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Reducing the activity of specific SAC genes diminishes the responsiveness <t>of</t> <t>PANC-1</t> cells to Abraxene and other drugs that target microtubules. <t>ShRNA-transfected</t> PANC-1 cells were treated with (A) Nab-paclitaxel, (B) Paclitaxel, (C) Docetaxel, and (D) Vincristine for 96 hrs. Relative viable cell numbers were measured using the Cell Titer-Glo assay. Data represent three independent experiments with mean ± SEM.
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1) Product Images from "A whole-genome CRISPR screen identifies the spindle accessory checkpoint as a locus of nab-paclitaxel resistance in pancreatic cancer cells"

Article Title: A whole-genome CRISPR screen identifies the spindle accessory checkpoint as a locus of nab-paclitaxel resistance in pancreatic cancer cells

Journal: bioRxiv

doi: 10.1101/2024.02.15.580539

Reducing the activity of specific SAC genes diminishes the responsiveness of PANC-1 cells to Abraxene and other drugs that target microtubules. ShRNA-transfected PANC-1 cells were treated with (A) Nab-paclitaxel, (B) Paclitaxel, (C) Docetaxel, and (D) Vincristine for 96 hrs. Relative viable cell numbers were measured using the Cell Titer-Glo assay. Data represent three independent experiments with mean ± SEM.
Figure Legend Snippet: Reducing the activity of specific SAC genes diminishes the responsiveness of PANC-1 cells to Abraxene and other drugs that target microtubules. ShRNA-transfected PANC-1 cells were treated with (A) Nab-paclitaxel, (B) Paclitaxel, (C) Docetaxel, and (D) Vincristine for 96 hrs. Relative viable cell numbers were measured using the Cell Titer-Glo assay. Data represent three independent experiments with mean ± SEM.

Techniques Used: Activity Assay, shRNA, Transfection, Glo Assay

Blocking specific SAC genes leads to a decrease in PANC-1 cell proliferation and induces aneuploidy. Cell proliferation curves of shRNA-transfected PANC-1 cells cultured with (A) DMSO or (B) Nab-paclitaxel for 5 days. Data represent three biological replicates with mean ± SD. (C) Cell cycles of shRNA-transfected PANC-1 cells were measured with or without nab-paclitaxel for 24 hrs. The image represents one out of three biological replicates. (D) shRNA-transfected PANC-1 cells were treated with nab-paclitaxel and then immunostained for DAP1 (blue), Histone H3p28 (White), and α-tubulin (red). Scale bars: 1 μM.
Figure Legend Snippet: Blocking specific SAC genes leads to a decrease in PANC-1 cell proliferation and induces aneuploidy. Cell proliferation curves of shRNA-transfected PANC-1 cells cultured with (A) DMSO or (B) Nab-paclitaxel for 5 days. Data represent three biological replicates with mean ± SD. (C) Cell cycles of shRNA-transfected PANC-1 cells were measured with or without nab-paclitaxel for 24 hrs. The image represents one out of three biological replicates. (D) shRNA-transfected PANC-1 cells were treated with nab-paclitaxel and then immunostained for DAP1 (blue), Histone H3p28 (White), and α-tubulin (red). Scale bars: 1 μM.

Techniques Used: Blocking Assay, shRNA, Transfection, Cell Culture


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Santa Cruz Biotechnology mouse sirt2 short hairpin rna lentiviral particles
<t>Sirt2</t> expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.
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1) Product Images from "Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model"

Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

Journal: Immunity, Inflammation and Disease

doi: 10.1002/iid3.1160

Sirt2 expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.
Figure Legend Snippet: Sirt2 expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.

Techniques Used: Expressing, Flow Cytometry

The effect of Sirt2 silencing on regulatory T cell (Treg)/T helper 17 (Th17) generation without stimulation. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction. 24 h, 24 h after transduction; 48 h, 48 h after transduction; Ctrl, control lentivirus encoding a scrambled siRNA; Fresh, freshly sorted Tregs; Sirt2 siRNA, Sirt2 siRNA‐encoding lentivirus. (B and C) The frequencies of Foxp3‐tdTomato + , Foxp3‐tdTomato − , and IL‐17‐EGFP + cells in Tregs 24 h (B) or 48 h (C) after lentiviral transduction. Ctrl, cells transfected with control lentivirus; Si‐S, cells transfected with Sirt2 siRNA‐encoding lentivirus. (D) Messenger RNA (mRNA) levels of indicated cytokines 48 h after lentiviral transduction. Fresh: freshly sorted Tregs. (E) Sirt2 mRNA levels after transduction. (F) Representative histograms showing the dilution of CellTrace Violet in activated conventional CD4 + T cells after 5‐day coculture with Tregs. Alone: conventional CD4 + T cells alone. Ctrl Tregs: conventional CD4 + T cells co‐cultured with control Tregs. Sirt2‐silenced Tregs: conventional CD4 + T cells co‐cultured with Sirt2‐silenced Tregs. (G) Statistics of CellTrace Violet intensities. N = three to six per group. * p < .05; ** p < .01; *** p < .001. Student's t test for (B), (C), and (E). One‐way analysis of variance for (D) and (G).
Figure Legend Snippet: The effect of Sirt2 silencing on regulatory T cell (Treg)/T helper 17 (Th17) generation without stimulation. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction. 24 h, 24 h after transduction; 48 h, 48 h after transduction; Ctrl, control lentivirus encoding a scrambled siRNA; Fresh, freshly sorted Tregs; Sirt2 siRNA, Sirt2 siRNA‐encoding lentivirus. (B and C) The frequencies of Foxp3‐tdTomato + , Foxp3‐tdTomato − , and IL‐17‐EGFP + cells in Tregs 24 h (B) or 48 h (C) after lentiviral transduction. Ctrl, cells transfected with control lentivirus; Si‐S, cells transfected with Sirt2 siRNA‐encoding lentivirus. (D) Messenger RNA (mRNA) levels of indicated cytokines 48 h after lentiviral transduction. Fresh: freshly sorted Tregs. (E) Sirt2 mRNA levels after transduction. (F) Representative histograms showing the dilution of CellTrace Violet in activated conventional CD4 + T cells after 5‐day coculture with Tregs. Alone: conventional CD4 + T cells alone. Ctrl Tregs: conventional CD4 + T cells co‐cultured with control Tregs. Sirt2‐silenced Tregs: conventional CD4 + T cells co‐cultured with Sirt2‐silenced Tregs. (G) Statistics of CellTrace Violet intensities. N = three to six per group. * p < .05; ** p < .01; *** p < .001. Student's t test for (B), (C), and (E). One‐way analysis of variance for (D) and (G).

Techniques Used: Flow Cytometry, Expressing, Transduction, Transfection, Cell Culture

The effect of Sirtuin2 (Sirt2) silencing on regulatory T cells (Tregs) after T helper 17 (Th17) induction. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐enhanced green fluorescent protein (EGFP) and and Forkhead box P‐3 (Foxp3)‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction and Th17 induction. Ctrl, cells transduced with control lentivirus; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) The frequencies of IL‐17‐EGFP + cells and Foxp3‐tdTomato + cells in Tregs after lentiviral transduction and Th17 induction. (C, D) Messenger RNA levels of IL‐22 (C) and retinoic‐acid‐receptor‐related orphan nuclear receptor gamma (D) in Tregs after lentiviral transduction and Th17 induction. Fresh: freshly sorted Tregs as the negative control. (E) Representative flow cytometry histograms showing Ki67 stain in Tregs after lentiviral transduction and Th17 induction. The data represent two independent experiments. N = 6 per group. ** p < .01; *** p < .001. Student's t test.
Figure Legend Snippet: The effect of Sirtuin2 (Sirt2) silencing on regulatory T cells (Tregs) after T helper 17 (Th17) induction. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐enhanced green fluorescent protein (EGFP) and and Forkhead box P‐3 (Foxp3)‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction and Th17 induction. Ctrl, cells transduced with control lentivirus; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) The frequencies of IL‐17‐EGFP + cells and Foxp3‐tdTomato + cells in Tregs after lentiviral transduction and Th17 induction. (C, D) Messenger RNA levels of IL‐22 (C) and retinoic‐acid‐receptor‐related orphan nuclear receptor gamma (D) in Tregs after lentiviral transduction and Th17 induction. Fresh: freshly sorted Tregs as the negative control. (E) Representative flow cytometry histograms showing Ki67 stain in Tregs after lentiviral transduction and Th17 induction. The data represent two independent experiments. N = 6 per group. ** p < .01; *** p < .001. Student's t test.

Techniques Used: Flow Cytometry, Expressing, Transduction, Negative Control, Staining

Involvement of the Sirtuin3 (Sirt3) signaling in Sirtuin2 (Sirt2)‐mediated effect. (A) Representative flow cytometry histograms showing the expression of phosphorylated Stat3 (left panel) and total Sta3 (right panel) in regulatory T cells (Tregs) after lentiviral transduction and T helper 17 (Th17) induction. Ctrl, cells transduced with control lentivirus; Isotype, isotype antibody control; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) Statistics of the mean fluorescent intensities of phosphorylated Stat3 (p‐Stat3) and total Stat3 in Tregs in (A). (C) The ratios of phosphorylated Stat3 (p‐Stat3) to total Stat3 in Tregs in (A). (D) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in Tregs after lentiviral transduction and Th17 induction in the presence or absence of C188‐9. V: DMSO. (E) Statistics of the frequencies of IL‐17‐EGFP + cells in (D). (F) Sirt2 messenger RNA levels in Tregs after lentiviral transduction and Th17 induction. (G) Immunoblotting images showing Stat3 proteins in Tregs on Day 0 (d0) and Day 3 (d3) after Th17 induction. (H) Statistics of normalized Stat3 protein levels in (G). N = five to six per group. ***: p < .001. Student's t test. DMSO, dimethyl sulfoxide.
Figure Legend Snippet: Involvement of the Sirtuin3 (Sirt3) signaling in Sirtuin2 (Sirt2)‐mediated effect. (A) Representative flow cytometry histograms showing the expression of phosphorylated Stat3 (left panel) and total Sta3 (right panel) in regulatory T cells (Tregs) after lentiviral transduction and T helper 17 (Th17) induction. Ctrl, cells transduced with control lentivirus; Isotype, isotype antibody control; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) Statistics of the mean fluorescent intensities of phosphorylated Stat3 (p‐Stat3) and total Stat3 in Tregs in (A). (C) The ratios of phosphorylated Stat3 (p‐Stat3) to total Stat3 in Tregs in (A). (D) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in Tregs after lentiviral transduction and Th17 induction in the presence or absence of C188‐9. V: DMSO. (E) Statistics of the frequencies of IL‐17‐EGFP + cells in (D). (F) Sirt2 messenger RNA levels in Tregs after lentiviral transduction and Th17 induction. (G) Immunoblotting images showing Stat3 proteins in Tregs on Day 0 (d0) and Day 3 (d3) after Th17 induction. (H) Statistics of normalized Stat3 protein levels in (G). N = five to six per group. ***: p < .001. Student's t test. DMSO, dimethyl sulfoxide.

Techniques Used: Flow Cytometry, Expressing, Transduction, Western Blot

Overview of the findings. Foxp3, Forkhead box P‐3; Sirt2, Sirtuin2; Sirt3, Sirtuin3; RORγt, retinoic‐acid‐receptor‐related orphan nuclear receptor gamma T; Th17, T helper 17; Treg, regulatory T cell.
Figure Legend Snippet: Overview of the findings. Foxp3, Forkhead box P‐3; Sirt2, Sirtuin2; Sirt3, Sirtuin3; RORγt, retinoic‐acid‐receptor‐related orphan nuclear receptor gamma T; Th17, T helper 17; Treg, regulatory T cell.

Techniques Used:


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Santa Cruz Biotechnology control short hairpin sh rna plasmid
Control Short Hairpin Sh Rna Plasmid, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    <t>Sirt2</t> expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.
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    Santa Cruz Biotechnology control short hairpin sh rna plasmid
    <t>Sirt2</t> expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.
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    Sirt2 expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.

    Journal: Immunity, Inflammation and Disease

    Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

    doi: 10.1002/iid3.1160

    Figure Lengend Snippet: Sirt2 expression in regulatory T cells (Tregs) and Treg/T helper 17 (Th17) cells. (A) Representative flow cytometry zebra plots showing Foxp3‐tdTomato + interleukin (IL)‐17‐EGFP − Tregs and Foxp3‐tdTomato + IL‐17‐EGFP + Treg/Th17 in the spleens (SP), mesenteric lymph nodes (mLN), and lamina propria (LP). (B) The frequencies of IL‐17‐EGFP + Treg/Th17 cells in indicated tissues. (C) Sirt2 messenger RNA levels in IL‐17‐EGFP − Tregs and IL‐17−EGFP + Treg/Th17 cells that were sorted from colitic mice. (D) Sirt2 protein levels in IL‐17‐EGFP − Tregs and IL‐17‐EGFP + Treg/Th17 cells that were sorted from colitic mice. N = five to six per group. ** p < .01; *** p < .001. Student's t test. DSS, dextran sulfate sodium‐treated colitic mice; IL‐17 − , IL‐17‐EGFP − Tregs; IL‐17 + , IL‐17‐EGFP + Treg/Th17 cells; V, vehicle‐treated mice.

    Article Snippet: For the lentiviral transduction of lamina propria Tregs, the mouse SIRT2 short hairpin RNA lentiviral particles and control lentivirus were purchased from Santa Cruz Biotechnology.

    Techniques: Expressing, Flow Cytometry

    The effect of Sirt2 silencing on regulatory T cell (Treg)/T helper 17 (Th17) generation without stimulation. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction. 24 h, 24 h after transduction; 48 h, 48 h after transduction; Ctrl, control lentivirus encoding a scrambled siRNA; Fresh, freshly sorted Tregs; Sirt2 siRNA, Sirt2 siRNA‐encoding lentivirus. (B and C) The frequencies of Foxp3‐tdTomato + , Foxp3‐tdTomato − , and IL‐17‐EGFP + cells in Tregs 24 h (B) or 48 h (C) after lentiviral transduction. Ctrl, cells transfected with control lentivirus; Si‐S, cells transfected with Sirt2 siRNA‐encoding lentivirus. (D) Messenger RNA (mRNA) levels of indicated cytokines 48 h after lentiviral transduction. Fresh: freshly sorted Tregs. (E) Sirt2 mRNA levels after transduction. (F) Representative histograms showing the dilution of CellTrace Violet in activated conventional CD4 + T cells after 5‐day coculture with Tregs. Alone: conventional CD4 + T cells alone. Ctrl Tregs: conventional CD4 + T cells co‐cultured with control Tregs. Sirt2‐silenced Tregs: conventional CD4 + T cells co‐cultured with Sirt2‐silenced Tregs. (G) Statistics of CellTrace Violet intensities. N = three to six per group. * p < .05; ** p < .01; *** p < .001. Student's t test for (B), (C), and (E). One‐way analysis of variance for (D) and (G).

    Journal: Immunity, Inflammation and Disease

    Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

    doi: 10.1002/iid3.1160

    Figure Lengend Snippet: The effect of Sirt2 silencing on regulatory T cell (Treg)/T helper 17 (Th17) generation without stimulation. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction. 24 h, 24 h after transduction; 48 h, 48 h after transduction; Ctrl, control lentivirus encoding a scrambled siRNA; Fresh, freshly sorted Tregs; Sirt2 siRNA, Sirt2 siRNA‐encoding lentivirus. (B and C) The frequencies of Foxp3‐tdTomato + , Foxp3‐tdTomato − , and IL‐17‐EGFP + cells in Tregs 24 h (B) or 48 h (C) after lentiviral transduction. Ctrl, cells transfected with control lentivirus; Si‐S, cells transfected with Sirt2 siRNA‐encoding lentivirus. (D) Messenger RNA (mRNA) levels of indicated cytokines 48 h after lentiviral transduction. Fresh: freshly sorted Tregs. (E) Sirt2 mRNA levels after transduction. (F) Representative histograms showing the dilution of CellTrace Violet in activated conventional CD4 + T cells after 5‐day coculture with Tregs. Alone: conventional CD4 + T cells alone. Ctrl Tregs: conventional CD4 + T cells co‐cultured with control Tregs. Sirt2‐silenced Tregs: conventional CD4 + T cells co‐cultured with Sirt2‐silenced Tregs. (G) Statistics of CellTrace Violet intensities. N = three to six per group. * p < .05; ** p < .01; *** p < .001. Student's t test for (B), (C), and (E). One‐way analysis of variance for (D) and (G).

    Article Snippet: For the lentiviral transduction of lamina propria Tregs, the mouse SIRT2 short hairpin RNA lentiviral particles and control lentivirus were purchased from Santa Cruz Biotechnology.

    Techniques: Flow Cytometry, Expressing, Transduction, Transfection, Cell Culture

    The effect of Sirtuin2 (Sirt2) silencing on regulatory T cells (Tregs) after T helper 17 (Th17) induction. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐enhanced green fluorescent protein (EGFP) and and Forkhead box P‐3 (Foxp3)‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction and Th17 induction. Ctrl, cells transduced with control lentivirus; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) The frequencies of IL‐17‐EGFP + cells and Foxp3‐tdTomato + cells in Tregs after lentiviral transduction and Th17 induction. (C, D) Messenger RNA levels of IL‐22 (C) and retinoic‐acid‐receptor‐related orphan nuclear receptor gamma (D) in Tregs after lentiviral transduction and Th17 induction. Fresh: freshly sorted Tregs as the negative control. (E) Representative flow cytometry histograms showing Ki67 stain in Tregs after lentiviral transduction and Th17 induction. The data represent two independent experiments. N = 6 per group. ** p < .01; *** p < .001. Student's t test.

    Journal: Immunity, Inflammation and Disease

    Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

    doi: 10.1002/iid3.1160

    Figure Lengend Snippet: The effect of Sirtuin2 (Sirt2) silencing on regulatory T cells (Tregs) after T helper 17 (Th17) induction. (A) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐enhanced green fluorescent protein (EGFP) and and Forkhead box P‐3 (Foxp3)‐tdTomato in sorted lamina propria CD4 + tdTomato + EGFP − Tregs after lentiviral transduction and Th17 induction. Ctrl, cells transduced with control lentivirus; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) The frequencies of IL‐17‐EGFP + cells and Foxp3‐tdTomato + cells in Tregs after lentiviral transduction and Th17 induction. (C, D) Messenger RNA levels of IL‐22 (C) and retinoic‐acid‐receptor‐related orphan nuclear receptor gamma (D) in Tregs after lentiviral transduction and Th17 induction. Fresh: freshly sorted Tregs as the negative control. (E) Representative flow cytometry histograms showing Ki67 stain in Tregs after lentiviral transduction and Th17 induction. The data represent two independent experiments. N = 6 per group. ** p < .01; *** p < .001. Student's t test.

    Article Snippet: For the lentiviral transduction of lamina propria Tregs, the mouse SIRT2 short hairpin RNA lentiviral particles and control lentivirus were purchased from Santa Cruz Biotechnology.

    Techniques: Flow Cytometry, Expressing, Transduction, Negative Control, Staining

    Involvement of the Sirtuin3 (Sirt3) signaling in Sirtuin2 (Sirt2)‐mediated effect. (A) Representative flow cytometry histograms showing the expression of phosphorylated Stat3 (left panel) and total Sta3 (right panel) in regulatory T cells (Tregs) after lentiviral transduction and T helper 17 (Th17) induction. Ctrl, cells transduced with control lentivirus; Isotype, isotype antibody control; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) Statistics of the mean fluorescent intensities of phosphorylated Stat3 (p‐Stat3) and total Stat3 in Tregs in (A). (C) The ratios of phosphorylated Stat3 (p‐Stat3) to total Stat3 in Tregs in (A). (D) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in Tregs after lentiviral transduction and Th17 induction in the presence or absence of C188‐9. V: DMSO. (E) Statistics of the frequencies of IL‐17‐EGFP + cells in (D). (F) Sirt2 messenger RNA levels in Tregs after lentiviral transduction and Th17 induction. (G) Immunoblotting images showing Stat3 proteins in Tregs on Day 0 (d0) and Day 3 (d3) after Th17 induction. (H) Statistics of normalized Stat3 protein levels in (G). N = five to six per group. ***: p < .001. Student's t test. DMSO, dimethyl sulfoxide.

    Journal: Immunity, Inflammation and Disease

    Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

    doi: 10.1002/iid3.1160

    Figure Lengend Snippet: Involvement of the Sirtuin3 (Sirt3) signaling in Sirtuin2 (Sirt2)‐mediated effect. (A) Representative flow cytometry histograms showing the expression of phosphorylated Stat3 (left panel) and total Sta3 (right panel) in regulatory T cells (Tregs) after lentiviral transduction and T helper 17 (Th17) induction. Ctrl, cells transduced with control lentivirus; Isotype, isotype antibody control; Si‐S, cells transduced with Sirt2 siRNA‐encoding lentivirus. (B) Statistics of the mean fluorescent intensities of phosphorylated Stat3 (p‐Stat3) and total Stat3 in Tregs in (A). (C) The ratios of phosphorylated Stat3 (p‐Stat3) to total Stat3 in Tregs in (A). (D) Representative flow cytometry zebra plots showing the expression of interleukin (IL)‐17‐EGFP and Foxp3‐tdTomato in Tregs after lentiviral transduction and Th17 induction in the presence or absence of C188‐9. V: DMSO. (E) Statistics of the frequencies of IL‐17‐EGFP + cells in (D). (F) Sirt2 messenger RNA levels in Tregs after lentiviral transduction and Th17 induction. (G) Immunoblotting images showing Stat3 proteins in Tregs on Day 0 (d0) and Day 3 (d3) after Th17 induction. (H) Statistics of normalized Stat3 protein levels in (G). N = five to six per group. ***: p < .001. Student's t test. DMSO, dimethyl sulfoxide.

    Article Snippet: For the lentiviral transduction of lamina propria Tregs, the mouse SIRT2 short hairpin RNA lentiviral particles and control lentivirus were purchased from Santa Cruz Biotechnology.

    Techniques: Flow Cytometry, Expressing, Transduction, Western Blot

    Overview of the findings. Foxp3, Forkhead box P‐3; Sirt2, Sirtuin2; Sirt3, Sirtuin3; RORγt, retinoic‐acid‐receptor‐related orphan nuclear receptor gamma T; Th17, T helper 17; Treg, regulatory T cell.

    Journal: Immunity, Inflammation and Disease

    Article Title: Sirtuin2 suppresses the polarization of regulatory T cells toward T helper 17 cells through repressing the expression of signal transducer and activator of transcription 3 in a mouse colitis model

    doi: 10.1002/iid3.1160

    Figure Lengend Snippet: Overview of the findings. Foxp3, Forkhead box P‐3; Sirt2, Sirtuin2; Sirt3, Sirtuin3; RORγt, retinoic‐acid‐receptor‐related orphan nuclear receptor gamma T; Th17, T helper 17; Treg, regulatory T cell.

    Article Snippet: For the lentiviral transduction of lamina propria Tregs, the mouse SIRT2 short hairpin RNA lentiviral particles and control lentivirus were purchased from Santa Cruz Biotechnology.

    Techniques: