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. 2018 Jul 27;9:1714. doi: 10.3389/fimmu.2018.01714

Figure 4.

Figure 4

Effect of changes in AMP-activated protein kinase (AMPK) activity on rheumatoid arthritis (RA) fibroblast-like synoviocyte (FLS) function. (A) Effect of the AMPK agonist and inhibitor on the expression of pro-inflammatory cytokines and matrix metalloproteinases (MMPs). RA FLSs were treated with TNF-α (10 ng/ml) for 12 h in the presence or absence of AICAR (1 mM) or Compound C (Comp C, 1 µM). mRNA expression was determined by quantitative real-time PCR. (B) Effect of AMPK inhibition on the expression of pro-inflammatory cytokines and MMPs. RA FLSs transfected with scramble (scr) or glycogen synthase 1 (GYS1) shRNA (shG) and AMPKα shRNA (shA) were treated with TNF-α (10 ng/ml) for 24 h in the presence or absence of Compound C (Comp C, 1 µM) or AICAR (1 mM). (C) Effect of AMPK activation or inhibition on the proliferation of RA FLSs. Cell proliferation was determined by EdU assay. The data are presented as the mean ± SD of five independent experiments. (D,E) Effect of AMPK activation or inhibition on the migration (D) and invasion (E) of RA FLSs. Cells transfected or not transfected with scramble control (Scr) or GYS1 shRNA (shG) or AMPKα shRNA (shA) were serum starved overnight, seeded in a Boyden chamber, allowed to migrate for 8 h, fixed, and stained with crystal violet. TNF-α (10 ng/ml) was used as a chemoattractant. An in vitro invasion assay was performed using inserts coated with a Matrigel basement membrane matrix in Boyden chambers. The data are presented as the mean ± SD of five independent experiments. *P < 0.05, **P < 0.01 vs. DMSO or scramble; #P < 0.05, ##P < 0.01 vs. TNF-α or scr + TNF-α or shG; &P < 0.05 vs. shG + TNF-α.