Figure 5. Malat1 knockdown promotes alternative activation of macrophages.

(A) BMDMs were transfected with 20 nM control (con) GapmeR or Malat1 GapmeR. Forty-eight hours after transfection, the cells were treated with or without 5 ng/ml IL-4 for 24 hours. Levels of the indicated genes were determined by real-time PCR. n = 4; mean ± SD. (B) Experiments were performed as in A. Levels of Arg-1 and YM-1 were determined by Western blotting or ELISA. n = 3; mean ± SD. (C) Malat1^fl/fl and Malat1 mye^–/– mice were i.t. instilled with IL-4 (1 μg)/anti–IL-4 antibody (5 μg) immunocomplex (IL-4c) in 50 μl saline. Twenty-four hours after administration, alveolar macrophages were harvested and levels of the indicated genes determined. n = 2, 3, 3 mice for each group; mean ± SE. (D) BMDMs were transfected with 20 nM con GapmeR or Malat1 GapmeR. Twenty-four hours after transfection, the cells were trypsinized and plated on Seahorse XF-24 microplates. Twenty-four hours after plating, the cells were treated without (top) or with (bottom) 5 ng/ml IL-4 for 24 hours. The media were then replaced with OCR assay media and cultured for 1 hour, followed by sequential treatments with 3 μg/ml oligomycin (Oligo), 6 μM FCCP, and 1 μM rotenone (Rot) plus 0.5 μM antimycin A (Ant). Real-time OCR was recorded. n = 5 for each condition; mean ± SE. Representative of 2 to 4 independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001 by 1-way ANOVA with Bonferroni’s post hoc test.