Microarray analysis of innate immune response induced by immunization with the adjuvant QS-21 in lymph node and muscle in mice.. Mus musculus

The goal of this study was to identify the transcriptional mechanisms involved in the activation of the immune system by QS-21, a triterpene glycoside purified from the bark of Quillaja saponaria which has adjuvant activity in vivo. Saponins represent a promising class of vaccine adjuvant. Together with the TLR4-ligand MPL, QS-21 is part of the Adjuvant System AS01, a key component of the Malaria and Zoster candidate vaccines that display demonstrated clinical efficacy. However, the mechanism of action of QS-21 in this liposomal formulation is poorly understood. Upon intra-muscular immunisation, we observed that QS-21 rapidly accumulated in CD169+ resident macrophages of the draining lymph node where it elicited a local innate immune response. Depletion of these cells abrogated QS-21-mediated innate cell recruitment to the lymph node, dendritic cell (DC) phenotypic maturation as well as the adjuvant effect on T cell and antibody responses to co-administered antigens. DCs rather than lymph node-resident macrophages were directly involved in T cell priming by QS-21 as revealed by the decrease in antigen-specific T cell response in Batf3−/− mice. Further analysis showed that the adjuvant effect of QS-21 depended on the integration of Caspase-1 and MyD88 pathways, at least in part through the local release of HMGB1. Taken together, this work unravels the key role of lymph node sentinel macrophage in controlling the adjuvant effect of a molecule proven to improve vaccine response in humans Overall design: C57BL/6 mice (6-8 weeks old) were immunized in both hind limbs, in the gastrocnemius (gcm) muscles and in a volume of 50 µl/muscle with QS-21 (100mg/ml) formulated in liposomes or PBS. Ileac lymph nodes were identified as draining in setup experiments. Each tissue sample consisted of a pool of tissues of 2 mice ( either dLN or muscle). Total RNA was isolated by homogenizing tissues in Tripure reagent (1 ml/100 mg tissue; Roche Applied Science) and then extracted with chloroform followed by RNeasy™ Minikit (Qiagen) according to the manufacturer’s protocol. A DNAse treatment was applied on the RNeasy column to avoid genomic DNA contamination. RNA was concentrated by ethanol precipitation, and quantified by RiboGreen™ (Life Technologies). 1 µg of each RNA sample was used for target preparation,using a one-cycle cDNA synthesis kit, and hybridized to GeneChip_ Whole Mouse Genome 430 2.0 arrays (Affymetrix). Data acquisition was performed using GeneChip Operating Software (Affimetrix). For each condition, 6 mice were treated. dLN from 2 mice were pooled for each hybridization. 3 replicate chips were analysed per condition. Muscle_QS-21_4hpost_2 and Muscle_PBS_2hpost_3 have been excluded since they did not pass QC.