Journal: Frontiers in Oncology

Article Title: Primary extraovarian adult granulosa cell tumor of the greater omentum: a case report and literature review

doi: 10.3389/fonc.2025.1689815

Figure Lengend Snippet: Pathological characteristics of the extraovarian adult granulosa cell tumor of the greater omentum. (A–D) Photomicrograph of the tumor (H&E stain). (A) The tumor is encapsulated (H&E stain, 40×magnification, Scale bar: 625 μm). (B) Neoplastic cells are arranged in cord-like and ribbon-like patterns (H&E stain, 100×magnification, Scale bar: 200 μm). (C) Call-Exner bodies are visible (H&E stain, 200×magnification, Scale bar: 100 μm). (D) The tumor is composed of round, oval, or polygonal cells with ill-defined cytoplasmic borders imparting a syncytial appearance. The scant cytoplasm ranges from pale to eosinophilic. Nuclei are round, oval, or angular with finely dispersed chromatin and inconspicuous nucleoli. Longitudinal nuclear grooves and coffee-bean shaped nuclei are present(H&E stain, 400×magnification, Scale bar: 50 μm). (E–I) Photomicrograph of immunohistochemical staining. Immunohistochemistry staining is positive for α-inhibin (E) , SF-1 (F) , FOXL2 (G) , WT-1 (H) . (I) Ki-67 Proliferation Index was 20% (IHC stain, 200× magnification, Scale bar: 100 μm).

Article Snippet: WT-1 , Positive , Strong,diffuse nucleus , Supports diagnosis , OTIRIH(Rabbit Monoclonal), Zhongshan Golden Bridge.

Techniques: Staining, Immunohistochemical staining, Immunohistochemistry

Journal: International journal of molecular sciences

Article Title: Porcine Kidney Organoids Derived from Naïve-like Embryonic Stem Cells.

doi: 10.3390/ijms25010682

Figure Lengend Snippet: Figure 5. Gene expression in 3D porcine kidney organoids. (A) Expression of renal progenitor cell markers in the organoids on day 21 and fetal porcine kidney on day 45. Renal progenitor cells markers (EYA1, SIX1) were measured by immunofluorescence test (EYA1 was localized in the cytoplasm, SIX1 was localized in the nucleus). (B) Expression of mature renal cell markers in the organoids on day 21 and fetal porcine kidney on day 45. Mature nephron components markers (PAX2, E-CAD, PODO) were measured by immunofluorescence test and immunohistochemistry test (the circular dotted lines are tubule-like structures; PAX2 was localized in the nucleus; and PODO and ECAD were localized to the cell membrane). (C) Whole-mount co-staining of the organoids for WT1 and CD31 on day 21 (WT1 was localized in the nucleus and CD31 was localized to the cell membrane) (scale bar: 50 µm).

Article Snippet: The primary antibodies included the following: EYA1 (Abcam), SIX1 (Proteintech), PAX2 (Santa Cruz Biotechnology), WT1 (Abcam), E-CAD (Abcam), PODO (Absin), CD31 (Santa Cruz Biotechnology), AQP1 (Proteintech), GLUT1 (proteintech), and KI67 (Abcam); the secondary antibodies included goat anti-rabbit IgG Alexa Fluor 546 (ABclonal), goat anti-rabbit IgG Alexa Fluor 488 (Thermo Fisher Scientific), goat anti-mouse IgG Alexa Fluor 546 (Thermo Fisher Scientific), goat anti-mouse IgG Alexa Fluor 488 (Thermo Fisher Scientific), goat anti-rabbit IgG H&L(HRP) (Abcam), and goat anti-mouse IgG(H + L) (KeyGEN BioTECH, Nanjing, China).

Techniques: Gene Expression, Expressing, Immunohistochemistry, Membrane, Staining

Journal: Open Medicine

Article Title: WTAP affects intracranial aneurysm progression by regulating m6A methylation modification

doi: 10.1515/med-2023-0818

Figure Lengend Snippet: Roles of WTAP overexpression in the growth of the TNF-α-induced rBMVECs. (a) Cell viability of the rBMVECs, with different treatments, following 24, 48, and 72 h of culture. (b) Apoptosis of the rBMVECs, with different treatments, determined using flow cytometry. (c) Cleaved-caspase3 protein expression detected using western blotting. (d) WTAP protein expression detected using western blotting. * P < 0.05, as compared to the samples from the control cells; # P < 0.05, as compared to those from the TNF-α cells.

Article Snippet: Following 2 h of incubation, the primary antibodies, including the anti-WTAP antibody (1:5,000, Proteintech Group, Inc.), anti-cleaved caspase3 (1:1,000, Proteintech Group, Inc.), and anti-GAPDH antibody (1:1,000, Proteintech Group, Inc.), were added to the membranes.

Techniques: Over Expression, Flow Cytometry, Expressing, Western Blot, Control

Journal: Journal of Clinical Investigation

Article Title: Profilin1 is required to prevent mitotic catastrophe in murine and human glomerular diseases

doi: 10.1172/jci171237

Figure Lengend Snippet: Figure 1. Generation of podocyte-specific Pfn1-KO mice results in severe proteinuria and kidney failure. (A) Representative immunoblot images of profilin1 expression in primary podocytes freshly isolated from littermate control (Ctrl) and Pfn1-KO mice (age P7). (B) Representative immunofluorescence images of profilin1 (green) and WT1 (red) in control and Pfn1-KO primary podocytes. Scale bar: 10 μm. (C) Representative immunofluorescence images of profilin1 (green) and nephrin (red) on kidney sections of control and Pfn1-KO mice (age 3 weeks). Scale bars: 10 μm. (D) Pfn1-KO mice (red) failed to gain body weight by 8 weeks of age compared with control mice (green). n = 9 mice. *P < 0.05 vs. control. (E) The survival curve of Pfn1-KO mice (red) demon- strates approximately 90% death by 12 weeks of age. n = 8 mice. (F) SDS-PAGE (Coomassie blue staining) of standard BSA and of urine samples from Pfn1-KO mice at 4 weeks of age demonstrates albuminuria. Equal volumes of standard BSA and urine (4 μL) were loaded in each lane. (G) Quantification of urine albumin normalized to creatinine at 2, 6, and 10 weeks of age. n = 6 mice. *P < 0.05 vs. control. (H) Elevated plasma creatinine in Pfn1-KO mice at 3, 6, and 10 weeks of age. *P < 0.05 vs. control. Statistics were analyzed via a 2-tailed t test.

Article Snippet: Rabbit anti-profilin1 monoclonal antibody (used for immunofluorescence, Thermo Fisher Scientific, catalog MA5-32683); rabbit anti-profilin1 polyclonal antibody (used for Western blotting, Cell Signaling Technology, catalog 3237S); guinea pig anti-nephrin polyclonal antibody (Progen, catalog GP-N2); rabbit anti-Wilms tumor 1 (WT1) monoclonal antibody (Abcam, catalog ab89901); mouse anti-WT1 monoclonal antibody (Novus Biologicals, catalog NB11-60011); rabbit anti-p21 monoclonal antibody (Abcam, catalog ab188224); mouse anti-p53 monoclonal antibody (Cell Signaling Technology, catalog 2524S); rabbit anti-cyclin monoclonal D1 antibody (Cell Signaling Technology, catalog 2978S); mouse anti-cyclin polyclonal B1 antibody (Cell Signaling Technology, catalog 4138S); mouse anti–Ser 319–phosphorylated γH2AX monoclonal antibody (EMD Millipore, catalog 05-636); rat anti-mCherry monoclonal antibody (Invitrogen, catalog M11217); goat anti-mVenus polyclonal antibody (MyBioSource, catalog MBS448126); Hoechst (Thermo Fisher Scientific, catalog 62249); rabbit anti-GAPDH monoclonal antibody (Cell Signaling Technology, catalog 2118S); mouse anti-GFP monoclonal antibody (Roche, catalog 11814460001); Alexa Fluor 488–conjugated phalloidin (Invitrogen, catalog A12379); Alexa Fluor 594–conjugated phalloidin (Invitrogen, catalog A12381); Alexa Fluor 488–conjugated tubulin (Abcam, catalog ab195883); Alexa Fluor 488 goat anti-mouse IgG antibody (Invitrogen, catalog A11029); Alexa Fluor 488 goat anti-rabbit IgG antibody (Invitrogen, catalog A11034); Alexa Fluor 488 donkey anti-goat IgG antibody (Invitrogen, catalog A11055); Alexa Fluor 594 goat anti-rabbit IgG antibody (Invitrogen, catalog A21207); Alexa Fluor 594 goat anti-mouse (Invitrogen, catalog A11032); Alexa Fluor 594 goat anti-guinea pig (Invitrogen, catalog A11076); Alexa Fluor 594 goat rat (Invitrogen, catalog A11007); Alexa Fluor 647 goat anti-rabbit (Invitrogen, catalog A21245); mouse anti– mouse IgG HRP-conjugated antibody (Rockland, catalog 18-8817- 31); and rabbit anti–mouse IgG HRP-conjugated antibody (Millipore Sigma, catalog AP160P) were purchased commercially.

Techniques: Western Blot, Expressing, Isolation, Control, Immunofluorescence, SDS Page, Staining, Clinical Proteomics

Journal: Journal of Clinical Investigation

Article Title: Profilin1 is required to prevent mitotic catastrophe in murine and human glomerular diseases

doi: 10.1172/jci171237

Figure Lengend Snippet: Figure 3. Loss of podocyte Pfn1 results in morphologic MC appearance, chromosomal instability, and dsDNA damage. (A) Representative images of transmission electron micrography demonstrate abnormal MC podocytes in Pfn1-KO mice (arrow) compared with control glomeruli at 4 weeks of age. Scale bar: 1 μm. (B) Immunofluorescence images of primary podocytes isolated from control and Pfn1-KO mice at P7 stained with WT1 (red, podocyte marker) and Hoechst (blue, DNA marker) showing abnormal MC podocytes in Pfn1-KO mice (arrow) compared with control podocytes. Scale bar: 20 μm. (C) Immunofluorescence images of primary culture podocytes isolated from control and Pfn1-KO mice stained with Hoechst and anti-tubulin antibody showing the chromosome bridge (arrow) in a MC Pfn1-KO podocyte. Scale bar: 20 μm. (D) Quantification of the percentage of MC podocytes per field of view in B. Total of 100 fields of view in 5 independent experiments. (E) Quantification of the percentage of chromosome bridge per field of view in C. n = 5 independent experiments. *P < 0.05 vs. control. (F) Immunofluorescence images of primary culture podocytes stained with γ-H2AX (green, double-strand breaks [DSBs] marker) and WT1 (red) showing abnormal MC podocytes in Pfn1-KO mice, as indicated by the arrows. Scale bar: 20 μm. (G) Quantification of γH2AX foci per podocyte nucleus (left) and per podocyte nuclear area (right) in F. Total of 400 cells in 5 independent experiments. *P < 0.05 vs. control. Statistics were analyzed via a 2-tailed t test.

Article Snippet: Rabbit anti-profilin1 monoclonal antibody (used for immunofluorescence, Thermo Fisher Scientific, catalog MA5-32683); rabbit anti-profilin1 polyclonal antibody (used for Western blotting, Cell Signaling Technology, catalog 3237S); guinea pig anti-nephrin polyclonal antibody (Progen, catalog GP-N2); rabbit anti-Wilms tumor 1 (WT1) monoclonal antibody (Abcam, catalog ab89901); mouse anti-WT1 monoclonal antibody (Novus Biologicals, catalog NB11-60011); rabbit anti-p21 monoclonal antibody (Abcam, catalog ab188224); mouse anti-p53 monoclonal antibody (Cell Signaling Technology, catalog 2524S); rabbit anti-cyclin monoclonal D1 antibody (Cell Signaling Technology, catalog 2978S); mouse anti-cyclin polyclonal B1 antibody (Cell Signaling Technology, catalog 4138S); mouse anti–Ser 319–phosphorylated γH2AX monoclonal antibody (EMD Millipore, catalog 05-636); rat anti-mCherry monoclonal antibody (Invitrogen, catalog M11217); goat anti-mVenus polyclonal antibody (MyBioSource, catalog MBS448126); Hoechst (Thermo Fisher Scientific, catalog 62249); rabbit anti-GAPDH monoclonal antibody (Cell Signaling Technology, catalog 2118S); mouse anti-GFP monoclonal antibody (Roche, catalog 11814460001); Alexa Fluor 488–conjugated phalloidin (Invitrogen, catalog A12379); Alexa Fluor 594–conjugated phalloidin (Invitrogen, catalog A12381); Alexa Fluor 488–conjugated tubulin (Abcam, catalog ab195883); Alexa Fluor 488 goat anti-mouse IgG antibody (Invitrogen, catalog A11029); Alexa Fluor 488 goat anti-rabbit IgG antibody (Invitrogen, catalog A11034); Alexa Fluor 488 donkey anti-goat IgG antibody (Invitrogen, catalog A11055); Alexa Fluor 594 goat anti-rabbit IgG antibody (Invitrogen, catalog A21207); Alexa Fluor 594 goat anti-mouse (Invitrogen, catalog A11032); Alexa Fluor 594 goat anti-guinea pig (Invitrogen, catalog A11076); Alexa Fluor 594 goat rat (Invitrogen, catalog A11007); Alexa Fluor 647 goat anti-rabbit (Invitrogen, catalog A21245); mouse anti– mouse IgG HRP-conjugated antibody (Rockland, catalog 18-8817- 31); and rabbit anti–mouse IgG HRP-conjugated antibody (Millipore Sigma, catalog AP160P) were purchased commercially.

Techniques: Transmission Assay, Control, Immunofluorescence, Isolation, Staining, Marker

Journal: Journal of Clinical Investigation

Article Title: Profilin1 is required to prevent mitotic catastrophe in murine and human glomerular diseases

doi: 10.1172/jci171237

Figure Lengend Snippet: Figure 5. Loss of podocyte Pfn1 results in podocytopenia and cell cycle entry. (A) A schematic representing cell cycle phase visualization by different colors in podocytes from the R26Fucci2aR Pfn1-KO mice. G1 phase nuclei (mCherry, red); S phase nuclei (both mCherry and mVenus, yellow); and G2 phase nuclei (mVenus, green). (B) Representative immunofluorescence images of podocytes in control-FUCCI and Pfn1-KO-FUCCI glomeruli at 4 weeks of age stained with Cherry (red), Venus (green), and WT1 (blue). Scale bar: 20 μm. (C) Quantification of the distribution of podocytes in cell cycle phase in B. Total of 100 glomeruli in 5 different mice. (D) Primary podocytes isolated from control and Pfn1-KO mice demonstrated a significant decrease in adhesion after plating for 5 minutes and 10 minutes on the collagen type I–coated plates. n = 6 independent experiments. (E) Representative immunofluorescence images of podocytes in control and Pfn1-KO glomeruli at 7 weeks of age stained with WT1 (red). Scale bar: 20 μm. (F) Quantification of podocyte density in glomeruli in E. Total of 40 glomeruli from 5 different mice. (G) Representative immunofluorescence images of urinary podocytes from control and Pfn1-KO mice at 4 weeks of age stained with mCherry (red), mVenus (green), and WT1 (blue). G1 phase podocyte nuclei (red arrow), S phase podocyte nuclei (yellow arrow), and G2 phase podocyte nuclei (green arrow) were depicted. Scale bar: 20 μm. *P < 0.05 vs. control. Statistics were analyzed via a 2-tailed t test.

Article Snippet: Rabbit anti-profilin1 monoclonal antibody (used for immunofluorescence, Thermo Fisher Scientific, catalog MA5-32683); rabbit anti-profilin1 polyclonal antibody (used for Western blotting, Cell Signaling Technology, catalog 3237S); guinea pig anti-nephrin polyclonal antibody (Progen, catalog GP-N2); rabbit anti-Wilms tumor 1 (WT1) monoclonal antibody (Abcam, catalog ab89901); mouse anti-WT1 monoclonal antibody (Novus Biologicals, catalog NB11-60011); rabbit anti-p21 monoclonal antibody (Abcam, catalog ab188224); mouse anti-p53 monoclonal antibody (Cell Signaling Technology, catalog 2524S); rabbit anti-cyclin monoclonal D1 antibody (Cell Signaling Technology, catalog 2978S); mouse anti-cyclin polyclonal B1 antibody (Cell Signaling Technology, catalog 4138S); mouse anti–Ser 319–phosphorylated γH2AX monoclonal antibody (EMD Millipore, catalog 05-636); rat anti-mCherry monoclonal antibody (Invitrogen, catalog M11217); goat anti-mVenus polyclonal antibody (MyBioSource, catalog MBS448126); Hoechst (Thermo Fisher Scientific, catalog 62249); rabbit anti-GAPDH monoclonal antibody (Cell Signaling Technology, catalog 2118S); mouse anti-GFP monoclonal antibody (Roche, catalog 11814460001); Alexa Fluor 488–conjugated phalloidin (Invitrogen, catalog A12379); Alexa Fluor 594–conjugated phalloidin (Invitrogen, catalog A12381); Alexa Fluor 488–conjugated tubulin (Abcam, catalog ab195883); Alexa Fluor 488 goat anti-mouse IgG antibody (Invitrogen, catalog A11029); Alexa Fluor 488 goat anti-rabbit IgG antibody (Invitrogen, catalog A11034); Alexa Fluor 488 donkey anti-goat IgG antibody (Invitrogen, catalog A11055); Alexa Fluor 594 goat anti-rabbit IgG antibody (Invitrogen, catalog A21207); Alexa Fluor 594 goat anti-mouse (Invitrogen, catalog A11032); Alexa Fluor 594 goat anti-guinea pig (Invitrogen, catalog A11076); Alexa Fluor 594 goat rat (Invitrogen, catalog A11007); Alexa Fluor 647 goat anti-rabbit (Invitrogen, catalog A21245); mouse anti– mouse IgG HRP-conjugated antibody (Rockland, catalog 18-8817- 31); and rabbit anti–mouse IgG HRP-conjugated antibody (Millipore Sigma, catalog AP160P) were purchased commercially.

Techniques: Immunofluorescence, Control, Staining, Isolation

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: Induced by PMA for 48 hours, the THP-1 cells were collected, the expression of WT1 cells was detected by western blotting assay, and the CD11b and CD14 of THP-1 cells was detected by flow cytometry. A. WT1 expression of THP-1 cells induced by PMA; B: Mean WT1 protein expression of THP-1 cells induced by PMA. C: WT1 expression of THP-1 cells after exposure to PMA or PMA+empty vector; D:Mean WT1 expression of THP-1 cells after exposure to PMA or PMA+empty vector; E: WT1 expression of THP-1 cells transfected with WT1 vector as compared with control group or PMA group; F: Mean relative expression of WT1 in THP-1 cells transfected with WT1 vector as compared with control group or PMA group; G. Inhibiting percent of WT1 expression by PMA and the restoration percent of WT1 expression by PMA + WT1 vector to that of control group, which were based on the results of and ; H. The WT1 expression after exposure to PMA or PMA + WT1 vector by Western blot assay; I. Mean relative expression of WT1 in THP-1 cells after exposure to PMA or PMA + WT1 vector; J. The CD11b expression by flow cytometry assay; K. Expression of CD11b antigen after exposure to PMA or PMA + WT1 vector; L. The CD14 expression after exposure to PMA or PMA + WT1 vector by flow cytometry assay; M: Expression of CD14 antigens after exposure to PMA or PMA + WT1 vector. Each experiment was repeated three times. (* P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Expressing, Western Blot, Flow Cytometry, Plasmid Preparation, Transfection, Control

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: TargetScan and miRTarBase were used to predict potential upstream associated miRNAs of WT1. RNA seq technology to identify differentially altered genes (greater than 2 and a P < 0.05) as the boundary. TargetScan and MirDB databases were used to predict the miRNAs associated with monocyte/macrophage differentiation, and the intersection of the two databases was tried. Further validation of positively related miRNAs was demonstrated by qRT-PCR assay. In order to further verify whether miRNA-132-3p could bind WT1 3’UTR and modulate its promoter activity, a dual luciferase reporter assay was used. The relative expression of miRNAs was detected by qRT-PCR assay, and the correlation analysis between expression of miRNA-132-3p and WT1was indicated by Pearson analysis. A. TargetScan database predicts the possible miRNAs that target and bind to WT1; B. The number of differential miRNA species before and after differentiation and the number of macrophage differentiation-associated miRNA species analyzed by database; C. Target gene prediction, differential detection before and after differentiation, and database analysis macrophage-associated miRNA intersection analysis; D. miRNA-132-3p and miR-212-3p related to WT1 expression by intersection analysis; E: related expression of miRNA-132-3p before and after differentiation. F: miR-212-3p expression before and after differentiation; G: miRNA-132-3p targets WT1 3̛ UTR sequence prediction; H. Luciferase activity assay of miRNA-132-3p targeting WT1; I. The relative expression level of miRNA-132-3p in THP-1 cells before and after PMA induction; J. Relative expression levels of WT1 in THP-1 cells before and after PMA induction; K. Correlation analysis between miRNA-132-3p and WT1 expression. (ns: no significant, * P < 0.05, * *** P < 0.0001).

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: RNA Sequencing, Biomarker Discovery, Quantitative RT-PCR, Activity Assay, Luciferase, Reporter Assay, Expressing, Sequencing

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: Rescue experiment about miRNA-132-3p to regulate differentiation of THP-1 cells by targeting WT1 was tried. WT1 protein expression was detected by Western blotting assay, the expression level of CD11b and CD14 differentiation antigen were detected by flow cytometry assay. A. The transfection efficiency of miRNA-132-3p mimics was detected by qRT-PCR; B. WT1 expression after exposure to mimics NC, miR-132-3p mimics or miR-132-3p mimics+empty vector; C.mean WT1 expression after exposure to mimics NC, miR-132-3p mimics or miR-132-3p mimics+empty vector; D. WT1 protein expression was detected by Western blot; E. Mean WT1 expression level; F. CD11b expression of mimics NC group by flow cytometry; G.CD11b expression of miR-132-3p mimics group by flow cytometry; H. CD11b expression of miR-132-3p mimics+WT1 vector group by flow cytometry; I.Mean expression level of CD11b differentiation antigen; J. CD14 expression of mimics NC group by flow cytometry; K.CD14 expression of miR-132-3p mimics group by flow cytometry; L.CD14 expression of miR-132-3p mimics+WT1 vector group by flow cytometry M. Mean expression levels of CD14 differentiation antigens. Each experiment was repeated three times.(** P < 0.01, *** P < 0.001).

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Expressing, Western Blot, Flow Cytometry, Transfection, Quantitative RT-PCR, Plasmid Preparation

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: β 1 expression by WT1 on committed differentiation of monocytes/macrophages. WT1 protein expression was detected by Western blotting assay, and the expression level of CD11b and CD14 differentiation antigen were detected by flow cytometry assay. A. Western blot was used to detect the protein level of TGF-β1 in THP-1 cells after PMA treatment; B. Changes in the mean level of TGF-β1 protein in THP-1 cells after PMA induction; C. The expression of WT1 or TGF-β1 protein in THP-1 cells transfected with si-WT1 scramble NC or si-WT1; D. Mean WT1 protein expression level in THP-1 cells transfected with si-WT1 scramble NC or si-WT1; E. Mean TGF-β1 protein expression in THP-1 cells transfected with si-WT1 scramble NC or si-WT1; F. Expression of CD11b antigens of control group by flow cytometry; G. Expression of CD11b antigens of si-WT1 scramble NC group by flow cytometry; H. Expression of CD11b antigens of si-WT1 group by flow cytometry; I. Mean CD11b antigens of above-mentioned indicated group; J. Expression of CD14 antigens of control group by flow cytometry; K. Expression of CD14 antigens of si-WT1 scramble NC group by flow cytometry; L. Expression of CD14 antigens of si-WT1 group by flow cytometry; M. Mean CD14 antigens of above-mentioned indicated group;. Each experiment was repeated three times, (* P < 0.05, *** P < 0.001, **** P < 0.0001).

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Expressing, Western Blot, Flow Cytometry, Transfection, Control

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: β 1 expression. To further verify whether WT1 targets binding and regulates expression of TGF-β1, Chromosome immunoprecipitation (ChIP) assay for targeting regulation of TGF-β1 by WT1. A. Prediction of the binding region of WT1-targeted TGF-β1 promoter; B. Primer sequence design for ChIP assay of TGF-β1; C. ChIP assay results of WT1 targeting TGF-β1.

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Expressing, Binding Assay, Immunoprecipitation, Sequencing

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: β 1 to induce differentiation of THP-1 cells. miRNA-132-3p mimics-NC group, miRNA-132-3p mimics group, and miRNA-132-3p mimics+si-TGF-β1 group three groups were used to try rescue assay about miRNA-132-3p modulating expression of TGF-β1. Western blotting assay was used for detecting expression of TGF-β1 protein. A. TGF-β1 expression in the indicated mimics NC, miR-132-3p mimics or miR-132-3p mimics+si-TGF-β1 scramble NC groups by Western blot assay; B.Mean expression of TGF-β1 protein in the above indicated groups; C. TGF-β1 expression in the indicated mimics NC, miR-132-3p mimics or miR-132-3p mimics+si-TGF-β1 groups by Western blot assay; D. Mean expression of TGF-β1 protein in the above indicated groups; E. Expression of CD11b antigens of mimics NC group by flow cytometry;.F. Expression of CD11b antigens of miR-132-3p mimics group by flow cytometry; G. Expression of CD11b antigens of miR-132-3p mimics+WT1 vector group by flow cytometry; H.Mean expression of CD11b in the indicated groups; I. Expression of CD14 antigens of mimics NC group by flow cytometry;.J. Expression of CD14 antigens of miR-132-3p mimics group by flow cytometry; K. Expression of CD14 antigens of miR-132-3p mimics+WT1 vector group by flow cytometry; L.Mean expression of CD14 antigens in the indicated groups;.Each experiment was repeated three times, (* P < 0.05, ** P < 0.01).

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Rescue Assay, Expressing, Western Blot, Flow Cytometry, Plasmid Preparation

Journal: PLOS One

Article Title: The up-regulation of TGF-β1 by miRNA-132-3p/WT1 is involved in inducing leukemia cells to differentiate into macrophages

doi: 10.1371/journal.pone.0306150

Figure Lengend Snippet: β1 axis contributed to inducing differentiation of THP-1 cells into macrophages. In THP-1 cells, PMA could induce the upregulation of miRNA-132-3p, the upregulation of miRNA-132-3p results in the downregulation of WT1 by sponge action with binding on its 3’UTR, the down regulation of WT1 contributes to the upregulation of TGF-β1, then the polarization of macrophages is induced, which is indicated by the upregulation of CD11b and CD14 expression. Therefore, the miRNA-132-3p/WT1/TGF-β1 axis is involved in the committed differentiation of THP-1 leukemia cells into macrophages.

Article Snippet: Regarding the antibodies, WT1 antibody (cat no. 83535S) was purchased from CST Biological Reagents Co., Ltd., β-actin (cat. no. 20536–1-AP) primary antibody was from Proteintech Group, Inc., TGF-β1 antibody (cat. no. YT4632) was purchased from Immunoway Biotechnology, Inc., and HRP-labeled rabbit secondary antibody (cat. no. ZB-2301) was provided by OriGene Technologies, Inc. Sangon Biotech Co., Ltd. designed and synthesized the primers.

Techniques: Binding Assay, Expressing

Journal: iScience

Article Title: FEZ1 participates in human embryonic brain development by modulating neuronal progenitor subpopulation specification and migrations

doi: 10.1016/j.isci.2023.108497

Figure Lengend Snippet:

Article Snippet: Rabbit anti-Nanog (IF: 1:500) , Cell Signaling , Cat#3580S, RRID: AB_2150399.

Techniques: Transduction, Virus, Recombinant, Membrane, Proliferation Assay, CyQUANT Assay, Viability Assay, DNA Purification, SYBR Green Assay, Imaging, cDNA Synthesis, Gene Expression, Software

Journal: Pakistan Journal of Medical Sciences

Article Title: Differentiation of CD117 + Amniotic Fluid Stem Cells towards Nephron Progenitors

doi: 10.12669/pjms.38.6.4887

Figure Lengend Snippet: Characterization of NPCs colonies for WT1 by immunofluorescence staining as performed on three different amniotic fluid derived NPCs. This figure shows immunofluorescence analysis for positive nuclear marker WT1 (a) Bright field image of NPCs at 40x showed irregular with ill-defined borders having tightly packed cells for the three different amniotic fluid derived NPCs. (b) Counter DAPI nuclear stain showed blue color of Nucleus. (c) Characteristic of NPCs, WT1 is shown in nuclear compartment of cells.

Article Snippet: The amniotic fluid samples were washed thoroughly with Phosphate Buffered Saline then fixed at 1:1 acetone: methanol for twenty minutes at 25°C, after washing with Buffer Saline containing tween 20 (PBST) three times Triton X-100 was use for permeabilization of cell membrane then again washed with PBST, it was clogged by bovine serum albumin (BSA) 1% plus PBS (without Calcium &Magnesium) for 30 minutes at Room Temperature and incubated with primary monoclonal unconjugated antibody WT1 (WT1/857 + 6F-H2 clone, Novus Biologicals, Biotechne, USA) as positive and CD117 (CD117 APC, Thermofisher, 104D2) as a negative control for an hour.

Techniques: Immunofluorescence, Staining, Derivative Assay, Marker