Apolipoprotein E (ApoE) orchestrates adipose tissue inflammation and metabolic disorders through NLRP3 inflammasome

Figures

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  Fig. 1. Downregulation of APOE expression in adipose tissue during obesity and inflammation. a Previous human samples data was collected from the GEO dataset (GSE9624, n = 5–6). b The reads count of ApoE in indicated groups from RNA-Seq of human OAT (n = 30, BMI < 24; n = 17, BMI ≥ 24). c Individual correlation between ApoE mRNA level and BMI value. d-g QPCR analysis of ApoE gene expression in eWAT (d, f) and iWAT (e, g) from WT with HFD feeding (n = 6) and db/db mice (n = 7). eWAT, epididymal WAT. iWAT, inguinal WAT. HFD, high-fat diet. h Previous human samples data of ApoE expression were collected from the GEO dataset (GSE54350, n = 6). i Plasma LPS levels in mice after 10-week HFD feeding (n = 6). j, k QPCR analysis of ApoE gene expression in LPS-stimulated PM (i) and BMDM (j) (100 ng/mL, 4 h, n = 3). VAT, visceral adipose tissue. OAT, omental adipose tissue. PM, peritoneal macrophages. BMDM, bone marrow-derived macrophages. All data represent mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001. Significance is performed with unpaired 2-tailed Student’s t-test (a, b, d-k) and Spearman’s correlation (c)

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  Fig. 2. Loss of APOE modulates lipid metabolism during diet-induced obesity. a Body weight in chow-fed WT and ApoE-/- mice (n = 9). b Representative images of WT and ApoE-/- mice fed with chow for 12 weeks. c, d Body weight progression and representative pictures of WT and ApoE-/- mice fed with HFD (n = 6–7). e-h Representative images of eWAT (e), iWAT (f), iBAT (g), and muscles (h) of WT and ApoE-/- mice fed with HFD. i, j Tissue weight following HFD-fed for 10 weeks. k, l Averaged daily food intake of mice from chow (k) and HFD-fed (l) mice (n = 6–7). m, n Triglyceride in plasma and liver from HFD-fed WT and ApoE-/- mice. o-q Plasma total cholesterol (TC), HDL cholesterol (HDL-C), and LDL cholesterol (LDL-C) in WT and ApoE-/- mice fed with HFD (n = 8). All data represent mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ns, no significance

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  Fig. 3. ApoE abrogation ameliorates HFD-induced glucose clearance. a, b Different during fasting blood glucose from chow and HFD-fed mice at the indicated time (n = 8). c Basal plasma insulin levels after overnight fasting (n = 8). d, e GTT (d) and AUC (e) in WT and ApoE-/- mice fed with HFD for 12 weeks. f, g ITT (f) and AUC (g) in WT and ApoE-/- mice fed with HFD for 12 weeks (n = 6). h, i Transcription analysis of Glut4, IRS1, IRS2, and INSR in SVF of eWAT (h) and iWAT (i) from WT and ApoE-/- mice. j Transcription analysis of IRS2 and INSR in in LPS-stimulated PM (100 ng/mL, 24 h, n = 3). PM, peritoneal macrophages. Data represent mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ns, no significance

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  Fig. 4. ApoE deficiency aggravates HFD-induced metabolic inflammation in adipose tissues. a, b Representative H&E-stained sections (a) and quantitative results of adipocytes area and CLS (b) of eWAT from WT and ApoE-/- HFD feeding mice (n = 6). Scale bars, 100 μm. c, d Representative H&E staining images (c) and quantification of adipocytes area and CLS (d) of iWAT from WT and ApoE-/- HFD feeding mice (n = 6). Scale bars, 100 μm. e, f QPCR analysis of proinflammatory genes in eWAT (e) and iWAT (f) from WT and ApoE-/- mice on HFD (n = 6). g, h TNF-α and IL-1β levels in supernatants of eWAT cultures determined by ELISA (n = 3). i, j TNF-α and IL-1β levels in supernatants of iWAT cultures determined by ELISA (n = 3). Data represent mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001

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  Fig. 5. Depletion of ApoE expedites the transcriptional initiation of proinflammatory gene expression in SVF and macrophages. a, b mRNA levels of indicated proinflammatory genes in eWAT and iWAT SVF from indicated mice fed HFD (n = 5). c mRNA levels of classically activated macrophage marker in PM following exposure to LPS (100 ng/mL) for 24 h (n = 3). d mRNA levels of alternatively activated macrophage marker in PM following exposure to IL-4 (20 ng/mL) for 24 h (n = 3). e IL-1β and TNF-α levels in culture supernatants from LPS-primed (500 ng/mL) PM followed with ATP (2 mM) for 45 min (n = 3)

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  Fig. 6. NLRP3 inflammasome is required for proinflammatory macrophage activation in mice. a, b The FPKM value of NLRP3 and NLRP1 in indicated groups from RNA-Seq of human OAT (n = 30, BMI < 24; n = 17, BMI ≥ 24). c Previous human samples data of Il1b expression were collected from the GEO dataset (GSE54350, n = 6). d-g Transcription analysis of ApoE in Ewat and iWAT under chow diet (d, f) or HFD (e, g) from WT and Nlrp3-/- mice (n = 6). h, i Transcription analysis of ApoE in (i) or absent (h) LPS-stimulated PM (100 ng/mL, 24 h, n = 3). j Schematic depicting PM treatment with LPS&ATP-stimulated conditioned medium harvested from PM of WT and Nlrp3-/- mice. k NLRP3 expression in PM and eWAT from WT and Nlrp3-/- mice were determined by Western blot (n = 3, each lane represents one independent biological sample). l QPCR analysis of inflammatory gene expression in PM of ApoE-/- mice treated with CM from indicated groups. m Schematic model depicting the ApoE manipulates inflammation and metabolism. OAT, omental adipose tissue. CD, chow diet. HFD, high-fat diet. PM, peritoneal macrophages. Data represent mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ns, no significance