Modulation of the ATP/Adenosine axis by ectonucleotidases CD39 and CD73 in the context of sepsis

• Applicant: Dr. Anne Rissiek
• Subject Area: Anaesthesiology
• Term: from 2018 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 390917730

• Final Report Year: 2023

Final Report Abstract

Despite years of research, sepsis still accounts to one of the most severe conditions in the clinic with a high mortality rate. During sepsis progression, a massive, uncontrolled inflammatory response of the host to the invading microorganisms occurs, evolving in an immune paralytic state. As a consequence of cell activation, adenosine triphosphate (ATP) is released from cells, resulting in accumulation of the inflammatory molecule ATP in the extracellular space, inducing purinergic signaling and inflammation. However, extracellular ATP can be hydrolyzed by the ectonucleotidases CD39 and CD73 to anti-inflammatory adenosine (ADO). While CD39 expression on T cells is upregulated during immune cell activation, CD73 expression at the cell surface decreases, and a soluble form of CD73 can be found at sites of inflammation. Thus, pro-inflammatory ATP is degraded in the close proximity of CD39-expressing immune cells, downregulation of CD73 at the same time abrogates the local generation of anti-inflammatory ADO. To gain a general view of changes in the leukocyte compartment caused by sepsis, we performed flow cytometric analyses on blood samples from sepsis patients. To this end we collected samples form sepsis patients admitted to the ICU and two control groups: healthy donors (HD) and non-septic ICU patients. We confirmed and could analyze in detail the general loss of lymphocytes in the blood of septic patients, while at the same time blood neutrophil numbers increase. Taken together, we found that ICU sepsis patients were in general lymphopenic, and their T cells had an activated phenotype, as demonstrated by the upregulation of CD39 and the decrease of CD73 surface expression. In the T cell compartment, we found that CD8+ T cells displayed a significantly lower percentage of naïve and increased percentage of exhausted cells in ICU sepsis patients when compared to both controls. The phenotyping analyses suggests that the CD8+ T cell compartment of sepsis patients converts towards an activated/exhausted state. Overall, we collected a lot of data from the individual sepsis patients, the corresponding ICU controls and HDs. Consequently, we selected a fraction of the obtained flow cytometric parameters for a dimensionality reduction analyses (UMAP) and clustering analyses. We performed the analyses with or without the expression data of CD39 and CD73 on the subsets. Here, ICU sepsis patients formed an individual cluster, regardless if the CD39/CD73 expression data was included or not. However, when including subset CD39 and CD73 expression data the nonseptic ICU patients and HD segregate into two clusters. Our findings suggest that multiparameter flow cytometric analyses in combination with UMAP and clustering analyses has the potential to identify patients that might develop a sepsis within the next days, providing the opportunity for early preventive measures.

Publications

• CD39 is upregulated during activation of mouse and human T cells and attenuates the immune response to Listeria monocytogenes. PLOS ONE, 13(5), e0197151.
  Raczkowski, Friederike; Rissiek, Anne; Ricklefs, Isabell; Heiss, Kirsten; Schumacher, Valéa; Wundenberg, Kira; Haag, Friedrich; Koch-Nolte, Friedrich; Tolosa, Eva & Mittrücker, Hans-Willi
  
• Generation and Function of Non-cell-bound CD73 in Inflammation. Frontiers in Immunology, 10.
  Schneider, Enja; Rissiek, Anne; Winzer, Riekje; Puig, Berta; Rissiek, Björn; Haag, Friedrich; Mittrücker, Hans-Willi; Magnus, Tim & Tolosa, Eva
  
• Imaging flow cytometry facilitates multiparametric characterization of extracellular vesicles in malignant brain tumours. Journal of Extracellular Vesicles, 8(1).
  Ricklefs, Franz L.; Maire, Cecile L.; Reimer, Rudolph; Dührsen, Lasse; Kolbe, Katharina; Holz, Mareike; Schneider, Enja; Rissiek, Anne; Babayan, Anna; Hille, Claudia; Pantel, Klaus; Krasemann, Susanne; Glatzel, Markus; Heiland, Dieter Henrik; Flitsch, Jörg; Martens, Tobias; Schmidt, Nils Ole; Peine, Sven; Breakefield, Xandra O. ... & Lamszus, Katrin
  
• CD73-mediated adenosine production by CD8 T cell-derived extracellular vesicles constitutes an intrinsic mechanism of immune suppression. Nature Communications, 12(1).
  Schneider, Enja; Winzer, Riekje; Rissiek, Anne; Ricklefs, Isabell; Meyer-Schwesinger, Catherine; Ricklefs, Franz L.; Bauche, Andreas; Behrends, Jochen; Reimer, Rudolph; Brenna, Santra; Wasielewski, Hauke; Lauten, Melchior; Rissiek, Björn; Puig, Berta; Cortesi, Filippo; Magnus, Tim; Fliegert, Ralf; Müller, Christa E.; Gagliani, Nicola & Tolosa, Eva
  
• Increased frequency of TIGIT+CD73-CD8+T cells with a TOX+TCF-1low profile in patients with newly diagnosed and relapsed AML. OncoImmunology, 10(1).
  Brauneck, F.; Haag, F.; Woost, R.; Wildner, N.; Tolosa, E.; Rissiek, A.; Vohwinkel, G.; Wellbrock, J.; Bokemeyer, C.; Schulze, zur Wiesch J.; Ackermann, C. & Fiedler, W.
  
• P2X7 is expressed on human innate‐like T lymphocytes and mediates susceptibility to ATP‐induced cell death. European Journal of Immunology, 52(11), 1805-1818.
  Winzer, Riekje; Serracant‐Prat, Arnau; Brock, Valerie J.; Pinto‐Espinoza, Carolina; Rissiek, Björn; Amadi, Miriam; Eich, Niklas; Rissiek, Anne; Schneider, Enja; Magnus, Tim; Guse, Andreas H.; Diercks, Björn‐Philipp; Koch‐Nolte, Friedrich & Tolosa, Eva