Omics analyses are commonly used for identifying pathways and genes responsible for physiologic and pathologic processes. Though sex is considered a biological variable in rigorous assessments of pulmonary responses to oxidant exposures, the contribution of the murine strain is largely ignored. This study utilized an unbiased integrated assessment of high-resolution metabolomic profiling and RNA-sequencing to explore sex- and strain-dependent pathways in adult mouse lungs. The results indicated that strain exhibited a greater influence than sex on pathways associated with inflammatory and oxidant/antioxidant responses and that interaction metabolites more closely resembled those identified as differentially expressed by strain. Metabolite analyses revealed that the components of the glutathione antioxidant pathway were different between strains, specifically in the formation of mixed disulfides. Additionally, selenium metabolites such as selenohomocystiene and selenocystathionine were similarly differentially expressed. Transcriptomic analysis revealed similar findings, as evidenced by differences in glutathione peroxidase, peroxiredoxin, and the inflammatory transcription factors RelA and Jun. In summary, an multi-omics integrated approach identified that murine strain disproportionately impacts baseline expression of antioxidant systems in lung tissues. We speculate that strain-dependent differences contribute to discrepant pulmonary responses in preclincal mouse models, with deleterious effects on clinical translation.

Extremely preterm infants are often treated with supraphysiological oxygen, which contributes to the development of bronchopulmonary dysplasia (BPD). These same infants exhibit compromised antioxidant capacities due in part to selenium (Se) deficiency. Se is essential for basal and inducible antioxidant responses. The present study utilized a perinatal Se deficiency (SeD) mouse model to identify the combined effects of newborn hyperoxia exposure and SeD on alveolarization and antioxidant responses, including the identification of affected developmental pathways. Se-sufficient (SeS) and SeD C3H/HeN breeding pairs were generated, and pups were exposed to room air or 85% O₂ from birth to 14 d. Survival, antioxidant protein expression, and RNA seq analyses were performed. Greater than 40% mortality was observed in hyperoxia-exposed SeD pups. Surviving SeD pups had greater lung growth deficits than hyperoxia-exposed SeS pups. Gpx2 and 4 protein and Gpx activity were significantly decreased in SeD pups. Nrf2-regulated proteins, Nqo1 and Gclc were increased in SeD pups exposed to hyperoxia. RNA seq revealed significant decreases in the Wnt/β-catenin and Notch pathways. Se is a biologically relevant modulator of perinatal lung development and antioxidant responses, especially in the context of hyperoxia exposure. The RNA seq analyses suggest pathways essential for normal lung development are dysregulated by Se deficiency.

Human milk (HM) is rich in bioactive factors promoting postnatal small intestinal development and maturation of the microbiome. HM is also protective against necrotizing enterocolitis (NEC), a devastating inflammatory condition predominantly affecting preterm infants. The HM glycosaminoglycan, hyaluronan (HA), is present at high levels in colostrum and early milk. Our group has demonstrated that HA with a molecular weight of 35 kDa (HA35) promotes maturation of the murine neonatal intestine and protects against two distinct models of NEC. However, the molecular mechanisms underpinning HA35-induced changes in the developing ileum are unclear. CD-1 mouse pups were treated with HA35 or vehicle control daily, from P7 to P14, and we used network and functional analyses of bulk RNA-seq ileal transcriptomes to further characterize molecular mechanisms through which HA35 likely influences intestinal maturation. HA35-treated pups separated well by principal component analysis, and cell deconvolution revealed increases in stromal, Paneth, and mature enterocyte and progenitor cells in HA35-treated pups. Gene set enrichment and pathway analyses demonstrated upregulation in key processes related to antioxidant and growth pathways, such as nuclear factor erythroid 2-related factor-mediated oxidative stress response, hypoxia inducible factor-1 alpha, mechanistic target of rapamycin, and downregulation of apoptotic signaling. Collectively, pro-growth and differentiation signals induced by HA35 may present novel mechanisms by which this HM bioactive factor may protect against NEC.