Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). Scale bars, 500μm. (B, C) Representative pictures of the abdominal aorta areas (B), maximal abdominal aorta diameter and abdominal aorta area (C) measurements of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). (D) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection. (E-G) Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–13/group). (E) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). Scale bars, 500μm. (F, G) Representative pictures of the abdominal aorta areas (F), maximal abdominal aorta diameter, and abdominal aorta area measurements (G) of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). (H) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection (n=5/group). (I) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in heart tissues from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=5 to 7/group). (J) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in abdominal aortas from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=4 to 5–8/group). Data are presented as mean ± SEM. Pooled from 2 experiments (A-J). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A, C, D, E, G, H, I, J). FLICA, Fluorochrome Labeled Inhibitors of Caspases.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection, Labeling

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A, B) Immunofluorescent staining and mean fluorescence intensity (MFI) of SIRT1 (red) in heart (A) and abdominal aorta (B) tissues of PBS and LCWE-injected mice at 2 weeks post-LCWE injection (n=4 to 5/group). DAPI (Blue) used to stain nuclei. Scale bars, 50μm. (C) mRNA levels of Sirt1 measured by qRT-PCR in heart and abdominal aorta tissues of PBS and LCWE-injected mice at 2 weeks post-LCWE injection (n=9/group). Data are presented as mean ± SEM, representative of one experiment (A, B) or pooled from 2 independent experiments (C). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by unpaired t-test (B, C, D, E).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Fluorescence, Injection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Representative H&E-stained heart sections and heart vessel inflammation scores of PBS or LCWE-injected WT and Sirt1 super mice, at 2 weeks post-injection. (n=5 to 10/group). Scale bars, 500μm. (B, C) Representative pictures of the abdominal aorta areas (B), maximal abdominal aorta diameter, and abdominal aorta area measurements (C) of PBS or LCWE-injected WT and Sirt1 super mice, at 2 weeks post-injection. (n=5 to 10/group). (D) IL-1β measurements in the peritoneal lavage of WT and Sirt1 super mice injected with PBS or LCWE 24 hours post-injection. (E) Schematic of the experimental design. (F-H) WT and Sirt1 ER/ER mice were injected with either PBS or LCWE, and one week later, injected with tamoxifen for 5 consecutive days. Heart and abdominal aorta tissues were collected at day 18 post-LCWE injection for analysis. (F) Representative H&E-stained heart sections and heart vessel inflammation scores of PBS or LCWE-injected, tamoxifen-treated WT and Sirt1 super mice, at day 18 post-injection. (n=5 to 14/group). Scale bars, 500μm. (G, H) Representative pictures of the abdominal aorta areas (G), maximal abdominal aorta diameter and abdominal aorta area measurements (H) of PBS or LCWE-injected, tamoxifen-treated WT and Sirt1 ER/ER mice, at day 18 post-injection (n=5 to 14/group). Data are presented as mean ± SEM, representative of at least 2 independent experiments (A-I). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A, C, D, F, H).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Principal component (PC) analysis of the proteome from the abdominal aortas of WT and Sirt1 super mice injected with either PBS or LCWE, at 2 weeks post-injection (n=5/group). (B) Venn diagram of differentially expressed proteins (DEPs, p-value <0.05; fold change [FC] >2) between the indicated groups. A set of 443 proteins associated with the development of LCWE-induced KD was identified (highlighted in red). (C) Selected pathways and functional annotation terms of proteins increased in LCWE-injected WT mice compared with PBS-injected WT mice and decreased in LCWE-injected Sirt1 super mice compared with LCWE-injected WT mice (n=5/group). (D) Selected pathways and functional annotation terms of proteins decreased in LCWE-injected WT mice compared with PBS-injected WT mice and increased in LCWE-injected Sirt1 super mice compared with LCWE-injected WT mice (n=5/group). (E) Ingenuity pathway analysis (IPA) performed on the set of DEPs associated with LCWE-induced KD highlighted in red from (B) between the indicated groups. (F) Mitochondrial respiration was analyzed in isolated mitochondria from heart tissues of LCWE-injected WT and Sirt1 super mice. Measurements of acute response (left), coupling efficiency (middle), and ATP production (right). Data are presented as mean ± SEM, representative of one experiment. Each symbol represents one individual mouse. * p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by unpaired t-test (F).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Injection, Functional Assay, Isolation

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Quantification of dihydroethidium (DHE) staining of heart (top) and abdominal aorta (bottom) tissues from WT and Sirt1 super mice injected with either PBS or LCWE at 2 weeks post-injection (n=4 to 5/group). (B) Heatmap of differentially expressed proteins related to autophagy/ROS, fatty acid oxidation, and oxidative phosphorylation pathways between abdominal aortas of PBS and LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=5/group). (C) Immunofluorescent staining and quantification of p62 (red) and LC3B puncta (red) in abdominal aorta tissues from PBS or LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=9,10/group). DAPI (blue) is used to stain nuclei. Scale bars, 50μm. (D) Immunofluorescent staining and quantification of p62 (red) and LC3B puncta (green) from hearts of PBS or LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=9,10/group). DAPI (blue) is used to stain nuclei. Scale bars, 50μm. (E) Western blot analysis of SIRT1, pUbiquitin S65 and β-actin in protein lysates from heart tissues from WT and Sirt1 super mice injected with LCWE at 2 weeks post-injection (n=4, 5/group). Data are presented as mean ± SEM. Pooled from 2 experiments (A, C, D, E), representative of one experiment (B). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A) or unpaired t-test (C, D). MFI, mean fluorescence intensity.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection, Phospho-proteomics, Western Blot, Fluorescence

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Heatmap of differentially expressed proteins (DEPs) related to VSMCs type I and synthetic VSMCs type II in the abdominal aortas of PBS and LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=5/group). (B) Primary coronary artery SMCs were treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours). Representative Western blots of LC3-I/II, pS6 S235/236 , S6, pULK1 S555 , pAMPK T172 and β-actin from whole cell lysate (n=4/group). (C) Representative Western blots of p62, LC3-I/II, TOM70 and Rho-GDI from Mitochondrial (Mito) and cytosolic (Cyto) fractions of primary coronary artery SMCs treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours) and CQ (5μM, last 4 hours). Ponceau S. was used as loading control. (D) Representative Western blots of LC3-I/II, TOM70, COXIV and β-actin from whole cell lysate of primary coronary artery SMCs treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours) and CQ (5μM, last 4 hours). (E) Quantification of CytoID staining in WT primary coronary artery SMCs treated with SRT1720 (5μM, 24 hours) or EX527 (5μM, 24 hours) alone or in combination with rIL-1β (10ng/ml, 24 hours) and CQ (5μM, last 4 hours) (n=3/group). (F) Quantification of mitoSOX/mitoTracker ratio from WT Primary coronary artery SMCs treated with SRT1720 (5μM, 24 hours) or EX527 (5μM, 24 hours) alone or in combination with rIL-1β (10ng/ml, 24 hours) (n=8/group). (G-H) Bone marrow-derived macrophages (BMDMs) differentiated from WT and Sirt1 super mice treated with LCWE (60μg/ml, 24 hours). (G) IL-1β and TNFα levels were measured from the supernatants by ELISA (n=6/group). (H) Representative Western blots for SIRT1 and pS6 S235/236 from whole cell lysates (n=4/group). Ponceau S. was used as loading control. Data are presented as mean ± SEM. Representative of 2 experiments (B, C, D, E, F, G, H), representative of one experiment (A). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by one-way ANOVA (E, F) or two-way ANOVA with Tukey post hoc test (H). rIL-1β, recombinant IL-1β.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Injection, Western Blot, Control, Staining, Derivative Assay, Enzyme-linked Immunosorbent Assay, Recombinant

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). Scale bars, 500μm. (B, C) Representative pictures of the abdominal aorta areas (B), maximal abdominal aorta diameter and abdominal aorta area (C) measurements of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). (D) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection. (E-G) Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–13/group). (E) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). Scale bars, 500μm. (F, G) Representative pictures of the abdominal aorta areas (F), maximal abdominal aorta diameter, and abdominal aorta area measurements (G) of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). (H) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection (n=5/group). (I) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in heart tissues from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=5 to 7/group). (J) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in abdominal aortas from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=4 to 5–8/group). Data are presented as mean ± SEM. Pooled from 2 experiments (A-J). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A, C, D, E, G, H, I, J). FLICA, Fluorochrome Labeled Inhibitors of Caspases.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection, Labeling

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A, B) Immunofluorescent staining and mean fluorescence intensity (MFI) of SIRT1 (red) in heart (A) and abdominal aorta (B) tissues of PBS and LCWE-injected mice at 2 weeks post-LCWE injection (n=4 to 5/group). DAPI (Blue) used to stain nuclei. Scale bars, 50μm. (C) mRNA levels of Sirt1 measured by qRT-PCR in heart and abdominal aorta tissues of PBS and LCWE-injected mice at 2 weeks post-LCWE injection (n=9/group). Data are presented as mean ± SEM, representative of one experiment (A, B) or pooled from 2 independent experiments (C). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by unpaired t-test (B, C, D, E).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Fluorescence, Injection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Representative H&E-stained heart sections and heart vessel inflammation scores of PBS or LCWE-injected WT and Sirt1 super mice, at 2 weeks post-injection. (n=5 to 10/group). Scale bars, 500μm. (B, C) Representative pictures of the abdominal aorta areas (B), maximal abdominal aorta diameter, and abdominal aorta area measurements (C) of PBS or LCWE-injected WT and Sirt1 super mice, at 2 weeks post-injection. (n=5 to 10/group). (D) IL-1β measurements in the peritoneal lavage of WT and Sirt1 super mice injected with PBS or LCWE 24 hours post-injection. (E) Schematic of the experimental design. (F-H) WT and Sirt1 ER/ER mice were injected with either PBS or LCWE, and one week later, injected with tamoxifen for 5 consecutive days. Heart and abdominal aorta tissues were collected at day 18 post-LCWE injection for analysis. (F) Representative H&E-stained heart sections and heart vessel inflammation scores of PBS or LCWE-injected, tamoxifen-treated WT and Sirt1 super mice, at day 18 post-injection. (n=5 to 14/group). Scale bars, 500μm. (G, H) Representative pictures of the abdominal aorta areas (G), maximal abdominal aorta diameter and abdominal aorta area measurements (H) of PBS or LCWE-injected, tamoxifen-treated WT and Sirt1 ER/ER mice, at day 18 post-injection (n=5 to 14/group). Data are presented as mean ± SEM, representative of at least 2 independent experiments (A-I). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A, C, D, F, H).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Principal component (PC) analysis of the proteome from the abdominal aortas of WT and Sirt1 super mice injected with either PBS or LCWE, at 2 weeks post-injection (n=5/group). (B) Venn diagram of differentially expressed proteins (DEPs, p-value <0.05; fold change [FC] >2) between the indicated groups. A set of 443 proteins associated with the development of LCWE-induced KD was identified (highlighted in red). (C) Selected pathways and functional annotation terms of proteins increased in LCWE-injected WT mice compared with PBS-injected WT mice and decreased in LCWE-injected Sirt1 super mice compared with LCWE-injected WT mice (n=5/group). (D) Selected pathways and functional annotation terms of proteins decreased in LCWE-injected WT mice compared with PBS-injected WT mice and increased in LCWE-injected Sirt1 super mice compared with LCWE-injected WT mice (n=5/group). (E) Ingenuity pathway analysis (IPA) performed on the set of DEPs associated with LCWE-induced KD highlighted in red from (B) between the indicated groups. (F) Mitochondrial respiration was analyzed in isolated mitochondria from heart tissues of LCWE-injected WT and Sirt1 super mice. Measurements of acute response (left), coupling efficiency (middle), and ATP production (right). Data are presented as mean ± SEM, representative of one experiment. Each symbol represents one individual mouse. * p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by unpaired t-test (F).

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Injection, Functional Assay, Isolation

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Quantification of dihydroethidium (DHE) staining of heart (top) and abdominal aorta (bottom) tissues from WT and Sirt1 super mice injected with either PBS or LCWE at 2 weeks post-injection (n=4 to 5/group). (B) Heatmap of differentially expressed proteins related to autophagy/ROS, fatty acid oxidation, and oxidative phosphorylation pathways between abdominal aortas of PBS and LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=5/group). (C) Immunofluorescent staining and quantification of p62 (red) and LC3B puncta (red) in abdominal aorta tissues from PBS or LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=9,10/group). DAPI (blue) is used to stain nuclei. Scale bars, 50μm. (D) Immunofluorescent staining and quantification of p62 (red) and LC3B puncta (green) from hearts of PBS or LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=9,10/group). DAPI (blue) is used to stain nuclei. Scale bars, 50μm. (E) Western blot analysis of SIRT1, pUbiquitin S65 and β-actin in protein lysates from heart tissues from WT and Sirt1 super mice injected with LCWE at 2 weeks post-injection (n=4, 5/group). Data are presented as mean ± SEM. Pooled from 2 experiments (A, C, D, E), representative of one experiment (B). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A) or unpaired t-test (C, D). MFI, mean fluorescence intensity.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection, Phospho-proteomics, Western Blot, Fluorescence

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Heatmap of differentially expressed proteins (DEPs) related to VSMCs type I and synthetic VSMCs type II in the abdominal aortas of PBS and LCWE-injected WT and Sirt1 super mice at 2 weeks post-injection (n=5/group). (B) Primary coronary artery SMCs were treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours). Representative Western blots of LC3-I/II, pS6 S235/236 , S6, pULK1 S555 , pAMPK T172 and β-actin from whole cell lysate (n=4/group). (C) Representative Western blots of p62, LC3-I/II, TOM70 and Rho-GDI from Mitochondrial (Mito) and cytosolic (Cyto) fractions of primary coronary artery SMCs treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours) and CQ (5μM, last 4 hours). Ponceau S. was used as loading control. (D) Representative Western blots of LC3-I/II, TOM70, COXIV and β-actin from whole cell lysate of primary coronary artery SMCs treated with SRT1720 (5μM) or EX527 (5μM) alone or in combination with LCWE (60μg/ml, 24 hours) and CQ (5μM, last 4 hours). (E) Quantification of CytoID staining in WT primary coronary artery SMCs treated with SRT1720 (5μM, 24 hours) or EX527 (5μM, 24 hours) alone or in combination with rIL-1β (10ng/ml, 24 hours) and CQ (5μM, last 4 hours) (n=3/group). (F) Quantification of mitoSOX/mitoTracker ratio from WT Primary coronary artery SMCs treated with SRT1720 (5μM, 24 hours) or EX527 (5μM, 24 hours) alone or in combination with rIL-1β (10ng/ml, 24 hours) (n=8/group). (G-H) Bone marrow-derived macrophages (BMDMs) differentiated from WT and Sirt1 super mice treated with LCWE (60μg/ml, 24 hours). (G) IL-1β and TNFα levels were measured from the supernatants by ELISA (n=6/group). (H) Representative Western blots for SIRT1 and pS6 S235/236 from whole cell lysates (n=4/group). Ponceau S. was used as loading control. Data are presented as mean ± SEM. Representative of 2 experiments (B, C, D, E, F, G, H), representative of one experiment (A). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by one-way ANOVA (E, F) or two-way ANOVA with Tukey post hoc test (H). rIL-1β, recombinant IL-1β.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Injection, Western Blot, Control, Staining, Derivative Assay, Enzyme-linked Immunosorbent Assay, Recombinant

Journal: bioRxiv

Article Title: Sirtuin 1 Activation Mitigates Murine Vasculitis Severity by Promoting Autophagy and Mitophagy

doi: 10.1101/2025.09.04.671113

Figure Lengend Snippet: (A) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). Scale bars, 500μm. (B, C) Representative pictures of the abdominal aorta areas (B), maximal abdominal aorta diameter and abdominal aorta area (C) measurements of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=9 to 10/group). (D) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Myh11 Cre/ERT2 Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection. (E-G) Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–13/group). (E) Representative H&E-stained heart sections and heart vessel inflammation scores of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). Scale bars, 500μm. (F, G) Representative pictures of the abdominal aorta areas (F), maximal abdominal aorta diameter, and abdominal aorta area measurements (G) of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice were injected with PBS or LCWE and analyzed at 2 weeks post-injection (n=5 to 11–12/group). (H) Levels of IL-1β in the peritoneal lavage of Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice injected with PBS or LCWE 24 hours post-injection (n=5/group). (I) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in heart tissues from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=5 to 7/group). (J) Quantification of FLICA + F4/80 + NLRP3 + cell numbers in abdominal aortas from PBS or LCWE injected Sirt1 fl/fl and Csf1r Cre Sirt1 Δ/Δ mice at 2 weeks post-injection (n=4 to 5–8/group). Data are presented as mean ± SEM. Pooled from 2 experiments (A-J). Each symbol represents one individual mouse. *p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001, and **** p-value < 0.001 by two-way ANOVA with Tukey post hoc test (A, C, D, E, G, H, I, J). FLICA, Fluorochrome Labeled Inhibitors of Caspases.

Article Snippet: Wild-type (WT) C57BL/6J, 129S1/SvImJnd (#002448), Sirt1 super (#024510), Sirt1 ER/ER (#043520), Myh11 Cre /ERT2 (#019079), and Csfr1 cre (#029206) mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA).

Techniques: Staining, Injection, Labeling