Neddylation inhibition attenuates LPS-induced TNF-α and IL-6 release and gene expression in BMDC. Protein analysis by western blot of (A, top panel) neddylated Cullin proteins and (A, middle panel) Cul1 protein using whole cell lysate of BMDC stimulated with LPS in the presence or absence of MLN4924 or dexamethasone at the indicated doses. Cells receiving treatment were preincubated with MLN4924 or dexamethasone for 2 hours followed by concurrent LPS stimulation (0.5 µg/mL) for 4 hours. α-tubulin protein levels were analyzed as an equal loading control. Quantification via ELISA of cytokines TNF-α (B) and IL-6 (C) and qPCR analysis of TNF-α (D) and IL-6 (E) transcripts in BMDC stimulated with LPS in the presence or absence of MLN4924 or dexamethasone at the indicated doses. Cells receiving treatment were preincubated with MLN4924 or dexamethasone for 2 hours followed by concurrent LPS stimulation (0.5 µg/mL) for 4 hours. One representative experiment of 3 is shown. ***P < .0001.