Journal: Genes

Article Title: Activation of Steroidogenesis, Anti-Apoptotic Activity, and Proliferation in Porcine Granulosa Cells by RUNX1 Is Negatively Regulated by H3K27me3 Transcriptional Repression

doi: 10.3390/genes11050495

Figure Lengend Snippet: The influence of RUNX1 on steroidogenesis, cell apoptosis and proliferation in pGCs. ( a ) The expression pattern of steroidogenesis related genes when RUNX1 was overexpressed or knocked down. The * indicates p < 0.05 and ** indicates p < 0.01. ( b ) Protein translation of RUNX1, PTGS2, ER1, ER2, HSD17β1, HSD17β4, CYP11A1, and CYP19A1 when pGCs were with/without human chorionic gonadotropin (hCG) in the situation of siRUNX1 or siNC. The numbers 1 and 2 indicate that pGCs were treated without/with hCG. ( c ) The hormone synthesis of androgen, estrogen, prostaglandin, and progesterone in the situation of upregulation and knockdown of RUNX1 in pGCs. ( d ) The effect of RUNX1 on pGC apoptosis. The pGCs apoptotic proportion in that column plot is a visualization from the sum of the upper and lower right quadrants in the scatter plot. ( e ) The effect of RUNX1 on pGC proliferation. The proportion of proliferated cells in the column equal to the ratio of the numbers of proliferated cells to total cells.

Article Snippet: Antibodies against RUNX1 (Abcam), PTGS2 (bs-0732R, BioSS ANTIBODIES, Woburn, USA), CYP11A1 (bs-10099R, BioSS ANTIBODIES, Woburn, MA, USA), CYP19A1 (bs-1292R, BioSS ANTIBODIES, Woburn, USA), HSD17β1 (bs-3855R, BioSS ANTIBODIES, Woburn, MA, USA), HSD17β4 (bs-11296R, BioSS ANTIBODIES, Woburn, USA), ER1 (bs-2098R, BioSS ANTIBODIES, Woburn, USA), and ER2 (bs-0116R, BioSS ANTIBODIES, Woburn, USA) were used to measure the corresponding protein levels by WB.

Techniques: Expressing

Journal: Genes

Article Title: Activation of Steroidogenesis, Anti-Apoptotic Activity, and Proliferation in Porcine Granulosa Cells by RUNX1 Is Negatively Regulated by H3K27me3 Transcriptional Repression

doi: 10.3390/genes11050495

Figure Lengend Snippet: The influence of the H3K27me3-RUNX1 signal on steroidogenesis, cell apoptosis and proliferation in pGCs. ( a ) The expression patterns of steroidogenesis-related genes when RUNX1 was over-expressed and knocked down, and also when treated with antagonist GSK-126 or agonist GSK-J4 at the same time. The * indicates p < 0.05 and ** indicates p < 0.01. ( b ) Protein translation of RUNX1, PTGS2, ER1, ER2, HSD17β1, HSD17β4, CYP11A1, and CYP19A1. The numbers 1, 2, 3, and 4 indicate treatments with antagonist GSK-126, antagonist-NC, agonist GSK-J4, and agonist-NC. ( c ) The concentrations of androgen, estrogen, prostaglandin, and progesterone in the situation of interfering with the H3K27-RUNX1 signal in pGCs. ( d ) The effect of RUNX1 along with antagonist GSK-126 or agonist GSK-J4 on pGC apoptosis. The pGCs apoptotic proportion in the column plot is a visualization from the sum of the upper and lower right quadrants in the scatter plot. ( e ) The effect of RUNX1 along with antagonist GSK-126 or agonist GSK-J4 on pGCs proliferation. The proportion of proliferated cells in the column is equal to the ratio of total proliferated cells/ total cells.

Article Snippet: Antibodies against RUNX1 (Abcam), PTGS2 (bs-0732R, BioSS ANTIBODIES, Woburn, USA), CYP11A1 (bs-10099R, BioSS ANTIBODIES, Woburn, MA, USA), CYP19A1 (bs-1292R, BioSS ANTIBODIES, Woburn, USA), HSD17β1 (bs-3855R, BioSS ANTIBODIES, Woburn, MA, USA), HSD17β4 (bs-11296R, BioSS ANTIBODIES, Woburn, USA), ER1 (bs-2098R, BioSS ANTIBODIES, Woburn, USA), and ER2 (bs-0116R, BioSS ANTIBODIES, Woburn, USA) were used to measure the corresponding protein levels by WB.

Techniques: Expressing

Journal: Cells

Article Title: The Male-Biased Expression of miR-2954 Is Involved in the Male Pathway of Chicken Sex Differentiation.

doi: 10.3390/cells12010004

Figure Lengend Snippet: Figure 3. The morphology and gene expression in gonads at E10 chicken after AI treatment. (A–C) Gonads on top of the mesonephros in male, female, and AI-treated female chicken, respectively. (R) indicates right gonad, (L) indicates left gonad. (D–F) HE staining of gonads in male, female, and AI-treated female, respectively. In all HE staining images, the left gonad was selected. Scale bar as is shown in pictures, thickness = 6 µm. (G–L) Q-PCR expression of DMRT1, SOX9, AMH, CYP19A1, FOXL2, and miR-2954, respectively. Error bars show standard error of the mean (SEM) from at least three biological replicates in each group. Asterisks indicate statistical significance, * p < 0.05, ** p < 0.01, *** p < 0.0001. (M1–M3,N1–N3,O1–O3,P1–P3) Immunofluorescence results of DMRT1, SOX9, FOXL2, and CYP19A1 in control and AI-treated gonads. (Q1–Q3) In situ hybridization of miR-2954 in male, female, and AI-treated female. Blue fluorescence shows DAPI, which marks the nucleus; red fluorescence indicates expression of sex markers or miR-2954. Scale bar = 200 µm, thickness = 10 µm.

Article Snippet: Then, the sections were fixed in 4% paraformaldehyde for 15 min, permeabilized in PBS containing 1% Triton X-100 at room temperature for 10 min, and blocked at 37 ◦C in PBS containing 5% BSA for 1 h. The primary antibody solution was diluted with 1% PBS, and DMRT1 antibody (1:100, AbClone, WG-03818, Wuhan, China), SOX9 antibody (1:100, Abcam, EPR12755, Cambridge, UK), FOXL2 antibody (1:200, Cells 2023, 12, 4 5 of 15 Novus, NB100-1277s), and CYP19A1 antibody (1:50, Bio-Rad, MCA2077S) were added and incubated overnight at 4 ◦C.

Techniques: Gene Expression, Staining, Expressing, Control, In Situ Hybridization

Journal: Cells

Article Title: The Male-Biased Expression of miR-2954 Is Involved in the Male Pathway of Chicken Sex Differentiation.

doi: 10.3390/cells12010004

Figure Lengend Snippet: Figure 5. Expression of genes related to gonad differentiation at E10 after miR-2954 inhibition. (A–E), Q-PCR results of DMRT1, SOX9, AMH, CYP19A1, and FOXL2, respectively. Error bars show standard error of the mean (SEM) from at least three biological replicates in each group. Asterisks indicate statistical significance: * p < 0.05; ns means no significant difference. (F–Q) Immunofluores- cence results of DMRT1, SOX9, FOXL2, and CYP19A1, respectively, in male, female, and VMO-treated male gonads. Blue fluorescence shows DAPI, which marks the nucleus; Red fluorescence indicates expression of sex markers. Scale bar as is shown in picture, thickness = 10 µm. M-MO indicates male chicken embryo treated by VMO-miR-2954.

Article Snippet: Then, the sections were fixed in 4% paraformaldehyde for 15 min, permeabilized in PBS containing 1% Triton X-100 at room temperature for 10 min, and blocked at 37 ◦C in PBS containing 5% BSA for 1 h. The primary antibody solution was diluted with 1% PBS, and DMRT1 antibody (1:100, AbClone, WG-03818, Wuhan, China), SOX9 antibody (1:100, Abcam, EPR12755, Cambridge, UK), FOXL2 antibody (1:200, Cells 2023, 12, 4 5 of 15 Novus, NB100-1277s), and CYP19A1 antibody (1:50, Bio-Rad, MCA2077S) were added and incubated overnight at 4 ◦C.

Techniques: Expressing, Inhibition

Journal: International Journal of Molecular Sciences

Article Title: Single-Cell Transcriptomics Analysis of Human Small Antral Follicles

doi: 10.3390/ijms222111955

Figure Lengend Snippet: Identification of oocytes and granulosa cells from small human antral follicles. ( A ) Schematic illustration of the study design; ( B ) Uniform manifold approximation and projection (UMAP) plots showing 4 clusters of cells (left panel), colored by donors (right panel); ( C ) Expression of oocyte markers on the UMAP plot; ( D ) Expression of general granulosa cell (GCs) markers on the UMAP plot; ( E ) Expression of cumulus GC markers on the UMAP plot; ( F ) Expression of mural GC markers on the UMAP plot; ( G ) Immunofluorescence for DDX4 and CYP19A1 (top panels); and pan-cytokeratin (pKRT) and FOXL2 (bottom panels) on small human antral follicles showing the follicle area close to the oocyte. Orange arrows indicate the oocyte. Orange dashed lines indicate the follicular basement membrane. ø, follicle diameter. Scale bars are 50 μm; ( H ) Combined expression of selected immune markers on the UMAP plot; ( I ) Percentage of oocytes (CL3) and GCs (CL0) per small antral follicle.

Article Snippet: The primary antibodies used were mouse anti-PCNA (1:100, SC-56, Santa Cruz, Santa Cruz, CA, USA), mouse anti-CD68 (1:50, M087629-2, DAKO, Glostrup, Denmark), rabbit anti-C1Q (1:200, A0136, DAKO), goat anti-FOXL2 (1:250, ab5096, Abcam, Cambridge, United Kingdom), mouse anti-StAR (1:100, sc166821, Santa Cruz), rabbit anti-Collagen Type IV (1:50, AB748, Merck, Kenilworth, NJ, USA), mouse anti-CYP19A1 (1:50, sc-374176, Santa Cruz), mouse anti-Cytokeratin (1:100, M351501, DAKO), mouse anti-Connexin-43/GJA1 (1:50, 13–8300, Zymed, San Francisco, CA, USA), rabbit anti-S100B (1:100, AB52642, Abcam), mouse anti-β-Catenin/CTNNB1 (1:200, 610154, BD Biosciences, Franklin Lakes, NJ, USA), goat anti-DPPA5 (1/100, AF3125, R&D Systems, Minneapolis, MN, USA), rabbit anti-ZO-1/TJP1 (1:100, 61–7300, eBioscience, Hatfield, UK), mouse anti-AMH (1:30, MCA2246T, BioRad, Hercules, CA, USA), mouse anti-N-Cadherin/CDH2 (GC-4) (1:100, C3865, Sigma-Aldrich, St. Louis, MO, USA), rabbit anti-phospho-SMAD2 (Ser465/467) (1:200, 3108, Cell Signaling, Danvers, MA, USA), rabbit anti-phospho-Smad1 (Ser463/465)/Smad5 (Ser463/465)/Smad9(Ser465/467) (1:200, 13820, Cell Signaling), rabbit anti-NOTCH2 (1:200, 5732, Cell Signaling), goat anti-JAG1 (1:50, sc-6011, Santa Cruz).

Techniques: Expressing, Immunofluorescence

Journal: PLoS ONE

Article Title: ETA-mediated anti-TNF-α therapy ameliorates the phenotype of PCOS model induced by letrozole

doi: 10.1371/journal.pone.0217495

Figure Lengend Snippet: (A-D) The mRNA and protein expressions of CYP19A1 in KGN cells were significantly inhibited after 36 h of persistent treatment with different doses of TNF-α (a,b,c indicate comparison of control with 10, 50, and 100 ng/mL of ETA). However, the influence of TNF-α on CYP17A1 expression could not be determined due to the different expressions of CYP17A1 at different time points compared to that of the control (All groups, * P <0.05 and ** P <0.01 by one-way ANOVA; CTL versus TNF-α, # P <0.05 and ## P <0.01 by Dunnett’s multiple comparison test). (E, F) ETA ameliorated the downregulation of CYP19A1 induced by TNF-α. However, ETA-mediated regulation of CYP17A1 was still undetermined (All groups, * P <0.05 and ** P <0.01 by one-way ANOVA; CTL versus TNF-α, # P <0.05 and ## P <0.01 by Dunnett’s multiple comparison test; TNF-α versus TNF-α+ETA, & P <0.05 and && P <0.01 by Dunnett’s multiple comparison test).

Article Snippet: The primary antibody for P65 (1:100 dilution) was purchased from Proteintech Group (Chicago, USA) and for CYP19A1 (1:100 dilution) from Boster Biotech Co. Ltd. (Wuhan, China).

Techniques: Comparison, Control, Expressing

Journal: PLoS ONE

Article Title: ETA-mediated anti-TNF-α therapy ameliorates the phenotype of PCOS model induced by letrozole

doi: 10.1371/journal.pone.0217495

Figure Lengend Snippet: Persistent exposure to androgen promoted the secretion of TNF-α and other inflammatory factors. TNF-αsignificantly upregulated CYP17A1 and downregulated CYP19A1, the key enzymes involved in the synthesis and conversion of androgens, leading to the increased levels of androgens. Furthermore, etanercept (ETA)-mediated anti-TNF-α therapy in PCOS model significantly alleviated the abnormal hyperandrogenism, lipid recruitment, and follicular development.

Article Snippet: The primary antibody for P65 (1:100 dilution) was purchased from Proteintech Group (Chicago, USA) and for CYP19A1 (1:100 dilution) from Boster Biotech Co. Ltd. (Wuhan, China).

Techniques:

Journal: Journal of Biomedical Research

Article Title: Hydroxylation and sulfation of sex steroid hormones in inflammatory liver

doi: 10.7555/JBR.31.20170031

Figure Lengend Snippet: Primers used for qRT-PCR analysis

Article Snippet: Cyp19a1 antibody (#14528) was purchased from Cell Signaling Technology (Beverly, MA, USA).

Techniques:

Journal: Drug Design, Development and Therapy

Article Title: Semaglutide Alleviates Ovary Inflammation via the AMPK/SIRT1/NF‑κB Signaling Pathway in Polycystic Ovary Syndrome Mice

doi: 10.2147/dddt.s484531

Figure Lengend Snippet: Figure 3 (A) Immunohistochemical images of CYP19A1; (B) CYP19A1 mean density; (C) Immunofluorescence images of CYP17A1 (green), StAR (red), and DAPI (blue); (D) CYP17A1 expression rate; (E) StAR expression rate. Data are presented as mean ± SD. vs NC, ##P <0.01; vs PCOS, **P <0.01; n = 6 per group.

Article Snippet: Ovarian tissue sections were treated with bovine serum albumin for 10 minutes and then incubated overnight with IL-1β rabbit antibodies (dilution 1:50; No. 516288, ZENBIOSCIENCE, China), IL-6 antibodies (dilution 1:50; No. BA4339, BOSTER, China), TNF-α antibodies (dilution 1:50; No. BA0103, BOSTER, China), CYP19A1 antibodies (dilution 1:50; No. BA3704, BOSTER, China), and NF-κB antibodies (dilution 1:50; No. BA0610, BOSTER, China).

Techniques: Immunohistochemical staining, Immunofluorescence, Expressing