1120

Abstract

Introduction: Recent preclinical data in murine models, peripheral blood mononuclear cells (PBMCs), and neutrophils isolated from healthy human subjects demonstrate that triterpenoid analogs such as methyl 2-cyano-3,12-dioxoolean-1,9-dien-28-oate (CDDO-Me), are potent Nrf2 activators. However, the possibility of activating the Nrf2 pathway in critically ill patients to restore redox balance has not been studied. Methods: We prospectively enrolled 18 adult patients (≥18 years old) who were hospitalized in participating ICUs and had onset of septic shock within the previous 24–48 h. Blood samples were collected in cell preparation tubes with sodium citrate. PBMCs and monocytes were isolated and treated with vehicle or CDDO-Me (20 nM) for 20 h. Expression of Nrf2-regulated antioxidant genes (NQO1, HO-1, GCLM, GCLC, and FTL) and inflammatory gene IL-6 was measured with quantitative real-time PCR. Cells pretreated with CDDO-Me were stimulated with LPS (100 ng/ml) for an additional 1 h and ROS measured in culture media. A paired, two-tailed t-test was used to compare Nrf2 targets and inflammatory markers between the groups. Results: CDDO-Me treatment increased the expression of NQO1 (p=0.04) and decreased the expression of HO-1 (p=0.03) in PBMCs from patients with septic shock. Purified monocytes exhibited significant increases in the expression of NQO1 (p=0.01) and GCLM (p=0.003) after CDDO-Me treatment. Levels of other Nrf2 target genes (HO-1 and FTL) remained similar to those of vehicle-treated cells. PBMCs showed a trend toward increased IL-6 expression after CDDO-Me treatment, whereas purified monocytes showed a trend toward decreased IL-6. CDDO-Me treatment significantly increased ROS production in PBMCs (p=0.04). Although CDDO-Me pretreatment significantly attenuated ROS production to subsequent LPS exposure (p=0.03), the change was comparable to the change observed in vehicle-treated PBMCs. CDDO-Me pretreatment followed by LPS exposure had no significant effect on ROS levels in purified monocytes, though levels tended to increase. Conclusions: The Nrf2 pathway is differentially responsive to CDDO-Me activation in PBMCs from critically ill patients. Furthermore, CDDO-Me treatment results in increased ROS production. These findings may suggest a suppressed Nrf2 pathway in patients with septic shock.