The wide array of molecules carried by plasma regulates critical immune functions and constitutes valuable biomarkers and therapeutic targets. In recent years the introduction of “systems approaches” has provided investigators with powerful means for assessing immune responses in patient samples on a global scale. However, while the use of genome-wide profiling technologies has become widespread, measuring the plasma proteome still presents considerable challenges. An alternative approach that consists in measuring transcriptome responses in reporter cells exposed in vitro to patient plasma has been successfully employed in a limited number of studies. Here we devised such a “Transcriptomic Reporter Assay” system to assess the immunogenicity of plasma from septic patients and evaluate its potential for biomarker discovery. Sepsis is a common, severe systemic infectious process for which physicians still lack efficient diagnostic or prognostic tools. Of the three different cell reporter systems tested, neutrophils were identified as the most capable “plasma sensor”. Compared to peripheral blood mononuclear cells and dendritic cell preparations neutrophils were best able to discriminate between plasma from septic and control subjects and responded by upregulating a robust immune transcriptional program. Additionally, the amplitude of the neutrophil transcriptomic response was shown to be associated with disease severity in two additional sets of patients. Overall, our results demonstrate both the suitability and potential clinical relevance of a neutrophil reporter assay for assessing immunopathogenic processes in a complex and severe condition such as sepsis.

Monocytes were isolated from two healthy donors. Monocytes were cultured with interleukin 4 (IL4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) to generate monocyte derived dendritic cells (MoDCs). Plasma samples were obtained from patients with culture-confirmed sepsis (n=12) and from uninfected controls (n=12). MoDCs were cultured for 6 h in medium alone, plasma from patients with sepsis, plasma from uninfected controls, and LPS using a final concentration of 20%. Transcriptional profiles were acquired using Illumina HumanHT12 V4 BeadChips.

Septic Plasma Exposure (2021 ICD-10-CM code* = Not Applicable)

The data were preprocessed using R, which included the following steps: quantile normalization, flooring all intensities <10 to 10, log2 transformation, and PALO (present at least once) filter. PALO, which selects only transcripts with detection p-values < 0.01 in at least one sample, was performed to reduce background noise.