29 - Nrf2 independent heme oxygenase-1 induction by auranofin is in lung macrophages

Abstract

Inhibition of thioredoxin reductase-1 (TXNRD1) activates nuclear factor (erythroid-derived 2)-like 2 (Nrf2) responses, attenuates experimental lung injury, and disproportionally increases heme oxygenase-1 (HO-1) levels. HO-1 has been investigated as a potential therapeutic target to prevent lung injury; however, the mechanisms by with TXNRD1 inhibition increases of HO-1 levels in macrophages is unclear. We recently demonstrated that the TXNRD1 inhibitor auranofin (AFN) induces HO-1 in lung epithelial cells via Nrf2-dependent mechanisms. TXNRD1 is abundantly expressed in both lung epithelia and macrophages. The present studies used murine alveolar macrophages (MH-S) to test the hypothesis that AFN-mediated HO-1 induction is Nrf2 dependent. Nrf2 expression was decreased in MH-S cells using siRNA. Knockdown was confirmed by qPCR and western blotting. Within 24 h of transfection, Nrf2 mRNA levels were 56% lower than in scramble siRNA transfected controls. In control cells, treatment with 0.5 µM AFN for 1 h decreased TXNRD1 activity by 90% and increased HO-1 mRNA levels by 2-fold when compared to vehicle-treated cells. In Nrf2 siRNA-treated cells, NQO1 mRNA levels were 80% less than in scramble siRNA transfected controls which is consistent with Nrf2 knockdown. Conversely, HO-1 mRNA levels were not different between scramble and Nrf2 siRNA-treated cells. Treatment with 0.5 µM AFN for 2h did not increase Nqo1 mRNA levels in Nrf2 siRNA-treated cells; however, HO-1 mRNA levels were 3-fold greater when compared to vehicle-treated knockdown cells. In conclusion, AFN induces HO-1 expression in MH-S cells which is consistent with our findings in previous studies. In contrast to lung epithelia, AFN-mediated HO-1 induction in lung macrophages is Nrf2 independent. The mechanisms by which TXNRD1 inhibition elicits HO-1 induction in alveolar macrophages are currently being investigated.