Fig. 1: Fasting drives a lysosomal program in murine adipose tissue.
From: Non-canonical lysosomal lipolysis drives mobilization of adipose tissue energy stores with fasting
a Murine model of fasting-induced lipolysis. Mice were fasted for 24 h and refed for 6 h (C57Bl6 male n = 5; female mice n = 6 cont, 6 fast, 5 refed, 8-weeks-old). Statistical significance was assessed by one-way ANOVA/Tukey’s test. Left: male serum glycerol: *p = 0.03; male serum non-esterified fatty acids (NEFA): ****p < 0.0001. Right: female glycerol: ***p = 0.0001, **p = 0.007; NEFA: ****p < 0.0001. When present error bars indicate s.d.m. (a). b Inguinal (iWAT) and gonadal (gWAT) adipose tissue immunoblots for canonical lipase (ATGL), lysosomal transcriptional regulators (TFEB, TFE3, MITF), and the lysosomal lipase (LAL). The control, fasting, refeeding protocol was the same as in ‘a’. Note: tubulin controls run on different gels. c qPCR of adipocytes isolated from inguinal adipose tissue, including canonical lipolytic genes and lysosomal genes. Male n = 6 con, 5 fast, 6 refed; Female n = 6 con, 6 fast, 5 refed: *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001, two-way ANOVA/Dunnett’s test. The control, fasting, refeeding protocol was the same as in ‘a’. d Heat-map comparing isolated adipocytes (from C) to nutrient-restricted adipocyte cultures derived from primary adipocyte progenitor (AP) cells (n = 3 biological replicates) or 3T3L1 cells (n = 3 biological replicates). e. Whole mount immunofluorescence staining for marker of lysosomes (LAMP1) in adipose tissue. Arrow=putative LAMP1+ puncta indicating lysosomes. LD=lipid droplet. N = nucleus. Scale = 10 micron. N = 5 control, 7 fasted stained in this manner and used for blinded counting shown in ‘f’. f Blinded observer counted perinuclear LAMP1+ puncta in perilipin+ adipocytes: n = 5 control, 7 fasted; two-sided T-test; error bars S.D.M. g Perilipin levels were measured by ELISA in lysosome preparations from isolated adipocytes. Perilipin was undetectable in the lysosome preparations from fed mice; therefore the lower-limit of the assay (0.055 ng/mL) was used for the purpose of the graph. Each dot represents one mouse, n = 8, significance assessed by two-tailed t-test; error bars S.D.M.
