Journal: Journal of Molecular Endocrinology

Article Title: CRTC2 and Nedd4 ligase involvement in FSH and TGFβ1 upregulation of connexin43 gap junction

doi: 10.1530/jme-15-0076

Figure Lengend Snippet: Figure 3 FSH and TGFb1 regulation of CRTC2, CREB and CBP binding to Cx43 gene (Gja1) in rat ovarian granulosa cells. Cells were treated with control vehicle, FSH (10 ng/ml) and/or TGFb1 (0.5 ng/ml) for the indicated time periods. At the end of culture, cell lysates were prepared for ChIP analysis using CRTC2, CREB or CBP antibody with normal IgG serving as a negative control and input DNA as an internal control. Ethidium bromide-stained intensity of the PCR products was determined and normalized to the input DNA. Relative density ratios were calculated using the FSHCTGFb1-treated group value as one. Each bar represents the mean (GS.E.M.) relative density (nZ3). Different lowercase letters indicate significant differences among treatment groups (P!0.05). C, control; F, FSH; FT, FSHCTGFb1.

Article Snippet: Recombinant human TGFb1 was purchased from R&D Systems, Inc. (Minneapolis, MN, USA).

Techniques: Binding Assay, Control, Negative Control, Staining

Journal: Journal of Molecular Endocrinology

Article Title: CRTC2 and Nedd4 ligase involvement in FSH and TGFβ1 upregulation of connexin43 gap junction

doi: 10.1530/jme-15-0076

Figure Lengend Snippet: Figure 6 FSH and TGFb1 regulation of Cx43 association with ubiquitin ligase Nedd4 and Cx43 ubiquitination in rat ovarian granulosa cells. Cells were treated with control vehicle or FSH (10 ng/ml) plus TGFb1 (0.5 ng/ml) for 48 h. At the end of culture, cell lysates were prepared for co-immunoprecipitation analysis using Cx43 with normal IgG serving as a negative control. Due to control, cells had much less Cx43 protein than the FSHCTGFb1-treated cells; similar total content of Cx43 for both groups was loaded during immunoblotting analysis. (A) To reveal Cx43 association with ubiquitina- tion, and (B) Cx43 association with Nedd4. Relative density ratios were calculated using the control value as one. Each bar represents the mean (GS.E.M.) relative density (nZ3). Asterisk indicates a significant difference compared with the control group (P!0.05). IP, immunoprecipitation; C, control; FCT, FSHCTGFb1.

Article Snippet: Recombinant human TGFb1 was purchased from R&D Systems, Inc. (Minneapolis, MN, USA).

Techniques: Ubiquitin Proteomics, Control, Immunoprecipitation, Negative Control, Western Blot

Journal: Annals of Translational Medicine

Article Title: MMP2/9 downregulation is responsible for hepatic function recovery in cirrhotic rats following associating liver partition and portal vein ligation for staged hepatectomy

doi: 10.21037/atm-22-1312

Figure Lengend Snippet: Detection of liver hypertrophy in rats after the operation. ELISA assay showing the changes in expression of HGF (A), TGF-β (B), TNF-α (C), and IL-6 (D) in rats. (E) The PCNA levels in rat livers were assessed by immunohistochemistry (×10 magnification, ×10: 1 cm: 40 µm). (F) Hematoxylin and eosin staining of rat liver tissues (×10 magnification, ×10: 1 cm: 40 µm). All quantitative variables are presented as mean ± standard deviation and compared using two-way ANOVA. P<0.05. #, A or P groups vs. S groups, P<0.05, marked as #NA, #NP, #CP, #CA; Δ, CA group vs. NA group, P<0.05; *, CP groups vs. CA groups. ELISA, enzyme-linked immunosorbent assay; HGF, hepatocyte growth factor; TGF, transforming growth factor; TNF, tumor necrosis factor; IL, interleukin; PCNA, proliferating cell nuclear antigen; ANOVA, analysis of variance; PVL, portal vein ligation; ALPPS, associating liver partition and portal vein ligation for staged hepatectomy; A, ALPPS; P, PVL; S, sham operated; CA, cirrhotic rats that underwent ALPPS; NA, normal rats that underwent ALPPS; CP, cirrhotic rats that underwent PVL.

Article Snippet: The serum levels of recombinant human hepatocyte growth factor (HGF), transforming growth factor-β (TGF-β), tumor necrosis factor (TNF)-α, and interleukin (IL)-6 in rats were detected by ELISA kits (Elabscience Biotechnology Company, Co., Ltd., China).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Immunohistochemistry, Staining, Standard Deviation, Ligation

Journal: REPRODUCTION

Article Title: Transforming growth factor-β1 inhibits luteinization and promotes apoptosis in bovine granulosa cells

doi: 10.1530/rep-08-0365

Figure Lengend Snippet: Figure 6 Western blot analysis of cleaved caspase-3 in granulosa cells treated with TGFB1 in the presence or absence of FSH. The 17 and 19 kDa cleaved caspase-3 bands were normalized to the housekeeping protein b-actin. Asterisk (*) indicate that cleaved caspase-3 is significantly different from 0 dose treatment of TGFB1. Data are meansGS.E.M. of three separate culture replicates. (P!0.05, one-way ANOVA, with Dunnett’s test).

Article Snippet: Cells were seeded into 24-well tissue culture plates (Corning Glass Works, Corning, NY, USA) at a density of 106 viable cells (tested by Trypan blue exclusion) in 1 ml of a-MEM with L-glutamine containing sodium bicarbonate (10 mM), Hepes (20 mM), non-essential amino acid mix (1.1 mM), penicillin (100 IU/ml) and streptomycin (100 mg/ml), protease-free BSA (0.1%), sodium selenite (4 ng/ml), transferrin (2.5 mg/ml), A4 (100 nM; all from Sigma), insulin (10 ng/ml; Invitrogen) and graded doses of recombinant active human TGFB1 (R&D Systems, Minneapolis, MN, USA), with or without ovine FSH (1 ng/ml; oFSH, AFP-5332B; NIDDKD, Torrance, CA, USA) starting on the first day of culture.

Techniques: Western Blot

Journal: The Journal of biological chemistry

Article Title: Analysis of small latent transforming growth factor-beta complex formation and dissociation by surface plasmon resonance. Absence of direct interaction with thrombospondins.

doi: 10.1074/jbc.272.26.16329

Figure Lengend Snippet: FIG. 1. LAP forms a complex with immobilized TGFb1. TGFb1 was immobilized (around 600 RU), and LAP was then injected to form small latent TGFb1 complex. A, sensorgrams (relative response in RU after background subtraction versus time in sec) of LAP binding (bottom to top curve: 50, 125, 150, 175, and 200 nM of LAP). B, plots of ks versus LAP concentration.

Article Snippet: Ultrapure human TGFb1 was from Genzyme (Cambridge, MA), recombinant human LAP (TGFb1), and recombinant latent TGFb1 were from R&D Systems (Abingdon, UK).

Techniques: Injection, Binding Assay, Concentration Assay

Journal: The Journal of biological chemistry

Article Title: Analysis of small latent transforming growth factor-beta complex formation and dissociation by surface plasmon resonance. Absence of direct interaction with thrombospondins.

doi: 10.1074/jbc.272.26.16329

Figure Lengend Snippet: FIG. 2. TGFb1 forms a complex with immobilized LAP. LAP was immobilized (around 800 RU) and then TGFb1 was injected to form small latent TGFb1 complex. Sensorgrams (relative response in RU after background subtraction versus time in sec) of TGFb1 binding (bottom to top curve: 50, 80, and 100 nM of TGFb1).

Article Snippet: Ultrapure human TGFb1 was from Genzyme (Cambridge, MA), recombinant human LAP (TGFb1), and recombinant latent TGFb1 were from R&D Systems (Abingdon, UK).

Techniques: Injection, Binding Assay

Journal: The Journal of biological chemistry

Article Title: Analysis of small latent transforming growth factor-beta complex formation and dissociation by surface plasmon resonance. Absence of direct interaction with thrombospondins.

doi: 10.1074/jbc.272.26.16329

Figure Lengend Snippet: FIG. 3. Activation of latent TGFb1 complex. TGFb1 was immo- bilized (around 600 RU), and then LAP (50 nM) was injected to form small latent TGFb1 complex. A, over this complex, 25 ml of 6 M guani- dine chloride (GnCl) was injected at a flow of 5 ml/min, followed by another injection of LAP (50 nM) and then a second injection of 6 M GnCl. B, over this complex, bottom to top curve: 50 ml of HBS (CTL), pH 8.6; *, plasmin (0.1 units/ml in HBS, pH 8.6); or **, heat-inactivated plasmin (0.1 units/ml in HBS, pH 8.6) was injected at a flow of 2 ml/min.

Article Snippet: Ultrapure human TGFb1 was from Genzyme (Cambridge, MA), recombinant human LAP (TGFb1), and recombinant latent TGFb1 were from R&D Systems (Abingdon, UK).

Techniques: Activation Assay, Injection

Journal: The Journal of biological chemistry

Article Title: Analysis of small latent transforming growth factor-beta complex formation and dissociation by surface plasmon resonance. Absence of direct interaction with thrombospondins.

doi: 10.1074/jbc.272.26.16329

Figure Lengend Snippet: FIG. 4. Absence of association of TSP1 and TSP2 with LAP, TGFb1, or small latent TGFb1 forms. A, TSP1 (50 nM) was injected over immobilized LAP, TGFb1, and a control (CTL) immobilized flow cell. B, TSP2 (50 nM) was injected over immobilized LAP, TGFb1, and a control immobilized flow cell. C, LAP was immobilized and then TGFb1 (50 nM) was injected to form small latent TGFb1 complex, and over this complex, TSP2 (50 nM) or TSP1 (50 nM) was injected. D, vice versa of C. TGFb1 was immobilized and then LAP (50 nM) was injected to form small latent TGFb1 complex, and over this complex, TSP1 (50 nM) or TSP2 (50 nM) was injected.

Article Snippet: Ultrapure human TGFb1 was from Genzyme (Cambridge, MA), recombinant human LAP (TGFb1), and recombinant latent TGFb1 were from R&D Systems (Abingdon, UK).

Techniques: Injection, Control