P026 IGG/IGM pre-incubation of whole blood may augment TNF-a production after LPS stimulation ex-vivo

Abstract

Introduction Polyclonal intravenous immunoglobulins (IVIGs) can modulate the host immune response. The purpose of the study was to investigate the effect of different immunoglobulin solutions on whole blood cytokine production capacity after Lipopolysaccharide (LPS) stimulation ex-vivo . Methods Peripheral heparinized blood was collected from 9 healthy volunteers for complete blood count, and ex-vivo stimulation with LPS. One hundred microliters of blood were added in 900 μl of RPMI 1640 for final volume 1 ml and placed to plastic culture dishes. Samples were added to wells and maintained at 37 °C in a 5% CO 2 atmosphere and then LPS was added (500 pg). After incubation of 8 h the culture plate was centrifuged (1800 rpm, 5 min) and supernatants were collected and stored at – 70 °C until cytokines level measurements using the ELISA method. In addition, samples were incubated either with IgG (20 mg IgG/ml) or IgG/IgM (15,2 mg IgG/ml and 2.4 mg/ml IgM) prior (pre-incubation, 2hrs) or simultaneously to LPS stimulation. Data are presented as mean ± SEM and comparisons were performed using Kruskal–Wallis test, with post hoc analysis using Dunn’s Multiple Comparison Test. Results Baseline TNF-a levels after LPS stimulation alone and incubation for 8-h was 460.0 ± 75.1 pg/ml. TNF-a levels after simultaneous IgG or IgG/IgM solution administration and LPS stimulation, were 553.9 ± 70.7 and 784.0 ± 152.6 respectively ( p > 0.05). TNF-a levels after pre-incubation with IgG or IgG/IgM and subsequent LPS stimulation were 773.2 ± 95.1 and 900.4 ± 123.0 respectively and the difference was statistically significant compared to baseline only for IgG/IgM ( p Conclusion Pre-incubation of whole blood with IgG/IgM solution and subsequent administration of LPS may augment whole blood TNF-a production capacity and thus express a pro-inflammatory effect. The study was partially funded by the Special Account for Research Grants of the National and Kapodistrian University of Athens, Grant 11103.