed that L-PFOS and T-PFOS affected genes involved in lipid metabolism, hepatic system development and cellular growth and proliferation. Pathway and interactome analysis suggested that genes may be affected through the RXR receptor, oxidative stress response, TP53 signaling, MYC signaling, Wnt/?-catenin signaling and PPAR? and SREBP receptors. In all functional categories and pathways examined, the response elicited by T-PFOS was greater than L-PFOS. These data show that T-PFOS elicits a greater transcriptional response in CEH than L-PFOS alone and demonstrates the importance of considering the isomer-specific toxicological properties of PFOS when assessing exposure risk. Study 2: In many bird populations, concentrations of perfluoroundanoic acid (PFUdA) are second only to perfluorooctane sulfonate (PFOS) among perfluoroalkyl compounds. Here, we used microarrays to characterize the transcriptional response of cultured chicken embryonic hepatocytes (CEH) to PFUdA and compared it to the response induced by PFOS. At non-cytotoxic doses, PFUdA (1 or 10 ?M) disrupted the expression of more genes (854) than PFOS (447, at 10 or 40 ?M) in CEH. Using functional, pathway and interactome analysis we identified several potentially important modes-of-action (MoAs) for PFUdA and some associated key events, including the suppression of the acute-phase response (APR) through peroxisome proliferator activated receptor activation. We then measured the expression of five APR genes, fibrinogen alpha (fga), fibrinogen gamma (fgg), thrombin (f2), plasminogen (plg), and protein C (proC), in the liver of chicken embryos exposed in ovo to PFUdA. The expression of fga, f2, and proC were down-regulated in embryo livers (100 or 1000 ng/g, p<0.1) as predicted from microarray analysis, whereas fibrinogen gamma (fgg) was up-regulated and plg was not significantly affected. Our results demonstrate PFUdA is more transcriptionally disruptive than PFOS in CEH. Additionally, we identified APR suppression as a potentially important and environmentally relevant MoA. These findings suggest in ovo exposure of birds to PFUdA could lead to post-hatch developmental deficiencies, such as impaired immune response.