Effect of Nrf2 deletion on developmental lung transcriptomics in mice

Background: NRF2 is an essential cytoprotective transcription factor inducing antioxidant response element (ARE)-bearing genes. However, association of NRF2 with lung development has not been examined. Human lungs are not fully developed until 2-3 years of and they are fully matured at about 8 years. Murine lungs at birth are immature (at saccular stage of lung development) and have been used to study developmental lung disorders. Methods: To investigate (1) the transcriptome changes during lung development and (2) the role of NRF2 in lung development and maturation in mice, lungs were harvested from Nrf2-deficient (Nrf2-/-) and wild-type (Nrf2+/+) mouse embryos, neonates and adults. Microarray and pathway analysis determined NRF2-directed mechanisms underlying lung development and maturation. Results: Nrf2 mRNA expression was peack at embryonic days E17.5-E18.5 (immediately before birth) probably to increase antioxidant apparatus to prepare against high O2 environment after birth. The pseudoglandular phase lungs (E13-E15) are undergoing vigorous cell proliferation under the control of high-fidelity DNA damage repair system. Fetal lungs (E13.5-E17.5) are lack in immune system, xenobiotic metabolism, and tissue damage genes. After birth at postnatal day 1 (PND1), lung cell division is quiescent but transporters and lipid metabolism are activated. When lung enters alveolar phase (PND4), cell proliferation is resumed. Mature lungs (PND14-P42) have heightened networks of host defense systems (immunity, antioxidants) and cellular injury and abnormality (e.g., glucose metabolism disorder). Nrf2 deletion in fetal lung (E13.5-E17.5) altered developmental, immunity, and metabolism genes, and it may have affected lung branching. Nrf2 deletion affected lung transcriptome changes the most at E17.5 when Nrf2 message level is maximum (E17.5-E18.5). Nrf2 deletion in newborn lung (PND0) decreased cell cycle progress and DNA damage repair. Nrf2 deletion in neonatal lung (PND1-4) enhanced tissue injury/cell death and inhibited developmental cell differentiation. Nrf2 deletion in matured mouse lung (PND42) affected not only antioxidant pathway but also immune responses and connective tissue cell migrations. Conclusion: Overall, NRF2 plays multiple roles in underdeveloped lungs and associated with lung morphogenesis, immunity, cell cycle progress, tissue differentiation and metabolism as well as cellular defense. Results provide putative molecular mechanisms of NRF2-directed lung morphogenesis and maturation. PARALLEL study design with 66 samples comparing 22 groups of different age (E13.5, E15.5, E17.5, E18.5, PND0, PND1, PND2, PND3, PND4, PND14, PND42) and genotype (Nrf2+/+ or WT, Nrf2-/- or KO): 11 groups Nrf2+/+ WT from E13.5 to PND42, 11 groups Nrf2-/- KO from E13.5 to PND42. Biological replicates: 3 per group