Carbon Monoxide Protects against Hepatic Ischemia/Reperfusion Injury via ROS-Dependent Akt Signaling and Inhibition of Glycogen Synthase Kinase 3β
Figure 4
CO-induced PI3 K/Akt-dependent GSK3 inactivation regulates TLR4 responses and affects the ability of CREB and NF-κB-p65 to associate with CBP. RAW264.7 cells were stimulated with 100 ng/mL of LPS for 1 hour in the absence or presence of CORM2 and the PI3 K inhibitor (LY294002). (a) Total cell lysates were analyzed for phosphorylated GS, GSK3β, Akt, and CREB as well as total GS, GSK3β, Akt, CREB, and β-actin protein levels by Western immunoblot analysis. (b) The mRNA expression of TNF-α and IL-10 was determined by semiquantitative RT-PCR. GAPDH was used as an internal control. ((c) and (d)) RAW264.7 cells were stimulated with 0.1 μg/mL of LPS for 1 hour in the absence or presence of CORM2 (50 μM) and nuclear extracts were obtained. Interaction of CBP with NF-κB p65 and CREB was assessed by immunoprecipitation of CBP followed by immunoblotting for NF-κB or CREB. Total CBP served as the input standard.
