Expression of <i>Coup-tfII</i> and <i>Hey1</i> in adipose tissue and isolated cell fractions.

<p>Expression of <i>Coup-tfII</i> (A) or <i>Hey1</i> (B) in gonadal (GN) and subcutaneous (SC) adipose tissues, as well as in isolated adipocytes and stromal vascular fractions (SVF) and in microvascular endothelial cells (MEC) derived from SC and GN adipose tissues obtained from obese mice, is shown relative to samples from lean mice (dotted line). Data are means ± SEM of at least 4 samples; * p < 0.05, ** p < 0.01, *** p < 0.001.</p

Body weight and fat depots of NUDE mice injected with 3T3-F442A preadipocytes with or without Hey1 gene silencing and kept on HFD for 4 weeks.

<p>Data are means ± SEM of n experiments. SC, subcutaneous; GN, gonadal; Hey1 kd (Hey1 gene silencing).</p

Effect of <i>Coup-tfII</i> gene silencing on <i>in vitro</i> differentiation of 3T3-F442A preadipocytes.

<p>(A) Western blotting of COUP-TFII protein in extracts of cells without (control) or with (clone #1 and <i>#2 CouptfII kd</i>) gene silencing; lane M represents the protein marker (50 kDa) and Pos represents a positive control. (B) Time course of <i>Coup-tfII</i> expression during differentiation without (⚫, black circles) or with (△, open triangles) knockdown (kd). (C-D) Oil Red O staining (C) and quantification; OD at 490 nm (D) at day 12 of differentiation. (E-H) Time course of expression of <i>Pref-1</i> (E), <i>PPAR-γ</i> (F), <i>CD36</i> (G) and <i>GLUT4</i> (H) during differentiation without (⚫, black circles) or with (△, open triangles) gene silencing. (I) Time course of <i>Hey1</i> expression without (⚫, black circles) or with (△, open triangles) <i>Coup-tfII</i> knockdown (kd). (J) Correlation between expression (delta CT levels) of <i>Coup-tfII</i> and <i>Hey1</i> with or without <i>Coup-tfII</i> knockdown. (K-M) Time course of expression of C/EBPα (K), <i>C/EBPβ</i> (L) and <i>C/EBPδ</i> (M) during differentiation without (⚫, black circles) or with (△, open triangles) gene silencing. Data are means ± SEM of 3 independent experiments; * p < 0.05; ** p<0.01; *** p<0.001; **** p<0.0001 versus control. The scale bar in panel C corresponds to 100 μm</p

Body weight and fat depots of NUDE mice injected with 3T3-F442A preadipocytes with or without COUP-TFII gene silencing and kept on HFD for 4 weeks.

<p>Data are means ± SEM of n experiments. SC, subcutaneous; GN, gonadal; COUP-TFII kd (COUP-TFII gene silencing).</p

Effect of bitter agonists on differentiation of 3T3-F442A preadipocytes.

<p>(a) Representative Oil-Red O stained 3T3-F442A cells at day 12 of differentiation in the presence of vehicle, 150 μM DB or 100 μM Q. (b) Quantification of the Oil-Red O uptake at differentiation day 6 (n = 4) and 12 (n = 8). (c-d) Relative mRNA expression of the markers leptin and adiponectin during differentiation in the absence and presence of DB or Q (n = 6). (e) Trypan blue cell viability assay, presented as the amount of dead cells (% cells stained) after the 12-day differentiation period in the absence and presence of DB or Q (n = 3). **: P<0.01, ***: P<0.001 vehicle vs bitter.</p

Comparison of the energy balance during treatment with bitter agonists of obese WT and α-gust^-/- mice.

<p>(a-b) Changes in body weight during daily intra-gastric administration of water, DB (60 μmol/kg) or Q (160 μmol/kg) for 4 weeks in high-fat diet (15 weeks) obese (a) WT (n = 9–12) and (b) α-gust^-/- mice (n = 8–12). Results are expressed as percentage change from baseline, defined as the mean body weight measured during one week before the treatment. (c) Combined weight of gonadal, subcutaneous and mesenteric fat pads as percentage of total body weight of control or bitter treated WT (n = 9–12) and α-gust^-/- mice (n = 9–12), at sacrifice. (d) Relative mRNA expression of UCP1 in gonadal WAT of control or bitter treated WT (n = 7–9) and α-gust^-/- (n = 7–8) mice. (e-f) Changes in energy intake during the 4-week treatment period in (e) WT (n = 6–8) and (f) α-gust^-/- mice (n = 7–8), expressed as percentage change from baseline, defined as the mean energy intake measured during 9 days before the treatment. **: P<0.01; ***: P<0.001 water vs bitter; ##: P<0.01 treatment (water vs DB) x genotype.</p

Hypothalamic neuropeptide mRNA expression after treatment with bitter agonists of obese WT and α-gust^-/- mice.

<p>Relative hypothalamic AgRP (a), NPY (b) and POMC (c) mRNA expression levels in HFD-obese WT (n = 7–10) and α-gust^-/- (n = 6–9) mice after 4 weeks of daily gavage with water, DB or quinine. *: P<0.05 water vs DB or Q; #: P<0.05; ##<0.01 treatment (water vs DB) x genotype.</p

Comparison of the energy balance between WT and α-gust^-/- mice on a high fat diet.

<p>(a) Time course of body weight of WT (n = 16) and α-gust^-/- (n = 16) mice, on a high-fat diet for 19 weeks post-weaning. (b) Energy intake (kcal/day) of WT (n = 16) and α-gust^-/- (n = 16) mice during the last 4 weeks before sacrifice. (c-d) Relative hypothalamic AgRP and POMC mRNA levels in WT (n = 10) and α-gust^-/- (n = 9) mice after 19 weeks on a HFD. (e-f) Respiratory quotient and heat production, measured continuously during 1 week in WT (n = 8) and α-gust^-/- (n = 8) mice. *: P<0.05; **: P<0.01, ***: P<0.001 WT vs α-gust^-/-.</p

Difference in adiposity between high-fat diet (19 weeks) induced obese WT and α-gust^-/- mice.

<p>(a) The sum of gonadal, subcutaneous and mesenteric white adipose tissue mass as percentage of total body weight in WT (n = 16) and α-gust^-/- mice (n = 16). (b) Plasma leptin levels in WT (n = 10) and α-gust^-/- mice (n = 11). (c-e) Relative gonadal, subcutaneous and mesenteric fat UCP1 mRNA levels in WT (n = 6–9) and α-gust^-/- mice (n = 6–9). (f) Relative intrascapular brown adipose tissue UCP1 protein level expression in WT (n = 4) and α-gust^-/- mice (n = 4) as determined by Western blot. *: P<0.05, **: P<0.01, ***: P<0.001 WT vs α-gust^-/-.</p