Journal: Journal of Lipid Research

Article Title: An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples [S]

doi: 10.1194/jlr.D119000591

Figure Lengend Snippet: Multiple reaction monitoring parameters for steroids using LC/MS/MS

Article Snippet: Pregnenolone (Preg), pregnenolone-17,21,21,21-d4 (Preg-d4), 17-hydroxypregnenolone (17OHPreg), 17-hydroxypregnenolone-21,21,21-d3 (17OHPreg-d3), 17-hydroxyprogesterone (17OHP), 17-hydroxyprogesterone-2,2,4,6,6,21,21,21-d8 (17OHP-d8), corticosterone (CORT), corticosterone-2,2,4,6,6,17,21,21-d8 (CORT-d8), cortisol (COR), 11-deoxycortisol (DOC), estradiol (E2), estriol (E3), estriol-2,4,17-d3 (E3-d3), and progesterone-2,2,4,6,6,17,21,21,21-d9 (P-d9) were purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques: Targeted Proteomics

Journal: Journal of Lipid Research

Article Title: An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples [S]

doi: 10.1194/jlr.D119000591

Figure Lengend Snippet: Intra- and interassay precisions of steroid metabolome analysis

Article Snippet: Pregnenolone (Preg), pregnenolone-17,21,21,21-d4 (Preg-d4), 17-hydroxypregnenolone (17OHPreg), 17-hydroxypregnenolone-21,21,21-d3 (17OHPreg-d3), 17-hydroxyprogesterone (17OHP), 17-hydroxyprogesterone-2,2,4,6,6,21,21,21-d8 (17OHP-d8), corticosterone (CORT), corticosterone-2,2,4,6,6,17,21,21-d8 (CORT-d8), cortisol (COR), 11-deoxycortisol (DOC), estradiol (E2), estriol (E3), estriol-2,4,17-d3 (E3-d3), and progesterone-2,2,4,6,6,17,21,21,21-d9 (P-d9) were purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques:

Journal: Journal of Lipid Research

Article Title: An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples [S]

doi: 10.1194/jlr.D119000591

Figure Lengend Snippet: Certified and measured values of steroids in the MassCheck® Steroid Panel Serum Control

Article Snippet: Pregnenolone (Preg), pregnenolone-17,21,21,21-d4 (Preg-d4), 17-hydroxypregnenolone (17OHPreg), 17-hydroxypregnenolone-21,21,21-d3 (17OHPreg-d3), 17-hydroxyprogesterone (17OHP), 17-hydroxyprogesterone-2,2,4,6,6,21,21,21-d8 (17OHP-d8), corticosterone (CORT), corticosterone-2,2,4,6,6,17,21,21-d8 (CORT-d8), cortisol (COR), 11-deoxycortisol (DOC), estradiol (E2), estriol (E3), estriol-2,4,17-d3 (E3-d3), and progesterone-2,2,4,6,6,17,21,21,21-d9 (P-d9) were purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques:

Journal: Journal of Lipid Research

Article Title: An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples [S]

doi: 10.1194/jlr.D119000591

Figure Lengend Snippet: Baseline characteristics and steroid concentrations of pregnant women

Article Snippet: Pregnenolone (Preg), pregnenolone-17,21,21,21-d4 (Preg-d4), 17-hydroxypregnenolone (17OHPreg), 17-hydroxypregnenolone-21,21,21-d3 (17OHPreg-d3), 17-hydroxyprogesterone (17OHP), 17-hydroxyprogesterone-2,2,4,6,6,21,21,21-d8 (17OHP-d8), corticosterone (CORT), corticosterone-2,2,4,6,6,17,21,21-d8 (CORT-d8), cortisol (COR), 11-deoxycortisol (DOC), estradiol (E2), estriol (E3), estriol-2,4,17-d3 (E3-d3), and progesterone-2,2,4,6,6,17,21,21,21-d9 (P-d9) were purchased from Toronto Research Chemicals (Toronto, Canada).

Techniques:

Journal: Scientific Reports

Article Title: Estrogen replacement therapy-induced neuroprotection against brain ischemia-reperfusion injury involves the activation of astrocytes via estrogen receptor β

doi: 10.1038/srep21467

Figure Lengend Snippet: An immunofluorescence assay revealed that in CA1 region ( A ), CA2 and CA3 region ( B ) of hippocampus, the number of GFAP-positive cells in the OVX mice was significantly lower than that in the normal mice; and 50 μg/kg E2 and 8 mg/kg DPN treatment significantly increased the number of GFAP-positive cells. ( C ) Data are shown as the mean ± S.D; **p < 0.01 vs. Con group; # p < 0.05 and ## p < 0.01 vs. OVX group. Bar: 50 μm.

Article Snippet: These mice were randomly divided into 4 groups: (A) control group (Con), (B) OVX without 17β-estradiol (E2) (Cayman, USA) replacement group (OVX), (C) OVX and 50 μg/kg/day E2 replacement group (E2), (D) OVX and 8 mg/kg/day DPN replacement group (DPN).

Techniques: Immunofluorescence

Journal: Scientific Reports

Article Title: Estrogen replacement therapy-induced neuroprotection against brain ischemia-reperfusion injury involves the activation of astrocytes via estrogen receptor β

doi: 10.1038/srep21467

Figure Lengend Snippet: ( A ) Data are shown as the mean ± S.D; **p < 0.05 vs. Con group; # p < 0.05 and ## p < 0.01 vs. OVX group. ( B ) Cropped gels and blots showing the expression of GFAP in 50 μg/kg E2 and 8 mg/kg DPN replacement group. The samples derive from the same experiment and that gels/blots were processed in parallel. Full-length blots/gels are presented in .

Article Snippet: These mice were randomly divided into 4 groups: (A) control group (Con), (B) OVX without 17β-estradiol (E2) (Cayman, USA) replacement group (OVX), (C) OVX and 50 μg/kg/day E2 replacement group (E2), (D) OVX and 8 mg/kg/day DPN replacement group (DPN).

Techniques: Expressing

Journal: Scientific Reports

Article Title: Estrogen replacement therapy-induced neuroprotection against brain ischemia-reperfusion injury involves the activation of astrocytes via estrogen receptor β

doi: 10.1038/srep21467

Figure Lengend Snippet: ( A ) Data are shown as the mean ± S.D; *p < 0.05 vs. Con group; # p < 0.05 vs. OVX group. ( B ) Cropped gels and blots showing the expression of cleaved-Caspase-3 protein in Con, OVX, E2 treatment or DPN treatment group. The samples derive from the same experiment and that gels/blots were processed in parallel. Full-length blots/gels are presented in .

Article Snippet: These mice were randomly divided into 4 groups: (A) control group (Con), (B) OVX without 17β-estradiol (E2) (Cayman, USA) replacement group (OVX), (C) OVX and 50 μg/kg/day E2 replacement group (E2), (D) OVX and 8 mg/kg/day DPN replacement group (DPN).

Techniques: Expressing

Journal: eLife

Article Title: Heat shock factor 1 (HSF1) cooperates with estrogen receptor α (ERα) in the regulation of estrogen action in breast cancer cells

doi: 10.7554/eLife.69843

Figure Lengend Snippet: ( A ) Overlap of differentially expressed genes and ( B, C ) heatmaps with hierarchical clustering of normalized read counts from RNA-seq (row z-score) for genes stimulated or repressed after the E2 treatment in cells with different levels of HSF1: unmodified (WT), stably transduced with nonspecific shRNA (SCR), stably transduced with HSF1-specific shRNA (shHSF1), the combination of control clones arisen from single cells following CRISPR/Cas9 gene targeting (MIX), HSF1-negative single clones obtained by CRISPR/Cas9 gene targeting (KO#1, KO#2). Ctr, untreated cells; E2, 17β-estradiol treatment (10 nM, 4 hr). ( D ) Principal component analysis (PCA) of normalized RNA-seq read counts (dimensions 1 and 2 were responsible for 41.7% of variability). ( E ) Gene set enrichment analysis showing significant terms from the Hallmark, BioCarta, PID, and Reactome subsets of the canonical pathways collection detected in E2-stimulated HSF1-proficient and HSF1-deficient cells. Blue, a fraction of downregulated genes; red, a fraction of upregulated genes. ( F ) Comparison of the response to E2 stimulation (E2 versus Ctr) in HSF1-proficient and HSF1-deficient cells. Genes shown in ( B ) are sorted from the highest to the lowest difference between average fold changes in both cell variants decreased by the standard deviation (SD). Upregulation, fold change >1.0; downregulation, fold change <1.0. ( G ) Comparison of the expression level (normalized RNA-seq read counts; mean ± SD) of the same set of genes in unstimulated (Ctr) and E2-stimulated HSF1-proficient and HSF1-deficient cells.

Article Snippet: 17β-estradiol (E2; #E4389, Merck KGaA) was added 48 hr later to a final concentration of 10 nM unless otherwise stated for the indicated time.

Techniques: RNA Sequencing, Stable Transfection, Transduction, shRNA, Control, Clone Assay, CRISPR, Comparison, Standard Deviation, Expressing

Journal: eLife

Article Title: Heat shock factor 1 (HSF1) cooperates with estrogen receptor α (ERα) in the regulation of estrogen action in breast cancer cells

doi: 10.7554/eLife.69843

Figure Lengend Snippet: ( A ) Gene set enrichment analysis showing significant (false discovery rate [FDR] ≤ 0.001) terms from the Hallmark, Reactome, BioCarta, and KEGG gene sets collection detected in E2-stimulated HSF1+ and HSF1− cells, as well as differentiating untreated cells. Blue, a fraction of downregulated genes; red, a fraction of upregulated genes. ( B ) Analyses of the expression by RT-qPCR using total RNA. The upper panels show E2-stimulated changes, lower panels show differences between untreated WT, HSF1+, and HSF1− cells (fold change). Ctr, untreated cells; E2, 17β-estradiol treatment (10 nM, 4 hr). ***p<0.0001, **p<0.001, *p<0.05 (above the bar – versus the corresponding control, between the bars – between cell variants).

Article Snippet: 17β-estradiol (E2; #E4389, Merck KGaA) was added 48 hr later to a final concentration of 10 nM unless otherwise stated for the indicated time.

Techniques: Expressing, Quantitative RT-PCR, Control

Journal: eLife

Article Title: Heat shock factor 1 (HSF1) cooperates with estrogen receptor α (ERα) in the regulation of estrogen action in breast cancer cells

doi: 10.7554/eLife.69843

Figure Lengend Snippet: ( A ) Interactions between ERα and HSP90 assessed by Proximity Ligation Assay (PLA; red spots) in HSF1-positive (HSF1+) and HSF1-negative (HSF1−) MCF7 cells created using DNA-free CRISPR/Cas9 system. Ctr, untreated cells; E2, 17β-estradiol treatment (10 nM). DNA was stained with DAPI. Scale bar, 20 μm. The mean number of spots per cell is shown in the boxplots below, which represent the median, upper and lower quartiles, maximum and minimum; ***p<0.0001, **p<0.001, *p<0.05 – significance of differences versus the corresponding control (above the box) or between cell variants. ( B ) Western blot analysis of HSP90 and ( C ) ERα levels in MCF7 and T47D cells cultured in standard conditions: unmodified (WT), variants stably transduced with nonspecific shRNA (SCR) or with HSF1-specific shRNA (shHSF1), a combination of control clones (MIX, HSF1+), two HSF1-negative clones (KO#1, KO#2), and a combination of HSF1-negative (HSF1−) clones arisen from single cells following CRISPR/Cas9 gene targeting. Actin (ACTB) was used as a protein loading control. The graphs show the results of densitometric analyses (n = 3); **p<0.001, *p<0.05 – significance of differences versus the corresponding control. ( D ) HSP90 and ERα localization assessed by immunofluorescence in wild-type MCF7 cells. DNA was stained with DAPI. Scale bar, 20 μm.

Article Snippet: 17β-estradiol (E2; #E4389, Merck KGaA) was added 48 hr later to a final concentration of 10 nM unless otherwise stated for the indicated time.

Techniques: Proximity Ligation Assay, CRISPR, Staining, Control, Western Blot, Cell Culture, Stable Transfection, Transduction, shRNA, Clone Assay, Immunofluorescence

Journal: eLife

Article Title: Heat shock factor 1 (HSF1) cooperates with estrogen receptor α (ERα) in the regulation of estrogen action in breast cancer cells

doi: 10.7554/eLife.69843

Figure Lengend Snippet:

Article Snippet: 17β-estradiol (E2; #E4389, Merck KGaA) was added 48 hr later to a final concentration of 10 nM unless otherwise stated for the indicated time.

Techniques: Transfection, Construct, Recombinant, Plasmid Preparation, shRNA, Expressing, Sequencing, In Situ, SYBR Green Assay, Software, Staining, CRISPR, Protease Inhibitor