5.4 kb mouse ap2 promoter construct  (Addgene inc)

Journal: Cell Death & Disease

Article Title: Adipocyte-expressed SIRT3 manipulates carnitine pool to orchestrate metabolic reprogramming and polarization of macrophages

doi: 10.1038/s41419-025-07699-6

Figure Lengend Snippet: The mRNA expression of Tnf-α ( A ) Il-6 ( B ), Il-1β ( C ) and iNos ( D ) in BMDMs. The mRNA expression of Mrc1 ( E ) Mgl2 ( F ) Arg1 ( G ) and Chil3 ( H ) in BMDMs. n = 6 independent experiments. I The F4/80/CD11c double-positive and F4/80/CD206 double-positive macrophages were analyzed by flow cytometry. n = 5 independent experiments. J The mRNA expression of Mcp-1 , Mip-1α , Ccl5 , Ccl11 , Cxcl10 and Cxcl11 in BMDMs. n = 6 independent experiments. K Transwell migration of macrophages towards CM from vector and SIRT3OE adipocytes. n = 6 independent experiments. Data are expressed as means ± SEM. # P < 0.05, ## p < 0.01, ### p < 0.001, vector vs. control; * P < 0.05, ** P < 0.01, vector vs. SIRT3OE.

Article Snippet: Recombinant adeno-associated serotype 9 viruses with Ap2 promoter for SIRT3 overexpression in adipocytes (AAV- Ap2 -SIRT3) or the empty vector (AAV- Ap2 ) were acquired from GeneChem Co, Ltd (Shanghai, China).

Techniques: Expressing, Flow Cytometry, Migration, Plasmid Preparation, Control

Journal: Cell Death & Disease

Article Title: Adipocyte-expressed SIRT3 manipulates carnitine pool to orchestrate metabolic reprogramming and polarization of macrophages

doi: 10.1038/s41419-025-07699-6

Figure Lengend Snippet: A VIP of significantly altered metabolites. B Scatter diagram of PC, OC, HC, Pro-LC, Isobu-LC and LC, each dot shows a technical replicate. C Volcano plot of significantly altered metabolites between the vector and SIRT3OE cells. n = 6 replicates. D Contents of PC and LC in CM of vector and SIRT3OE adipocytes. n = 6 replicates. * P < 0.05, ** P < 0.01, *** P < 0.001, vector vs. SIRT3OE. E Contents of PC and LC in serum of mice. n = 4 mice per group. Data are expressed as means ± SEM. ## P < 0.01, ### P < 0.001, HFD-AT-NC vs. RD-AT-NC; * P < 0.05, ** P < 0.01, *** P < 0.001, HFD-AT-NC vs. HFD - AT-SIRT3OE.

Article Snippet: Recombinant adeno-associated serotype 9 viruses with Ap2 promoter for SIRT3 overexpression in adipocytes (AAV- Ap2 -SIRT3) or the empty vector (AAV- Ap2 ) were acquired from GeneChem Co, Ltd (Shanghai, China).

Techniques: Plasmid Preparation

Journal: Cell Death & Disease

Article Title: Adipocyte-expressed SIRT3 manipulates carnitine pool to orchestrate metabolic reprogramming and polarization of macrophages

doi: 10.1038/s41419-025-07699-6

Figure Lengend Snippet: A The acetylated CPT2 levels in vector and SIRT3OE adipocytes. n = 3 independent experiments. B The CPT2 activity in vector and SIRT3OE adipocytes. n = 6 replicates. C The CPT2 activity in Scramble and SIRT3KD adipocytes. * P < 0.05, Scramble vs. SIRT3KD. n = 3 replicates. D The substrate dependency in vector and SIRT3OE adipocytes was determined by Seahorse XF mitochondrial fuel flex test. n = 3 replicates. E The CPT2 activity was evaluated. n = 6 replicates. The contents of PC ( F ) and LC ( G ) in CM of adipocytes. n = 3 replicates. The mRNA expression of Tnf-α ( H ), Il-6 ( I ), Il-1β ( J ), iNos ( K ) and Mcp-1 ( L ) in BMDMs treated with CM of adipocytes. n = 6 replicates. M The protein expression of p-IKKα/β, IKKα, IKKβ, p-IκBα, IκBα, p-p65 and p65. β -actin was used as a loading control. n = 3 replicates. Data are expressed as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001, vector vs. SIRT3OE; # P < 0.05, ## P < 0.01, ### P < 0.001, vector vs. CPT2KD; & P < 0.05, & & P < 0.01, control vs vector; $ P < 0.05, $$$ P < 0.001, CPT2KD vs. SIRT3OE-CPT2KD.

Article Snippet: Recombinant adeno-associated serotype 9 viruses with Ap2 promoter for SIRT3 overexpression in adipocytes (AAV- Ap2 -SIRT3) or the empty vector (AAV- Ap2 ) were acquired from GeneChem Co, Ltd (Shanghai, China).

Techniques: Plasmid Preparation, Activity Assay, Expressing, Control

Journal: eLife

Article Title: Discovery and functional assessment of a novel adipocyte population driven by intracellular Wnt/β-catenin signaling in mammals

doi: 10.7554/elife.77740

Figure Lengend Snippet: Figure 5. Wnt+ adipocytes are essential for initiating adaptive thermogenesis. (A) Immunofluorescent staining of mitochondrial membrane potentials in Wnt+ adipocytes induced from iBAT-derived SVF cells of T/L-GFP mice. n = 5 independent experiments. Scale bar, 100 μm; close-up scale bar, 20 μm. (B) Quantification of staining in (A). AOD, average optical density. Data are mean ± s.e.m., unpaired Student’s t-test; ***p < 0.001. (C) OCR plots of four groups of adipocytes differentiated from GFPpos-1, GFPpos-2, GFPneg-1, and mBaSVF cell lines, respectively. n = 3 independent experiments. (D) Immunofluorescent staining of iWAT from T/L-GFP mice with 4-day thermal challenge showing close association of Wnt+ adipocytes with UCP1+ beige adipocytes. n = 5 mice. Scale bar, 100 μm. (E) Immunofluorescent staining of iWAT from tamoxifen-treated Tcf/LefCreERT2;Rosa26RmTmG mice after 4-day cold exposure. n = 8 mice. Scale bar, 50 μm. (F) Immunofluorescent staining of iWAT from tamoxifen-treated T/L-DTA mice after 4-day cold exposure. Before cold challenge, mice were rested for 48 hr after final tamoxifen treatment. n = 7 mice. Scale bar, 50 μm. (G) Quantitative RT- PCR analysis of gene expression in iWAT from control (Fabp4-Flex-DTA) and T/L-DTA mice in (F). n = 6 and 7 mice. Levels of mRNA expression are normalized to that of Adipoq. (H) Western blot analysis showing protein levels of UCP1, OXPHOS complexes, and β-actin in iWAT from tamoxifen- treated controls and T/L-DTA mice under cold (6 °C) and ambient (22 °C) temperatures. n = 4 mice. (I) Immunofluorescent staining for UCP1 (green) and Perilipin (red) in the iWAT from control and Akita mice. Scale bars, 50 μm. Data are mean ± s.e.m., *p < 0.05, ** p < 0.01, ***p < 0.001, n.s., not significant, two-way repeated ANOVA followed by Bonferroni’s test (C) or unpaired Student’s t-test (G).

Article Snippet: Fabp4- Flex- DTA transgenic construct was generated by inserting the coding sequence of diphtheria toxin A (DTA) into the pBS Fabp4 promoter (5.4 kb) polyA vector (Addgene, 11424), flanked by flip- excision (FLEX) switch.

Techniques: Staining, Membrane, Derivative Assay, Quantitative RT-PCR, Gene Expression, Control, Expressing, Western Blot

Journal: eLife

Article Title: Discovery and functional assessment of a novel adipocyte population driven by intracellular Wnt/β-catenin signaling in mammals

doi: 10.7554/elife.77740

Figure Lengend Snippet: Figure 6. Wnt+ adipocytes enhance systemic glucose homeostasis. (A) Glucose tolerance test (GTT) with calculated area under the curve (AUC) in tamoxifen-treated control (Fabp4-Flex-DTA) and T/L-DTA mice on regular chow diet. n = 7 mice each. (B) Schematic of Wnt+ adipocyte gain-of-function studies by cell implantation. (C) Photographs of fat pad formed by implanted cells. Blue agarose beads were included to locate the Matrigel pad. Black arrow shows benign vascularization of fat pad within two weeks. Scale bar, 50 μm. (D) Immunofluorescent staining for Perilipin showing mature adipocytes and accompanied agarose beads (marked by B) in the ectopically formed fat pad in (C). Scale bar, 50 μm. (E) GTT with calculated AUC in mice that received implantation of committed pre-adipocytes/adipocytes from mBaSVF (n = 8 mice) or GFPpos-1 (n = 7 mice) cell lines for 2 weeks. Data are mean ± s.e.m., *p < 0.05, **p < 0.01, two-way repeated ANOVA followed by Bonferroni’s test. AUC was analyzed by two-tailed t-test, p = 0.0012 (A), 0.0017 (E).

Article Snippet: Fabp4- Flex- DTA transgenic construct was generated by inserting the coding sequence of diphtheria toxin A (DTA) into the pBS Fabp4 promoter (5.4 kb) polyA vector (Addgene, 11424), flanked by flip- excision (FLEX) switch.

Techniques: Control, Staining, Two Tailed Test