Journal: Food Science & Nutrition

Article Title: Panax notoginseng Saponins Accelerate Diabetic Wound Repair by Minimizing Mitochondrial Dysfunction and Inhibiting Pyroptosis of Macrophage via Regulating Ephx2/11,12‐ EET Axis

doi: 10.1002/fsn3.72494

Figure Lengend Snippet: PNS inhibits pyroptosis and mitochondrial homeostasis in wound tissue of DW mice. (A–D) ELISA was used to measure serum levels of IL‐1β, IL‐6, IL‐18, and IL‐10. (E) Immunofluorescence staining of wound sections identified M1 macrophages using antibodies against F4/80 and iNOS. Scale bar: 200 μm. (F) Immunofluorescence staining of wound sections identified M2 macrophages using antibodies against F4/80 and Arg1. Scale bar: 200 μm. (G, H) Western blotting detected protein expression of iNOS and Arg1 in wound tissues, followed by quantitative analysis. (I–K) Western blotting detected protein expression of NLRP3, Casp1, Casp1 p20, FL‐GSDMD, and N‐GSDMD in wound tissues, followed by quantitative analysis. (L, M) Immunohistochemical staining and quantitative analysis of GSDMD expression in wound sections. (N, O) Western blotting detected protein expression of Mfn1, Mfn2, OPA1, p‐Drp1, and Drp1 in wound tissues, followed by quantitative analysis. Values were expressed as means ± SEM, n = 6–8. ** p < 0.01, vs. NW; # p < 0.05, ## p < 0.01 vs. DW.

Article Snippet: For double immunofluorescence staining, sections were incubated overnight at 4°C with primary antibodies against F4/80 (a macrophage marker; 1:100; Santa Cruz, sc‐377009) in combination with either iNOS (an M1 marker; 1:100; ABclonal, A0312) or Arg1 (an M2 marker; 1:100; ABclonal, A1847).

Techniques: Tissue, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Western Blot, Expressing, Analysis, Immunohistochemical staining

Journal: Science Advances

Article Title: SIK2 antagonizes postinfarction cardiac repair by driving macrophage proinflammatory and impairing efferocytosis

doi: 10.1126/sciadv.adw6624

Figure Lengend Snippet: ( A ) Volcano plot displaying the DEGs (fold change ≥ 2 and P .adjust ≤ 0.05) between Sik2 F/F ; LysM Cre cardiac macrophages and Sik2 F/F controls on day 3 post-MI. NS, not significant. ( B ) Heatmap of down-regulated DEGs relevant to inflammation in Sik2 F/F ; LysM Cre cardiac macrophages on day 3 post-MI. ( C ) Heatmap of up-regulated DEGs relevant to anti-inflammation in Sik2 F/F ; LysM Cre cardiac macrophages on day 3 post-MI. ( D and E ) mRNA expression levels of major proinflammatory and reparative factors in Sik2 F/F and Sik2 F/F ; LysM Cre infarct hearts at day 5 post-MI ( n = 6). ( F ) Coimmunostaining of CD68 (green), NOS2 (red), or ARG1 (red) and DAPI (blue) in MI hearts from Sik2 F/F and Sik2 F/F ; LysM Cre mice. Transverse heart sections were prepared at day 5 post-MI (scale bars, 50 μm). ( G ) Gating strategy for the flow cytometry analysis of leukocyte population isolated from Sik2 F/F and Sik2 F/F ; LysM Cre infarcts at day 5 post-MI. ( H ) Flow cytometry–based statistical analysis of neutrophils, monocytes/macrophages (Mos/Mps), and the percentage of Ly6C high or Ly6C low macrophages relative to the total macrophage population in Sik2 F/F and Sik2 F/F ; LysM Cre hearts at day 5 post-MI ( n = 6).

Article Snippet: Primary antibodies against the following targets were used in these experiments: SIK2 (6919s, 1:200; CST, Danvers, MA, USA), Ly6G (551459, 1:200; BD Biosciences, Franklin Lakes, NJ, USA), CD68 (ab53444, 1:200; Abcam, Cambridge, UK), TNNI3 (SAB2502170, 1:200; Sigma-Aldrich, St. Louis, MO, USA), α-SMA (A5228, 1:700; Sigma-Aldrich), Collagen I (NBP1-30054, 1:200; Novus Biologicals, Littleton, CO, USA), CD31 (ab222783, 1:200; Abcam), NOS2 (13120s, 1:200; CST), and Arginase-1 (93668s, 1:200; CST).

Techniques: Expressing, Flow Cytometry, Analysis, Isolation

Journal: Science Advances

Article Title: SIK2 antagonizes postinfarction cardiac repair by driving macrophage proinflammatory and impairing efferocytosis

doi: 10.1126/sciadv.adw6624

Figure Lengend Snippet: ( A ) KEGG enrichment analysis of down-regulated DEGs in Sik2 F/F ; LysM Cre cardiac macrophages on day 3 post-MI. The top 20 enriched pathways are listed. ( B ) DEGs (fold change ≥ 2 and P .adjust ≤ 0.05) overlapped in top 20 of enriched pathways. ( C ) Transcription factor prediction was performed using TRRUST databases. The top 10 transcription factors are listed. ( D ) Immunoblotting analysis and quantification of P65 in the cytoplasm and nucleus of WT and SIK2-knockout PMs and treated with TNF-α (20 ng/ml) or vehicle ( n = 4) for 1 hour. ( E ) qPCR analysis of Il-1β , Il-6 , Nos2 , Cxcl-1 , and Mmp-9 mRNA expression in WT and SIK2-knockout PMs stimulated with TNF-α for 24 hours ( n = 6). ( F ) qPCR analysis of Il-1β , Il-6 , Nos2 , Cxcl-1 , and Mmp-9 mRNA expression ( n = 6). PMs were pretreated with ARN3236 (3 μmol) and stimulated with TNF-α for 24 hours. ( G ) WikiPathways enrichment analysis of differentially phosphorylated proteins in PMs treated with ARN3236 (3 μmol) or control. The top 20 enriched pathways are listed. ( H ) Immunoblotting analysis and quantification of phosphorylated P65 (p-P65), P65, phosphorylated P38 (p-P38), and P38 expression in WT and SIK2-knockout PMs stimulated with TNF-α or vehicle ( n = 6). ( I and J ) Flow cytometry analysis of efferocytosis in WT or LeSik2 Raw264.7 cells [stimulated with SB 203580 (10 μmol) or not] treated with CFSE-prelabeled RBCs ( n = 6).

Article Snippet: Primary antibodies against the following targets were used in these experiments: SIK2 (6919s, 1:200; CST, Danvers, MA, USA), Ly6G (551459, 1:200; BD Biosciences, Franklin Lakes, NJ, USA), CD68 (ab53444, 1:200; Abcam, Cambridge, UK), TNNI3 (SAB2502170, 1:200; Sigma-Aldrich, St. Louis, MO, USA), α-SMA (A5228, 1:700; Sigma-Aldrich), Collagen I (NBP1-30054, 1:200; Novus Biologicals, Littleton, CO, USA), CD31 (ab222783, 1:200; Abcam), NOS2 (13120s, 1:200; CST), and Arginase-1 (93668s, 1:200; CST).

Techniques: Analysis, Western Blot, Knock-Out, Expressing, Control, Flow Cytometry, Significance Assay