Abstract Introduction The gas exchanging surface of human lung is also an active cite for immune surveillance against respiratory pathogens. Lung leukocytes, key to this surveillance, change with lung post-natal developmental stage and later with advanced age though details are incompletely known. Human lung development during early postnatal years involves major changes at structural, cellular and molecular levels that result in increased gas exchange capacity and more efficient immune surveillance against infectious pathogens. However, with advancing age, the fully developed lung experiences progressive functional decline not only associated with deteriorating gas exchange capacity but also linked with dysregulated immunity and increased susceptibility to infectious diseases. In this study, we examined lung leukocyte subpopulation cell-proportion, gene expression and spatial changes during two key human transition periods in lung function, childhood to early adulthood, and, early adulthood to older adulthood. Experimental Design Utilizing cellular indexing of transcriptomes and epitopes by sequencing (CITE Seq) we assessed gene expression at single cell level in lung resident leukocytes from healthy subjects with no history of smoking or lung disease. To study age associated differences, we divided these cases into three groups: pediatric/adolescent (10 to 18 years, PA), young adult (18-50 years, YA), and older adult (55-69 years, OA). Spatial location and transcriptomic profiles of lung leukocyte subsets identified were further studied by XeniumTM spatial transcriptomics assay on the same subjects using a novel custom immunity-focused gene probe panel. The CITE Seq and Xenium data were compared (Figure-1A). Result By CITE sequencing on dissociated lung cells, we identified 43 different cell types using Azimuth cell referencing tools. By comparison, by performing Xenium spatial transcriptomics on tissue sections, we identified 50 cell types in the same subjects (Figures-1B,1C). Both CITE Seq and Xenium dataset showed differences in cell proportionalities comparing different age groups. Differential gene expression analysis comparing same cells-subset from different age cohorts showed increased functional maturity in YA compared to OA groups with alveolar macrophages showed showing maximum number of differentially expressed genes (DEG). During YA to OA transition, non-classical monocytes had the highest numbers of DEG. In these cells, cell adhesion linked pathways were among the most enriched in OA while extracellular matrix organization and disassembly related pathways were among the top enriched. Conclusion Significantly altered age-dependent gene expression patterns were identified in several lung leukocyte subsets consistent with immune functional maturation. Grant support NHLBI U01HL122700 and U01HL148861. This abstract is funded by: NIH
Bandyopadhyay et al. (Fri,) studied this question.