(323c) Myeloid-Derived Suppressor Cells Impair Neutrophil Migration in an Engineered Model of the Infectious Microenvironment

Patients who recover from sepsis exhibit extended immunological impairment marked by a high incidence of and increased mortality from opportunistic bacterial pathogens. However, our current knowledge of the causes of this dysfunction is limited. One contributing factor may be myeloid-derived suppressor cells (MDSCs), an immature and heterogeneous population of innate immune cells that persist following sepsis. MDSCs are best studied in the context of cancer, where they contribute to a pro-tumor environment by secreting soluble signals that impair T cell proliferation and effector functions. MDSCs are also found in high numbers in patients recovering from chronic infections, including sepsis, but their role in these settings is poorly understood. One potential explanation is that the non-specific cytokines released by MDSCs are also detrimental to cells crucial to the innate immune response to infection, including neutrophils, a population of fast-responding phagocytes, and endothelial cells. In response to bacterial infection, endothelial cells lining blood vessels interact with neutrophils to orchestrate their activation and extravasation into the extracellular matrix. If communication between neutrophils and endothelial cells is altered by the removal of key signals or the addition of suppressive signals, however, this process is disrupted. MDSCs may be implicated in this disruption due to their known secretion of suppressive cytokines like IL-10 that have been shown to reduce neutrophil migration. While this phenomenon would be difficult to study in vivo due to differences between model organism and human physiology as well as the complexity of isolating single signals, we use an infection-on-a-chip microfluidic model that includes a model blood vessel lined with endothelial cells, surrounded by an extracellular matrix, and seeded with a live pathogen. In this project, we used this system to investigate the impacts of MDSCs on neutrophil migration and extravasation as well as endothelial permeability and chemokine secretion during bacterial infection.

We generated MDSCs from healthy human monocytes using GM-CSF and IL-6 and confirmed their appropriate phenotype via flow cytometry to investigate surface markers and a T cell suppression assay to demonstrate their well-established ability to reduce T cell proliferation. Following this, we investigated the impact of live MDSCs on neutrophil chemotaxis to a stable gradient of IL-8 and found that adding MDSCs resulted in shorter and less direct neutrophil migration. MDSCs were also added into the collagen of the infection-on-a-chip device and were found to significantly reduce neutrophil extravasation to Pseudomonas aeruginosa infection. We then used a multiplexed ELISA to investigate signals that may be altered by MDSCs and found that in the presence of MDSCs, anti-inflammatory proteins, including IL-10, and cytokines that lead to vasoconstriction, including MIP-1É‘, were increased. These findings will be confirmed with future blocking experiments. Investigation of endothelial lumen permeability using fluorescent dextran also showed decreased permeability in the presence of MDSCs. Overall, these conclusions suggest that MDSCs may contribute to immune dysregulation by impairing neutrophil chemotaxis and extravasation through the secretion of cytokines that reduce neutrophil migration and decrease endothelial permeability. This expanded understanding of MDSC biology may be important for the development of future treatments to combat immune dysregulation.