Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 4. PPT1 expression is upregulated in senescent macrophages. A) Volcano plot of differentially expressed proteins of Young and Aged groups in rats heart. B) KEGG enrichment top 20. C) Heat map of differentially expressed proteins in the lysosomal pathway. D,E) Representative images and statistical analysis of IHC staining with PPT1 antibody. Magnification: 400×, scale bar = 100 μm. F) Statistical analysis of PPT1 mRNA level. G) Representative images and statistical analysis of PPT1 protein level. H) UMAP plots of heart macrophage from 27 healthy donors. I) A heatmap showing the markers ≈7 types of macrophages. J) Stacked graph of cell proportions. K) Violin plots of inflammatory response scores in different subgroups. L) Violin plots of the M1 scores of different macrophage subsets. M) Violin plots of PPT1 expression in different monocyte-macrophage subsets. N) Inflammatory response score. O) M1 score. P) Representative figure of the immunofluorescence of CD68, PPT1, and DAPI in heart. Magnification: 200×, scale bar = 20 μm. Q,R) Representative immunofluorescence images and statistical analysis of PPT1 intensity. Magnification: 200×, scale bar = 20 μm. S) Statistical analysis of PPT1 mRNA level in BMDM. T) Representative graphs and statistical analysis of PPT1 at protein level in BMDM. (n = 3–6, data are expressed as mean ± SEM, *p < 0.05, **p < 0.01 vs the Young group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: Expressing, Immunohistochemistry

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 5. Transgenic knockout of macrophage PPT1 improves cardiac inflammatory infiltration and myocardial fibrosis in D-gal induce-aged mice. A) Schematic of the experimental design. B) Representative echocardiographic graphs. C–J) Echocardiographic measurements of LVEF, LVFS, LVIDd, LVIDs, LVPWd, LVPWs, IVSd, and IVSs. K) H&E staining of left ventricles. Magnification: 200×, scale bar = 50 μm. L) Masson’s staining of left ventricles. Magnification: 200×, scale bar = 50 μm. M,N) Representative images of IHC staining with MCP-1 and 𝛼-SMA antibody. Magnification: 200×, scale bar = 50 μm. O) Statistical graphs of Masson’s staining. P,Q) Statistical graphs of IHC staining with MCP-1 and 𝛼-SMA antibody. R) Representative images of the immunofluorescence of CD68, PPT1, and DAPI in mice heart. Magnification: 200×, scale bar = 20 μm. S) Statistical analysis of COL3A1, COL1A1, PPT1, IL-6, and TNF-𝛼mRNA level. T,U) Representative images and statistical analysis of COL3A1, COL1A1, PPT1, IL-6, and TNF-𝛼at protein level. (n = 5–6, data are expressed as mean ± SEM, **p < 0.01 vs the WT group; #p < 0.05, ##p < 0.01 vs the D-gal group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: Transgenic Assay, Knock-Out, Staining, Immunohistochemistry

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 6. IPA inhibits PPT1 expression and reduces the secretion of inflammatory factors in aging macrophages. A,B) Representative images and statistical graphs of the immunofluorescence of iNOS. Magnification: 200×, scale bar = 20 μm. C) Statistical analysis of IL-6 and TNF-𝛼at mRNA level. D,E) Representative images and statistical analysis of IL-6 and TNF-𝛼at protein level. F,G) Representative images and statistical graphs of the immunofluorescence of iNOS. Magnification: 200×, scale bar = 20 μm. H) Statistical analysis of IL-6 and TNF-𝛼at mRNA level. I,J) Representative images and statistical analysis of IL-6 and TNF-𝛼at protein level. (n = 3–6, data are expressed as mean ± SEM, **p < 0.01 vs the Young or Vector group; #p < 0.05, ##p < 0.01 vs the Aged or oe-PPT1 group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: Expressing, Plasmid Preparation

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 7. IPA inhibits PPT1 in aged macrophages and alleviates collagen deposition in fibroblasts. A) Schematic of the experimental design. B,C) Representative figures and statistical graphs of the immunofluorescence of 𝛼-SMA. Magnification: 200×, scale bar = 20 μm. D) Statistical analysis of COL3A1, COL1A1 and 𝛼-SMA at mRNA level. E,F) Representative images and statistical analysis of COL3A1, COL1A1 and 𝛼-SMA at protein level. G,H) Representative Figure and statistical graphs of the immunofluorescence of 𝛼-SMA. Magnification: 200×, scale bar = 20 μm. I) Statistical analysis of COL3A1, COL1A1 and 𝛼-SMA at mRNA level. J,K) Representative figures and statistical analysis of COL3A1, COL1A1 and 𝛼-SMA at protein level. (n = 3– 6, data are expressed as mean ± SEM, *p < 0.5, **p < 0.01 vs the Young–CM or Vector–CM group; #p < 0.5, ##p < 0.01 vs the Aged–CM or oe-PPT1–CM group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: Plasmid Preparation

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 8. IPA inhibits PPT1 in elderly macrophages in vitro and reduces myocardial fibrosis in vivo. A) Schematic of the experimental design. B,C) Rep- resentative graphs and statistical graph of flow cytometry. D) Representative echocardiographic graphs. E–L) Echocardiographic measurements of LVEF, LVFS, LVIDd, LVIDs, LVPWd, LVPWs, IVSd, and IVSs. M) H&E staining of left ventricles. Magnification: 200×, scale bar = 50 μm. N) Masson’s staining of left ventricles. Magnification: 200×, scale bar = 50 μm. O,P) Representative images of IHC staining with MCP-1 and 𝛼-SMA antibody. Magnification: 200×, scale bar = 50 μm. Q) Statistical graphs of Masson’s staining. R,S) Statistical graphs of IHC staining with MCP-1 and 𝛼-SMA antibody. T) Statistical analysis of COL3A1, COL1A1, IL-6, and TNF-𝛼at mRNA level. U,V) Representative images and statistical analysis of COL3A1, COL1A1, IL-6, and TNF-𝛼 at protein level. (n = 3–6, data are expressed as mean ± SEM, *p < 0.05, **p < 0.01 vs the Young group; #p < 0.05, ##p < 0.01 vs the Aged group; &p < 0.05, &&p < 0.01 vs the Aged+sh-PPT1 group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: In Vitro, In Vivo, Cytometry, Staining, Immunohistochemistry

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Gut Metabolite Indole-3-Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging-Related Myocardial Fibrosis.

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Figure 10. cGAS-STING pathway is involved in IPA-induced inhibition of PPT1. A) Statistical graph of cGAS and STING mRNA levels in macrophages. B,C) Representative images and statistical graph of cGAS and STING at protein levels in macrophages. D,E) Representative images and statistical graph of cGAS and STING at protein levels in vitro. F) Schematic diagram of IPA. G–J) Representative images and statistical graph of PPT1 immunofluorescence intensity in M1 macrophages after treated with RU.521 or C176. Magnification: 200×, scale bar = 20 μm. K,L) Representative images and statistical graph of PPT1 protein levels in M1 macrophages after treated with RU.521 or C176. M) Graphical abstract. (n = 3–6, data are expressed as mean ± SEM, *p < 0.05, **p < 0.01 vs the Young or Control group; #p < 0.05, ##p < 0.01 vs the Aged or LPS/IFN-𝛾group).

Article Snippet: The antibody resources are as follows: ANP primary antibody (Proteintech, 27426-1-Ig, USA), β-tubulin primary antibody (Proteintech, 10094-1-Ig, USA), STING primary antibody (Proteintech, 19851-1-Ig, USA), AKT primary antibody (Proteintech, 60203-2-Ig, USA), PPT1 primary antibody (Proteintech, 29653-1-Ig, USA), COL1A1 primary antibody (Proteintech, 66761-1-Ig, USA), TNF-α primary antibody (Proteintech, 60291-1-Ig, USA), cGAS primary antibody (ABclonal, A8335, China), PI3K primary antibody (Affinity Biosciences, AF6241, China), p-PI3K primary antibody (CST, 4228S, USA), p-AKT primary antibody (Affinity Biosciences, AF0016, China), MCP-1 primary Adv.

Techniques: Inhibition, In Vitro, Control

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: PPT1 expression is upregulated in senescent macrophages. A) Volcano plot of differentially expressed proteins of Young and Aged groups in rats heart. B) KEGG enrichment top 20. C) Heat map of differentially expressed proteins in the lysosomal pathway. D,E) Representative images and statistical analysis of IHC staining with PPT1 antibody. Magnification: 400×, scale bar = 100 µm. F) Statistical analysis of PPT1 mRNA level. G) Representative images and statistical analysis of PPT1 protein level. H) UMAP plots of heart macrophage from 27 healthy donors. I) A heatmap showing the markers ≈7 types of macrophages. J) Stacked graph of cell proportions. K) Violin plots of inflammatory response scores in different subgroups. L) Violin plots of the M1 scores of different macrophage subsets. M) Violin plots of PPT1 expression in different monocyte‐macrophage subsets. N) Inflammatory response score. O) M1 score. P) Representative figure of the immunofluorescence of CD68, PPT1, and DAPI in heart. Magnification: 200×, scale bar = 20 µm. Q,R) Representative immunofluorescence images and statistical analysis of PPT1 intensity. Magnification: 200×, scale bar = 20 µm. S) Statistical analysis of PPT1 mRNA level in BMDM. T) Representative graphs and statistical analysis of PPT1 at protein level in BMDM. ( n = 3–6, data are expressed as mean ± SEM, * p < 0.05, ** p < 0.01 vs the Young group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: Expressing, Immunohistochemistry, Immunofluorescence

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: Transgenic knockout of macrophage PPT1 improves cardiac inflammatory infiltration and myocardial fibrosis in D‐gal induce‐aged mice. A) Schematic of the experimental design. B) Representative echocardiographic graphs. C–J) Echocardiographic measurements of LVEF, LVFS, LVIDd, LVIDs, LVPWd, LVPWs, IVSd, and IVSs. K) H&E staining of left ventricles. Magnification: 200×, scale bar = 50 µm. L) Masson's staining of left ventricles. Magnification: 200×, scale bar = 50 µm. M,N) Representative images of IHC staining with MCP‐1 and α‐SMA antibody. Magnification: 200×, scale bar = 50 µm. O) Statistical graphs of Masson's staining. P,Q) Statistical graphs of IHC staining with MCP‐1 and α‐SMA antibody. R) Representative images of the immunofluorescence of CD68, PPT1, and DAPI in mice heart. Magnification: 200×, scale bar = 20 µm. S) Statistical analysis of COL3A1, COL1A1, PPT1, IL‐6, and TNF‐α mRNA level. T,U) Representative images and statistical analysis of COL3A1, COL1A1, PPT1, IL‐6, and TNF‐α at protein level. ( n = 5–6, data are expressed as mean ± SEM, ** p < 0.01 vs the WT group; # p < 0.05, ## p < 0.01 vs the D‐gal group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: Transgenic Assay, Knock-Out, Staining, Immunohistochemistry, Immunofluorescence

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: IPA inhibits PPT1 expression and reduces the secretion of inflammatory factors in aging macrophages. A,B) Representative images and statistical graphs of the immunofluorescence of iNOS. Magnification: 200×, scale bar = 20 µm. C) Statistical analysis of IL‐6 and TNF‐α at mRNA level. D,E) Representative images and statistical analysis of IL‐6 and TNF‐α at protein level. F,G) Representative images and statistical graphs of the immunofluorescence of iNOS. Magnification: 200×, scale bar = 20 µm. H) Statistical analysis of IL‐6 and TNF‐α at mRNA level. I,J) Representative images and statistical analysis of IL‐6 and TNF‐α at protein level. ( n = 3–6, data are expressed as mean ± SEM, ** p < 0.01 vs the Young or Vector group; # p < 0.05, ## p < 0.01 vs the Aged or oe‐PPT1 group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: Expressing, Immunofluorescence, Plasmid Preparation

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: IPA inhibits PPT1 in aged macrophages and alleviates collagen deposition in fibroblasts. A) Schematic of the experimental design. B,C) Representative figures and statistical graphs of the immunofluorescence of α‐SMA. Magnification: 200×, scale bar = 20 µm. D) Statistical analysis of COL3A1, COL1A1 and α‐SMA at mRNA level. E,F) Representative images and statistical analysis of COL3A1, COL1A1 and α‐SMA at protein level. G,H) Representative Figure and statistical graphs of the immunofluorescence of α‐SMA. Magnification: 200×, scale bar = 20 µm. I) Statistical analysis of COL3A1, COL1A1 and α‐SMA at mRNA level. J,K) Representative figures and statistical analysis of COL3A1, COL1A1 and α‐SMA at protein level. ( n = 3–6, data are expressed as mean ± SEM, * p < 0.5, ** p < 0.01 vs the Young–CM or Vector–CM group; # p < 0.5, ## p < 0.01 vs the Aged–CM or oe‐PPT1–CM group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: Immunofluorescence, Plasmid Preparation

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: IPA inhibits PPT1 in elderly macrophages in vitro and reduces myocardial fibrosis in vivo. A) Schematic of the experimental design. B,C) Representative graphs and statistical graph of flow cytometry. D) Representative echocardiographic graphs. E–L) Echocardiographic measurements of LVEF, LVFS, LVIDd, LVIDs, LVPWd, LVPWs, IVSd, and IVSs. M) H&E staining of left ventricles. Magnification: 200×, scale bar = 50 µm. N) Masson's staining of left ventricles. Magnification: 200×, scale bar = 50 µm. O,P) Representative images of IHC staining with MCP‐1 and α‐SMA antibody. Magnification: 200×, scale bar = 50 µm. Q) Statistical graphs of Masson's staining. R,S) Statistical graphs of IHC staining with MCP‐1 and α‐SMA antibody. T) Statistical analysis of COL3A1, COL1A1, IL‐6, and TNF‐α at mRNA level. U,V) Representative images and statistical analysis of COL3A1, COL1A1, IL‐6, and TNF‐α at protein level. ( n = 3–6, data are expressed as mean ± SEM, * p < 0.05, ** p < 0.01 vs the Young group; # p < 0.05, ## p < 0.01 vs the Aged group; & p < 0.05, && p < 0.01 vs the Aged+sh‐PPT1 group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: In Vitro, In Vivo, Flow Cytometry, Staining, Immunohistochemistry

Journal: Advanced Science

Article Title: Gut Metabolite Indole‐3‐Propionic Acid Regulates Macrophage Autophagy Through PPT1 Inhibiting Aging‐Related Myocardial Fibrosis

doi: 10.1002/advs.202501070

Figure Lengend Snippet: cGAS‐STING pathway is involved in IPA‐induced inhibition of PPT1. A) Statistical graph of cGAS and STING mRNA levels in macrophages. B,C) Representative images and statistical graph of cGAS and STING at protein levels in macrophages. D,E) Representative images and statistical graph of cGAS and STING at protein levels in vitro. F) Schematic diagram of IPA. G–J) Representative images and statistical graph of PPT1 immunofluorescence intensity in M1 macrophages after treated with RU.521 or C176. Magnification: 200×, scale bar = 20 µm. K,L) Representative images and statistical graph of PPT1 protein levels in M1 macrophages after treated with RU.521 or C176. M) Graphical abstract. ( n = 3–6, data are expressed as mean ± SEM, * p < 0.05, ** p < 0.01 vs the Young or Control group; # p < 0.05, ## p < 0.01 vs the Aged or LPS/IFN‐γ group).

Article Snippet: The PPT1 conditional KO mice were generated by employing the CRISPR/Cas9 technology to disrupt macrophage PPT1 and purchased from Cyagen Biosciences (Suzhou) Inc.

Techniques: Inhibition, In Vitro, Immunofluorescence, Control