hsf1  (Bioss)

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: HN-001 activates the HSF1/PGC-1α axis in C3H10-T1/2 cells. (A) Schematic diagram of identification of HN-001 as a novel HSF1/PGC-1α axis activator. (B, C) Dose- and time-dependent effects on HSF1 and its phosphorylated level (Ser326) and quantification. (D) Examination of HSF1 and its phosphorylation level in the cytosol and nucleus, and quantification. (E) Enrichment of HSF1 in the promoter region of Pgc-1α DNA. N = 3 independent biological experiments. (F) Examination of PGC-1α luciferase activity in cells transfected with the HSE-PGC-1α-Luc or HSE Del -PGC-1α-Luc plasmid. (G) mRNA levels of Hsf1 in adipocytes after 24 h treatment of HN-001. * p < 0.05, ** p < 0.01, *** p < 0.001, compared with control cells. HN-001 inhibits adipocyte maturation and improves the thermogenic program.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Luciferase, Activity Assay, Transfection, Plasmid Preparation, Control

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: HN-001 enhances mitochondrial oxidation and inhibits adipocyte maturation alongside browning induction. (A) Mitochondrial copy numbers as indicated by mtDNA/nDNA. (B) Imaging mitochondria in adipocytes by using the probe Mito-Tracker ® Green FM. Scale bar, 50 μm. (C) Oxygen consumption ratio determination after 24 h treatment. (D) Cellular TG level assay. (E) Oil red O and Nile red staining. Scale bar, 100 μm. (F) Glycerol level determination. (G) mRNA levels of metabolism-related genes. (H) Expression levels of HSF1/PGC-1α axis- and metabolism-related proteins, and quantification. N = 3 independent biological experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, compared with control cells.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Imaging, Staining, Expressing, Control

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: HSF1 deficiency abolishes the beneficial metabolic effects of HN-001. (A) mRNA level of HSF1 after 24 h of transfection. (B) Mitochondrial copy numbers after 9 days of adipocyte differentiation. (C) Cellular TG quantification after 9 days of adipocyte differentiation. (D) Cellular glycerol quantification after 9 days of adipocyte differentiation. (E) Images of lipids and mitochondria in adipocytes after 3 days of differentiation. Scale bar, 50 μm. (F–I) mRNA levels of Pgc-1α , Ucp1 , Cpt-1β , and Fasn after 3 days of adipocyte differentiation. (J) Expression levels of HSF1/PGC-1α axis- and metabolism-related proteins, and quantification. N = 3 independent biological experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, compared with control cells; # p < 0.05, compared with Hsf1 −/− control cells.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Transfection, Expressing, Control

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: HN-001 induces a thermogenic program in WAT of mice. (A) Fat mass content. (B) H&E staining of WATs. Scale bar, 200 μm. (C) Adipocyte size measurement. (D) Expression of lipogenesis-related genes in eWAT and sWAT. (E) Mitochondrial content. (F) Enrichment of HSF1 in the promoter of the Pgc-1α gene . (G) mRNA levels of metabolic and browning regulatory genes. (H, I) Protein levels of HSF1/PGC-1α and browning markers in eWAT and sWAT, and quantification. (J) Immunohistochemistry analysis of UCP1 in eWAT and sWAT. Scale bar, 200 μm. N = 10 mice/group. * p < 0.05, ** p < 0.01, *** p < 0.001, compared with chow diet-fed mice; # p < 0.05, ## p < 0.01, ### p < 0.001, compared with HFC control mice. HN-001 enhances the thermogenic capacity of BAT.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Staining, Expressing, Immunohistochemistry, Control

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: HN-001 enhances thermogenic ability in the BAT of mice. (A) BAT mass measurement. (B) H&E staining of BAT. Scale bar, 200 μm. (C) Mitochondrial content. (D) mRNA levels of metabolic and browning regulatory genes. (E) Immunohistochemistry analysis of UCP1 and p-HSF1 Ser326 . Scale bar, 200 μm. N = 10 mice/group. # p < 0.05, ## p < 0.01, ### p < 0.001, compared with HFC control mice.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Mass Measurement, Staining, Immunohistochemistry, Control

Journal: Frontiers in Pharmacology

Article Title: Marine fungus Aspergillus c1. sp metabolite activates the HSF1/PGC-1α axis, inducing a thermogenic program for treating obesity

doi: 10.3389/fphar.2024.1320040

Figure Lengend Snippet: Induction of the thermogenic program in adipose tissue in HN-001-treated DIO mice. As exemplified by HN-001, activation of the HSF1/PGC-1α axis by enriching the phosphorylated HSF1 to the promoter of PGC-1α then activated the expression of UCP1, leading to mitogenesis and thermogenesis in adipose tissue, thus inhibiting adipocyte maturation. As a result, obesity is alleviated in mice. Abbreviations: BAT, brown adipose tissue; sWAT, subcutaneous white adipose tissue; eWAT, subcutaneous white adipose tissue; HSE, heat shock element; HSF1, heat shock Factor 1; PGC-1α, PPARγ coactivator 1α. UCP1, uncoupling protein 1.

Article Snippet: For immunoblot, protein lysates from cells or tissues were prepared as previously described for immunoblotting using specific antibodies described as follows: HSF1 (Bioss, Cat#bs-3757R, China), pHSF1 Ser326 (Abcam, Cat#ab76076, China), UCP1 (Abcam, Cat#ab234430, China), PGC-1α (Affinity, Cat#AF5395, China), CPT-1β (Abcam, Cat#ab134988, China), pLKB1 Ser431 (Santa Cruz, Cat#sc-271924, China), pAMPKα Thr172 (Affinity, Cat#AF3423, China), FASN (Affinity, Cat#DF6106, China), ACC (Affinity, Cat#AF6421, China).

Techniques: Activation Assay, Expressing

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: Promotion of Treg Induction by Heat Shock via Transcription Factor HSF1. Heat shock treatment at 39 °C for 30 minutes increased the expression of (A) foxp3 and (B) il-10 mRNA in naïve mouse CD4 + T cells. This transient heat shock also increased the frequency of CD4 + Foxp3 + Treg induction over a 3-day period depicted in representative contour plots (C) and bar graphs (D). In the chronically inflamed intestine (TNF ΔARE/+ mice), HSF1 was preferentially upregulated in CD4 + T cells and Tregs at both the mRNA (E) and protein (F) levels, as determined by RT-PCR and flow cytometry. Representative histograms and (G) bar charts display the MFI of HSF1 on CD4 + cells in the SP, MLN, and LP. The results are presented as mean ± SEM, n ≥ 4. * p < 0.05, ** p < 0.01, and *** p < 0.001. CD = clusters of differentiation; HSF1 = heat shock factor 1; LP = lamina propria; MFI = mean fluorescence intensity; MLN = mesenteric lymph nodes; RT-PCR = Reverse transcription polymerase chain reaction; SP = spleen.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Flow Cytometry, Fluorescence

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: HSF1-deficient Tregs exhibit impaired function and reduced Foxp3 protein. The frequency of CD4 + FoxP3 + CD25 + regulatory T cells (Treg) HSF1 −/− mice was analyzed by flow cytometry. The results showed a significant decrease in Treg frequency in HSF1 −/− mouse LP depicted in representative contour plots (A) and an increase in SP and MLN expression seen in the bar chart (B). Culturing naïve CD4 + T cells from wild-type (WT) and HSF1 −/− mice under Treg converting conditions (anti-CD3/CD28, IL-2, ±TGFβ) revealed decreased Treg induction in HSF1 −/− cells (C) compared to WT. (D) Heat shock for 30 minutes at 39 °C had no effect on Treg conversion in naïve HSF1 −/− T cells. To assess Treg suppressive function Isolated CD4 + CD25 + Tregs from WT and HSF1 −/− mice were co-cultured with irradiated CD90 Neg antigen-presenting cells and fluorescently labeled effector cells at increasing ratios (E). Proliferation was stimulated with soluble anti-CD3 and measured over 72 hours. The percentage of proliferating labeled effector lymphocytes served as a surrogate marker for Treg suppressive function. Tregs from HSF1 −/− mice displayed significantly weaker suppression of proliferation compared to WT Tregs. Results represent mean ± SEM, n = 3 from three independent studies. ** p < 0.01. CD = clusters of differentiation; HSF1 = heat shock factor 1; SEM = standard error of the mean; Treg = regulatory T cells; WT = wild-type.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Flow Cytometry, Expressing, Isolation, Cell Culture, Irradiation, Labeling, Marker

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: Evaluation of Conditional HSF1 −/− mice and HSF1 Tg mice. Flow cytometric analysis revealed a decrease in CD4 + Foxp3 + Treg frequency (A) in the colonic LP, MLN, and spleen of HSF1 Fl/Fl CD4 Cre/+ . This coincided with reduced mucosal Foxp3 protein indicated by (D) reduced mean fluorescence intensity in LP and MLN. CD4 + Foxp3 + Tregs frequency increased in the LP of HSF1 Fl/Fl FoxP3 Cre/+ mice relative to controls but decreased in MLN and spleen (B). HSF1 Fl/Fl FoxP3 Cre/+ mice displayed a similar reduction in Foxp3 protein by MFI (E). In contrast, HSF1 overexpression in HSF1 Tg mice coincided with increased mucosal CD4 + Foxp3 + Treg frequency (C) and a concomitant increase in HSF1 MFI in both LP and MLN (F). Conversion of naïve CD4 + T cells in vitro demonstrated that HSF1 Fl/Fl /CD4 Cre/+ and HSF1 Fl/Fl /FoxP3 Cre/+ had decreased conversion to CD4 + FoxP3 + cells and increased conversion in HSF1 Tg CD4 + T cells (G, H). Heat shock treatment increased the conversion of naïve CD4 + cells to CD4 + FoxP3 + T cells in control mice (CD4 Cre/+ or FoxP3 Cre/+ ), while HSF1-deficiency in either (I) HSF1 Fl/Fl CD4 Cre/+ or (J) HSF1 Fl/Fl FoxP3 Cre/+ T cells coincided in decreased Treg conversion. This was reversed in HSF1 Tg CD4 + T cells, which exhibited increased conversion to CD4 + FoxP3 + T cells after heat shock (K). Treg suppression assays indicated reduced suppressive activity in (L) HSF1 Fl/Fl CD4 Cre/+ and (M) HSF1 Fl/Fl FoxP3 Cre/+ Tregs compared to control mice. Conversely, HSF1 Tg Tregs (N) demonstrated increased suppressive function. Results represent mean ± SEM, n = 3 from three independent studies. * p < 0.05, ** p < 0.01, *** p < 0.001. CD = clusters of differentiation; HSF1 = heat shock factor 1; LP = lamina propria; MFI = mean fluorescence intensity; MLN = mesenteric lymph nodes; SEM = standard error of the mean; SP = spleen; Treg = regulatory T cells.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Fluorescence, Over Expression, In Vitro, Activity Assay

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: Assessment of HSF1 activation in Tregs. Naïve CD4 + T cells were stimulated in vitro with IL-2 and either plate-bound anti-CD3 or anti-CD28 antibodies for 4 hours. (A) Western blot analysis revealed increased HSF1 expression with TCR stimulation expressed relative to GAPDH as a reference. (B) Phosphorylation of HSF1 at serine 326 (s326) indicating HSF1 activation was observed with heat shock (HS), TCR stimulation, and combined TCR/TGFβ stimulation after 4 hours. (C) Western blotting of HSF1 in nuclear extracts showed increased HSF1 expression with IL-2 and TGFβ treatment for 4 hours. (D) Non-denatured Western blotting of nuclear extracts of CD4 + T cells treated with IL-2, anti-CD3, anti-CD28 with or without TGFβ demonstrated increased dimerization and trimerization of HSF1 after 4 hours. (E) Jurkat cells transfected with a heat shock element dual luciferase promoter reporter construct showed HSF1 binding at the HSE in response to Treg activation and conversion conditions. (F) Immunofluorescent evaluation of CD4 + CD25 Neg T cytospins demonstrates an increase in nuclear (4’6-diamidine-2-phenylindole (DAPI) Blue) expression of HSF1 (Red) after 4 hours Treg converting conditions. (G) ChIP assay assessed HSF1 binding to the foxp3 gene in naive CD4 + CD25 Neg T cells incubated with different culture conditions for 4 hours. DNA analysis was performed using EpiTect ChIP qPCR Primer Assay for mouse foxp3 (Qiagen). Data represent mean ± SEM from three mice per group from three independent studies. * p < 0.05, ** p < 0.01, *** p < 0.005. CD = clusters of differentiation; HSF1 = heat shock factor 1; SEM = standard error of the mean; TCR = T cell receptor; TGFβ = transforming growth factor β

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Activation Assay, In Vitro, Western Blot, Expressing, Transfection, Luciferase, Construct, Binding Assay, Incubation

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: In vivo suppression of inflammation by Tregs from HSF1 Fl/Fl CD4 Cre Tregs. To evaluate the effect of Tregs on inflammation, Rag1 −/− immunodeficient mice were injected with naïve CD4 + CD45RB High cells from WT C57BL/6 mice along with CD4 + CD25 + T cells isolated from either WT CD4 Cre/+ or HSF1 Fl/Fl CD4 Cre/+ mice at a ratio of1:4 (Treg to CD45RB High cells). Mice receiving HSF1 Fl/Fl CD4 Cre/+ Tregs experienced rapid weight loss (A), a surrogate marker of colitis and histological evidence of inflammation. Tissue histology (B) was independently scored by a pathologist blinded to the study (C). ELISA evaluation (D) was performed on 24-hour explant cultures using colonic tissue isolated from the aforementioned experimental mice and secretion of IL-6, IL-10, IL-17, and IFNγ were assessed. Flow cytometric analysis quantified the frequency (E) of CD4 + cells, (F) CD4 + Foxp3 + Tregs, (G) IL-10-secreting CD4 + Foxp3 + T and (H) IL-17-secreting CD4 + cells from the colonic lamina propria, mesenteric lymph node, and spleen. The results demonstrated a significant decrease in Treg frequency in the HSF1 Fl/Fl CD4 Cre/+ mice. Results represent the mean ± SEM, n = 6 mice per group from three independent studies. * p < 0.05, ** p < 0.01, *** p < 0.001. CD = clusters of differentiation; ELISA = enzyme-linked immunosorbent assay; HSF1 = heat shock factor 1; IFN = interferon; IL = interleukin; SEM = standard error of the mean; Treg = regulatory T cells.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: In Vivo, Injection, Isolation, Marker, Enzyme-linked Immunosorbent Assay

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: Evaluating the impact of HSF1 overexpression in an ileitis model. TNF ΔARE/+ HSF1 Tg mice were used to investigate the effects of HSF1 overexpression on ileitis in the TNF ΔARE/+ model. Assessment of ileitis in mice overexpressing HSF1 reveals decreased inflammation, as indicated by (A) decreased inflammatory scoring conducted by a blinded pathologist and (B) representative histology. Flow cytometry analysis of isolated ileal lamina propria cells identifies an increase in CD4 + Foxp3 + T cells (C, D) IL-10-secreting CD4 + Foxp3 + cells (E), and IL-17-secreting CD4 + cells in the TNF ΔARE/+ HSF1 Tg ileum (F). HSF1 overexpression (G) reduced 4kd FITC dextran transport across the intestinal epithelial barrier, a surrogate marker of intestinal inflammation. Tregs from TNF ΔARE/+ mice exhibit decreased suppressive function in vitro (H). However, Tregs from TNF ΔARE/+ HSF1 TG mice demonstrate improved suppressive function compared to Tregs from TNF ΔARE/+ mice (I). Results represent mean ± SEM, n ≥ 5 mice per group from three independent studies. * p < 0.05, ** p < 0.01, and *** p < 0.001. CD = clusters of differentiation; FITC = Fluorescein isothiocyanate; HSF1 = heat shock factor 1; IL = interleukin; SEM = standard error of the mean; TNF = tumor necrosis factor; Treg = regulatory T cells.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Over Expression, Flow Cytometry, Isolation, Marker, In Vitro

Journal: Mucosal immunology

Article Title: Heat shock factor 1 drives regulatory T-cell induction to limit murine intestinal inflammation

doi: 10.1016/j.mucimm.2023.11.003

Figure Lengend Snippet: Adoptive transfer of Tregs from HSF1 Tg mice improves inflammation. Rag1 −/− immunodeficient mice were co-injected with CD4 + CD45RB High cells from WT C57BL/6 mice and CD4 + CD25 + T cells isolated from either WT CD4 Cre/+ or HSF1 Tg mice at a ratio of 1:8 (Treg to CD45RB High cell) or no Tregs (A). Weight monitoring identified decreased weight loss in the mice treated with Tregs from the HSF1 Tg mice consistent with attenuation of colitis. (B) Tissue histology identified a decrease in acute and chronic inflammation in the HSF1 Tg Treg treated mice relative to controls resulting in improved overall inflammation scores in the HSF Tg Treg treated mice. (C) Representative H&E-stained micrographs demonstrate the architectural distortion and lymphocytic expansion in the lamina propria of the no Treg or WT Treg-treated mice but significantly less inflammation in the colon of the HSF1 Tg Treg-treated mice. Flow cytometric analysis of the frequency of (D) CD4 + , (E) IFNγ-secreting, (F) IL-17-secreting, (G) CD4 + FoxP3 + cells and (H) CD4 + FoxP3 + IL-10 secreting T cells from the colonic LP and mesenteric MLN demonstrate a significant increase in Treg frequency in the HSF1 Tg Treg treated mice across both tissues. Results represent mean ± SEM of 4–6 mice per group and from three independent studies. * p < 0.05. **** p < 0.001. CD = clusters of differentiation; HSF1 = heat shock factor 1; IL = interleukin; LP = lamina propria; MLN = mesenteric lymph nodes; SEM = standard error of the mean; Treg = regulatory T cells.; WT = Wild type.

Article Snippet: For [pSer 326 ]HSF1 WB cells were lysed in RIPA buffer after treatment for either 30 minutes at 39 °C (heat shock) or directly placed for 4 hours with or without anti-CD4 and TGFβ and anti [pSer 326 ]HSF1 polyclonal antibody (Enzo Life Sciences, Farming-dale, NY, USA) was used to probe for phosphorylated protein.

Techniques: Adoptive Transfer Assay, Injection, Isolation, Staining