human bladder cancer cell lines 5637  (ATCC)

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Long noncoding RNA MALAT1 is dynamically regulated in leader cells during collective cancer invasion.

doi: 10.1073/pnas.2305410120

Figure Lengend Snippet: Fig. 1. A dual double-stranded LNA nanobiosensor for probing lncRNA dynamics during collective cancer invasion. (A), Schematics of the FRET-based LNA nanobiosensor for detecting lncRNA in leader and follower cells. (B), Intracellular distributions of β-actin, MALAT1, and UCA1 RNA expressions detected by the nanobiosensors in live cancer cells (5,637). (Scale bars, 10 µm.) Images are representative of eight experiments. (C), Time-lapse images for MALAT1 dynamics in leader cells (marked by white arrows) during collective cell migration. (Scale bars, 30 µm.) Images are representative of five experiments. (D), Detection of MALAT1 expression in 3D tumor spheroids derived from patients with muscle-invasive bladder cancer. Blue dashed squares indicate the zoom-in regions (Right). Black arrows indicate protruding structures with leader cells sprouting from the spheroids. [Scale bars, 200 µm (Left) and 40 µm (Right).] Images are representative of six (Stage II) and ten (Stage IIIB) tumor spheroids. (E), 3D rendering of a cancer spheroid with leader cells (white arrows) and dissociated cancer cells (white arrowheads). (Scale bar, 200 µm.)

Article Snippet: The human bladder cancer cell line 5,637 from American Type Culture Collection (Manassas, VA) was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 10 mg/mL Gentamicin (Fisher Scientific, Hampton, NH).

Techniques: Migration, Expressing, Derivative Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Long noncoding RNA MALAT1 is dynamically regulated in leader cells during collective cancer invasion.

doi: 10.1073/pnas.2305410120

Figure Lengend Snippet: Fig. 2. LNA nanobiosensors detect TGF-β-induced MALAT1 upregulation in cell nuclei and cytoplasm. (A–C), Confocal images of β-actin mRNA, MALAT1, and UCA1 in live bladder cancer cells (5,637). The cells were treated with buffer control and TGF-β. FRET channels (Left), merged FRET and brightfield channels (Middle), and zoom-in views of single cells (Right) are shown to illustrate the expression distributions. Images are representative of five experiments. [Scale bars, 50 μm (Left and Middle), 10 μm (Right).] (D and E), Relative expression of β-actin, MALAT1, and UCA1 RNA (D) in whole cells and (E) colocalized with the nuclei. One-way ANOVA followed by Tukey’s post hoc test was used to compare transcript numbers between control and TGF-β treatment (n = 5, NS not significant, *P < 0.05, ****P < 0.0001).

Article Snippet: The human bladder cancer cell line 5,637 from American Type Culture Collection (Manassas, VA) was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 10 mg/mL Gentamicin (Fisher Scientific, Hampton, NH).

Techniques: Control, Expressing

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Long noncoding RNA MALAT1 is dynamically regulated in leader cells during collective cancer invasion.

doi: 10.1073/pnas.2305410120

Figure Lengend Snippet: Fig. 5. MALAT1 expression is associated with the clinical stage of bladder cancer patient-derived cancer cells. (A), β-actin mRNA and MALAT1 expressions in cancer cells derived from bladder cancer patients. (Scale bars, 300 µm.) (B and C), MALAT1 is up-regulated in the stage IIIB sample compared to the stage II sample. (D), The portion of cells with a high level of MALAT1. Student’s t test was used to compare between samples (n ≥ 1,148 for gene expression and n = 5 for cell portions with an enhanced MALAT1 expression).

Article Snippet: The human bladder cancer cell line 5,637 from American Type Culture Collection (Manassas, VA) was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 10 mg/mL Gentamicin (Fisher Scientific, Hampton, NH).

Techniques: Expressing, Derivative Assay, Gene Expression

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Long noncoding RNA MALAT1 is dynamically regulated in leader cells during collective cancer invasion.

doi: 10.1073/pnas.2305410120

Figure Lengend Snippet: Fig. 6. MALAT1 is associated with leader cells in tumor spheroids derived from muscle-invasive bladder cancer patients. (A–G), MALAT1 expression in 3D human tumor spheroids derived from bladder cancer patients. Clinical samples of stage II (DT2101 and DT2334), Stage IIIA (DT2092 and DT2148), Stage III (DT2153 and DT2115), and Stage IV (DT2296) were included to cover the spectrum of muscle-invasive bladder cancer. Blue dashed squares highlight the zoom-in views (Bottom) with leader cells (black arrows) or dissociated cells (black arrowheads). [Scale bars, 200 µm (Top) and 100 µm (Bottom).] Images are representative of at least six spheroids. (H and I), The number of detached cells and protrusions per spheroid on day 3 (n = 5). (J–L), Correlation of the spheroid volume, the number of detached cells, and the number of leader cells with MALAT1 expression on day 3. At least five spheroids were analyzed for each sample, and all detectable leader cells and detached cells were analyzed.

Article Snippet: The human bladder cancer cell line 5,637 from American Type Culture Collection (Manassas, VA) was cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 10 mg/mL Gentamicin (Fisher Scientific, Hampton, NH).

Techniques: Derivative Assay, Expressing

Journal: Cancer research

Article Title: A urinary drug-disposing approach as an alternative to intravesical chemotherapy for treating non-muscle invasive bladder cancer

doi: 10.1158/0008-5472.CAN-21-2897

Figure Lengend Snippet: A, Bdd does not trigger any innate immune response. No increase in the inflammatory cytokines concentrations was detected in the plasma of female BALB/cJ mice (n=3/group) 4 h after i.v. administration of the Bdd peptide (5 mg/kg, 150 μL). LPS was used as a positive control and concentrations of each cytokine were measured by ELISA kit. B, Cellular uptake of Cyanine5.5-labeled Bdd (Cy-Bdd). Representative fluorescence microscopic images of human UMUC-3 BC cells and murine Renca renal adenocarcinoma cells incubated for 6 and 24 h with Cy-Bdd (0.5 nmol). Dapi (9 μM) and LysoTracker-GFP (1 μM) were used for nuclear (blue) and organelle (green) staining, respectively, and were added to the cells 30 min prior to imaging. Scale bar is 25 μm. C, Comparing the potency of different chemotherapeutics (DM1, GEM, MIT, CIS, and DOX). UMUC-3 and Renca cells were incubated with the drugs at various concentrations for 72 h prior to measuring the cell viability. The dose response curves were plotted and the half maximal inhibitory concentrations (IC50 values) of each drug calculated using Graph Pad Prism 6.0 software. D, Conjugation of DM1 to Bdd. The cleavable linker SPDP was first conjugated to the peptide N-terminal in solid phase. DM1 was then added to the cleaved peptide in a solution mixture of PBS and NMP. E, Plot showing the percentage of accumulated DM1 released from the DM1-Bdd over time in PBS in the absence and presence of GSH (1 mM). The amount of drug released was quantified using rp-HPLC analysis (absorbance detected at 254 nm). F, Conjugation of aldox to Bdd. The peptide, supplemented with a N-terminal cysteine, was incubated with aldox in PBS (pH = 7.4) for 30 min prior to purification by rp-HPLC in neutral conditions. G, Plots showing the percentage of the accumulated DOX active metabolite released from aldox-Bdd (100 μM) over time in PBS buffers with different pH values. The amount of drug released was quantified using rp-HPLC analysis (absorbance detected at 480 nm). H, DM1-Bdd displays a similar cytotoxicity compared to free drug against murine bladder (MB49), human bladder (UMUC-3 and T24), and murine kidney (Renca) cancer cell lines. Plots of relative cell viability against the drug concentration. I, Aldox-Bdd is more potent than free aldox. Plots of the relative cell viability against the drug concentration.

Article Snippet: UMUC-3 and T24 human urinary bladder cancer cell lines (Cat# CRL-1749 and Cat# HTB-4), and Renca murine kidney adenocarcinoma cell line (Cat# CRL-2947), were obtained from ATCC (Manassas, VA).

Techniques: Clinical Proteomics, Positive Control, Enzyme-linked Immunosorbent Assay, Labeling, Fluorescence, Incubation, Staining, Imaging, Software, Conjugation Assay, Purification, Concentration Assay