Untargeted Proteomics to Assess the Impact of Moderate Hyperoxia on Patient Sample-derived Neonatal Airway Epithelial Cells in 3D Culture

ABSTRACT: Neonatal Tracheal Aspirate-derived Airway Epithelial Cells (nTAECs) were isolated utilizing fresh tracheal suction aspirates from preterm (born at 23-28wks gestational age) and term infants (born at 37-42wks gestation). Included infants (especially term infants) did not have significant lung disease at the time of collection. Isolated cells were plated in 804g-cell-derived matrix coated T25, then expanded to a T75 flask and subsequently cultured in transwell inserts (0.4μm size pore) at 105 live cells/insert under air-liquid interface (ALI). Basal nTAECs have the capacity to differentiate into the full repertoire of mature airway epithelial lineages (ciliated, secretory club and mucus secreting goblet cells) during ALI culture. Cells were exposed to 60% O2 or room air control (21% O2) for 7 days (ALI day 7 to 14). On ALI day 14, cells will be harvested from the transwells and prepared for proteomic analysis using Data-independent acquisition (DIA) proteomics analysis. The samples will undergo several processing steps including chloroform/methanol extraction with trypsin digestion, Orbitrap Exploris 480 mass spectrometry analysis and eventual library construction and bioinformatics analysis. Utilizing 5 donor samples for each group, the proteomic workflow compared 4 exposure groups: a) Term nTAECs in Normoxia, b) Term nTAECs Hyperoxia, c) Preterm nTAECs in Normoxia, d) Preterm nTAECs in Hyperoxia.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Lung

SUBMITTER: Dennis Province  

LAB HEAD: Abhrajit Ganguly

PROVIDER: PXD064170 | Pride | 2026-03-02

REPOSITORIES: Pride