Lipidomic Profiling of Host‐Pathogen Interactions Using Untargeted and Targeted Lipidomics Approaches in a Sepsis Cell Culture Model

Abstract

Sepsis is a life‐threatening organ dysfunction caused by a dysregulated host response to an infection. When pathogens disseminate into the systemic blood circulation from a localized infection, the body initiates a complex series of immune and biochemical reactions. Neutrophils are the main leukocytes that are recruited to an infection site to eliminate bacteria. In sepsis, systemic neutrophil activation is also evident. One of the main pathologies of sepsis is the damage of the endothelial cell integrity by immune stimuli leading to leaky blood vessels. Although numerous research has been conducted on sepsis, the characteristic heterogeneity of sepsis has challenged the identification of specific biomarkers and treatments. We have designed an in vitro co‐culture model of endothelial cells, human neutrophils and, E.coli to identify lipid biomarkers of sepsis. The three cell types in the model allow us to dissect lipidome of individual cell systems under a cell culture sepsis‐like state. EA.hy926 cells were initially incubated with urosepsis E.Coli strain CFT073 and further incubated after adding neutrophils. We compared the lipidome of co‐culture samples with non‐co‐culture samples to identify altered lipid profiles. In addition, we have performed similar experiments with non‐pathogenic JM109 E.Coli strain. We first used an untargeted lipidomics platform with high‐resolution mass spectrometry in both positive and negative ion modes. Potential lipid biomarkers identified in untargeted lipidomics were furthered confirmed by a targeted lipidomics approach using shotgun lipidomics. In CFT073 co‐cultures, lipids were changed mainly in 4 lipid classes; phosphatidylcholine (PC), phosphatidylethanolamine (PE), lysophosphatidylcholine and triglycerides. Minor changes were observed in sphingomyelin, phosphatidylserine and phosphatidylglycerol classes. Several of the changes in molecular species in each lipid class included significant increases in PE 32:1, PE 34:1, PE 32:2, PC 36:0, PC 40:8 and PC 32:0. Lipid profiles were also altered in non‐pathogenic JM109 co‐cultures. Interestingly, there are marked species variations depending on E.coli strain used in co‐cultures such as PC 32:1, PC 36:1 are only elevated in CFT073 co‐cultures. Collectively, our studies demonstrate that lipid profiles are altered during inflammation initiated by bacterial exposure and subsequent neutrophil activation. We have also shown that pathogenicity of bacteria may lead to distinct alterations in lipidomes. These studies represent initial insights into the complex lipidomic alterations during host‐pathogen interactions and may provide new targets to examine as biomarkers and mediators during in vivo sepsis.Support or Funding InformationR01 GM115553, R01 GM129508, U01 ES026458, U01 ES027697, U01 ES028182