De novo adipocyte differentiation from Pdgfrβ+ preadipocytes protects against pathologic visceral adipose expansion in obesity

Figures

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  Fig. 1. Mural Pparg overexpression drives healthy visceral WAT expansion in obesity. a PdgfrbrtTA; TRE-Pparg2 (Mural-PpargTG) mice are generated by breeding the Pdgfrb rtTA transgenic mice to animals expressing Pparg2 under the control of the tet-response element (TRE- Pparg2). Littermates carrying only PdgfrbrtTA or TRE-Pparg2 alleles were used as the control animals for Mural-PpargTG. PdgfrbrtTA; TRE-Cre; Rosa26RmT/mG; TRE-Pparg2 (MuralChaser-PpargTG) mice are generated by breeding the “MuralChaser” (PdgfrbrtTA; TRE-Cre; Rosa26RmT/mG) mice to animals carrying TRE-Pparg2 transgene. MuralChaser mice were used as control animals for MuralChaser-PpargTG. b Control and Mural-PpargTG mice were fed a standard chow diet until 6 weeks of age before being switched to dox-containing high-fat diet (HFD). Body weights were measured weekly following the onset of HFD feeding. Control-HFD, n = 8; Mural-PpargTG-HFD, n = 9. Data points represent mean + s.e.m. c Fat mass and lean mass (normalized to body weight) of control and Mural-PpargTG mice after 16 weeks of dox-HFD feeding. Control-HFD, n = 8; Mural-PpargTG -HFD, n = 9. Bars represent mean + s.e.m. d Fat pad weight (normalized to body weight) of…

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  Fig. 2. Mural Pparg overexpression leads to metabolic benefits in obesity. a Glucose tolerance tests of control and Mural-PpargTG mice after 16 weeks of dox-diet feeding. Two-way ANOVA, *p < 0.05; Control-Chow, n = 7; Mural-PpargTG-Chow, n = 7; Control-HFD, n = 8; Mural-PpargTG-HFD, n = 9. Data points represent mean + s.e.m. b Insulin tolerance tests of control and Mural-PpargTG mice after 16 weeks of dox-HFD feeding. Student’s t-test, *p < 0.05; Control-HFD, n = 8; Mural-PpargTG-HFD, n = 9. Data points represent mean + s.e.m. c Six-hour fasting serum insulin levels in control and Mural-PpargTG mice after 16 weeks of dox-diet feeding. Two-way ANOVA, *p < 0.05; Control-Chow, n = 7; Mural-PpargTG-Chow, n = 7; Control-HFD, n = 8; Mural-PpargTG-HFD, n = 9. Bars represent mean + s.e.m. d The glucose infusion rate needed to maintain euglycemia (~137 mg/dl) during hyperinsulinemic–euglycemic clamp assays of conscious unrestrained control and Mural-PpargTG mice after 16 weeks of dox-HFD feeding. Student’s t-test, *p < 0.05; Control-HFD, n = 7; Mural-PpargTG-HFD, n = 5. Bars represent mean + s.e.m. e Basal and clamped rates of endogenous glucose production during hyperinsulinemic–euglycemic clamp…

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  Fig. 3. The metabolic benefits of mural Pparg overexpression depend on adiponectin. a Adiponectin-deficient (Adiponectin−/−) mice and Adiponectin−/−; PdgfrbrtTA; TRE-Pparg2 (Adiponectin−/−; Mural-PpargTG) mice were fed a standard chow diet until 6 weeks of age before being switched to dox-containing HFD. Body weights were measured weekly following the onset of HFD feeding. n = 6 per genotype. Data points represent mean + s.e.m. b Fat mass and lean mass (normalized to body weight) of Adiponectin−/− and Adiponectin−/−; Mural-PpargTG mice after 16 weeks of dox-HFD feeding. n = 6 per genotype. Bars represent mean + s.e.m. c Average adipocyte size in indicated fat depots from control and Mural-PpargTG mice after 16 weeks of dox-diet feeding. Bars represent mean + s.e.m. d Fat pad weight (normalized to body weight) of Adiponectin−/− and Adiponectin−/−; Mural-PpargTG mice after 16 weeks of dox-HFD feeding. * denotes p < 0.05 from Student’s t-test. n = 6 per genotype. Bars represent mean + s.e.m. e–l Representative immunofluorescence staining of Perilipin (green) and Mac-2 (red) in e, f gonadal, g, h perirenal WAT, i, j inguinal WAT, and k–l mesenteric WAT paraffin sections obtained from…

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  Fig. 4. Healthy visceral WAT expansion in obesity depends on mural Pparg expression. a PdgfrbrtTA; TRE-Cre; PpargloxP/loxP (Mural-PpargKO) mice are generated by breeding the PdgfrbrtTA transgenic mice to animals expressing Cre recombinase under the control of the tet-reponse element (TRE-Cre) and carrying floxed Pparg alleles (PpargloxP/loxP). Littermates carrying only PdgfrbrtTA and Pparg−loxP/loxP alleles (i.e. Cre−) were used as the control animals. b Control and Mural-PpargKO mice were fed a standard chow diet until 8 weeks of age before being switched to dox-containing HFD for another 20 weeks. Body weights were measured weekly. Control-HFD, n = 14; Mural-PpargKO-HFD, n = 13. Data points represent mean + s.e.m. c Fat mass and lean mass (normalized to body weight) of control and Mural-PpargKO mice after 20 weeks of dox-diet feeding. n = 7 per genotype. Bars represent mean + s.e.m. d Fat pad weight (normalized to body weight) of control and Mural-PparγKO mice after 20 weeks of dox-HFD feeding. * denotes p < 0.05 from Student’s t-test. Control-HFD, n = 14; Mural-PpargKO-HFD, n = 13. Bars represent mean + s.e.m. e–h Representative immunofluorescence staining of Perilipin (green) and…

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  Fig. 5. Pdgfrβ+ precursors contribute to rosiglitazone-induced adipocyte hyperplasia. a MuralChaser mice were fed on standard chow diet until 6 weeks old before being switched to HFD for 10 weeks. Mice were then administrated with dox-containing HFD (600 mg/kg) for 7 days (“pulse”). Following the pulse-labeling period, mice were switched back to regular HFD for another 4 weeks during which vehicle (1% methylcellulose) or rosiglitazone (Rosi) (10 mg/kg/day) was delivered by gavage (“chase”). Paraffin sections of WAT from these animals were stained with anti-GFP (green) and anti-perilipin (red), then counterstained with DAPI (blue; nuclei). b–e Representative ×63 magnification confocal immunofluorescence images of gonadal WAT sections after rosiglitazone treatment (“chase”). * indicates GFP-labeled perilipin-positive cells. Scale bar, 50 μm. f–i Representative ×63 magnification confocal immunofluorescence images of perirenal WAT sections after rosiglitazone treatment (“chase”). * indicates GFP-labeled perilipin-positive cells. Scale bar, 50 μm. j–m Representative ×63 magnification confocal immunofluorescence images of inguinal WAT sections after rosiglitazone treatment (“chase”). *…

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  Fig. 6. Mural cell Pparg is required for TZD-driven visceral WAT remodeling. a, b Control and Mural-PpargKO mice were treated as described in Supplemental Figure 10a. Body weights were measured weekly. Control + Vehicle, n = 8; Mural-PpargKO + Vehicle, n = 9; Control + Rosi, n = 11; Mural-PpargKO + Rosi, n = 19. n.s. denotes not statistically significant from two-way ANOVA. Data points represent mean + s.e.m. c Fat mass and lean mass (normalized to body weight) of control and Mural-PpargKO mice after 4 weeks of vehicle or rosiglitazone treatment. Control + Vehicle, n = 8; Mural-PpargKO + Vehicle, n = 8; Control + Rosi, n = 8; Mural-PpargKO + Rosi, n = 9. * denotes p < 0.05 from Student’s t-test. n.s. denotes not statistically significant. Bars represent mean + s.e.m. d Fat pad weight (normalized to body weight) of control and Mural-PpargKO mice after 4 weeks of vehicle or rosiglitazone treatment. Control + Vehicle, n = 8; Mural-PpargKO + Vehicle, n = 8; Control + Rosi, n = 8; Mural-PpargKO + Rosi, n = 9. * denotes p < 0.05 from Student’s t-test. n.s. denotes not statistically significant. Bars represent mean + s.e.m. e–l Representative immunofluorescence staining of Perilipin (green) and…