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TLR4 genetic variation is associated with inflammatory responses in Gram-positive sepsis.

ABSTRACT: To identify important pathogen recognition receptor (PRR) pathways regulating innate immune responses and outcome in Staphylococcus aureus sepsis.We analysed whether candidate PRR pathway genetic variants were associated with killed S. aureus-induced cytokine responses ex vivo and performed follow-up in vitro studies. We tested the association of our top-ranked variant with cytokine responses and clinical outcomes in a prospective multicentre cohort of patients with staphylococcal sepsis.An intronic TLR4 polymorphism and expression quantitative trait locus, rs1927907, was highly associated with cytokine release induced by stimulation of blood from healthy Thai subjects with S. aureus ex vivo. S. aureus did not induce TLR4-dependent NF-?B activation in transfected HEK293 cells. In monocytes, tumor necrosis factor (TNF)-? release induced by S. aureus was not blunted by a TLR4/MD-2 neutralizing antibody, but in a monocyte cell line, TNF-? was reduced by knockdown of TLR4. In Thai patients with staphylococcal sepsis, rs1927907 was associated with higher interleukin (IL)-6 and IL-8 levels as well as with respiratory failure. S. aureus-induced responses in blood were most highly correlated with responses to Gram-negative stimulants whole blood.A genetic variant in TLR4 is associated with cytokine responses to S. aureus ex vivo and plasma cytokine levels and respiratory failure in staphylococcal sepsis. While S. aureus does not express lipopolysaccharide or activate TLR4 directly, the innate immune response to S. aureus does appear to be modulated by TLR4 and shares significant commonality with that induced by Gram-negative pathogens and lipopolysaccharide.

SUBMITTER: Chantratita N

PROVIDER: S-EPMC5218870 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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TLR4 genetic variation is associated with inflammatory responses in Gram-positive sepsis.

Chantratita N N Tandhavanant S S Seal S S Wikraiphat C C Wongsuvan G G Ariyaprasert P P Suntornsut P P Teerawattanasook N N Jutrakul Y Y Srisurat N N Chaimanee P P Mahavanakul W W Srisamang P P Phiphitaporn S S Mokchai M M Anukunananchai J J Wongratanacheewin S S Chetchotisakd P P Emond M J MJ Peacock S J SJ West T E TE

Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases 20160908 1

<h4>Objectives</h4>To identify important pathogen recognition receptor (PRR) pathways regulating innate immune responses and outcome in Staphylococcus aureus sepsis.<h4>Methods</h4>We analysed whether candidate PRR pathway genetic variants were associated with killed S. aureus-induced cytokine responses ex vivo and performed follow-up in vitro studies. We tested the association of our top-ranked variant with cytokine responses and clinical outcomes in a prospective multicentre cohort of patients ...[more]

PMID: 27615723

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Project description:Objective: It is unclear whether the host response of gram-positive sepsis differs from gram-negative sepsis at a transcriptome level. Using microarray technology, we compared the gene-expression profiles of gram-positive sepsis and gram-negative sepsis in critically ill patients. Design: A prospective cross-sectional study. Setting: A 20-bed general intensive care unit of a tertiary referral hospital. Patients: Seventy-two patients admitted to the intensive care unit. Interventions: Intravenous blood was collected for leukocyte separation and RNA extraction. Microarray experiements were then performed examing the expression level of 19,232 genes in each sample. Measurements and Main Results: There was no difference in the expression profile between gram-positive and gram-negative sepsis. The finding remained unchanged even when genes with lower expression level were included or after statistical stringency was lowered. There were, however, ninety-four genes differentially expressed between sepsis and control patients. These genes included those involved in immune regulation, inflammation and mitochondrial function. Hierarchical cluster analysis confirmed that the difference in gene expression profile existed between sepsis and control patients, but not between gram-positive and gram-negative patients. Conclusion: Gram-positive and gram-negative sepsis share a common host response at a transcriptome level. These findings support the hypothesis that the septic response is non-specific and is designed to provide a more general response that can be elicited by a wide range of different micro-organisms. The study included seventy-two critically ill patients admitted to the intensive care unit (ICU) of Nepean Hospital, Sydney, Australia. Of these, fifty-five patients were diagnosed to have sepsis, as confirmed by microbiological culture. The remaining seventeen patients did not have sepsis and were therefore used as controls. The study was approved by the hospital ethics committee and informed consent was obtained from all patients or their relatives. Patient Samples. Whole blood was taken from each patient on admission to ICU. Neutrophils were separated from whole blood using density-gradient separation with Ficoll-PaqueP P(Amersham). Subsequent neutrophil RNA extraction was performed using guanidinium thiocyanate (Ambion). Microarray Experiment. The neutrophil RNA was converted to cDNA, fluorescently labeled and hybridized to its complimentary sequences on the microarray (Invitrogen). The fluorescent signals on each micrroarray were captured using the GenePix 4000B laser scanner (Axon Instruments). Expression level of each gene was represented by the intensity of its fluorescent signal. Data Extraction. All signal intensity values were processed using background-subtraction method. Prior to analysis, all values were log-transformed and normalized by fitting a print-tip group Lowess curve. Normalization minimizes bias due to dye chemistry, signal intensity or location of a gene on the array. It ensures the detection of genes that are truly differentially expressed, instead of those caused by experimental artifacts or variation in the hybridization process. After normalization, genes that had more than 50% of data missing were removed. We then selected genes that had at least 80% of the data showing two-fold changes from the geneâs median values. After filtering, 1617 genes were available for further analysis.

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TLR4 genetic variation is associated with inflammatory responses in Gram-positive sepsis.

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TLR4 genetic variation is associated with inflammatory responses in Gram-positive sepsis.

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