Outcome prediction value of circulating plasma DNA concentration in sepsis and stroke patients depends on rs352162 TLR9 gene polymorphism

Circulating extracellular DNA (cexDNA) in plasma represents a pathogenically related candidate biomarker for prediction of multiple organ failure development and sepsis outcome. One of key receptors that recognizes cexDNA as pathogen- and damage-associated molecular pattern is TLR9. The aim of the study was to assess whether the allelic variants of the TLR9 gene might contribute to predictive value of cexDNA in Intensive Care Unit (ICU) patients. The study included ICU patients (n = 187) with critical illness (post-surgery infection complications and stroke). CexDNA was isolated from plasma of patients using organic solvents and the concentration was determined on a spectrophotometer by SYBR Green. Genotyping of allelic variants of the TLR9 rs352162 gene was performed using a PCR and designed allele-specific tetraprimers followed by electrophoretic separation of the products. Calculations of medians, odds ratio and statistical significance between groups of patients were performed by the Fisher test, Mann-Whitney test and One-way ANOVA (GraphPad and SigmaStat software, USA). Data demonstrated that in sepsis, plasma cexDNA concentration served as a prognostic biomarker, and as such exhibited increased AUC values for fatal outcome in homozygous carriers of TLR9 CC genotype versus group of patients of TLR9 CT and TT genotypes (0.795 vs. 0.652, respectively, P<0.05). In neuroresuscitation ICU setting, increased levels of cexDNA in stroke patients on admission associated with development of health-associated infection and 30-days lethality (P<0.001). Plasma cexDNA >300 ng/ml associated with increased OR values for 30-day lethality in TLR9CC-positive stroke patients versus stroke patients with TLR9CT and TLR9TT genotypes (OR=63 vs. OR=6,145, P