Oxidant/Antioxidant Status Is Impaired in Sepsis and Is Related to Anti-Apoptotic, Inflammatory, and Innate Immunity Alterations

Abstract

Oxidative stress is considered pivotal in the pathophysiology of sepsis. Oxidants modulate heat shock proteins (Hsp), interleukins (IL), and cell death pathways, including apoptosis. This multicenter prospective observational study was designed to ascertain whether an oxidant/antioxidant imbalance is an independent sepsis discriminator and mortality predictor in intensive care unit (ICU) patients with sepsis (n = 145), compared to non-infectious critically ill patients (n = 112) and healthy individuals (n = 89). Serum total oxidative status (TOS) and total antioxidant capacity (TAC) were measured by photometric testing. IL-6, -8, -10, -27, Hsp72/90 (ELISA), and selected antioxidant biomolecules (Ζn, glutathione) were correlated with apoptotic mediators (caspase-3, capsase-9) and the central anti-apoptotic survivin protein (ELISA, real-time PCR). A wide scattering of TOS, TAC, and TOS/TAC in all three groups was demonstrated. Septic patients had an elevated TOS/TAC, compared to non-infectious critically ill patients and healthy individuals (p = 0.001). TOS/TAC was associated with severity scores, procalcitonin, IL-6, -10, -27, IFN-γ, Hsp72, Hsp90, survivin protein, and survivin isoforms -2B, -ΔΕx3, -WT (p < 0.001). In a propensity probability (age-sex-adjusted) logistic regression model, only sepsis was independently associated with TOS/TAC (Exp(B) 25.4, p < 0.001). The AUC_TOS/TAC (0.96 (95% CI = 0.93-0.99)) was higher than AUC_TAC (z = 20, p < 0.001) or AUC_TOS (z = 3.1, p = 0.002) in distinguishing sepsis. TOS/TAC, TOS, survivin isoforms -WT and -2B, Hsp90, IL-6, survivin protein, and repressed TAC were strong predictors of mortality (p < 0.01). Oxidant/antioxidant status is impaired in septic compared to critically ill patients with trauma or surgery and is related to anti-apoptotic, inflammatory, and innate immunity alterations. The unpredicted TOS/TAC imbalance might be related to undefined phenotypes in patients and healthy individuals.

Keywords: TAC; TOS; antioxidant status; apoptosis; heat shock proteins; innate immunity; interleukins; oxidative stress.

Figure 1

Serum median levels of (A) total oxidative stress (TOS) and (B) total antioxidant capacity (TAC) in septic patients in comparison to patients with non-infectious (trauma/surgery) and healthy controls. The bold black line in box plots indicates the median per group, the bottom of the box indicates the 25th percentile, and the top of the box represents the 75th percentile; the T-bars (whiskers) and horizontal lines show minimum and maximum values of the calculated non-outlier values. Connectors indicate significantly higher levels in sepsis (post hoc Dunn’s pairwise tests with Bonferroni corrections).

Figure 2

Boxplots of serum median levels of (A) apoptotic caspase-9 (intrinsic pathway inducible caspase), (B) caspase-3 (effector caspase), (C) survivin (antiapoptotic protein) in septic patients in comparison to patients with non-infectious critical illness (trauma/surgery) and healthy controls. The bold black line in box plots indicates the median per group, the bottom of the box indicates the 25th percentile and the top of the box represents the 75th percentile; the T-bars (whiskers) and horizontal lines show minimum and maximum values of the calculated non-outlier values. Solid circles represent outliers, stars extremes. Connectors indicate significantly higher levels in sepsis (post hoc Dunn’s pairwise tests with Bonferroni corrections).

Figure 3

ROC curve for discriminating sepsis among critically ill patients. Independently, TOS/TAC, TOS, Hsp72 and Hsp90, SOFA score, survivin protein, and repressed TAC were independent discriminators of sepsis (AUROC > 60%, p < 0.05). The TOS/TAC ratio achieved the best AUROC (0.96 (95% CI = 0.93–0.99), p < 0.001).

Figure 4

Recordings of individual TOS/TAC ratios (open blue dots) in the sepsis and the two control groups (logarithmic scale). The TOS/TAC cut off point is depicted by the dotted line. Horizontal lines indicate post hoc differences between groups.

Figure 5

ROC curve for predicting mortality in septic patients. Independently, TAC (inverse prediction), TOS, survivin protein, IL-6, survivin transcript variants -2B and -WT (positive prediction) achieved significant receiver operating characteristic curves—AUROC of >0.70 (p < 0.004). The TOS/TAC ratio achieved the best AUROC (0.80, p < 0.001).