R. Marwan1, S. Al-Juboori1, E. Dobrinskikh2, J. Haines3, R. Marwan1 1University Of Colorado Denver,Pediatric Surgery,Aurora, CO, USA 2University Of Colorado Denver,Medicine,Aurora, CO, USA 3University Of Colorado Denver,Biochemistry And Molecular Genetics,Aurora, CO, USA
Introduction: Infants with severe congenital diaphragmatic hernia (CDH) offered experimental tracheal occlusion (TO) have inconsistent clinical outcomes. Understanding such clinical variability is crucial to advancing neonatal care but the reasons for such deviations are currently unknown. Our laboratory has proposed a novel concept of heterogeneous topological zones formation in fetal lungs following TO. However, the temporal pattern of airspace morphometry and metabolic landscape changes following TO is not fully elucidated. Our objective is to explore the natural history of heterogeneous changes in fetal lungs following TO.
Methods: We evaluated fetal lungs after 1 and 4 days following TO; global and local metabolic changes, as well as morphometric indices were examined using mass spectrometry-based metabolomics, Fluorescence Lifetime Imaging Microscopy (FLIM) and Tissue Airspace Ratio (TAR), respectively.
Results: 1 day (D1) after TO, there is an appearance of two distinct morphometric areas varying between small (high TAR values) and large (small TAR values) airspaces, with an overall shift toward smaller airspaces compared to controls. In contrast, by day 4 (D4) TO lungs have a higher frequency of large airspaces (small TAR values) compared to control. D4 TO lungs global metabolomics data demonstrate a significant decrease in glucose and hexose phosphate, and an increase in lactate production, compared to D1 TO lungs. Moreover, there is a suppression of the tricarboxylic acid cycle (TCA) with a significant decrease in citrate and fumarate at D4. These changes are accompanied by significant increase in spermidine and spermine at D4, which are important for cellular growth and proliferation. Locally on FLIM, D1 TO lungs demonstrate two types of heterogenous zones. Namely, zones which have free/bound NADH ratio similar to control lungs, and zones with decreased free/bound NADH ratio- indicating increased OXPHOS in these regions. Similar to the histological findings, FLIM on D4 TO lungs demonstrates appearance of zones with enlarged airspaces, which have metabolic shift towards glycolysis, accompanied by decreased FLIM-surfactant signal only in those areas.
Conclusion: In normal fetal lungs, we report a novel unique natural history of heterogeneous changes; TO leads to heterogeneous pulmonary response. Initially, there is formation of zones with small airspaces, followed by airspace enlargement over time. While FLIM of D1 TO lungs reveals zones with increased OXPHOS, FLIM in D4 TO lungs demonstrate a shift toward glycolysis in the enlarged airspaces with decreased surfactant production. We speculate that the “best responders” to tracheal occlusion should have bigger lungs with small airspaces and normal surfactant production.
