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. 2020 May 29;10(4):673–697. doi: 10.1016/j.jcmgh.2020.05.007

Figure 7.

Figure 7

IL23 promotes induction of autophagy pathways in human macrophages. (A–E) MDMs were treated with 10 ng/mL IL23 or IL12 for 48 hours. (A and B) LC3II was assessed by (A) flow cytometry with a representative histogram plot with mean fluorescence intensity (MFI) values and a summary graph with MFI (n = 10 from 2 independent experiments). (B) Western blot. (C and D) ATG5 and ATG16L1 were assessed by (C) flow cytometry (MFI) (n = 10 from 2 independent experiments for ATG5; n = 6 for ATG16L1) or (D) Western blot. (E) Fold increase in cathepsin D activity (n = 12 from 2 independent experiments). Pepstatin A (peps.) was used as a control for reduced lysosomal activity. (F–L) MDMs were transfected with scrambled or the indicated siRNAs, alone or in combination (comb). (F and G) Knockdown efficacy per (F) flow cytometry (n = 4) and (G) Western blot. (H) LC3II expression (MFI) (n = 12 from 2 independent experiments). (I) Intracellular bacterial clearance (CFU) (n = 12 from 2 independent experiments). (J and K) MDMs then were treated with IL23 or IL12 for 48 hours. (J) LC3II expression by flow cytometry (MFI) (n = 6). (K) Intracellular clearance bacterial clearance (CFU) (n = 10 from 2 independent experiments). (L) Cell death per annexin V+ cells (n = 4). A total of 50–100 J/m2 UV-treated cells served as a positive control. Means + SEM. ∗∗∗ P < .001, †P <1 × 10-4, and ††P <1 × 10-5. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NT, no treatment; scr, scrambled; Tx, treatment.