human cancer cell lines ht1080  (ATCC)

Journal: Oncogene

Article Title: LRP6 expression promotes cancer cell proliferation and tumorigenesis by altering beta-catenin subcellular distribution.

doi: 10.1038/sj.onc.1208123

Figure Lengend Snippet: Figure 2 Western blot and flow cytometric analyses of LRP6- transduced HT1080 cells. (a) Western blot analysis of LRP6- transduced HT1080 cells and control cells. Cell lysates were analysed via 6% SDS–PAGE under reducing conditions and Western blotted with anti-LRP5/6 or anti-HA antibodies as indicated. (b) Flow cytometry analysis of LRP6-transduced HT1080 cells and control cells. The negative controls without the primary antibody are indicated with light lines, whereas the signals from receptor staining are drawn with dark lines

Article Snippet: RT–PCR Human cancer cell lines HT1080, HCT116, DLD-1, MDAMB-231, MDA-MB-468, H441, and H520 were obtained from the American Type Culture Collection (Manassas, VA, USA), and cultured in DMEM with 10% fetal calf serum.

Techniques: Western Blot, Control, SDS Page, Flow Cytometry, Staining

Journal: Oncogene

Article Title: LRP6 expression promotes cancer cell proliferation and tumorigenesis by altering beta-catenin subcellular distribution.

doi: 10.1038/sj.onc.1208123

Figure Lengend Snippet: Figure 3 LRP6-transduced cells display increased levels of cytosolic b-catenin, and TCF/LEF transcriptional activity. (a) LRP6-transduced HT1080 cells and control cells were fractionated into membrane and cytosolic fractions, and the b-catenin levels in these fractions and the whole-cell extract were examined by Western blotting using b-catenin antibody. Loading in each lane was normalized to protein content. (b) LRP6-transduced HT1080 cells and control cells were labeled with anti-b-catenin antibody, followed by detection with Alexafluor488 goat anti-mouse IgG and confocal microscopy. (c) LRP6-transduced HT1080 cells and control cells were co-transfected with a TCF/LEF transcriptional activity reporter plasmid (TOP-FLASH), and the luciferase activity was measured 48 h after transfection. This experiment is a representative of two such experiments performed with similar data. *Po0.01 compared to pLNCX2 control cells

Article Snippet: RT–PCR Human cancer cell lines HT1080, HCT116, DLD-1, MDAMB-231, MDA-MB-468, H441, and H520 were obtained from the American Type Culture Collection (Manassas, VA, USA), and cultured in DMEM with 10% fetal calf serum.

Techniques: Activity Assay, Control, Membrane, Western Blot, Labeling, Confocal Microscopy, Transfection, Plasmid Preparation, Luciferase

Journal: Oncogene

Article Title: LRP6 expression promotes cancer cell proliferation and tumorigenesis by altering beta-catenin subcellular distribution.

doi: 10.1038/sj.onc.1208123

Figure Lengend Snippet: Figure 4 LRP6-transduced HT1080 cells display greater Wnt1 or b-catenin-induced TCF/LEF transcriptional activity than control cells. (a) LRP6-transduced HT1080 cells and control cells were transiently transfected with Wnt1, b-catenin, or empty pcDNA3 vector. The cells were fractionated into membrane and cytosolic fractions 48 h after transfection. b-Catenin levels in these fractions and the whole-cell extract, as well as Wnt1 levels in whole-cell extract, were examined by Western blotting using b-catenin antibody or Wnt1 antibody. (b) LRP6-transduced HT1080 cells and control cells were co-transfected with Wnt1, b-catenin, or empty pcDNA3 vector, and a TCF/LEF transcriptional activity reporter plasmid (TOP-FLASH). The luciferase activity was measured 48 h after transfection. This experiment is a representa- tive of three such experiments performed with similar data. *Po0.01 compared to pcDNA3 control

Article Snippet: RT–PCR Human cancer cell lines HT1080, HCT116, DLD-1, MDAMB-231, MDA-MB-468, H441, and H520 were obtained from the American Type Culture Collection (Manassas, VA, USA), and cultured in DMEM with 10% fetal calf serum.

Techniques: Activity Assay, Control, Transfection, Plasmid Preparation, Membrane, Western Blot, Luciferase

Journal: Oncogene

Article Title: LRP6 expression promotes cancer cell proliferation and tumorigenesis by altering beta-catenin subcellular distribution.

doi: 10.1038/sj.onc.1208123

Figure Lengend Snippet: Figure 5 Cell proliferation in culture and colony formation on soft agar by LRP6-transduced HT1080 cells. (a) Effect of LRP6 expression on HT1080 cell proliferation in culture. LRP6- transduced HT1080 cells and control cells were plated in six-well plates, and counted with trypan blue at days 3, 5, and 7. The mean cell numbers were plotted with s.d. given as error bars. (b) Effect of LRP6 expression on HT1080 cell colony formation on soft agar. LRP6-transduced HT1080 cells and control cells were suspended in 0.3% agar with complete DMEM and then layered on top of 0.5% bottom agar in six-well plates. Colonies were counted after 3 weeks. This experiment is a representative of three such experiments performed with similar data. *Po0.01 compared to pLNCX2 control cells

Article Snippet: RT–PCR Human cancer cell lines HT1080, HCT116, DLD-1, MDAMB-231, MDA-MB-468, H441, and H520 were obtained from the American Type Culture Collection (Manassas, VA, USA), and cultured in DMEM with 10% fetal calf serum.

Techniques: Expressing, Control

Journal: Oncogene

Article Title: LRP6 expression promotes cancer cell proliferation and tumorigenesis by altering beta-catenin subcellular distribution.

doi: 10.1038/sj.onc.1208123

Figure Lengend Snippet: Figure 6 Subcutaneous tumor growth after s.c. implantation of LRP6-transduced HT1080 cells and control cells into mice. For each implantation, cells were injected subcutaneously into the back of each 4-week-old, female athymic mouse. Nine mice were used in each group. Tumor size was measured at the indicated times. Each point represents the average volume of the nine tumors with s.d. given as error bars

Article Snippet: RT–PCR Human cancer cell lines HT1080, HCT116, DLD-1, MDAMB-231, MDA-MB-468, H441, and H520 were obtained from the American Type Culture Collection (Manassas, VA, USA), and cultured in DMEM with 10% fetal calf serum.

Techniques: Control, Injection