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. 2012 Apr 2;109(16):6130–6135. doi: 10.1073/pnas.1201351109

Fig. 5.

Fig. 5.

Chemical inhibition of iNOS activity enhances IL-1β and iNOS levels, whereas overexpression of CLIC4 down-regulates IL-1β and iNOS. (A–C) Primary mouse macrophages from wild-type mice were treated with LPS and IFNγ (1 μg/mL and 10 ng/mL, respectively) for 24 h (A) or for 48 h (B and C). Cells were pretreated with 2.5 mM l-NAME for 1 h where indicated. (A) Nuclear and cytosolic lysates were prepared and immunoblotted for CLIC4 and β-actin antibodies. (B) Whole-cell lysates were immunoblotted for IL-1β, iNOS, CLIC4, CLIC1, and β-actin. (C) Media from treated samples was assayed for cleaved IL-1β by ELISA. Bars represent the mean ± SEM of three replicates. (D) RAW macrophages were transduced with adenoviruses expressing nuclear-targeted CLIC4 (NUC-CLIC4), HA-tagged CLIC4 (HA-CLIC4), or GFP proteins overnight followed by treatment with LPS and IFNγ (1 μg/mL and 10ng/mL, respectively) for 24 h. Whole-cell lysates were immunoblotted for IL-1β, iNOS, CLIC4, GFP, and β-actin.