Journal: Frontiers in Endocrinology

Article Title: Regulatory Actions of LH and Follicle-Stimulating Hormone on Breast Cancer Cells and Mammary Tumors in Rats

doi: 10.3389/fendo.2018.00239

Figure Lengend Snippet: Modulatory effects of gonadotropins on breast cancer (BC) cell expression/activation of follicular-stimulating hormone receptor (FSHR), LHR, moesin, and focal adhesion kinase (FAK). Protein extracts from (A) T-47D (estrogen-receptor positive) and (E) MDAMB-468 (estrogen-receptor negative) BC cells treated for 48 h with increasing concentration of LH and follicle-stimulating hormone (FSH) (5, 5 + 50, and 50 mUI/ml) were assayed using western blot analysis for their overall content of LHR, FSHR, moesin, p-Moesin T558 , FAK, p-FAKY 397 , and actin. (B–D,F–H ) LHR, FSHR, moesin, p-Moesin T558 , FAK, p-FAKY 397 densitometry values were adjusted to actin intensity, respectively, then normalized to the control sample. * P < 0.05 vs. corresponding control. The experiments were performed in triplicates and representative images are shown.

Article Snippet: Antibodies used were: FAK (dilution 1:1,000), p-FAK (Y397, 1:1,000), and moesin (clone 38, 1:1,000) (BD Transduction Laboratories, Lexington, KY, USA); Thr558-p-Moesin (sc-12895, 1:750), p-FAK-Tyr397 (sc-11765, 1:500), LHR (H-50, 1:500), FSHR (N-20, 1:500), and ACTIN (C-11, 1:1,000) (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Expressing, Activation Assay, Concentration Assay, Western Blot

Journal: Frontiers in Endocrinology

Article Title: Regulatory Actions of LH and Follicle-Stimulating Hormone on Breast Cancer Cells and Mammary Tumors in Rats

doi: 10.3389/fendo.2018.00239

Figure Lengend Snippet: LH/follicle-stimulating hormone (FSH) signal to moesin and focal adhesion kinase (FAK) via LHR/follicle-stimulating hormone receptor (FSHR). (A–C) T-47D breast cancer cells were transfected with shRNA vs. LHR and FSHR (shRNA LHR and shRNA FSHR) or with vehicle, and protein analysis for LHR, FSHR, actin, wild-type (FAK and moesin), or p-FAK and p-moesin was performed on cell lysates after treatment with LH and FSH in different concentrations (5, 5 + 50, and 50 mUI/ml) for 48 h. (B,D) The graphs display the quantitative analysis of the intensity of the bands, obtained as number of photons measured by the ChemiDoc digital imaging system and evaluated with the Quantity One Software (Bio-Rad).

Article Snippet: Antibodies used were: FAK (dilution 1:1,000), p-FAK (Y397, 1:1,000), and moesin (clone 38, 1:1,000) (BD Transduction Laboratories, Lexington, KY, USA); Thr558-p-Moesin (sc-12895, 1:750), p-FAK-Tyr397 (sc-11765, 1:500), LHR (H-50, 1:500), FSHR (N-20, 1:500), and ACTIN (C-11, 1:1,000) (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Transfection, shRNA, Imaging, Software

Journal: Frontiers in Endocrinology

Article Title: Regulatory Actions of LH and Follicle-Stimulating Hormone on Breast Cancer Cells and Mammary Tumors in Rats

doi: 10.3389/fendo.2018.00239

Figure Lengend Snippet: Gonadotrophins’ signaling to moesin and focal adhesion kinase (FAK) increase T-47D cell migration and invasion. Breast cancer cells were transfected with 100 nM target shRNA for LHR and follicular-stimulating hormone receptor (FSHR), siRNA for FAK and 75 nM target phosphorothioate oligonucleotides antisense vs. moesin for 48 h and then treated with LH and/or follicle-stimulating hormone (FSH) (5 + 50 mUI/ml) for 48 h. (A,B) Cell migration distances were measured as mean migration distance (μm) ± SD. * P ≤ 0.05 vs. control. The experiments were performed in triplicates and representative images are shown. (C–H) Cell invasion was assayed using invasion chambers. Average of invading cells observed, photographed under the microscope at 100× magnification and counted in three different central fields of triplicate membranes. * P ≤ 0.05 vs. control. The experiments were performed in triplicates. Invasion indexes and representative images are shown.

Article Snippet: Antibodies used were: FAK (dilution 1:1,000), p-FAK (Y397, 1:1,000), and moesin (clone 38, 1:1,000) (BD Transduction Laboratories, Lexington, KY, USA); Thr558-p-Moesin (sc-12895, 1:750), p-FAK-Tyr397 (sc-11765, 1:500), LHR (H-50, 1:500), FSHR (N-20, 1:500), and ACTIN (C-11, 1:1,000) (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Migration, Transfection, shRNA, Microscopy

Journal: Breast Cancer Research : BCR

Article Title: LIN7A is a major determinant of cell-polarity defects in breast carcinomas

doi: 10.1186/s13058-016-0680-x

Figure Lengend Snippet: Polarity abnormalities in invasive micropapillary carcinoma (IMPC). a Representative views of cell-division control protein 42 (CDC42), cis-Golgi marker (GM130) and occludin (OCLN) immunostainings in three normal ducts ( left panel ) and in three IMPC ( right panel ). Scale bars 10 μm, L lumen. G × 400 magnification. b Analysis of polarity protein expression and subcellular localization in 24 IMPC. Expression and cellular localization (sub-apical, apical, cytoplasmic, membranous, cell/cell: lateral membrane staining between two cells and basolateral) of polarity proteins was compared to that in normal cells. p-ERM phospho-ezrin-radixin-moesin, p-aPKCζ phospho-atypical PKC, PALS1 protein associated with Lin seven 1, SCRIB Scribble, ZO-1 zonula occludens 1

Article Snippet: Antibodies used were directed against phospho-ezrin-radixin-moesin (p-ERM,1:200 dilution, pH6.1, BD Biosciences), GM130 (clone 35, 1:100 dilution, pH9, BD Biosciences), cell division control protein 42 (CDC42, 1:300 dilution, pH6.1, Lifespan Biosciences, Seattle, WA, USA), phospho-atypical PKCζ (p-aPKCζ, clone190D10, 1:250 dilution, pH6.1, Cell Signaling), protein associated with Lin seven 1 (PALS1, 1:50 dilution, pH9, Lifespan Biosciences), occludin (OCLN, 1:200 dilution, pH6.1, Lifespan Biosciences), Scribble (SCRIB, 1:50 dilution, pH6.1, Santa Cruz Biotechnology, Dallas, TX, USA), zonula occludens 1 (ZO-1, 1:500 dilution, pH6.1, BD Biosciences), β-catenin (clone 14/Beta-catenin, 1:200 dilution, pH6.1, BD Biosciences), E-cadherin (clone 4A2C7, 1:100 dilution, pH9, Invitrogen, Camarillo, CA, USA), KI-67 (clone MIB1, 1:100 dilution, pH6.1, Dako A/S) or active-caspase-3 (1:250 dilution, pH6.1, Cell Signaling).

Techniques: Marker, Expressing, Staining