Fig. 3. Obesity-induced alterations of PVAT phenotype and secretory profile.

a Representative pictures of heart, aorta and PVAT dissected from wt and ob/ob mice. b H&E stainings (from 1 out of 3 replicates) on wt and ob/ob AT depots sections. Scale bar: 100 µm. c Principal Component Analysis of the 293 secreted proteins consistently detected by MS in the conditioned media of wt and ob/ob PVAT explants. d Volcano plot showing proteins differentially secreted by wt and ob/ob PVAT samples (N = 18 vs. 18, with N = 6 biological replicates per depot). The limma package was used to compare different groups using the Ebayes algorithm (unpaired analysis) followed by Benjamini–Hochberg adjustment. Green spots: well-established adipokines. Blue spots: proteins involved in lipid transport. Red spots: proteins involved in axonal guidance. e Heatmap showing the 107 secreted proteins significantly up- or downregulated in the secretomes of ob/ob PVAT explants with Gene Ontology analysis showing the Biological Processes categories enriched in the two groups. Fisher’s Exact test p-values are shown. Hierarchical clustering was performed using the complete agglomeration method, Z-score-scaled abundances are displayed. N = 6 biological replicates per group. f and g Western blot on wt and ob/ob PVAT pooled conditioned media (f) and corresponding pooled lysates (g) (N = 5 biological replicates per pool) stained for cell-adhesion molecules Ncam1, Chl1 and L1cam and for the intracellular sympathetic marker TYR.H. AR PVAT aortic arch perivascular AT, TH PVAT thoracic PVAT, AB PVAT abdominal PVAT, VI WAT visceral white AT, SC WAT subcutaneous WAT, BAT interscapular brown AT, TYR.H. tyrosine hydroxylase, Cfd adipsin, Retn resistin, Negr1 neuronal growth regulator 1, Ncam1 neural cell-adhesion molecule 1, Chl1 close homologue of L1, L1cam neural cell-adhesion molecule L1, Lgals3 galectin 3, C1qtnf9 complement C1q and tumour necrosis factor-related protein 9, Rbp4 retinol-binding protein 4, M molecular weight marker.