Sarabhai, Theresia Peter, Christoph Bär, Anne-Kathrin Windolf, Joachim Relja, Borna Wesselborg, Sebastian Wahlers, Thorsten Paunel-Görgülü, Adnana STS resistance does not depend on immunoglobulins or Fc receptor-triggered pathways. <p><b>A.</b> Pooled patient serum was subjected to serial depletion of immunoglobulins (Ig) using Protein G Plus/Protein A Agarose. Beads supernatant was collected after each depletion step. After 10 depletion steps, neutrophils from healthy volunteers (2 × 10<sup>6</sup>/ml) were cultured in the presence of the Ig-depleted fractions and tested for intrinsic apoptosis resistance. Cells were treated with STS (0.2 μM) for 18 h and apoptosis was determined by flow cytometry. Data are presented as means ± SEM of four to eight independent experiments. *p<0.05, **p<0.01, ***p<0.001 (t-test). <b>B.</b> Serum depleted for Ig for 8 times was subjected to SDS-PAGE and the gel was stained with Coomassie Brillant Blue (<i>left</i>). In addition, immunoblot analysis using Ig-specific antibody was performed (<i>right</i>). Arrows indicate the light and heavy chain of Ig. <b>C.</b> Beads used for the 8<sup>th</sup> depletion step were washed with Tris-HCl (25 mM, pH 7.5) and Tris-NaCl (25 mM Tris-HCl pH 7.5, 1 M NaCl), respectively, and supernatants were collected. Neutrophils were cultured in the presence of Ig-depleted serum, Tris-eluted supernatant (each 1%) and STS (0.2 μM) as already described. The percentage of cells displaying fragmented DNA (% subG1) is depicted. Graphs show means ± SEM of four independent experiments. *p<0.05 (t-test). <b>D.</b> Neutrophils were cultured in the presence of three different concentrations of Fc receptor blocker, using a peptide based technology, and patient serum (1%) and further treated with STS (0.2 μM). Relative apoptosis rate vs. control sample without Fc receptor blocker and STS is displayed. Data (means ± SEM) from five independent experiments are presented. No significant differences in apoptosis could be found when compared to the STS-treated control without Fc blocker (t-test).</p> serum levels;proteasomal degradation;neutrophil cell death;AAT-triggered antiapoptotic pathways;response syndrome;proapoptotic action;neutrophils;apoptosis induction;Previous work;serum factors;SIRS;AAT depletion;apoptosis resistance;AAT augmentation therapies;PKC inhibitor STS;apoptotic cell death;mechanism implicating caspases-independent pathways;STS treatment;response syndrome Excessive neutrophil activation;conditions causes;trauma patients;antiapoptotic Mcl;organ dysfunction syndrome;STS-induced cell death;ERK;antiapoptotic mediators 2017-05-11
    https://plos.figshare.com/articles/figure/STS_resistance_does_not_depend_on_immunoglobulins_or_Fc_receptor-triggered_pathways_/4999850
10.1371/journal.pone.0177450.g003