P51 Variation in the Ex Vivo Response to LPS Stimulation in Healthy Volunteers: Identification of Extremes of Monocyte Function

Abstract

Introduction Numerous epidemiological and clinical studies have established a firm link between infection, the inflammatory response and cardiovascular disease. Furthermore, it is known that individuals differ in their responses to immune stimuli such as Lipopolysaccharide (LPS) that stimulates monocytes through Toll‐like receptor 4 (TLR4). In this study we set out to identify healthy individuals with extremes of monocyte response to LPS stimulation. Once identified these individuals will be studied further to identify genes responsible for the observed differences.Methods Small volumes of whole blood from 208 healthy blood donors were incubated with 0, 1 or 5 ng mL−1E. coli LPS for 4 hr at 37°C. Cell free plasma was collected both pre‐ and post‐incubation and the production of the pro‐inflammatory cytokine IL‐6 measured by ELISA. In addition, TaqMan genotyping was performed on two TLR4 SNPs, (rs4986790 and rs4986791, encoding D299G and T399I respectively) to determine whether these SNPs correlated with the observed functional responses.Results The range of IL‐6 produced was 0–26 ng mL−1 in response to 1 ng mL−1 LPS, and 2–56 ng mL−1 in response to 5 ng mL−1 LPS. Frequency distributions of IL‐6 levels show one‐tailed distributions for both stimuli with more outlying high extremes than low extremes. Quality control assays showed that both inter‐ and intra‐assay variability had a CV <10% and that the response correlates weakly with monocyte counts. Furthermore, repeat testing on 20 individuals demonstrated that the response to LPS is reproducible over time. No correlation was observed between TLR4 genotype and IL‐6 production following LPS stimulation.Conclusions Our stimulation results demonstrate that there is substantial inter‐individual variation in response to both 1 ng mL−1 and 5 ng mL−1 doses of LPS. We now have identified a number of individuals with either a high or low response to LPS stimulation. These individuals will be recalled for further functional testing and gene expression profiling to identify differences between the two extremes of monocyte function.