Multiomic Investigations into Lung Health and Disease.

Diseases of the lung account for more than 5 million deaths worldwide and are a healthcare burden. Improving clinical outcomes, including mortality and quality of life, involves a holistic understanding of the disease, which can be provided by the integration of lung multi-omics data. An enhanced understanding of comprehensive multiomic datasets provides opportunities to leverage those datasets to inform the treatment and prevention of lung diseases by classifying severity, prognostication, and discovery of biomarkers. The main objective of this review is to summarize the use of multiomics investigations in lung disease, including multiomics integration and the use of machine learning computational methods. This review also discusses lung disease models, including animal models, organoids, and single-cell lines, to study multiomics in lung health and disease. We provide examples of lung diseases where multi-omics investigations have provided deeper insight into etiopathogenesis and have resulted in improved preventative and therapeutic interventions.

Integrative metabolomics and transcriptomics analysis reveals novel therapeutic vulnerabilities in lung cancer.

BACKGROUND: Non-small cell lung cancer (NSCLC) comprises the majority (~85%) of all lung tumors, with lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) being the most frequently diagnosed histological subtypes. Multi-modal omics profiling has been carried out in NSCLC, but no studies have yet reported a unique metabolite-related gene signature and altered metabolic pathways associated with LUAD and LUSC. METHODS: We integrated transcriptomics and metabolomics to analyze 30 human lung tumors and adjacent noncancerous tissues. Differential co-expression was used to identify modules of metabolites that were altered between normal and tumor. RESULTS: We identified unique metabolite-related gene signatures specific for LUAD and LUSC and key pathways aberrantly regulated at both transcriptional and metabolic levels. Differential co-expression analysis revealed that loss of coherence between metabolites in tumors is a major characteristic in both LUAD and LUSC. We identified one metabolic onco-module gained in LUAD, characterized by nine metabolites and 57 metabolic genes. Multi-omics integrative analysis revealed a 28 metabolic gene signature associated with poor survival in LUAD, with six metabolite-related genes as individual prognostic markers. CONCLUSIONS: We demonstrated the clinical utility of this integrated metabolic gene signature in LUAD by using it to guide repurposing of AZD-6482, a PI3Kß inhibitor which significantly inhibited three genes from the 28-gene signature. Overall, we have integrated metabolomics and transcriptomics analyses to show that LUAD and LUSC have distinct profiles, inferred gene signatures with prognostic value for patient survival, and identified therapeutic targets and repurposed drugs for potential use in NSCLC treatment.

Sex-specific epigenetic development in the mouse hypothalamic arcuate nucleus pinpoints human genomic regions associated with body mass index.

Recent genome-wide association studies corroborate classical research on developmental programming indicating that obesity is primarily a neurodevelopmental disease strongly influenced by nutrition during critical ontogenic windows. Epigenetic mechanisms regulate neurodevelopment; however, little is known about their role in establishing and maintaining the brain's energy balance circuitry. We generated neuron and glia methylomes and transcriptomes from male and female mouse hypothalamic arcuate nucleus, a key site for energy balance regulation, at time points spanning the closure of an established critical window for developmental programming of obesity risk. We find that postnatal epigenetic maturation is markedly cell type and sex specific and occurs in genomic regions enriched for heritability of body mass index in humans. Our results offer a potential explanation for both the limited ontogenic windows for and sex differences in sensitivity to developmental programming of obesity and provide a rich resource for epigenetic analyses of developmental programming of energy balance.

Vitamin D actions in neurons require the PI3K pathway for both enhancing insulin signaling and rapid depolarizing effects.

Despite correlations between low vitamin D levels and diabetes incidence/severity, supplementation with vitamin D has not been widely effective in improving glucose parameters. This may be due to a lack of knowledge regarding how low vitamin D levels physiologically affect glucose homeostasis. We have previously shown that the brain may be a critical area for vitamin d-mediated action on peripheral glucose levels. However, the mechanisms for how vitamin D acts in the brain are unknown. We utilized a multimodal approach to determine the mechanisms by which vitamin D may act in the brain. We first performed an unbiased search (RNA-sequencing) for pathways affected by vitamin D. Vitamin D (125-dihydroxyvitamin D3; 1,25D3) delivered directly into the third ventricle of obese animals differentially regulated multiple pathways, including the insulin signaling pathway. The insulin signaling pathway includes PI3K, which is important in the brain for glucose regulation. Since others have shown that vitamin D acts through the PI3K pathway in non-neuronal cells (muscle and bone), we hypothesized that vitamin D may act in neurons through a PI3K-dependent pathway. In a hypothalamic cell-culture model (GT1-7 cells), we demonstrate that 1,25D3 increased phosphorylation of Akt in the presence of insulin. However, this was blocked with pre-treatment of wortmannin, a PI3K inhibitor. 1,25D3 increased gene transcription of several genes within the PI3K pathway, including Irs2 and p85, without affecting expression of InsR or Akt. Since we had previously shown that 1,25D3 has significant effects on neuronal function, we also tested if the PI3K pathway could mediate rapid actions of vitamin D. We found that 1,25D3 increased the firing frequency of neurons through a PI3K-dependent mechanism. Collectively, these data support that vitamin D enhances insulin signaling and neuronal excitability through PI3K dependent processes which involve both transcriptional and membrane-initiated signaling events.

DNA methylation in AgRP neurons regulates voluntary exercise behavior in mice.

DNA methylation regulates cell type-specific gene expression. Here, in a transgenic mouse model, we show that deletion of the gene encoding DNA methyltransferase Dnmt3a in hypothalamic AgRP neurons causes a sedentary phenotype characterized by reduced voluntary exercise and increased adiposity. Whole-genome bisulfite sequencing (WGBS) and transcriptional profiling in neuronal nuclei from the arcuate nucleus of the hypothalamus (ARH) reveal differentially methylated genomic regions and reduced expression of AgRP neuron-associated genes in knockout mice. We use read-level analysis of WGBS data to infer putative ARH neural cell types affected by the knockout, and to localize promoter hypomethylation and increased expression of the growth factor Bmp7 to AgRP neurons, suggesting a role for aberrant TGF-ß signaling in the development of this phenotype. Together, these data demonstrate that DNA methylation in AgRP neurons is required for their normal epigenetic development and neuron-specific gene expression profiles, and regulates voluntary exercise behavior.

Transcriptomic and microRNA analyses of gene networks regulated by eicosapentaenoic acid in brown adipose tissue of diet-induced obese mice.

Brown adipose tissue (BAT) dissipates chemical energy as heat via thermogenesis and protects against obesity by increasing energy expenditure. However, regulation of BAT by dietary factors remains largely unexplored at the mechanistic level. We investigated the effect of eicosapentaenoic acid (EPA) on BAT metabolism. Male C57BL/6J (B6) mice were fed either a high-fat diet (HF, 45% kcal fat) or HF diet supplemented with EPA (HF-EPA, 6.75% kcal EPA) for 11 weeks. RNA sequencing (RNA-Seq) and microRNA (miRNA) profiling were performed on RNA from BAT using Illumina HiSeq and Illumina Genome Analyzer NextSeq, respectively. We conducted pathway analyses using ingenuity pathway analysis software (IPA®) and validated some genes and miRNAs using qPCR. We identified 479 genes that were differentially expressed (2-fold change, n = 3, P ≤ 0.05) in BAT from HF compared to HF-EPA. Genes negatively correlated with thermogenesis such as hypoxia inducible factor 1 alpha subunit inhibitor (Hif1an), were downregulated by EPA. Pathways related to thermogenesis such as peroxisome proliferator-activated receptor (PPAR) were upregulated by EPA while pathways associated with obesity and inflammation such as nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) were downregulated by EPA. MiRNA profiling identified nine and six miRNAs that were upregulated and downregulated by EPA, respectively (log2 fold change > 1.25, n = 3, P ≤ 0.05). Key regulatory miRNAs which were involved in thermogenesis, such as miR-455-3p and miR-129-5p were validated using qPCR. In conclusion, the depth of transcriptomic and miRNA profiling revealed novel mRNA-miRNA interaction networks in BAT which are involved in thermogenesis, and regulated by EPA.

A new mild hyperthermia device to treat vascular involvement in cancer surgery.

Surgical margin status in cancer surgery represents an important oncologic parameter affecting overall prognosis. The risk of disease recurrence is minimized and survival often prolonged if margin-negative resection can be accomplished during cancer surgery. Unfortunately, negative margins are not always surgically achievable due to tumor invasion into adjacent tissues or involvement of critical vasculature. Herein, we present a novel intra-operative device created to facilitate a uniform and mild heating profile to cause hyperthermic destruction of vessel-encasing tumors while safeguarding the encased vessel. We use pancreatic ductal adenocarcinoma as an in vitro and an in vivo cancer model for these studies as it is a representative model of a tumor that commonly involves major mesenteric vessels. In vitro data suggests that mild hyperthermia (41-46 °C for ten minutes) is an optimal thermal dose to induce high levels of cancer cell death, alter cancer cell's proteomic profiles and eliminate cancer stem cells while preserving non-malignant cells. In vivo and in silico data supports the well-known phenomena of a vascular heat sink effect that causes high temperature differentials through tissues undergoing hyperthermia, however temperatures can be predicted and used as a tool for the surgeon to adjust thermal doses delivered for various tumor margins.