Identification of the key differential transcriptional responses of human whole blood following TLR2 or TLR4 ligation in-vitro.
The use of human whole blood for transcriptomic analysis has potential advantages over the use of isolated immune cells for studying the transcriptional response to pathogens and their products. Whole blood stimulation can be carried out in a laboratory without the expertise or equipment to isolate immune cells from blood, with the added advantage of being able to undertake experiments using very small volumes of blood. Toll like receptors (TLRs) are a family of pattern recognition receptors which recognise highly conserved microbial products. Using the TLR2 ligand (Pam3CSK4) and the TLR4 ligand (LPS), human whole blood was stimulated for 0, 1, 3, 6, 12 or 24 hours at which times mRNA was isolated and a comparative microarray was undertaken. A common NFκB transcriptional programme was identified following both TLR2 and TLR4 ligation which peaked at between 3 to 6 hours including upregulation of many of the NFκB family members. In contrast an interferon transcriptional response was observed following TLR4 but not TLR2 ligation as early as 1 hour post stimulation and peaking at 6 hours. These results recapitulate the findings observed in previously published studies using isolated murine and human myeloid cells indicating that in vitro stimulated human whole blood can be used to interrogate the early transcriptional kinetic response of innate cells to TLR ligands. Our study demonstrates that a transcriptomic analysis of mRNA isolated from human whole blood can delineate both the temporal response and the key transcriptional differences following TLR2 and TLR4 ligation.
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom.
Simon
S
Blankley
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom.
Christine M
CM
Graham
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom.
Ashleigh
A
Howes
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom.
Chloe I
CI
Bloom
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom; Department of Respiratory Medicine, Imperial College Healthcare NHS trust, London, United Kingdom.
Matthew P R
MP
Berry
Baylor Institute for Immunology Research/ANRS Center for Human Vaccines, INSERM, Dallas, Texas, United States of America; Systems Immunology, Benaroya Research Institute, Seattle, Washington, United States of America.
Damien
D
Chaussabel
Baylor Institute for Immunology Research/ANRS Center for Human Vaccines, INSERM, Dallas, Texas, United States of America.
Virginia
V
Pascual
The Jackson Laboratory for Genomic Medicine, Farmington, Connecticut, United States of America.
Jacques
J
Banchereau
Department of Respiratory Medicine, Royal Free London NHS Foundation Trust, University College London, London, United Kingdom.
Marc
M
Lipman
Division of Immunoregulation, MRC National Institute for Medical Research, London, United Kingdom; Department of Medicine, National Heart and Lung Institute, Imperial College, London, United Kingdom.
Anne
A
O'Garra
D001769:Blood / Q000378:metabolism*; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D055666:Lipopeptides / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism; D046228:Microarray Analysis; D016328:NF-kappa B / Q000378:metabolism; D013997:Time Factors; D051195:Toll-Like Receptor 2 / Q000378:metabolism*; D051197:Toll-Like Receptor 4 / Q000378:metabolism*
D055666:Lipopeptides; D008070:Lipopolysaccharides; D016328:NF-kappa B; C508075:Pam(3)CSK(4) peptide; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4