Multi-omics in nasal epithelium reveals three axes of dysregulation for asthma risk in the African Diaspora populations

Background: Asthma, a complex chronic lung disease affecting the airways, has striking disparities across ancestral groups, but the molecular underpinning of these differences is poorly understood and minimally studied. A major goal of the Consortium on Asthma among African-ancestry Populations in the Americas (CAAPA) is to understand multi-omics signatures of asthma risk in the nasal epithelium focusing on populations of African ancestry. Methods: RNASeq data were generated from nasal epithelium in subjects recruited from up to 7 sites (Baltimore, Washington DC, Chicago, Denver, Salvador Brazil, Barbados, and Nigeria). Current asthma cases (N=253) were compared to never-asthma controls (N=283) to identify differentially expressed genes (DEGs; q <0.05). Network analyses were performed with Ingenuity Pathway Analysis (IPA; DEGs with q<0.05) and weighted gene co-expression network analysis (WGCNA; DEGs with q<0.15). All models were fully adjusted for ancestry, sampling site, and appropriate latent factors. Findings: CAAPA represents diversity across the African Diaspora with a wide range of continental African ancestry (9%-100%). We identified 389 DEGs; the top DEG, FN1, was downregulated in asthma cases (q=3.26x10-9) and encodes fibronectin which plays a role in wound healing. Others in the top 10 DEGs have high relevance for asthma: SNTG2 (q = 1.12x10-4) is the target of multiple miRNAs related to asthma; PPP1R9A expression (q=7.60x10-5) was previously determined to be influenced by IL-13 in mouse lung; and SPTBN1 (q=1.12x10-4) plays a key role in mediating TGFß signaling. IPA revealed networks with upstream regulators relevant for immune response (IL4; p=7.25x10-10 and TGFß1; p=5.47x10-8) and drug response (dexamethasone; p=4.31x10-10 and fluticasone propionate; p=9.42x10-8). Among asthma cases, genes regulated by dexamethasone and fluticasone propionate were not associated with inhaled corticosteroid medication use. The top three WGCNA modules implicate networks related to immune response (CEACAM5; p=9.62x10-16 and CPA3; p=2.39x10-14) and wound healing (FN1; p=7.63x10-9). Multi-omic analysis identified FKBP5 as a key contributor to asthma risk, whereby the association between nasal epithelium gene expression is mediated through methylation and is associated with increased use of inhaled corticosteroids. FKBP5 is a co-chaperone of glucocorticoid receptor signaling and known to be involved in drug response in asthma. Interpretation: Our analyses reveal genes and networks in asthma that are differentially expressed in nasal epithelium of asthma cases of African ancestry in CAAPA. Importantly, this work reveals molecular dysregulation on three axes – increased Th2 inflammation, decreased capacity for wound healing, and impaired drug response – that may play a critical role in asthma within the African Diaspora. Overall design: RNA sequencing data was generated from nasal epithelium samples in newly recruited subjects representing 7 locations across the African Diaspora: Baltimore, Washington DC, Chicago and Denver in the United States; Salvador, Brazil; Barbados in the Caribbean; and Nigeria in West Africa. We examined transcriptomic differences between asthma cases and never-asthma controls to identify differentially expressed individual genes and networks of co-expressed genes.

背景：哮喘是一种累及气道的复杂慢性肺部疾病，不同祖先人群间的哮喘发病差异十分显著，但此类差异的分子机制仍知之甚少，相关研究也极为匮乏。美洲非洲血统人群哮喘研究联盟（Consortium on Asthma among African-ancestry Populations in the Americas, CAAPA）的核心目标之一，便是以非洲血统人群为研究对象，解析鼻上皮细胞中哮喘风险的多组学特征。
方法：本研究从7个招募点（美国巴尔的摩、华盛顿特区、芝加哥、丹佛，巴西萨尔瓦多，巴巴多斯以及尼日利亚）的受试者中采集鼻上皮样本，进行RNA测序（RNASeq）。以253例现症哮喘患者为病例组，283例从未罹患哮喘的个体为对照组，筛选差异表达基因（differentially expressed genes, DEGs；校正后P值q<0.05）。分别采用Ingenuity通路分析（Ingenuity Pathway Analysis, IPA；针对q<0.05的DEGs）和加权基因共表达网络分析（weighted gene co-expression network analysis, WGCNA；针对q<0.15的DEGs）开展网络分析。所有统计模型均对祖先背景、采样地点以及潜在混杂因素进行了全面校正。
研究结果：CAAPA队列涵盖了非洲散居人群的遗传多样性，其非洲大陆血统占比范围为9%至100%。本研究共筛选得到389个DEGs，其中排名首位的DEG为FN1，在哮喘患者中表达下调（q=3.26×10^-9），其编码的纤连蛋白参与伤口愈合过程。前十位DEGs中其余基因也与哮喘密切相关：SNTG2（q=1.12×10^-4）是多种哮喘相关miRNA的靶基因；PPP1R9A的表达（q=7.60×10^-5）此前被证实可受小鼠肺组织中IL-13的调控；SPTBN1（q=1.12×10^-4）在介导转化生长因子β（transforming growth factor β, TGFβ）信号通路中发挥关键作用。IPA分析显示，其上游调控因子网络涉及免疫应答相关分子（IL-4；p=7.25×10^-10以及TGFβ1；p=5.47×10^-8）与药物应答相关分子（地塞米松；p=4.31×10^-10以及丙酸氟替卡松；p=9.42×10^-8）。在哮喘患者亚组中，受地塞米松与丙酸氟替卡松调控的基因与吸入性糖皮质激素用药史无显著关联。排名前三的WGCNA模块所关联的网络分别涉及免疫应答（CEACAM5；p=9.62×10^-16以及CPA3；p=2.39×10^-14）与伤口愈合（FN1；p=7.63×10^-9）。多组学分析证实FKBP5是哮喘风险的关键调控因子：鼻上皮细胞中FKBP5的基因表达与甲基化介导的调控通路相关，且与吸入性糖皮质激素使用量增加存在关联。FKBP5作为糖皮质激素受体信号通路的辅助分子伴侣，已被证实参与哮喘的药物应答过程。
阐释与讨论：本研究分析揭示了CAAPA队列中非洲血统哮喘患者鼻上皮细胞内与哮喘相关的差异表达基因及共表达网络。尤为重要的是，本研究明确了非洲散居人群哮喘发病的三类分子调控异常——Th2炎症升高、伤口愈合能力下降以及药物应答受损，这些异常可能在该人群的哮喘发病过程中发挥关键作用。
总体研究设计：本研究从非洲散居人群分布的7个地区新招募受试者，采集其鼻上皮样本并进行RNA测序，这7个地区分别为美国的巴尔的摩、华盛顿特区、芝加哥与丹佛，巴西的萨尔瓦多，加勒比地区的巴巴多斯以及西非的尼日利亚。本研究通过对比哮喘病例组与从未罹患哮喘的对照组的转录组差异，筛选差异表达的单个基因及共表达基因网络。