Acute removal of common sepsis mediators does not explain the effects of extracorporeal blood purification in experimental sepsis

Abstract

The effect of extracorporeal blood purification on clinical outcomes in sepsis is assumed to be related to modulation of plasma cytokine concentrations. To test this hypothesis directly, we treated rats that had a cecal ligation followed by puncture (a standard model of sepsis) with a modest dose of extracorporeal blood purification that did not result in acute changes in a panel of common cytokines associated with inflammation (TNF-α, IL-1β, IL-6, and IL-10). Pre- and immediate post-treatment levels of these cytokines were unchanged compared to the sham therapy of extracorporeal circulation without blood purifying sorbent. The overall survival to 7 days, however, was significantly better in animals that received extracorporeal blood purification compared to those with a sham procedure. This panel of common plasma cytokines along with alanine aminotransferase and creatinine was significantly lower 72 h following extracorporeal blood purification compared to sham-treated rats. Thus, the effects of this procedure on organ function and survival do not appear to be due solely to immediate changes in the usual measured circulating cytokines. These results may have important implications for the design and conduct of future trials in sepsis including defining alternative targets for extracorporeal blood purification and other therapies.

Figure 1. Effects of extracorporeal blood purification (EBP) in cytokine removal

Plasma cytokine levels over time are shown for cecal ligation puncture (CLP) control, EBP, and sham-treated animals. All data were natural log transformed (ln pg/ml) and expressed as mean ± s.e.m. Each panel shows a separate cytokine: (a) tumor necrosis factor-α (TNF-α); (b) interleukin (IL)-6; (c) IL-1β; and (d) IL-10. *P < 0.05, vs. EBP at the same time points; ^#P < 0.05, vs. the baseline (at 18 h) in the same groups.

Figure 2. Effects of extracorporeal blood purification (EBP) on 1-week survival

Survival time (days) was observed from the start of cecal ligation puncture (CLP). P = 0.02, EBP vs. sham. P = 0.04, EBP vs. CLP control.

Figure 3. Effects of extracorporeal blood purification (EBP) on high-mobility group box 1 (HMGB-1)

Plasma HMGB-1 levels are shown over time (mean ± s.e.m., ng/ml). *P < 0.05, vs. EBP at the same time points; ^#P < 0.05, vs. the baseline (at 18 h) in the same groups.

Figure 4. Effects of extracorporeal blood purification (EBP) on organ function

Shown are plasma alanine aminotransferase (ALT; IU/l) and creatinine (Cr; mean ± s.e.m., mg/dl). *P < 0.05, vs. EBP at the same time points; ^#P < 0.05, vs. the baseline (at 18 h) in the same groups. (a) Plasma ALT. (b) Liver histology from EBP showing mild swelling of hepatocytes. (c) Liver histology slice from sham showing moderate to severe swelling of hepatocytes with focal piecemeal necrosis. (d) Plasma creatinine. (e) Kidney histology from EBP showing vacuolization in tubules; however, these changes are milder compared with sham (f). (f) Kidney histology from sham showing significant vacuolization in tubules.

Figure 5. Effects of exchange transfusion on interleukin-6 (IL-6)

Shown are plasma IL-6 levels over time for each of the four groups. Data were natural log transformed (ln pg/ml) and expressed as mean ± s.e.m. EBP, extracorporeal blood purification group; EBP to sham, sham animals received blood from the EBP animals; Sham, sham group; Sham to EBP, EBP animals received blood from the sham animals. IL-6 at 72 h in sham group was significantly higher than at baseline (18 h), and IL-6 at 72 h in the other three groups was lower than their baseline levels (P < 0.05).

Figure 6. Effects of exchange transfusion on survival

Survival time (day) was observed from the start of cecal ligation puncture (CLP) to 7 days. EBP, extracorporeal blood purification group; EBP to sham, sham animals received blood from the EBP animals; Sham, sham group; Sham to EBP, EBP animals received blood from the sham animals. P = 0.02, EBP vs. Sham; P = 0.045, Sham to EBP vs. Sham; P = 0.047, EBP to sham vs. Sham.

Figure 7. Effects of extracorporeal blood purification (EBP) on nuclear factor-κB (NF-κB) activation in circulating leukocytes

Data are expressed as median (range, n = 7). EBP, extracorporeal blood purification group; PMN, polymorphonuclear neutrophil; Sham, sham group. *P < 0.05, EBP vs. Sham.