Tilo Winkler

Background & Objectives: Mechanical ventilation of heterogeneously expanding lungs is associated with different mechanisms of regional mechanical injury such as lung hyperinflation, cyclic recruitment, and propagation of fluid-gas interfaces. Heterogeneity of regional blood flow could result in the regional exposure of lung parenchyma to different loads of circulating inflammatory cells and mediators. We hypothesize that these different regional mechanisms of injury result in distinct regional cellular responses to injury reflected in different regional patterns of gene expression. Our aim was to apply methods of gene functional classification to identify processes differentially active in ventral versus dorsal lung regions in supine sheep submitted to endotoxemia and mechanical ventilation for 16h.

Materials & Methods: Supine sheep (n=6) were mechanically ventilated for 16h (ARDS-NET protocol). Endotoxemia was produced with continuous infusion of lipopolysaccharide (10ng/kg/min) to model an extra-pulmonary inflammatory stimulus. After 16h, animals were euthanized and lung tissue samples harvested from ventral and dorsal regions. All Agilent platform G4813A-019921 arrays were processed and probesets annotated. Probes were subset to those with annotations recognized by g:profiler and differentially expressed genes identified using the linear model for microarray data analysis limma. Top 200 differentially expressed genes were examined for gene ontology category enrichment and statistics were calculated for the genes recognized by g;profiler.

Results: The data starts out with 15618 probes against O. aries RNAs and gprofiler recognizes 14343 genes from O.Aries. Of the probes on the Agilent platform G4813A-019921 data, 1104 are annotated with gene names recognized by gprofiler and of these 615 are against unique genes. Functional gene annotation clustering (Image) revealed significant gene ontology categories enrichment for biological processes of cell activation (p=1.86.10-2), cellular component of T receptor complex (p=3.2.10-2) and human phenotype ontology for imunodeficiency (p=2.75.10-3) and abnormality of cellular imune system (p= 1.56.10-2).

Conclusion: Genomic analysis of non-dependent (ventral) versus dependent (dorsal) pulmonary regions revealed the spatially differential expression of genes that encodes biological pathways relevant to early stages of lung injury related to cell activation and abnormality of cellular immune system. Our results suggest topographical heterogeneous functional gene expression consistent with spatially heterogeneous metabolic changes in early lung injury.