A Systems Biology Approach to Investigate Human Lung Cell Response to Air Pollutants

Exposure to air pollution is associated with many diseases, such as asthma, bronchitis, and lung cancer. Despite these adverse health effects, the cellular mechanisms underlying air pollution-associated diseases remain largely unknown. In this study we set out to investigate the genome-wide responses of human lung cells exposed to multiple air pollutant conditions. We first employ a toxicogenomic approach to compare transcripts and molecular networks modulated upon exposure to freshly emitted air pollutants and photochemically altered pollutant mixtures, containing secondary pollutants. The results demonstrate that secondary pollutants initiate a more robust genomic response. After identifying this trend, we investigate potential mechanisms underlying responses to individual secondary pollutants. Here, we evaluate global microRNA expression modifications resulting from formaldehyde exposure. Our analysis reveals that formaldehyde induces significant changes in microRNA levels, which may in turn, regulate genes associated with inflammation and cancer. Together, these investigations reveal novel mechanisms potentially underlying air pollutant-induced disease

A Systems Biology-Based Approach to Investigate Formaldehyde’s Effects on MicroRNA Expression Profiles

Formaldehyde is a common indoor and outdoor air pollutant that adversely impacts global public health. Many toxicological studies have shown that formaldehyde causes nasopharyngeal cancer, possibly through tissue damage, increased cell proliferation, and/or DNA damage. However, there is lack of knowledge regarding formaldehyde's effects at the systems biology level and whether epigenetic mechanisms may contribute to cellular responses. Furthermore, whether formaldehyde is capable of altering genomic and epigenomic processes throughout sites distal to the respiratory tract is unknown. This topic is of high interest, as the link between formaldehyde inhalation exposure and leukemia development is currently under heated debate. Epidemiological studies have shown evidence supporting a link between formaldehyde exposure and leukemia development, while toxicological investigations have yet to provide evidence supporting formaldehyde's ability to influence sites distant to the respiratory tract. Before this dispute is resolved, further evaluation of the biological mechanisms linking formaldehyde to disease is clearly necessary. In particular, formaldehyde-induced changes to epigenetic contributors to transcriptional programs are extremely understudied, where microRNA (miRNA) expression profiles have yet to be investigated in relation to formaldehyde. We set out to test the novel hypothesis that miRNAs have altered expression profiles within the respiratory and hematopoietic systems upon exposure to formaldehyde. Our studies were the first to show that formaldehyde significantly disrupts miRNA expression patterns in vitro, within cultured human lung cells, and in vivo, within the nasal epithelium of nonhuman primates. Using a rodent model, the impact of formaldehyde exposure on miRNA-related processes in direct contact and distant tissues, including the nasal mucosa, circulating white blood cells, and bone marrow, was evaluated. Formaldehyde was found to significantly alter miRNA expression profiles within the nose and blood, but not the bone marrow. Evaluating the epigenetic effects of formaldehyde exposure at the systems biology level, putative miRNA-mediated responses were mapped onto interacting networks. Signaling related to inflammation, cell death, and cancer was identified as enriched. Taken together, our research increases the knowledge of under-studied mechanisms linking formaldehyde exposure to disease, acting as an important foundation for future research in public health and toxicology.Doctor of Philosoph

A Toxicogenomic Comparison of Primary and Photochemically Altered Air Pollutant Mixtures

Background: Air pollution contributes significantly to global increases in mortality, particularly within urban environments. Limited knowledge exists on the mechanisms underlying health effects resulting from exposure to pollutant mixtures similar to those occurring in ambient air. In order to clarify the mechanisms underlying exposure effects, toxicogenomic analyses are used to evaluate genomewide transcript responses and map these responses to molecular networks

Systems Biology and Birth Defects Prevention: Blockade of the Glucocorticoid Receptor Prevents Arsenic-Induced Birth Defects

Background: The biological mechanisms by which environmental metals are associated with birth defects are largely unknown. Systems biology–based approaches may help to identify key pathways that mediate metal-induced birth defects as well as potential targets for prevention

A role for microRNAs in the epigenetic control of sexually dimorphic gene expression in the human placenta

Aim: The contribution of miRNAs as epigenetic regulators of sexually dimorphic gene expression in the placenta is unknown. Materials & methods: 382 placentas from the extremely low gestational age newborns (ELGAN) cohort were evaluated for expression levels of 37,268 mRNAs and 2,102 miRNAs using genome-wide RNA-sequencing. Differential expression analysis was used to identify differences in the expression based on the sex of the fetus. Results: Sexually dimorphic expression was observed for 128 mRNAs and 59 miRNAs. A set of 25 miRNA master regulators was identified that likely contribute to the sexual dimorphic mRNA expression. Conclusion: These data highlight sex-dependent miRNA and mRNA patterning in the placenta and provide insight into a potential mechanism for observed sex differences in outcomes

High-Throughput Screening Data Interpretation in the Context of In Vivo Transcriptomic Responses to Oral Cr(VI) Exposure

The toxicity of hexavalent chromium [Cr(VI)] in drinking water has been studied extensively, and available in vivo and in vitro studies provide a robust dataset for application of advanced toxicological tools to inform the mode of action (MOA). This study aimed to contribute to the understanding of Cr(VI) MOA by evaluating high-throughput screening (HTS) data and other in vitro data relevant to Cr(VI), and comparing these findings to robust in vivo data, including transcriptomic profiles in target tissues. Evaluation of Tox21 HTS data for Cr(VI) identified 11 active assay endpoints relevant to the Ten Key Characteristics of Carcinogens (TKCCs) that have been proposed by other investigators. Four of these endpoints were related to TP53 (tumor protein 53) activation mapping to genotoxicity (KCC#2), and four were related to cell death/proliferation (KCC#10). HTS results were consistent with other in vitro data from the Comparative Toxicogenomics Database. In vitro responses were compared to in vivo transcriptomic responses in the most sensitive target tissue, the duodenum, of mice exposed to ≤ 180 ppm Cr(VI) for 7 and 90 days. Pathways that were altered both in vitro and in vivo included those relevant to cell death/proliferation. In contrast, pathways relevant to p53/DNA damage were identified in vitro but not in vivo. Benchmark dose modeling and phenotypic anchoring of in vivo transcriptomic responses strengthened the finding that Cr(VI) causes cell stress/injury followed by proliferation in the mouse duodenum at high doses. These findings contribute to the body of evidence supporting a non-mutagenic MOA for Cr(VI)-induced intestinal cancer

Chemical Mixtures in Household Environments: In Silico Predictions and In Vitro Testing of Potential Joint Action on PPARγ in Human Liver Cells

There are thousands of chemicals that humans can be exposed to in their everyday environments, the majority of which are currently understudied and lack substantial testing for potential exposure and toxicity. This study aimed to implement in silico methods to characterize the chemicals that co-occur across chemical and product uses in our everyday household environments that also target a common molecular mediator, thus representing understudied mixtures that may exacerbate toxicity in humans. To detail, the Chemical and Products Database (CPDat) was queried to identify which chemicals co-occur across common exposure sources. Chemicals were preselected to include those that target an important mediator of cell health and toxicity, the peroxisome proliferator activated receptor gamma (PPARγ), in liver cells that were identified through query of the ToxCast/Tox21 database. These co-occurring chemicals were thus hypothesized to exert potential joint effects on PPARγ. To test this hypothesis, five commonly co-occurring chemicals (namely, benzyl cinnamate, butyl paraben, decanoic acid, eugenol, and sodium dodecyl sulfate) were tested individually and in combination for changes in the expression of PPARγ and its downstream target, insulin receptor (INSR), in human liver HepG2 cells. Results showed that these likely co-occurring chemicals in household environments increased both PPARγ and INSR expression more significantly when the exposures occurred as mixtures vs. as individual chemicals. Future studies will evaluate such chemical combinations across more doses, allowing for further quantification of the types of joint action while leveraging this method of chemical combination prioritization. This study demonstrates the utility of in silico-based methods to identify chemicals that co-occur in the environment for mixtures toxicity testing and highlights relationships between understudied chemicals and changes in PPARγ-associated signaling

Placental genomic and epigenomic signatures associated with infant birth weight highlight mechanisms involved in collagen and growth factor signaling

Birth weight (BW) represents an important clinical and toxicological measure, indicative of the overall health of the newborn as well as potential risk for later-in-life outcomes. BW can be influenced by endogenous and exogenous factors and is known to be heavily impacted in utero by the health and function of the placenta. An aspect that remains understudied is the influence of genomic and epigenomic programming within the placenta on infant BW. To address this gap, we set out to test the hypothesis that genes involved in critical placental cell signaling are associated with infant BW, and are likely regulated, in part, through epigenetic mechanisms based on microRNA (miRNA) mediation. This study leveraged a robust dataset based on 390 infants born at low gestational age (ranged 23-27 weeks) to evaluate genome-wide expression profiles of both mRNAs and miRNAs in placenta tissues and relate these to infant BW. A total of 254 mRNAs and 268 miRNAs were identified as associated with BW, the majority of which showed consistent associations across placentas derived from both males and females. BW-associated mRNAs were found to be enriched for important biological pathways, including glycoprotein VI (the major receptor for collagen), human growth, and hepatocyte growth factor signaling, a portion of which were predicted to be regulated by BW-associated miRNAs. These miRNA-regulated pathways highlight key mechanisms potentially linking endogenous/exogenous factors to changes in birth outcomes that may be deleterious to infant and later-in-life health

Metal mixtures modeling identifies birth weight-associated gene networks in the placentas of children born extremely preterm

Prenatal exposure to toxic metals is linked to numerous adverse birth and later-in-life outcomes. These outcomes are tied to disrupted biological processes in fetal-derived tissues including the placenta and umbilical cord yet the precise pathways are understudied in these target tissues. We set out to examine the relationship between metal concentrations in umbilical cord and altered gene expression networks in placental tissue. These novel relationships were investigated in a subset of the Extremely Low Gestational Age Newborn (ELGAN) cohort (n = 226). Prenatal exposure to 11 metals/metalloids was measured using inductively coupled plasma tandem-mass spectrometry (ICP-MS/MS) in cord tissue, ensuring passage through the placental barrier. RNA-sequencing was used to quantify >37,000 mRNA transcripts. Differentially expressed genes (DEGs) were identified with respect to each metal. Weighted gene co-expression analysis identified gene networks modulated by metals. Two innovative mixtures modeling techniques, namely principal components analysis and quantile-based g-computation, were employed to identify genes/gene networks associated with multi-metal exposure. Individually, lead was associated with the strongest genomic response of 191 DEGs. Joint lead and cadmium exposure was related to 657 DEGs, including DNA Methyl Transferase 1 (DNMT1). These genes were enriched for the Eukaryotic Initiation Factor 2 (EIF2) pathway. Four gene networks, each containing genes within a Nuclear Factor kappa-light-chain-enhancer of Activated B Cells (NF-kB)-mediated network, were significantly increased in average expression level in relation to increases in all metal concentrations. All four of these metal mixture-associated gene networks were negatively correlated with important predictors of neonatal health including birth weight, placenta weight, and fetal growth. Bringing together novel methodologies from epidemiological mixtures analyses and toxicogenomics, applied to a unique cohort of extremely preterm children, the present study highlighted critical genes and pathways in the placenta dysregulated by prenatal metal mixtures. These represent potential mechanisms underlying the developmental origins of metal-induced disease

Pre-pregnancy BMI-associated miRNA and mRNA expression signatures in the placenta highlight a sexually-dimorphic response to maternal underweight status

Pre-pregnancy body mass index (BMI) is associated with adverse pregnancy and neonatal health outcomes, with differences in risk observed between sexes. Given that the placenta is a sexually dimorphic organ and critical regulator of development, examining differences in placental mRNA and miRNA expression in relation to pre-pregnancy BMI may provide insight into responses to maternal BMI in utero. Here, genome-wide mRNA and miRNA expression levels were assessed in the placentas of infants born extremely preterm. Differences in expression were evaluated according to pre-pregnancy BMI status (1) overall and (2) in male and female placentas separately. Overall, 719 mRNAs were differentially expressed in relation to underweight status. Unexpectedly, no genes were differentially expressed in relation to overweight or obese status. In male placentas, 572 mRNAs were associated with underweight status, with 503 (70%) overlapping genes identified overall. Notably, 43/572 (8%) of the mRNAs associated with underweight status in male placentas were also gene targets of two miRNAs (miR-4057 and miR-128-1-5p) associated with underweight status in male placentas. Pathways regulating placental nutrient metabolism and angiogenesis were among those enriched in mRNAs associated with underweight status in males. This study is among the first to highlight a sexually dimorphic response to low pre-pregnancy BMI in the placenta