Journal: Oncotarget

Article Title: Stromal cells promote anti-estrogen resistance of breast cancer cells through an insulin-like growth factor binding protein 5 (IGFBP5)/B-cell leukemia/lymphoma 3 (Bcl-3) axis

doi:

Figure Lengend Snippet: Effect of 100 nM fulvestrant on A. colony size of MCF-7 cells in the clonogenic assays, B. expression of ERα, Ki67, P-ERK1/2 and P-AKT levels in Western blot analysis, C. spheroid formation and D. RNA expression of mesenchymal markers (ACTA, VIM, FN1) and stem cell markers (CD44, PROCR, ABCG2, ALDH3A1). E, F. Effects of MSCs, MSC-CM and CAF-CM on MCF-7 cell growth in the presence and absence of fulvestrant. Cell growth was either determined by measuring the sizes of individual clones in the clonogenic assay (E) or by the ATP-based growth assay (F). G, H. Effect of CAF-CM on (G) spheroid size in the presence and absence of fulvestrant and (H) on the RNA levels of mesenchymal and stem cell markers in the absence and presence of fulvestrant. In (A, E), the data of a representative experiment are shown, in (D, F–H), each bar represents the mean value ± S.D. of at least three independent experiments. Statistical analysis for the clonogenic assay was performed by the Wilcoxon test (A, E). Other statistical analyses were done by using the student's t -test. ACTA = α-smooth muscle actin, VIM = vimentin, FN1 = fibronectin-1, ABCG2 = ATP binding cassette subfamily G2), ALDH3A1 (aldehyde dehydrogenase 3 family, member A1), NE/CE = nuclear/cytosolic protein extract, RLU = relative light units.

Article Snippet: Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.

Techniques: Expressing, Western Blot, RNA Expression, Clone Assay, Clonogenic Assay, Growth Assay, Binding Assay

Journal: Oncotarget

Article Title: Stromal cells promote anti-estrogen resistance of breast cancer cells through an insulin-like growth factor binding protein 5 (IGFBP5)/B-cell leukemia/lymphoma 3 (Bcl-3) axis

doi:

Figure Lengend Snippet: A, C. Effect of siIGFBP5 (A), siBcl3 (C) and siLuc (A, C) on the size of individual colonies of MCF-7 cells in the clonogenic assay in the presence or absence of fulvestrant and in the presence or absence of CAF-CM. B. Clonogenic assays performed with insulin- or mock-treated MCF-7 cells. Statistical analyses were carried out by using the Wilcoxon test.

Article Snippet: Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.

Techniques: Clonogenic Assay

Journal: Oncotarget

Article Title: Stromal cells promote anti-estrogen resistance of breast cancer cells through an insulin-like growth factor binding protein 5 (IGFBP5)/B-cell leukemia/lymphoma 3 (Bcl-3) axis

doi:

Figure Lengend Snippet: A–D. After BT474 cells (A, B) or T47D cells (D, E. ) were treated with CAF-CM or left untreated for three days, RNA or proteins were isolated and analyzed for IGFBP5 and Bcl-3 RNA levels (A, D) or levels of certain proteins as indicated (B, E), respectively. To check for equal loading of plasma membrane proteins, blots were reprobed with an E-cadherin -specific antibody (α-E-cadh.). F. T47D cells were also tested for AKT phosphorylation and CAIX expression after 3-day-treatment with insulin or CoCl 2 , respectively. (C, G, H. ) Effect of CAF-CM on BT474 and T47D cell growth in the presence or absence of fulvestrant as measured either by an ATP based assay (C, G) or by measuring the sizes of single colonies (H). In (A, C, D, G), each bar represents the mean value ± S.D. of at least three independent experiments, in (H), the data of a representative experiment are shown. Statistical analysis was either performed by using the student's t -test (A, C, D, G) or the Wilcoxon test (H).

Article Snippet: Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.

Techniques: Isolation, Clinical Proteomics, Membrane, Phospho-proteomics, Expressing, ATP Assay

Journal: Oncotarget

Article Title: Stromal cells promote anti-estrogen resistance of breast cancer cells through an insulin-like growth factor binding protein 5 (IGFBP5)/B-cell leukemia/lymphoma 3 (Bcl-3) axis

doi:

Figure Lengend Snippet: Factor(s) secreted by MSCs and CAFs lead to a decline in the IGFBP5 expression. The reduced IGFBP5 level results in the release of IGFs from the IGF/IGFBP5 complex allowing IGF to activate IGF1R, thereby activating the PI3K/AKT signaling pathway. Besides blocking IGF activity by this IGF-dependent function, IGFBP5 also shows IGF-independent actions (e.g. as an intracellular protein). Through this IGF-independent action IGFBP5 likely keeps Bcl-3 expression down. Consequently, when stromal cells downregulate IGFBP5 expression the Bcl-3 expression raises. This leads to fulvestrant resistance and to changes in the expression of the Bcl-3 target genes IGF1R, KLHL4 and SEPP1.

Article Snippet: Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.

Techniques: Expressing, Blocking Assay, Activity Assay

Journal: Cancer Cell International

Article Title: The cholesterol metabolite 27-hydroxycholesterol stimulates cell proliferation via ERβ in prostate cancer cells

doi: 10.1186/s12935-017-0422-x

Figure Lengend Snippet: 27-OHC stimulates cell proliferation via ER. Cell proliferation assay in LNCaP ( a ) and PC3 ( b ) cells demonstrates an attenuation of 27-OHC-induced cell proliferation with the ER inhibitor ICI 182,780 (fulvestrant). Cells were treated with 1 µM 27-OHC, 2 nM of E2 and 10 µM ICI 182,780. Readings were recorded 48 h after treatment with 27-OHC. Data is expressed as mean ± SEM. **p < 0.01; ***p < 0.001 versus controls, ### p < 0.001 versus 27-OHC only treatment

Article Snippet: 27-Hydroxycholesterol was purchased from Santa Cruz Biotechnologies (Dallas, TX), docetaxel, 4-[2-phenyl-5,7-bis(trifluoromethyl) pyrazolo[1,5-a]pyrimidin-3-yl]phenol (PHTPP) and fulvestrant from Cayman Chemicals (Ann Arbor, MI) and β-estradiol from Sigma-Aldrich (St. Louis, MO).

Techniques: Proliferation Assay