Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 1. 17α-E2 reduces body mass and adiposity. Weekly changes in (A) total body mass, (B) fat mass, and (C) lean mass in CON (blue) and 17α-E2 (red) treatment groups from Study 1. Representative cross-sectional images of visceral (pink) and subcutaneous (gray) adiposity in the lumbar region of (D) CON and (E) 17α-E2 treatment groups at Week 12 of Study 1. (F) Quantitative analysis of regional adiposity in the lumbar region from CON and 17α-E2 treatment groups from Study 1. (G) Plasma leptin and (H) plasma leptin normalized to total, lean, and fat mass from CON and 17α-E2 treatment groups from Study 1. (I) Plasma adiponectin and (J) plasma adiponectin normalized to total, lean, and fat mass from CON and 17α-E2 treatment groups from Study 1. Daily changes in (K) total body mass and weekly changes in (L) lean:fat mass ratios in CON (yellow), 17α-E2 (orange), and calorie restriction (green) treatment groups from Study 2. All data are expressed as mean ± SEM of 7–10 mice per treatment group and were analyzed by repeated measures analysis of variance (A–C, K, L) or Student’s t test (F–J). *p < .05. #p < .01. $p < .005.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Clinical Proteomics

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 2. 17α-E2 reduces energy intake but not expenditure or excretion. (A) Oxygen consumption, (B) energy expenditure, and (C) voluntary activity during Weeks 0 and 12 from CON (blue) and 17α-E2 (red) treatment groups from Study 1. (D) Oxygen consumption, (E) energy expenditure, and (F) voluntary activity during Week 4 from CON (yellow), 17α-E2 (orange), and calorie restriction (CR; green) treatment groups from Study 2. (G) Fecal energy excretion during Weeks 0 and 5 from CON, 17α-E2, and CR treatment groups from Study 2. (I) Daily calorie intake per week and total calories consumed from CON, 17α-E2, and CR treatment groups from Study 2. (J) Daily calorie intake per week normalized to body mass and total calories consumed normalized to body mass from CON, 17α-E2, and CR treatment groups from Study 2. (K) Anorexigenic gene expression in the hypothalamus from CON, 17α-E2, and CR treatment groups from Study 2. All data are expressed as mean ± SEM of 7–10 mice per treatment group and were analyzed by Student’s t test (A–C) or one-way analysis of variance (D–K).

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Activity Assay, Gene Expression

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 3. 17α-E2 reduces inflammation in white adipose tissue (WAT) and the circulation. Pro-inflammatory gene expression in (A) subcutaneous (iWAT) and (B) visceral (eWAT) WAT from CON (blue) and 17α-E2 (red) treatment groups from Study 1. Representative images of SA-βgal+ in iWAT from (C) CON and (D) 17α-E2 treatment groups from Study 1. (E) Percent SA-βgal+ in iWAT from CON and 17α-E2 treatment groups from Study 1. (F) SA-mRNA expression in iWAT from CON and 17α-E2 treatment groups from Study 1. Representative images of SA-βgal+ in eWAT from (G) CON and (H) 17α-E2 treatment groups from Study 1. (I) Percent SA-βgal+ in eWAT from CON and 17α-E2 treatment groups from Study 1. (J) SA-mRNA expression in eWAT from CON and 17α-E2 treatment groups from Study 1. (K) Plasma pro-inflammatory cytokines and chemokines in CON and 17α-E2 treatment groups at Week 15 of Study 1. Arrows indicate SA-βgal+ activity in representative samples. All data are expressed as mean ± SEM of 8–10 mice per treatment group and were analyzed by Student’s t test. *p < .05. #p < .01. $p < .005. Scale bars: 50 µm.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Gene Expression, Expressing, Clinical Proteomics, Activity Assay

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 4. 17α-E2 reverses age-related lipid redistribution and improves glucose homeostasis. Representative images of total hepatic lipid content from (A) CON and (B) 17α-E2 treatment groups from Study 1. Triglyceride accumulation in (C) liver and (D) quadriceps from CON (blue) and 17α-E2 (red) treatment groups from Study 1. (E) Fasting glucose at Weeks 0 and 15 in CON and 17α-E2 treatment groups from Study 1. (F) Fasting insulin and (G) Glycosylated hemoglobin (HbA1c) at Week 15 in CON and 17α-E2 treatment groups from Study 1. (H) Glucose tolerance at Week 10 in CON and 17α-E2 treatment groups from Study 1. (I) Glucose tolerance area under the curve at Week 10 in CON and 17α-E2 treatment groups from Study 1. (J) Fasting glucose at Weeks 0, 2, and 4 in CON (yellow), 17α-E2 (orange), and calorie restriction (CR; green) treatment groups from Study 2. (K) HbA1c at Weeks 0 and 5 in CON, 17α-E2, and CR treatment groups from Study 2. All data are expressed as mean ± SEM of 8–10 mice per treatment group and were analyzed by Student’s t test (C–G), repeated measures analysis of variance (ANOVA; H), or one-way ANOVA (J–K). *p < .05. #p < .01. $p < .005. ♦p < .05 (different from nonlabeled groups); ♠p < .05 (different from nonlabeled and ♦-labeled groups). Scale bars: 200 µm.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Labeling

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 5. 17α-E2 improves systemic insulin sensitivity. (A) Glucose and (B) insulin under basal and clamped conditions from CON (blue) and 17α-E2 (red) treatment groups from Study 1. (C) Real-time and (D) average glucose infusion rates (GIRs) from CON and 17α-E2 treatment groups from Study 1. (E) Whole-body glucose disposal, (F) endogenous glucose production (EGP) under basal and clamped conditions, and (G) hyperinsulinemia-induced percent EGP suppression from CON and 17α-E2 treatment groups from Study 1. (H) Tissue-specific disposal of 2-deoxyglucose during hyperinsulinemia from CON and 17α-E2 treatment groups from Study 1. (I) Circulating free fatty acids (FFAs) under basal and clamped conditions and (J) percent lipolysis suppression during hyperinsulinemia from CON and 17α-E2 treatment groups from Study 1. All data are expressed as mean ± SEM of 7 mice per treatment group and were analyzed by Student’s t test (A, B, D–J) or repeated-measures analysis of variance (c). *p < .05. $p < .005.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques:

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 6. 17α-E2 selectively activates AMPKα and suppresses mTOR complex 1. Representative Western blots of p-AMPKα, AMPKα, p-mTOR, mTOR, p-S6, S6, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from (A) visceral adipose (eWAT), (B) liver, and (C) quadriceps from CON and 17α-E2 treatment groups from Study 1. Densitometry results for p-AMPKα:AMPKα, p-mTOR:mTOR, and p-S6:S6 ratios from (D) visceral (eWAT), (E) liver, and (F) quadriceps from CON (blue) and 17α- E2 (red) treatment groups from Study 1. All data are expressed as mean ± SEM of 7–10 mice per treatment group and were analyzed by Student’s t test (D–F). $p < .005.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Western Blot

Journal: The journals of gerontology. Series A, Biological sciences and medical sciences

Article Title: 17α-Estradiol Alleviates Age-related Metabolic and Inflammatory Dysfunction in Male Mice Without Inducing Feminization.

doi: 10.1093/gerona/glv309

Figure Lengend Snippet: Figure 7. 17α-E2 does not feminize male mice. (A) Gonadal, (B) seminiferous tubule, and (C) breast mass during necropsy from CON (blue) and 17α-E2 (red) treatment groups from Study 1. Plasma (D) testosterone, (E) 17β-E2, and (F) 17α-E2 in CON and 17α-E2 treatment groups at Week 15 of Study 1. (G) ERα and (H) ERβ mRNA expression in liver, quadriceps, subcutaneous and visceral WAT from CON and 17α-E2 treatment groups from Study 1. All data are expressed as mean ± SEM of 8–10 mice per treatment group and were analyzed by Student’s t test. *p < .05. $p < .005.

Article Snippet: Study 1 utilized LabDiet 5LG6 (65.4% CHO, 22.4% PRO, 12.2% FAT) as the control (CON) diet and was supplemented with 17α-E2 (14.4 ppm; Steraloids, Newport, RI) for the treatment diet.

Techniques: Clinical Proteomics, Expressing

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Kinetic profile of budesonide-modulated gene expression in BEAS-2B cells. BEAS-2B cells were either not treated or treated with budesonide (300 nM) prior to harvesting at 1, 2, 6, and 18 h post-treatment. RNA was prepared, and microarray analysis performed using Affymetrix Primeview arrays. Probe set intensities ( N = 4) from different samples were normalized using the Robust Multi-Array Average (RMA) method. Comparing the probe set intensities of the treated samples against their time-matched untreated controls identified 820 genes that were induced ≥2 fold ( P ≤ 0.05) or repressed ≤0.5 fold ( P ≤ 0.05) by budesonide at any time-point (Additional file ). a . Bar graph illustrating the number of genes meeting the induction or repression criteria at each time-point. b . Scatter plot showing log 2 fold change for genes that were induced or repressed at each time. The overall mean of log 2 fold change is depicted as a solid red line. c . Genes were grouped based on the time point that showed the greatest absolute log 2 fold change into; early (1–2 h; 93 genes), intermediate (6 h; 263 genes) or late (18 h; 463 genes). Heat maps show the relative expression of genes within each group. Data are represented as log 2 fold change relative to the corresponding untreated controls

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Expressing, Microarray

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Effect of budesonide and dexamethasone on gene expression in A549 cells. A549 cells were either not treated or treated with 300 nM budesonide (Bud) or 1 μM dexamethasone (Dex). After 6 h, RNA was prepared, and microarray analysis performed using Affymetrix Primeview arrays. Probe set intensities ( N = 3) from the different samples were normalized using the robust multi-array average (RMA) method and used to calculate the fold change compared to untreated (Additional file ). a . Fold change, compared to untreated, for the dexamethasone and budesonide treatments, is plotted for each gene on the x and y axes, respectively. b . The relationship between the fold change for each treatment is shown in respect of all genes induced ≥2 fold ( P ≤ 0.05) by either budesonide or dexamethasone treatment. Genes selected for qPCR validation are shown as circles (○) labelled with their gene symbol. c . A549 cells were either not treated or treated with 300 nM budesonide or 1 μM dexamethasone. After 6 h, RNA was extracted, and qPCR performed for the indicated genes and GAPDH. Data ( N = 6), normalized to GAPDH, were expressed as fold relative to untreated control and are plotted as means ± SE

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Expressing, Microarray

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Genes induced by budesonide in A549, BEAS-2B and primary human bronchial epithelial (HBE) cells. A549 ( N = 3), BEAS-2B ( N = 4), and primary HBE ( N = 4 donors) cells were not treated or treated with budesonide (300 nM for A549 and BEAS-2B, 100 nM for HBE) for 6 h. RNA was prepared, and microarray analysis performed using Affymetrix Primeview arrays. Probe set intensities from the different samples were normalized using the robust multi-array average (RMA) method. Comparing the probe set intensities of the treated samples against their untreated controls identified 410 genes showing significant induction ≥2 fold ( P ≤ 0.05) compared to untreated control in any of the cell variants (Additional file ). a . The genes meeting these criteria in each cell variant were used to produce a Venn diagram to illustrate commonality between the three cell variants. The 17 genes induced in common were subjected to GO analysis in DAVID and are shown along with GO terms showing significant enrichment (EASE score ≤ 0.1) with genes from this list. Genes representing each term are indicated. GO terms are categorized into 3 primary groups (transcription, proliferation/apoptosis or signaling) as indicated. b . The 410 genes were then grouped as defined by the lower stringency, ≥ 1.25 fold, cut-off (Additional file ). The genes within each group are listed in Additional file . Heat maps were constructed for each grouping. Data are presented as zero-mean log 2 intensity, centered around the mean of untreated (not stimulated; NS) and treated for each gene in each cell variant. Following GO analysis in DAVID, GO terms, showing significant enrichment (EASE score ≤ 0.1) and representing at least 5 genes of the 93 genes induced in common, are listed. The number of genes associated with each term are indicated. The GO terms were then categorized into 3 primary groups (transcription, proliferation/apoptosis or signaling), or other, as shown

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Microarray, Variant Assay, Construct

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Commonalities between GO terms enriched with budesonide-induced genes in A549, BEAS-2B and HBE cells. a . Budesonide-induced genes (≥ 2 fold, P ≤ 0.05; Additional file ) in A549 (187 genes), BEAS-2B (240 genes), and HBE (86 genes) were each subjected to GO analysis using DAVID. Biological process and molecular function terms showing significant enrichment (EASE score ≤ 0.1) with each list, and also represented by at least 5 genes, were obtained (Additional file ). The number of GO terms enriched in each cell variant is indicated and the Venn diagram illustrates the overlap in terms between each cell variant. Terms showing enrichment in HBE cells are provided to the right and are categorized (color coded) according to main function. b . This analysis was repeated using the lower stringency budesonide-induced gene lists, as defined in Fig. b, and the resultant GO terms are listed in Additional file . Total numbers of enriched GO terms for each cell variant are indicated and the Venn diagram illustrates overlap between terms in the 3 cell variants. To the right are the 24 GO terms that were enriched in common for all cell variants. These were categorized as in panel a

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Variant Assay

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Genes repressed by budesonide in A549, BEAS-2B and primary HBE cells. As detailed for Fig. , microarray analysis identified 425 genes as significantly repressed by budesonide (≤0.5 fold, P ≤ 0.05) in any of the three cell variants (Additional file ). a . Genes meeting these criteria were used to produce a Venn diagram to illustrate commonality between the three cell variants. The 8 genes repressed in common are listed and, following analysis in DAVID, GO terms showing significant enrichment (EASE score ≤ 0.1) with the 8 genes from this list are shown. Genes representing each GO term are indicated. Terms are categorized into 4 functional groups, as indicated. b . Genes within each group (A = A549, B = BEAS-2B and H = HBE), according to the Venn diagram in a , are summarized by log 2 intensity. Data are color-coded (Red = A549, yellow/orange = BEAS-2B, Blue = HBE) where open bars represent untreated (NS) and solid bars are budesonide (Bud) treatments. The box defines the upper and lower quartiles, and the line inside represents the median. Whiskers represent the 5th and 95th percentiles. c . The 425 genes were then grouped using the lower stringency, ≤0.8 fold, cut-off (Additional file ). Heat maps were constructed for each of the groups. Data are presented as zero-mean log 2 intensity, centered around the mean of untreated (NS) and treated for each gene in each cell variant. GO terms, showing enrichment (EASE score ≤ 0.1) and represented by at least 5 genes in the 82 gene repressed in common group, are listed. The number of genes associated with each term are indicated. Terms were then categorized into functional groups, as indicated

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Microarray, Functional Assay, Construct, Variant Assay

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Commonalities between GO terms enriched with the budesonide-repressed (≤0.5 fold, P ≤ 0.05) genes in A549, BEAS-2B and HBE cells. Budesonide-repressed genes (≤0.5 fold, P ≤ 0.05; Additional file ) in each of A549 (108 genes), BEAS-2B (236 genes), and HBE (146 genes) were subjected to GO analysis using DAVID. Biological process and molecular function terms showing significant enrichment (EASE score ≤ 0.1) with each list, and represented by at least 5 genes, were obtained (Additional file ). a . The number of GO terms enriched in the gene lists from each cell variant is indicated and overlaps are shown in the Venn diagram. The 14 GO terms enriched in common for all cell variants are shown. b . Three GO terms enriched in common in A549 and HBE cells, but not in BEAS-2B cells. c . Nineteen GO terms enriched in common in BEAS-2B and HBE cells, but not in A549 cells. d . Sixteen GO terms uniquely enriched in HBE cells. Each box (dotted lines in panels a – d ) also depicts the percentage of the genes associated with each term within the total number of genes repressed in HBE cells and the fold enrichment of each term in the HBE cell list. Significance, following correction for family-wide false discovery, in A549 (A), BEAS-2B (B), and HBE (H) cells, is indicated where: Benjamini ≤0.05 (*), Benjamini ≤0.01 (**), or Benjamini ≤0.001 (***)

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: Variant Assay

Journal: BMC Medical Genomics

Article Title: Glucocorticoid-driven transcriptomes in human airway epithelial cells: commonalities, differences and functional insight from cell lines and primary cells

doi: 10.1186/s12920-018-0467-2

Figure Lengend Snippet: Comparison of budesonide-regulated genes in the three epithelial cell variants with those in airway tissue in vivo. Microarray analysis of A549 ( N = 3), BEAS-2B ( N = 4) and primary HBE ( N = 4 donors) cells as well as airway tissue ( N = 11 individuals) following budesonide exposure for 6 h identified 451 and 442 genes showing significant induction (≥ 2 fold, P ≤ 0.05) or repression (≤ 0.5 fold, P ≤ 0.05), respectively, compared to untreated control/placebo, in any of the cell variants or airway tissue. a . The 451 genes significantly induced (≥ 2 fold, P ≤ 0.05) by budesonide in any cell variant or airway tissue produced 15 groups showing overlaps as defined by the lower stringency, ≥ 1.25 fold cut-off, for each group. b . The 441 genes significantly repressed (≤ 0.5 fold, P ≤ 0.05) by budesonide in any cell variant or in airway tissue produce 15 groups showing overlaps, as defined by the lower stringency, ≤0.8 fold cut-off, for each group. Bars represent the number of genes within each group and below the genes in selected groups are listed. Genes within all groups are listed in Additional file (for induced genes) and 13 (for repressed genes). Underlined genes represent genes that were in common between the cell variants using the stringent cut-off in Fig. (for induced genes) and Fig. (for repressed genes)

Article Snippet: Budesonide (AstraZeneca, Sweden) and dexamethasone (Steraloids, Newport, RI) were dissolved in dimethyl sulphoxide (DMSO) (Sigma-Aldrich) as stocks of 10 mM.

Techniques: In Vivo, Microarray, Variant Assay, Produced

Journal: The Journal of Biological Chemistry

Article Title: β-Arrestin-1 inhibits glucocorticoid receptor turnover and alters glucocorticoid signaling

doi: 10.1074/jbc.RA118.007150

Figure Lengend Snippet: β-arrestin-1 influences the GR transcriptional response. Quantitative real-time PCR to measure the expression levels of GR target genes TSC22D3 (A), IL-1β (B), and CXCL5 (C), in cells transfected with NTC or ARRB1-KD siRNA or that were treated with 100 nm dexamethasone for the indicated times. All data represent mean ± S.D. (error bars) from three independent experiments. ****, p < 0.0001 for Dex versus control. #, p < 0.05; ##, p < 0.01; ####, p < 0.0001 to compare ARRB1-KD with NTC at each time point.

Article Snippet: Dexamethasone (chemical name: 1,4-pregnadien-9α-fluoro-16α-methyl-11β,17,21-triol-3,20-dione) and mifeptistone (RU-486) were purchased from Steraloids (Newport, RI).

Techniques: Real-time Polymerase Chain Reaction, Expressing, Transfection, Control

Journal: The Journal of Biological Chemistry

Article Title: β-Arrestin-1 inhibits glucocorticoid receptor turnover and alters glucocorticoid signaling

doi: 10.1074/jbc.RA118.007150

Figure Lengend Snippet: PELI1 is differentially regulated by Dex in β-arrestin-1 KD cells. A, quantitative RT-PCR analysis of ARRB1 mRNA (left) and β-arrestin-1 protein (right) levels from A549 cells transfected with NTC or ARRB1-KD siRNA that were treated with vehicle or 100 nm Dex for 3 h. B, qRT-PCR analysis of GR mRNA (left) and protein (right) levels from NTC and ARRB1-KD cells that were treated with vehicle or 100 nm Dex for 3 h. C, NTC and ARRB1-KD cells were treated with vehicle (Ctrl) or 100 nm Dex for 3 h to evaluate GR cellular localization by immunofluorescence. D, quantitative real-time PCR to measure the expression levels of E3 ubiquitin ligase genes in cells transfected with NTC or ARRB1-KD siRNA that were treated with 100 nm dexamethasone for 3 h. Data represent the mean ± S.D. (error bars) from three independent experiments. *, p < 0.05; ***, p < 0.001; ****, p < 0.0001 for Dex versus control. #, p < 0.05; ##, p < 0.01; ###, p < 0.001 to compare ARRB1-KD with NTC.

Article Snippet: Dexamethasone (chemical name: 1,4-pregnadien-9α-fluoro-16α-methyl-11β,17,21-triol-3,20-dione) and mifeptistone (RU-486) were purchased from Steraloids (Newport, RI).

Techniques: Quantitative RT-PCR, Transfection, Immunofluorescence, Real-time Polymerase Chain Reaction, Expressing, Ubiquitin Proteomics, Control