Regulatory Genomics of Ozone Air Pollution Response in Vitro and In Vivo

体外和体内臭氧空气污染响应的监管基因组学

基本信息

批准号：
10467348
负责人：
Samir Kelada
金额：
$ 64.55万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-18 至 2027-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10467348
关键词：
ATAC-seq Acute Address Adverse effects Affect Air Air Pollutants Air Pollution Alleles Applications Grants Asthma Binding Biological Biological Models Black race CRISPR/Cas technology Candidate Disease Gene Cardiopulmonary Cell model Cells Cellular biology Chromatin Complex DNA-Binding Proteins Data Data Set Development Epithelial Cells Exhibits Exposure to GSTM1 gene Gene Expression Gene Expression Regulation Genes Genetic Genetic Determinism Genetic Transcription Genome Genomics Genotype Goals Human Human Volunteers IL8 gene In Vitro Individual Individual Differences Inflammation Inflammatory Link Lipid Peroxidation Lipid Peroxides Lipids Liquid substance Lung Lung diseases MUC5AC gene Maps Measures Mediating Mediation Modeling Molecular Morbidity - disease rate Multiomic Data Natural Immunity Neutrophil Infiltration Oxidative Stress Ozone Participant Persons Phenotype Predisposition Prevention Production Public Health Quantitative Trait Loci Reaction Reactive Oxygen Species Regulator Genes Regulatory Element Reporter Genes Reproducibility Research Personnel Respiratory Disease Respiratory System Rest Role Sampling Single Nucleotide Polymorphism Specificity Structure of parenchyma of lung Suggestion Testing Tissue Donors Toxicology Validation Variant Work adverse outcome air filter airway epithelium airway inflammation asthmatic bronchial epithelium causal model cell type chemokine cytokine cytotoxicity data integration epigenome epigenomics epithelial injury exposed human population gene environment interaction genetic variant in vivo innovation insight interest knock-down mortality multiple omics neutrophil novel ozone exposure pulmonary function response sex single-cell RNA sequencing trait transcription factor transcriptome sequencing

ATAC-seq Acute Address Adverse effects Affect Air Air Pollutants Air Pollution Alleles Applications Grants Asthma Binding Biological Biological Models Black race CRISPR/Cas technology Candidate Disease Gene Cardiopulmonary Cell model Cells Cellular biology Chromatin Complex DNA-Binding Proteins Data Data Set Development Epithelial Cells Exhibits Exposure to GSTM1 gene Gene Expression Gene Expression Regulation Genes Genetic Genetic Determinism Genetic Transcription Genome Genomics Genotype Goals Human Human Volunteers IL8 gene In Vitro Individual Individual Differences Inflammation Inflammatory Link Lipid Peroxidation Lipid Peroxides Lipids Liquid substance Lung Lung diseases MUC5AC gene Maps Measures Mediating Mediation Modeling Molecular Morbidity - disease rate Multiomic Data Natural Immunity Neutrophil Infiltration Oxidative Stress Ozone Participant Persons Phenotype Predisposition Prevention Production Public Health Quantitative Trait Loci Reaction Reactive Oxygen Species Regulator Genes Regulatory Element Reporter Genes Reproducibility Research Personnel Respiratory Disease Respiratory System Rest Role Sampling Single Nucleotide Polymorphism Specificity Structure of parenchyma of lung Suggestion Testing Tissue Donors Toxicology Validation Variant Work adverse outcome air filter airway epithelium airway inflammation asthmatic bronchial epithelium causal model cell type chemokine cytokine cytotoxicity data integration epigenome epigenomics epithelial injury exposed human population gene environment interaction genetic variant in vivo innovation insight interest knock-down mortality multiple omics neutrophil novel ozone exposure pulmonary function response sex single-cell RNA sequencing trait transcription factor transcriptome sequencing

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项目摘要

Project Summary Exposure to the ambient air pollutant ozone (O3) is associated with cardiopulmonary morbidity and mortality, rendering it an important public health issue. Controlled exposure studies show that acute O3 exposure causes airway inflammation, epithelial injury, and a transient decrease in lung function. These studies have also demonstrated that subjects exhibit highly reproducible differences in O3 response, suggestive of gene-by- environment interactions (GxE). Candidate gene studies have provided evidence of GxE for a handful of genes, however, the role of genetic variants in the rest of the genome is largely unknown. This data gap limits our ability to identify susceptible individuals and gain insight into mechanisms by which O3 causes adverse effects. Here, we put forth a proposal to address this data gap using human bronchial epithelial cells (hBECs) in vitro. hBECs are the first cells of the respiratory tract to interact with O3, and we have shown that hBECs exposed to O3 in vitro upregulate the expression of key pro-inflammatory genes (e.g., CXCL8), mirroring the in vivo response. We hypothesize that variation in O3-induced inflammation is associated with differences in hBEC gene expression, and that inter-individual differences in gene expression at baseline and after O3 have a genetic basis, i.e., are expression quantitative trait loci (eQTL). Further, we hypothesize that some eQTL are caused by single nucleotide polymorphisms (SNPs) that affect chromatin accessibility (caQTL). In Aim 1, we will establish well- differentiated hBEC cultures, grown at air-liquid interface, from 300 banked lung tissue donors of both sexes and diverse ancestries, then expose them to O3 vs. filtered air (FA) and measure key hBEC O3 response phenotypes (e.g. IL-8 production, oxidative stress, lipid peroxidation, barrier function, and cytotoxicity). We will profile gene expression in FA and O3-exposed hBECs using both bulk RNA-seq and single cell RNA-seq to identify O3- induced/repressed genes and their cell-type specificity. After genotyping, we will map eQTL at baseline (mRNAFA), response eQTL (mRNAO3-mRNAFA), and QTL for all hBEC O3 response phenotypes, then use mediation analyses to identify SNPs and genes fitting a putative causal model: O3+SNP → [mRNA] → hBEC O3 response phenotype. In Aim 2, we will perform ATAC-seq to characterize how O3 alters chromatin accessibility in hBECs, then map baseline and response caQTL. We will perform multi-omic data integration (eQTL, caQTL, QTL) to identify gene regulatory models of O3 response, i.e., O3+SNP→chromatin accessibility→[mRNA]→hBEC O3 response phenotype. Finally, in Aim 3, we will validate novel genes and gene regulatory mechanisms underlying variation in O3 response in vitro and in vivo. We will determine how key SNPs affect gene regulation and whether knocking down the corresponding genes alters O3 response in vitro. For in vivo validation, we will test for association between SNPs of interest and O3-induced neutrophil recruitment in a dataset of 191 human volunteers exposed to O3. In total, our work will identify genetic variants and gene regulatory mechanisms that influence susceptibility to O3-induced airway inflammation.

项目摘要暴露于环境空气污染物臭氧（O3）与心肺发病率和死亡率有关使其成为一个重要的公共卫生问题。受控暴露研究表明急性O3暴露原因气道炎症，上皮损伤和肺功能的瞬时降低。这些研究也已经证明受试者暴露了高度再现的O3反应差异，暗示了基因by- 环境相互作用（GXE）。候选基因研究为少数基因提供了GXE的证据，但是，遗传变异在基因组的其余部分中的作用在很大程度上是未知的。这个数据差距限制了我们的能力确定敏感的个体并深入了解O3会导致不良影响的机制。这里，我们提出了一项建议，以在体外使用人支气管上皮细胞（HBEC）来解决此数据差距。 HBEC 是呼吸道与O3相互作用的第一个细胞，我们已经证明了在体外暴露于O3的HBEC 上调关键促炎基因的表达（例如CXCL8），反映了体内反应。我们假设O3诱导的炎症的变化与HBEC基因表达的差异有关，并且在基线和O3之后基因表达的个体间差异具有遗传基础，即表达定量性状基因座（EQTL）。此外，我们假设某些EQTL是由单个eqtl引起的影响染色质可及性（CAQTL）的核苷酸多态性（SNP）。在AIM 1中，我们将建立良好的从300个男女的300个银行肺组织供体中生长的分化HBEC培养物，在空气界面生长潜水员的祖先，然后将它们暴露于O3与过滤空气（FA）并测量关键HBEC O3响应表型（例如IL-8产生，氧化应激，脂质过氧化，屏障功能和细胞毒性）。我们将介绍基因使用大量RNA-Seq和单细胞RNA-Seq鉴定O3-在FA和O3暴露的HBEC中表达诱导/抑制基因及其细胞类型的特异性。基因分型后，我们将在基线时绘制EQTL （mRNAFA），响应eqtl（mRNAO3-MRNAFA）和QTL用于所有HBEC O3响应表型，然后使用调解分析以识别适合推定因果模型的SNP和基因：O3+SNP→[mRNA]→HBEC O3 反应表型。在AIM 2中，我们将执行ATAC-SEQ，以表征O3如何改变染色质的可及性在HBEC中，然后映射基线和响应CAQTL。我们将执行多OMIC数据集成（EQTL，CAQTL， QTL）确定O3反应的基因调节模型，即O3+SNP→染色质访问性→[mRNA]→HBEC O3反应表型。最后，在AIM 3中，我们将验证新型基因和基因调节机制 O3反应在体外和体内的基本变化。我们将确定关键SNP如何影响基因调节以及击倒相应的基因在体外是否改变O3反应。对于体内验证，我们将在191人的数据集中，感兴趣的SNP与O3诱导的中性粒细胞募集之间的关联测试志愿者暴露于O3。总的来说，我们的工作将确定遗传变异和基因调节机制影响O3诱导的气道炎症的敏感性。