ABSTRACT
We describe methods to identify high and low responders in a whole-blood assay of lipopolysaccharide-stimulated cytokine responses. Two multivariate measures of the cytokine responses both captured high and low responses for each of the four individual cytokines that were assayed.
Subject(s)
Cytokines/drug effects , Lipopolysaccharides/pharmacology , Multivariate Analysis , Cytokines/blood , Female , Humans , MaleABSTRACT
The aim of this study was to determine if certain genotypes might be associated with variable responses to the candidate vaccine adjuvants RC529 and monophosphoryl lipid A (MPL), as well as to bacterial LPS, a structurally similar control stimulus. In this study, the +896 TLR4 polymorphism and selected cytokine polymorphisms were genotyped, and together with the donor sex, these factors were used to model the in vitro cytokine responses to RC529, MPL, and LPS. We show evidence that each of the three stimuli engage human TLR4, that each gave higher responses in men than women, and that TLR4 expression levels in blood monocytes were higher in men than women. From 74 to 92% of the response variation in the whole blood assay was between subjects, yet the multifactorial analyses accounted for only 3-18% of the variation within the study. Potentially there was a sex-dependent TLR4 effect since there was a significant sex-TLR4 interaction term in the multifactorial model for some responses. Since the genotypes we analyzed had such a modest impact in the study, we anticipate that +896 TLR4 genotype and the other factors analyzed in this study have only incremental effects in determining the overall response to TLR4-engaging stimuli.
Subject(s)
Adjuvants, Immunologic/pharmacology , Lipid A/analogs & derivatives , Lipid A/pharmacology , Adolescent , Adult , Alleles , Cytokines/biosynthesis , Cytokines/metabolism , DNA/genetics , DNA Primers , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Genotype , Humans , Kinetics , Lipopolysaccharides/pharmacology , Male , Membrane Glycoproteins/genetics , Membrane Glycoproteins/immunology , Middle Aged , Phenotype , Polymorphism, Genetic/genetics , Receptors, Cell Surface/genetics , Receptors, Cell Surface/immunology , Reverse Transcriptase Polymerase Chain Reaction , Sex Characteristics , Signal Transduction/genetics , Toll-Like Receptor 4 , Toll-Like ReceptorsABSTRACT
Immunization with native fusion (F) protein from respiratory syncytial virus (RSV) adsorbed to alum adjuvant generates greater than fourfold rises in serum neutralizing antibody titers in approximately 50% of seropositive humans. Using BALB/c mice we demonstrate herein that enhanced neutralization titers and accelerated clearance of virus from the lungs after challenge are possible if the attachment (G) glycoprotein is added to F protein-based vaccines. We further reveal for the first time that polarized type 2 T cell responses and immunopathology associated with G protein are inhibited by adjuvants recognized by toll-like receptors (TLR). Co-formulation with compounds that targeted TLR-2, TLR-3, TLR-4, or TLR-9 elicited significantly diminished type 2 T cell responses that caused granulocytic inflammation and eosinophilia in the airways after challenge. These results were not observed with recombinant IL-12 or QS-21. The data are important for improving combination vaccines for RSV.
