Proof of concept study of mass cytometry in septic shock patients reveals novel immune alterations

Abstract

Innovative single cell technologies such as mass cytometry (CyTOF) widen possibilities to deeply improve characterisation of immune alterations mechanisms in human diseases. So far, CyTOF has not been used in sepsis – a condition characterized by complex immune disorders. Here, we evaluated feasibility of CyTOF analysis in patients with septic shock. We designed a mass cytometry panel of 25 extracellular markers to study mononuclear cells from 5 septic shock patients and 5 healthy donors. We explored single-cell data with global and specific unsupervised approaches such as heatmaps, SPADE and viSNE. We first validated relevance of our CyTOF results by highlighting established immune hallmarks of sepsis, such as decreased monocyte HLA-DR expression and increased expressions of PD1 and PD-L1 on CD4 T cells and monocytes. We then showed that CyTOF analysis reveals novel aspects of sepsis-induced immune alterations, e.g. B cell shift towards plasma cell differentiation and uniform response of several monocyte markers defining an immune signature in septic patients. This proof of concept study demonstrates CyTOF suitability to analyse immune features of septic patients. Mass cytometry could thus represent a powerful tool to identify novel pathophysiological mechanisms and therapeutic targets for immunotherapy in septic shock patients.

How Clinical Flow Cytometry Rebooted Sepsis Immunology

“…) absent from controls. These cells might be circulating regulatory B cells never reported before in septic patients . Thus, the field of sepsis research is primed for similar translational studies that bring mass cytometry “to the bedside”.…”

“…Interestingly, remaining circulating B cells present with an altered phenotype and altered functions . While the presence of B cells with regulatory functions has been shown in mice models of infections and in some rare clinical contexts, preliminary data suggest that these cells might be induced after septic shock . This deserves to be further explored in sepsis.…”

“…However, recent mass cytometry analyses of patients undergoing major surgery showed that underlying the canonical immune response to traumatic injury were distinct immune phenotypes associated with surgical recovery outcomes . In sepsis, the use of mass cytometry in a preliminary study led to the description of novel immune alterations . For instance, it was observed that increased checkpoints inhibitor expressions were not uniformly co‐expressed on T lymphocyte population (e.g., TIM‐3 and PD‐1, Fig.…”

“…Illustrative example of unsupervised analysis of CyTOF results in healthy volunteers and septic patients. These results have been obtained from reference ( A ) SPADE clustering analysis was performed in order to group peripheral blood mononuclear cell subpopulations into “nodes” according to phenotype similarities. These nodes and their relationships are represented in a minimum spanning tree format (i.e., SPADE tree).…”

Immune Profiling Panel: a proof of concept study of a new multiplex molecular tool to assess the immune status of critically-ill patients

“…Nonetheless, personalized care is impeded by the absence of a comprehensive and fast diagnostic tool that would allow clinicians to precisely monitor the patient’s immune profile. For more than 20 years, researchers have described several biomarkers in different platforms to characterize the immune dysfunctions of sepsis (5-8). Transcriptomic gene signatures were identified to stratify septic patients according to the severity and worsening of outcomes that could be used to guide therapy (9-11).…”