human estrogen receptor positive luminal breast cancer cell line mcf7  (ATCC)

Journal: Cell

Article Title: The bone microenvironment invigorates metastatic seeds for further dissemination

doi: 10.1016/j.cell.2021.03.011

Figure Lengend Snippet: (A) Diagram of intra-iliac artery (IIA) injection and representative bioluminescent images (BLI) showing the in vivo distribution of tumor cells after IIA injection of 1E5 MDA-MB-231 fLuc-mRFP cells. . (B-C) Representative ex vivo BLI images (B), and PET-μCT (C) on hindlimbs and other tissues of the same animal with MDA-MB-231 cells inoculated in the right hindlimb after 8 weeks. R.H, Right Hindlimb; Lu, Lung; L.H, Left Hindlimb; Li, Liver; Ki, Kidney; Sp, Spleen; Br, Brain; Ve, Vertebrae; F.L, Forelimbs; Ri, Ribs; St, Sternum; Cr, Cranium. (D-E) Representative immunofluorescent images of tumor lesions in various bones (D) and other organs (E). To obtain complete views of entire organs, smaller fields were acquired in tiles by mosaic scanning and then stitched by Zen. Scale bar, 20 μm. (F-H) Representative BLI images of animals and tissues after IIA injection of 2E5 prostate cancer cells PC3 (F), 1E5 ER+ breast cancer cells MCF7 (G), and 1E5 murine mammary carcinoma cells AT-3 (H) at the indicated time. (I) Diagram of intra-femoral injection (IF) (Left) and representative ex vivo BLI images of tissues from animals received 1E5 MDA-MB-231 cells (Middle) or AT-3 cells (Right) via IF injection. (J-K) Heat map of ex vivo BLI intensity and status of metastatic involvement on various types of tissues from animals carried MDA-MB-231 (J) and AT-3 (K) bone tumors. Columns, individual animal; rows, various tissues or status of multi-site metastases; Gray, no detectable lesion. N (# of mice): MDA-MB-231, 16 (IIA), 11 (IF); AT-3, 10 (IIA), 10 (IF). P values were assessed by Fisher’s exact test on the ratio of metastasis while by Mann-Whitney test on the tumor burden. See also Figure S1.

Article Snippet: Cell lines and Cell Culture Human triple negative breast cancer cell lines MDA-MB-231(RRID: CVCL_0062), human estrogen receptor positive luminal breast cancer cell line MCF7 (RRID: CVCL_0031), human prostate cancer cell line PC-3 (RRID: CVCL_0035), and HEK293T cells (RRID: CVCL_0063) were obtained from ATCC.

Techniques: Injection, In Vivo, Ex Vivo, IF-P, MANN-WHITNEY

Journal: Cell

Article Title: The bone microenvironment invigorates metastatic seeds for further dissemination

doi: 10.1016/j.cell.2021.03.011

Figure Lengend Snippet: (A) Diagram of intra-iliac vein (IIV) injection, and representative BLI images of animals and tissues 8 weeks after IIV injection of 1E5 MDA-MB-231 cells. (B) Diagram of mammary fat pad (MFP) implantation, and representative BLI images of animals and tissues 8 weeks after MFP implantation of 1E5 MDA-MB-231 cells. (C-D) Comparison of metastatic pattern and tumor burden (C) and the ratio of multi-site metastasis (D) in animals with bone (IIA/IF), lung (IIV) or mammary (MFP) tumors of MDA-MB-231 cells. N (# of mice) = 27 (Bone); 18 (MFP); 10 (Lung). (E-F) Comparison of metastatic pattern and tumor burden (E) and the ratio of multi-site metastasis (F) in animals with bone (IIA/IF), lung (IIV) or mammary (MFP) tumors of AT-3 cells. N (# of mice) = 20 (Bone); 11 (MFP); 9 (Lung). (G-H) Comparison of metastatic pattern and tumor burden (G) and the ratio of multi-site metastasis (H) in animals with bone (IIA), lung (IIV) or mammary (MFP or MIND) tumors of MCF7 cells. N (# of mice) = 8 (Bone); 10 (MFP); 13 (MIND); 9 (Lung). P values were assessed by Chi-square test in C-H on the ratio of metastasis; by uncorrected Dunn’s test following Kruskal-Wallis test in C, E and H on the tumor burden. See also Figure S2.

Article Snippet: Cell lines and Cell Culture Human triple negative breast cancer cell lines MDA-MB-231(RRID: CVCL_0062), human estrogen receptor positive luminal breast cancer cell line MCF7 (RRID: CVCL_0031), human prostate cancer cell line PC-3 (RRID: CVCL_0035), and HEK293T cells (RRID: CVCL_0063) were obtained from ATCC.

Techniques: Injection, Comparison

Journal: Cell

Article Title: The bone microenvironment invigorates metastatic seeds for further dissemination

doi: 10.1016/j.cell.2021.03.011

Figure Lengend Snippet: (A) Experimental design (left) and representative BLI images (right) to test the metastatic capacity of mammary, lung, or bone-entrained SCP21s. (B) Normalized whole-body BLI intensity 7 days after intra-cardiac (IC) injection of same number of MFP-, LuM-, BoM- or Par-SCP21 cells. (C) Colonization kinetics of MFP-, LuM-, BoM- and Par-SCP21 cells after IC injection. N (# of mice) = 8 (Par); 10 (MFP); 15 (BoM); 10 (LuM). (D) Percentage of ALDH+ population in MFP-, LuM-, BoM- and Par-SCP21 cells by flow cytometry. (E) Histogram (left) and median fluorescent intensity (MFI) (right) of surface CD44 protein in MFP-, LuM-, BoM- and Par-SCP21 cells by flow cytometry. (F) Expression levels of proteins in Par- and organ-entrained SCP21s. Protein levels were quantified and converted into Z-score from three or four western blottings. (G-I) Representative BLI images (G), normalized BLI intensity at day 7 (H), and the colonization kinetics (I) of MFP-, BoM-, and Par- MCF7-SCP2 cells after I.C. injection. N (# of mice) = 10 (Par); 8 (MFP); 10 (BoM). (J-K) Percentage of ALDH+ population (J) and expression of surface CD44 (K) in MFP-, BoM-, and Par- MCF7-SCP2 cells by flow cytometry. N (# of repeats) = 3 (J); 2 (K). (L-M) Representative fluorescent images (L) and quantification (M) of CD44 and ALDH1A1 expression on CTCs from NRG mice bearing MDA-MB-231 cells derived mammary or bone tumors. CTCs were pooled from 5 blood samples. Scale bar, 10 μm. (N) Expression levels of CD44 mRNA in CTCs from breast cancer patients with bone metastases or other metastases (GSE86978). Data are represented as mean ± SEM in B, C, D, E, H, I and J. P values were assessed by Fisher’s LSD test following one-way ANOVA test in B, D, E, H, and J; by Fisher’s LSD test post two-way ANOVA test in C and I; by student t-test in F; by Mann-Whitney test in M and N. See also Figure S6.

Article Snippet: Cell lines and Cell Culture Human triple negative breast cancer cell lines MDA-MB-231(RRID: CVCL_0062), human estrogen receptor positive luminal breast cancer cell line MCF7 (RRID: CVCL_0031), human prostate cancer cell line PC-3 (RRID: CVCL_0035), and HEK293T cells (RRID: CVCL_0063) were obtained from ATCC.

Techniques: Injection, Flow Cytometry, Expressing, Western Blot, Derivative Assay, MANN-WHITNEY

Journal: Cell

Article Title: The bone microenvironment invigorates metastatic seeds for further dissemination

doi: 10.1016/j.cell.2021.03.011

Figure Lengend Snippet: (A) Levels of EZH2 signature genes (GSVA) in bone entrained and other SCP21 cells. (B) Levels of EZH2 signature genes in bone entrained-SCP21 cells after different passages in vitro. (C) Percentage of ALDH1+ population in bone entrained-SCP21 cells at different passages. (D) Representative western blotting of proteins in bone entrained-SCP21 cells after different passages. (E-G) The schematic diagram and representative BLI images (E), normalized BLI intensity at day 7 (F), and the colonization kinetics (G) of BoM-SCP21 cells with in vitro EPZ011989 (EPZ) treatment before IC injection. Non-treated BoM-SCP21 cells were used as control. N (# of mice) = 15 (-EPZ); 9 (+EPZ). (H) Comparison of ALDH1+ cells in EPZ treated and non-treated BoM-MCF7-SCP2 cells by flow cytometry. N (# of replicate) =3. (I-K) Representative BLI images (I), normalized BLI intensity at day 7 (J), and the colonization kinetics (K) of BoM-MCF7-SCP2 cells with in vitro treatment of EPZ before IC injection. Non-treated BoM-MCF7-SCP2 cells were used as control. N (# of mice) = 10 (-EPZ); 7 (+EPZ). (L) Experimental design assessing the multi-site metastases from bone lesions with inducible depletion of EZH2. (M) Growth kinetics of the primary bone lesions in mice receiving doxycycline or control water, assessed by in vivo BLI imaging. BLI intensities at right hindlimbs were normalized to the mean intensity at day 0. N (# of mice) = 10 for each arm. (N) Heat map of ex vivo BLI intensity and status of metastatic involvement in tissues from animals with EZH2 depleted or control bone metastases. Data are represented as mean ± SEM in F, G, J, K, and M. P values were assessed by student t-test in A, F, and J; by test for linear trend following repeat measure one-way ANOVA in B and C; by LSD test following two-way ANOVA in G, K and M; by ratio paired t-test in H; by Fisher’s exact test on the ratio of metastatic involvement and Mann-Whitney test on BLI intensity in N. See also Figure S7.

Article Snippet: Cell lines and Cell Culture Human triple negative breast cancer cell lines MDA-MB-231(RRID: CVCL_0062), human estrogen receptor positive luminal breast cancer cell line MCF7 (RRID: CVCL_0031), human prostate cancer cell line PC-3 (RRID: CVCL_0035), and HEK293T cells (RRID: CVCL_0063) were obtained from ATCC.

Techniques: In Vitro, Western Blot, Injection, Control, Comparison, Flow Cytometry, In Vivo, Imaging, Ex Vivo, MANN-WHITNEY

Journal: Cell

Article Title: The bone microenvironment invigorates metastatic seeds for further dissemination

doi: 10.1016/j.cell.2021.03.011

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Cell lines and Cell Culture Human triple negative breast cancer cell lines MDA-MB-231(RRID: CVCL_0062), human estrogen receptor positive luminal breast cancer cell line MCF7 (RRID: CVCL_0031), human prostate cancer cell line PC-3 (RRID: CVCL_0035), and HEK293T cells (RRID: CVCL_0063) were obtained from ATCC.

Techniques: In Vivo, Western Blot, Staining, Recombinant, Multiplex Assay, Sequencing, RNA Sequencing, Derivative Assay, Cloning, shRNA, Software