Neutrophils from chronic lymphocytic leukemia patients exhibit an increased capacity to release extracellular traps (NETs)

Abstract

Chronic lymphocytic leukemia (CLL) is characterized by immune defects that contribute to a high rate of infections and autoimmune cytopenias. Neutrophils are the first line of innate immunity and respond to pathogens through multiple mechanisms, including the release of neutrophil extracellular traps (NETs). These web-like structures composed of DNA, histones, and granular proteins are also produced under sterile conditions and play important roles in thrombosis and autoimmune disorders. Here we show that neutrophils from CLL patients are more prone to release NETs compared to those from age-matched healthy donors (HD). Increased generation of NETs was not due to higher levels of elastase, myeloperoxidase, or reactive oxygen species production. Instead, we found that plasma from CLL patients was able to prime neutrophils from HD to generate higher amounts of NETs upon activation. Plasmatic IL-8 was involved in the priming effect since its depletion reduced plasma capacity to enhance NETs release. Finally, we found that culture with NETs delayed spontaneous apoptosis and increased the expression of activation markers on leukemic B cells. Our study provides new insights into the immune dysregulation in CLL and suggests that the chronic inflammatory environment typical of CLL probably underlies this inappropriate neutrophil priming.

Chronic lymphocytic leukemia cells increase neutrophils survival and promote their differentiation into CD16highCD62Ldim immunosuppressive subset

“…showed that a higher percentage of CD16 high CD62L dim neutrophils in HNSCC correlates with better clinical outcome because of the antitumor activity exerted by these cells through the release of NETs. By contrast, we previously reported in CLL that NETs promote leukemic cells survival and activation, suggesting that an increase percentage of the CD16 high CD62L dim subset could be detrimental instead of beneficial to CLL patients. Neutrophils displaying a B‐helper phenotype have been previously described at the marginal zone of the spleen where IL‐10 released by activated endothelial cells enable neutrophils to produce BAFF, APRIL and NET‐like structures that promote B‐cell activation.…”

“…Millrud et al 19 showed that a higher percentage of CD16 high CD62L dim neutrophils in HNSCC correlates with better clinical outcome because of the antitumor activity exerted by these cells through the release of NETs. By contrast, we previously reported in CLL that NETs promote leukemic cells survival and activation, 18 suggesting that an increase percentage of the CD16 high CD62L dim subset could be viable neutrophils (mean AE SD, n = 5 or 7, respectively). Statistical analysis was performed by Friedman test and Dunn's posttest.…”

“…Leukemic cells and neutrophils were isolated as we previously described. 18 Neutrophils (2 × 10 6 /ml) were cocultured with CLL cells (2 × 10 6 /ml) in Roswell Park Memorial Institute medium (RPMI) medium supplemented with 10% fetal calf serum and antibiotics (Gibco-Thermo Fisher Scientific Waltham, USA), during 72 hr at 37 C. At 24, 48 and 72 hr, samples were collected and stained with Annexin-V-FITC (Biolegend San Diego, USA) to assess neutrophil apoptosis by flow cytometry. HD-neutrophil samples used in cocultures had <1% of lymphocytes and <0.5% of monocytes (CD14 + ).…”

Clinico-Biological Implications of Modified Levels of Cytokines in Chronic Lymphocytic Leukemia: A Possible Therapeutic Role

“…IL-8 is one of the first cytokines described to act on B-CLL cells. Its serum concentrations are augmented in B-CLL patients compared to healthy controls, and it has been demonstrated that IL-8 increases B-CLL cell survival [104][105][106][107][108]. IL-8 participates via an autocrine modality to the B-cell accrual typical of this type of leukemia.…”

Revisiting the role of interleukin-8 in chronic lymphocytic leukemia

“…Interleukin-8 (IL-8/CXCL8) is one of the first molecules reported to play a key role in CLL biology 4 . Its serum levels are increased in most patients compared to age-matched healthy donors 5 , 6 and it is widely accepted that IL-8 prolongs CLL cell survival in an autocrine fashion 5 , 7 – 9 . However, there is no reported evidence of the expression of IL-8 receptors, CXCR1 and CXCR2, on circulating CLL cells.…”