AFF4 controls adipogenesis via autophagy.

(A) GSEA enrichment of adipogenesis and autophagy related pathway in AFF4-deficient hMSCs. (B, C) Representative TEM images and quantification of autophagy vesicles in preadipocytes from CON and CKO mice. Scale bar, 500 nm. n = 3. (D) Western blot analysis of autophagy-related proteins from primary preadipocytes of CON and CKO mice. CQ, chloroquine. (E, F) Representative immunofluorescent images and quantification of LC3 puncta in siCtrl and siAFF4 hMSCs. Scale bar, 25 μm. n = 3. (G) Western blot analysis of autophagy related markers in AFF4 overexpressing and Ctrl 3T3-L1 cells. (H-K) Autophagy inhibitors corrected augmented adipogenic differentiation of AFF4-overexpressing cells. (H) Representative images of oil red o staining in the presence of 3-MA and CQ. Scale bar, 25μm. 3MA, 3-methyladenine. CQ, chloroquine. (I) Quantification of oil red o staining, absorbance at 518 nm. n = 3. (J) Quantification of triglyceride. n = 3. (K) Western blot of adipogenic markers in the presence of 3-MA and CQ. All quantified data are presented by as mean ± SD. The P values were calculated by two-tailed Student’s t-test (C, F) or one-way ANOVA followed by the Tukey’s post hoc test (I, J).