Table S1. General findings of the studies included in this review.

Reference/Year of Study design Main objective Main results publication Gurlevik G et al. Experimental To evaluate the effects of granulocyte - The chemotactic activity of neutrophils treated with G-CSF increased. 2007 (11) study colony-stimulating factor (G-CSF) in - Treated rats showed increased survival. the immune function of granulocytes - Neutrophils have impaired activity against peritoneal infection. during severe abdominal infection and - Treatment with G-CSF improved the immune function of neutrophils and in the survival of rats with peritonitis. . Martin EL et al. Experimental To analyze the specific role of PI3Kγ - The absence of PI3 kinase activity confers protection against mortality and 2010 (12) study activity in the pathogenesis of sepsis. multiple organ failure. - In severe CLP, PI3 kinase blockade-maintained blood pressure levels, protected against organ damage (liver and intestines) and increased tissue apoptosis. - There was a reduction in neutrophil chemotactic activity. - In severe CLP, there was a failure in neutrophil migration to the peritoneum, accompanied by an increase in GRK2 and a reduction in CXCR2. - PI3 kinase blocks the influx of neutrophils. - Mice treated with PI3 kinase blockade showed reduced mortality and improved neutrophil migration and bacterial control. Wang JF et al. Experimental and To evaluate the level of PD- - PD-L1 levels were increased in mice subjected to CLP with a 24-hour peak. 2015 (13) clinical expression in neutrophils of mice and - Low PD-L1+:CD11a, CD62L and CCR2 and high CD16 and CD64 neutrophils (prospective sepsis patients. To evaluate if showed impaired migration capacity. case-control) neutrophils obtained from mice would - Neutrophils may induce lymphocyte apoptosis via direct contact mediated by study induce lymphocyte apoptosis by PD- PD-L1. L1 through a Transwell system and - Low CD11a and CCR2 was associated with reduced neutrophil . anti-PDL1 antibody. - PD-L1+ neutrophils showed impaired migration. - Sepsis may induce positive PD-L1 regulation in neutrophils, an inhibitory subtype that can impair chemotaxis and induce lymphocyte apoptosis during sepsis. Schmidt EP et al. Experimental and To evaluate acute lung injury (ALI) - Exposure of endothelial layers after exposure to bacterial particles such as 2012 (14) clinical study associated with sepsis correlating with endotoxins leads to neutrophil recognition and adhesion to the vascular pulmonary endothelial glycocalyx membrane via ICAM-1 and VCAM-1. degradation, and - Heparinase may be a potential lung protector during sepsis. neutrophil adhesion. - The exposure of endothelial molecules is required for neutrophil adhesion, and low glycocalyx mechanisms may induce neutrophil extravasation. Souto FO et al. Experimental and To evaluate the role of the CCR2 - TLR2 and TLR4 participate in neutrophil chemotaxis. 2010 (15) clinical study receptor in the infiltration - Chemotaxis via TLR was abolished in neutrophils treated with CCR2 of neutrophils and tissue damage in antagonists or CCR2 knock-out mice. remote organs during sepsis. - The expression of CCR2 was increased in neutrophils treated with lipoteichoic acid (LTA) and LPS. - CHX ( synthesis) is necessary for CCR2 expression induced by LTA and LPS stimuli. - Neutrophil chemotaxis was completely abolished after NFkB inhibition. - TLRs directed the expression of CCR2 via NFkB. - The increased regulation of CCR2 in sepsis is mediated by TLR signaling. - TLR signaling induces CCR2 expression in neutrophils, granting these cells the capacity to migrate in response to CCR2 ligands. Xu R et al. Clinical study To evaluate C5aR and C5L2 levels in - Sepsis patients showed high neutrophil counts compared to SIRS patients. 2016 (16) neutrophils from patients with SIRS - The percentage expression of C5L2 was reduced in neutrophils from patients and sepsis. with sepsis. - C5aR and C5L2 expression may serve as a biomarker to predict survival in sepsis patients. - Neutrophil exposure to C5a causes lower expression of C5aR. - Neutrophils of sepsis patients showed reduced ability to secrete IL-8 in response to C5a in vitro compared to healthy controls and SIRS patients. - High levels of C5a and C5L2 in neutrophils were associated with survival in sepsis patients, while low levels of C5aR and C5L2 in neutrophils were a good predictor of mortality in sepsis patients. - C5a induces disturbances in neutrophil production of IL-8. This phenomenon can be associated with the “immune paralysis” that occurs in sepsis. Demaret J et al. Clinical study To evaluate the alterations in - Analyses were carried out in late-stage sepsis to study neutrophils in the 2015 (17) neutrophil immunosuppression in immunosuppressive stage of sepsis. sepsis patients using functional and - The number of circulating neutrophils was higher in sepsis patients than in phenotypic studies. control participants. - CXCR1 and CXCR2 expression was also reduced in sepsis patients. - Neutrophils stimulated by IL-8, fMLP and GRO showed diminished migration capacity in sepsis patients compared to healthy controls. - There was no difference in phagocytosis capacity between patients and controls. - Oxidative stress analysis showed that oxidative stress in patient cells was lower than that in cells from healthy volunteers. - The production of bactericidal agents was compromised after septic shock. - Apoptosis was diminished. - There was a reduction in neutrophil migration capacity. Blom C et al. Clinical study To evaluate inflammatory markers in - Stimulation of granulocytes with the compound did not increase the 2015 (18) the plasma of severe sepsis patients adhesion of naïve cells. in order to create an experimental - Stimulation of granulocytes with the cytokine compound reduced the adhesion compound of and capacity of these cells. investigate the activation and - In the first 24 hours of sepsis, patients showed granulocytes with greater interaction between granulocytes and regulated adhesion profiles. human cerebrovascular endothelial cells in vitro. Khaenam P et al. Clinical study To evaluate the transcriptomic profile - Neutrophils are important in identifying transcriptomes that may predict severity 2014 (28) of neutrophils, monocytes and in sepsis. dendritic cells in order to obtain - Neutrophils showed altered responses in sepsis patient samples, with the immune response profiles on a identification of 709 related to the immune response. systems scale. - A total of 30 transcripts showed high accuracy in the prediction of sepsis.

Parlato M et al. Clinical study To evaluate the apoptotic pathway in - A greater number of apoptotic cells was observed after usage of the anti-CD24 2014 (21) human neutrophils triggered by the antibody. surface molecule CD24. - An increase in CD16 expression was detected on the cellular membrane of neutrophils marked with anti-CD24. - The signal for apoptosis through CD24 in neutrophils was translated via caspase 3/caspase 9. - Inhibition of apoptosis via blockade of the NADPH system with diphenylene iodonium (DPI) and apocynin demonstrates that the use of DPI also blocks apoptosis induced by the CD24 pathway. - CD24 induces apoptosis in neutrophils via collapse of the mitochondrial membrane. - Oxidative stress contributes to neutrophil death via CD24 - Neutrophils from sepsis patients showed reduced CD24 expression - The binding of anti-CD24 antibody with CD24 induces apoptosis in neutrophils. - CD24 is a trigger for cell death via mitochondrial membrane depolarization, ROS generation and activation of procaspase-9 in human neutrophils. - Neutrophil apoptosis in critical patients took 8 to 10 hours, while the half-life of neutrophils from healthy persons is 7 to 10 hours. Ubagai T et al. Clinical study To evaluate the expression of the - Expression levels of the TREM1 in 4 out of 7 patients with pneumonia 2014 (19) triggering receptor expressed in were increased. myeloid cells (TREM)-1 gene and G- - Six patients showed low expression of TREM1 when compared to healthy protein coupled receptor kinase patients. (GRK)-2 in granulocytes isolated from - The levels of TREM1 were lower in sepsis patients. patients. - The incubation of granulocytes with the TREM1 oligopeptide increased the levels of TREM1 RNA. - The levels of GRK2 mRNA expression were reduced in patients with bacterial infection, indicating suppression of granulocyte migration. Daigo K et al. Clinical study To evaluate PTX3 ligands in sepsis by - High concentrations of PTX3 were detected in blood samples. 2012 (25) means of a proteomic study of - A total of 104 candidate that interact with PTX3 were identified; these circulating PTX3 complexes. proteins are involved in the activation of the complement system, opsonization of pathogens, regulation of inflammation and deposition of extracellular matrix. - The proteomic profile demonstrated that PTX3 forms complexes with several components of NETs. - PTX3 may have a defense function and is a putative therapeutic target. Santos SS et al. Clinical study To evaluate the generation of reactive - The production of NO by monocytes was higher in sepsis patients than in 2012 (26) oxygen species (ROS) and nitric heathy controls. oxide (NO) by monocytes and neutrophils in sepsis patients and correlate their levels with clinical outcomes.

Kothari N et al. Clinical study To evaluate the activity of the - SIRS and sepsis patients showed increased levels of myeloperoxidase, 2011 (27) myeloperoxidase (MPO) enzyme in although patients with septic shock showed lower counts of cells and MPO. the plasma of critical patients and - Cytokine levels were also elevated in SIRS, sepsis and septic shock patients. verify the hypothesis that concentrations in the plasma are higher in sepsis and systemic inflammatory conditions. Paunel-Görgülü A Clinical study To evaluate the role of soluble Fas - Apoptosis was reduced during the observed period when compared with control et al. 2011 (22) (sFas) in the regulation of extrinsic cells. neutrophil apoptosis in post-traumatic - Elevated levels of soluble Fas may inhibit apoptosis in circulating neutrophils patients and in the development of and prolong cell activity. sepsis. - Apoptosis was reduced in patients who developed sepsis following trauma. Danikas DD et al. Clinical study To evaluate the impact of neutrophil - Granulocyte phagocytic activity on the day of admission was reduced in severe 2008 (24) and phagocytic activity on sepsis patients who did not survive compared to healthy persons. the outcome of patients with severe - The expression of CD64 in granulocytes and monocytes was significantly sepsis. elevated in severe sepsis patients who survived compared to healthy controls. - Surviving patients showed negative regulation of CD64 expression in granulocytes and monocytes when compared to levels on the day of admission and at resolution. - A positive correlation between granulocyte activity and CD64 expression at admission was demonstrated. - In a univariate analysis, a significant positive correlation between clinical outcomes and granulocyte phagocytic activity as well as between clinical outcomes and CD64 expression in monocytes and granulocytes was shown. - The increase in phagocytic activity and CD64 expression in phagocytes had a favorable impact on survival. - Granulocyte phagocytic activity appears to be an independent predictor of survival in severe sepsis patients. Martins PS et al. Clinical study To evaluate cellular activation in the - There was lower TLR2 expression in the group of patients with septic shock 2008 (29) context of sepsis and its correlation than in the control group. with organic dysfunction. - A SOFA score of 7 was the cut-off point for patients who did or did not survive. Berger C et al. Double-blind To evaluate the effect of lidocaine on - Lidocaine strongly inhibited the increase in induced in neutrophils 2014 (20) randomized leucocyte recruitment in sepsis 4 and 24 hours after the initiation of medication. clinical trial patients. - Neutrophils showed increased adherence time for cell rolling in P- /ICAM-1/IL-8 medium compared to P-selectin/ICAM-1 medium. - In isolated neutrophils, lidocaine was shown to block rolling. - Pre-treatment with lidocaine and Ro 31 8220 significantly reduced adhesion. - Neutrophils incubated with plasma obtained from patients before administration of medication showed higher transmigration than neutrophils incubated with plasma from healthy volunteers. - Transmigration of neutrophils incubated with plasma from sepsis patients treated with lidocaine was significantly reduced compared to that of neutrophils incubated with plasma from sepsis patients treated with normal saline.

Celma IF et al. Double-blind To evaluate the effect of parenteral - levels in CD15+ neutrophils were reduced in patients treated with 2009 (23) randomized vitamin C administration in neutrophil vitamin C compared to healthy controls but compared to patients treated with clinical trial apoptosis by determining the placebo. expression of the Fas receptor and - The neutrophil count was high in both treated groups compared to the healthy levels of caspase-3, poly (ADP- control group. ribose) polymerase (PARP) and Bcl-2 - Caspase-3, PARP and Bcl-2 levels were significantly reduced in healthy in neutrophils from patients who had volunteers compared to the placebo and vitamin C-treated patients. undergone surgery for abdominal - Caspase 3 and PARP levels were lower in the group treated with vitamin C sepsis. than in the group treated with placebo. - Bcl-2 levels were higher in the group treated with vitamin C than in the group treated with placebo.

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