Fig. 1: TGM2 co-expression with adipose tissue macrophages (ATMs) in diet-induced obesity.
From: Transglutaminase 2-expressing macrophages modulate adipose tissue inflammation
C57BL/6 J mice were placed on chow diet (CD) or 60% high fat diet (HFD) at 6 weeks of age and remained on diet treatment until 16 weeks of age. A Total transglutaminase activity assay in eWAT from CD (n = 8) vs. HFD (n = 8) mice. B qPCR evaluation of Tgm2 mRNA expression in eWAT from CD (n = 10) or HFD (n = 10) mice. C Western blot for TGM2 protein expression in eWAT from CD control (n = 11) or HFD-treated mice (n = 16) normalized to B-ACTIN or VINCULIN expression. eWAT-derived single-nucleus RNA-seq from C57BL/6 J mice placed on a 60% HFD for 18 weeks (n = 3 pooled mice) showing Tgm2 expression in adipose tissue D global tissue populations or E immune cells in CD vs. HFD diet-treatment groups, where we observed a greater number of cells in the HFD sub-group as evidenced by the larger number of dots in the violin plot. Next, eWAT- derived from CD (n = 3) or HFD-treated mice (n = 3) was cryosectioned or paraffin sectioned and fluorescently stained for F Perilipin, F4/80 and intracellular TGM2 (or corresponding isotype control) for microscopy analysis. G CD (n = 12) vs. HFD (n = 12) eWAT-derived SVF was analyzed by flow cytometry for H number of macrophages and I Number of macrophages expressing TGM2 in CD vs. HFD diet treatment groups. I CD (n = 5) vs. HFD (n = 5) eWAT-derived SVF sorted preadipocytes, macrophages or monocytes evaluated for Tgm2 expression by qPCR assessment. J snRNAseq assessment of 60%HFD Tgm2- vs. Tgm2 + ATMs differential gene expression profile. Data are shown as bar graphs with SEM of five or twelve mice per control or treated group and are representative of two -four independent experiments. For all graphs, data normality status was used to determine statistical analysis. Normal data statistical significance was determined by parametric student’s unpaired t test, while not normally distributed data were analyzed using non-parametric Mann-Whitney test. ns = P > 0.05, *= P < 0.05, ** = P < 0.01, ***P < 0.001.
