cell culture bt474  (ATCC)

Journal: NPJ breast cancer

Article Title: LINC00589-dominated ceRNA networks regulate multiple chemoresistance and cancer stem cell-like properties in HER2 + breast cancer.

doi: 10.1038/s41523-022-00484-0

Figure Lengend Snippet: Fig. 2 LINC00589 promotes the sensitivity of breast cancer cells to trastuzumab and inhibits anchorage-independent growth. A–C TR SKBR3 cells were infected with NC or LINC00589-overexpression lentivirus and treated with 25 μg/ml trastuzumab. WT SKBR3 and BT474 cells were infected with sh-NC or sh-LINC00589 lentivirus and treated with 5 μg/ml trastuzumab treatment. The relative cell viabilities were determined by CCK-8 assays at the indicated times. D, E TR and WT SKBR3 cells were infected with overexpression or shRNA lentiviruses and then were treated with 25 μg/ml trastuzumab or 5 μg/ml trastuzumab for 48 h, followed by FITC-conjugated annexin V and PE-labeled PI staining. The apoptosis rate was analyzed by flow cytometry. F, G TR or WT SKBR3 cells were cultured in soft agar for 21 days after trastuzumab treatment and then subjected to crystal violet staining. The clones were observed, and the number of clones for each sample was calculated. Scale bar 100 μm. Data are represented as the mean ± SD of three replicates or are representative of three independent experiments. Two-way ANOVA were used to analyze the data in (A–C), and two-tailed t test was used to analyze the data in (E, G). *P < 0.05, **P < 0.01, and ***P < 0.001 versus negative control (NC).

Article Snippet: Cell lines and cell culture BT474 human breast cancer cells (HER2-overexpression) were obtained from the American Type Culture Collection (catalog number HTB-20, ATCC) and were cultured in RPMI 1640 supplemented with 10% FBS.

Techniques: Infection, Over Expression, CCK-8 Assay, shRNA, Labeling, Staining, Cytometry, Cell Culture, Clone Assay, Two Tailed Test, Negative Control

Journal: NPJ breast cancer

Article Title: LINC00589-dominated ceRNA networks regulate multiple chemoresistance and cancer stem cell-like properties in HER2 + breast cancer.

doi: 10.1038/s41523-022-00484-0

Figure Lengend Snippet: Fig. 7 LINC00589 represses trastuzumab resistance, cancer stem cell-like properties, and multiple chemoresistance via DLG5 and PRDM16. A–E WT SKBR3 and BT474 cells infected with sh-LINC00589 or sh-NC lentivirus were transfected with pCDNA (pC) -DLG5 or pC- PRDM16 for 48 h; or TR SKBR3 cells infected with Lv-LINC00589 or the Lv-NC were transfected with sh-DLG5 or sh-PRDM16 for 48 h. The relative cell viabilities of WT SKBR3 cells (A), BT474 cells (B) and TR SKBR3 cells (C) were quantified by CCK-8 assay. Apoptosis of TR cells were examined by flow cytometry assay (D) and the apoptosis rate was calculated (E). F–M LINC00589- or NC- overexpressing TR cells were infected with sh- NC, sh-DLG5 or sh-PRDM16 lentivirus. Cells were cultured in soft agar for 21 days, representative images of colony formation were observed (F), and the number of colonies were calculated (G). Scale bar 100 μm. Cells were seeded in an ultra-low-attachment culture system. Represent images of mammosphere formation were observed (H), and numbers (I) and volumes (J) of mammospheres were calculated. Scale bar 100 μm. Data are shown as the mean ± SD of five random high-power fields (HPF) and were analyzed by two-tailed t test. *P < 0.05, and **P < 0.01. mRNA and protein expression of molecular markers of breast cancer CSCs were determined by qRT-PCR assay (K) and western blotting (L). GAPDH and β-actin were used as internal controls. M Cells were exposed to 5-FU, Dox, Pac, Cis, Gem, or VCR for 48 h, and the cell viability was evaluated by CCK-8 assay. Data are shown as mean ± SD; two-tailed t test was used to analyze the data in (A, B, C, E, K, and M). *P < 0.05, **P < 0.01, and ***P < 0.001 versus negative control (NC).

Article Snippet: Cell lines and cell culture BT474 human breast cancer cells (HER2-overexpression) were obtained from the American Type Culture Collection (catalog number HTB-20, ATCC) and were cultured in RPMI 1640 supplemented with 10% FBS.

Techniques: Infection, Transfection, CCK-8 Assay, Cytometry, Cell Culture, Two Tailed Test, Expressing, Quantitative RT-PCR, Western Blot, Negative Control