Fig 1: Inhibition of autophagy inhibits interleukin (IL)-6 and IL-10 release by macrophages in vitro. Mouse RAW 264.7 macrophages were treated with different concentrations of 3-methyladenine (3-MA) in combination with (a, c) 100 ng mL-1 lipopolysaccharide (LPS) or (b, d) 1 µg mL-1 R-848 for (a, b) 6 h or (c, d) 16 h and IL-6 release was measured by ELISA. (e, f) RAW 264.7 cells were transfected with nontargeting (scrambled) or small interfering RNA (siRNA) targeting Becn1 and treated with different concentrations of (e) LPS or (f) R-848 for 16 h and IL-6 release was measured by ELISA. (g) Western blot confirmation of Beclin-1 protein knockdown in RAW 264.7 cells. The image is representative of three independent experiments. The full Western blot is shown in Supplementary figure 7. (h, i) RAW 264.7 macrophages were treated with different concentrations of 3-MA in combination with (h) 100 ng mL-1 LPS or (i) 1 µg mL-1 R-848 for 16 h and IL-10 release was measured by ELISA. (j, k) RAW 264.7 cells were transfected with nontargeting (scrambled) or Becn1 siRNA and treated with different concentrations of (e) LPS or (f) R-848 and IL-10 release was measured by ELISA. (l) RAW 264/7 cells were treated with LPS (100 ng mL-1) with or without 3-MA (10 mM) for 4 h and Il10 messenger RNA (mRNA) expression was measured by quantitative PCR (qPCR). (m) RAW 264.7 cells were transfected with scrambled or Becn1 siRNA and treated with LPS or R-848 for 4 h. Expression of Il10 mRNA was measured by qPCR. N = 3 independent experiments. **P < 0.01, ***P < 0.005, ****P < 0.001. [Colour figure can be viewed at wileyonlinelibrary.com]