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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and <t>Nrf2</t> pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.
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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and <t>Nrf2</t> pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.
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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and <t>Nrf2</t> pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.
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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and <t>Nrf2</t> pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.
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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and <t>Nrf2</t> pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.
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Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and Nrf2 pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Journal: The FASEB Journal

Article Title: 4‐Octyl Itaconate Attenuates Age‐Induced Parotid Gland Dysfunction Through the Suppression of Fibrosis and Lipid Deposition by AMPKα/PPARα and the Inhibition of Oxidative Stress via the AMPKα/Nrf2 Signaling Pathway

doi: 10.1096/fj.202602956RR

Figure Lengend Snippet: Effect of 4‐OI on the metabolomics and transcriptome profile of aged mice parotid glands and enhancement of AMPKα and Nrf2 pathway in the parotid gland of aged mice by 4‐OI. OPLS‐DA score plot of metabolites detected in (A) 13 M versus 2 M and (B) 13 M + 4‐OI versus 13 M. (C) The Venn diagram illustrates the differentially expressed metabolites that are particular or common within different comparison groups. (D) KEGG pathway classifications on differential metabolites. The vertical axis represents KEGG compound categories, while the horizontal axis shows the number of compounds annotated to each category. Bar colors indicate the primary classification level of the compounds. Volcano plots plot of differential metabolites detected in (E) 13 M versus 2 M and (F) 13 M + 4‐OI versus 13 M. (G) Histogram displaying the numbers of differential metabolites in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Heatmap showing the variable importance (VIP) scores of the top 30 differential metabolites in (H) 13 M versus 2 M and (I) 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary metabolic pathways responsible for the enrichment of differential metabolites in (J) 13 M versus 2 M and (K) 13 M + 4‐OI versus 13 M. Volcano plots plot of DEGs detected in (L) 13 M versus 2 M and (M) 13 M + 4‐OI versus 13 M. (N) Histogram of the numbers of up‐regulated and downregulated DEGs in 13 M versus 2 M and 13 M + 4‐OI versus 13 M. Bubble plots illustrate the primary KEGG pathways responsible for the enrichment of DEGs in (O) 13 M versus 2 M and (P) 13 M + 4‐OI versus 13 M. (Q, R) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, and PPARα in the parotid gland of different groups of mice ( n = 3). (S, T) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Article Snippet: Mice in the 13 M + 4‐OI + ML385 group received an i.p. injection of an Nrf2 inhibitor (ML385, 30 mg/kg; HY‐13418A; MedChemExpress, NJ, USA) daily for 4 weeks, whereas the mice in the 13 M group and 2 M group were given vehicle by i.p. injection.

Techniques: Comparison, Western Blot, Analysis

Nrf2 inhibitor reverses 4‐OI's restoration of function and structure impairments in parotid gland in aged mice. (A) Representative images and quantitative analysis of hematoxylin and eosin (H&E) staining in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm (top) and 20 μm (bottom). (B) Salivary flow rates in different groups of mice ( n = 6). (C, D) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). (E) Representative images of immunofluorescent staining for Nrf2 (red) and DAPI (blue) and quantitative analysis of relative fluorescence intensity of Nrf2 in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Journal: The FASEB Journal

Article Title: 4‐Octyl Itaconate Attenuates Age‐Induced Parotid Gland Dysfunction Through the Suppression of Fibrosis and Lipid Deposition by AMPKα/PPARα and the Inhibition of Oxidative Stress via the AMPKα/Nrf2 Signaling Pathway

doi: 10.1096/fj.202602956RR

Figure Lengend Snippet: Nrf2 inhibitor reverses 4‐OI's restoration of function and structure impairments in parotid gland in aged mice. (A) Representative images and quantitative analysis of hematoxylin and eosin (H&E) staining in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm (top) and 20 μm (bottom). (B) Salivary flow rates in different groups of mice ( n = 6). (C, D) Representative Western blot and quantitative analysis the protein levels of Nrf2, Keap1, NQO‐1, and HO‐1 in the parotid gland of different groups of mice ( n = 3). (E) Representative images of immunofluorescent staining for Nrf2 (red) and DAPI (blue) and quantitative analysis of relative fluorescence intensity of Nrf2 in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Article Snippet: Mice in the 13 M + 4‐OI + ML385 group received an i.p. injection of an Nrf2 inhibitor (ML385, 30 mg/kg; HY‐13418A; MedChemExpress, NJ, USA) daily for 4 weeks, whereas the mice in the 13 M group and 2 M group were given vehicle by i.p. injection.

Techniques: Analysis, Staining, Western Blot, Fluorescence

AMPKα inhibitor reverses 4‐OI's alleviation of apoptosis, inflammation, and mitochondrial dysfunction in parotid gland in aged mice. (A) Representative ROS‐induced fluorescence staining images labeled with DHE dye (red) and DAPI (blue), accompanied by (C) quantitative analysis of relative DHE fluorescence intensity in the parotid glands of mice from different groups ( n = 6). (B) Representative images and (D) quantitative analysis of JC‐1 staining in the parotid glands of mice from different groups ( n = 6). Scale bar: 50 μm (top) and 20 μm (bottom). (E) Representative transmission electron microscopy image of mitochondria in the parotid glands of 2 M, 13 M, 13 M + 4‐OI, and 13 M + 4‐OI + CC groups. Scale bars: 1 μm (top) and 500 nm (bottom). (F, G) Representative Western blot and quantitative analysis the protein levels of PGC‐1α, TFAM, and cytochrome c in the parotid gland of different groups of mice ( n = 3). (H, I) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, SIRT1, GSK3β, p‐GSK3β, Nrf2, and Keap1 in the parotid gland of different groups of mice ( n = 3). (J) Representative images of immunofluorescent staining for Nrf2 (red) and DAPI (blue) and HO‐1 (red) and DAPI (blue) in the parotid gland of different groups of mice. Scale bars: 50 μm. (K, L) Representative Western blot and quantitative analysis the protein levels of TLR4, NF‐κB p65, p‐NF‐κB p65, IκB‐α, and p‐IκB‐α in the parotid gland of different groups of mice ( n = 3). (M, N) Representative Western blot and quantitative analysis the protein levels of NLRP3, TNF‐α, P16 INK4A , P21, and P53 in the parotid gland of different groups of mice ( n = 3). (O) Representative images and quantitative analysis of senescence‐associated β‐galactosidase (SA‐β‐gal) staining in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. (P) Representative images of immunofluorescent staining for 8‐OHdG (red) and DAPI (blue) in the parotid gland of different groups of mice. Scale bars: 50 μm. (Q) Representative images of immunofluorescent staining for TUNEL (red) and DAPI (blue) and quantitative analysis of TUNEL positive cells in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. DHE, dihydroethidium; ns, not significant; ROS, reactive oxygen species.

Journal: The FASEB Journal

Article Title: 4‐Octyl Itaconate Attenuates Age‐Induced Parotid Gland Dysfunction Through the Suppression of Fibrosis and Lipid Deposition by AMPKα/PPARα and the Inhibition of Oxidative Stress via the AMPKα/Nrf2 Signaling Pathway

doi: 10.1096/fj.202602956RR

Figure Lengend Snippet: AMPKα inhibitor reverses 4‐OI's alleviation of apoptosis, inflammation, and mitochondrial dysfunction in parotid gland in aged mice. (A) Representative ROS‐induced fluorescence staining images labeled with DHE dye (red) and DAPI (blue), accompanied by (C) quantitative analysis of relative DHE fluorescence intensity in the parotid glands of mice from different groups ( n = 6). (B) Representative images and (D) quantitative analysis of JC‐1 staining in the parotid glands of mice from different groups ( n = 6). Scale bar: 50 μm (top) and 20 μm (bottom). (E) Representative transmission electron microscopy image of mitochondria in the parotid glands of 2 M, 13 M, 13 M + 4‐OI, and 13 M + 4‐OI + CC groups. Scale bars: 1 μm (top) and 500 nm (bottom). (F, G) Representative Western blot and quantitative analysis the protein levels of PGC‐1α, TFAM, and cytochrome c in the parotid gland of different groups of mice ( n = 3). (H, I) Representative Western blot and quantitative analysis the protein levels of AMPKα, p‐AMPKα, SIRT1, GSK3β, p‐GSK3β, Nrf2, and Keap1 in the parotid gland of different groups of mice ( n = 3). (J) Representative images of immunofluorescent staining for Nrf2 (red) and DAPI (blue) and HO‐1 (red) and DAPI (blue) in the parotid gland of different groups of mice. Scale bars: 50 μm. (K, L) Representative Western blot and quantitative analysis the protein levels of TLR4, NF‐κB p65, p‐NF‐κB p65, IκB‐α, and p‐IκB‐α in the parotid gland of different groups of mice ( n = 3). (M, N) Representative Western blot and quantitative analysis the protein levels of NLRP3, TNF‐α, P16 INK4A , P21, and P53 in the parotid gland of different groups of mice ( n = 3). (O) Representative images and quantitative analysis of senescence‐associated β‐galactosidase (SA‐β‐gal) staining in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. (P) Representative images of immunofluorescent staining for 8‐OHdG (red) and DAPI (blue) in the parotid gland of different groups of mice. Scale bars: 50 μm. (Q) Representative images of immunofluorescent staining for TUNEL (red) and DAPI (blue) and quantitative analysis of TUNEL positive cells in the parotid gland of different groups of mice ( n = 6). Scale bars: 50 μm. The data are presented as the means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001. DHE, dihydroethidium; ns, not significant; ROS, reactive oxygen species.

Article Snippet: Mice in the 13 M + 4‐OI + ML385 group received an i.p. injection of an Nrf2 inhibitor (ML385, 30 mg/kg; HY‐13418A; MedChemExpress, NJ, USA) daily for 4 weeks, whereas the mice in the 13 M group and 2 M group were given vehicle by i.p. injection.

Techniques: Fluorescence, Staining, Labeling, Analysis, Transmission Assay, Electron Microscopy, Western Blot, TUNEL Assay

Journal: iScience

Article Title: Pentachlorophenol enhances bladder cancer cell invasion through altered protein stability

doi: 10.1016/j.isci.2026.117357

Figure Lengend Snippet:

Article Snippet: Rabbit anti-NFE2L2 , ABclonal , Cat# A3577; RRID: AB_2863473.

Techniques: Recombinant, CCK-8 Assay, Bicinchoninic Acid Protein Assay, shRNA, Plasmid Preparation, Over Expression, Software, Transfection, Western Blot