Journal: Cancer research

Article Title: Development of novel aptamer-based targeted chemotherapy for bladder cancer

doi: 10.1158/0008-5472.CAN-21-2691

Figure Lengend Snippet: ( A ) Selection scheme of chemically modified RNA aptamers that can internalize into T24 human bladder cancer cells but not SV-huc-1 normal bladder urothelial cells. R: purine. Y: pyrimidine. ( B ) Predicted secondary structure of aptamer B1 using sfold. ( C ) Binding curve of aptamer B1 with T24 cells. An all-DNA version of B1 and a B1-derived scrambled 35-nt sequence were used as control. Representative data of three independent experiments are shown. ( D ) Confocal microscopy analysis showed that aptamer B1 internalization is cell type-specific. T24 and KU-7 are two bladder cancer cell lines, SV-huc-1 is a normal bladder urothelial cell line. Scale bars: 40 μm. Representative data of three independent experiments are shown. (E) Comparison of the binding curves of aptamer B1 with primary human tumor cells and normal urothelial cells generated from surgical specimens harvested from a NMIBC patient. Freshly collected bladder tumor tissue or normal para-tumor urothelium tissue was digested into single cells, incubated with Cy5.5-labeled aptamer B1 for 1 h and analyzed with flow cytometry. Raw data are shown in Supplementary Fig. 3B .

Article Snippet: Human bladder cancer cell lines T24 (CLS Cat# 300352/p619_T24, RRID: CVCL_0554) ( 27 ), TCCSUP (ATCC Cat# HTB-5, RRID: CVCL_1738), J82 (KCLB Cat# 30001, RRID: CVCL_0359), SW780 (ATCC Cat# CRL-2169, RRID: CVCL_1728), UM-UC-3 (KCB Cat# KCB 2014053YJ, RRID: CVCL_1783), RT4 (CLS Cat# 300326/p10281_RT-4, RRID: CVCL_0036) and Biu87 (KCB Cat# KCB 2006103YJ, RRID: CVCL_6881) were purchased from the National Collection of Authenticated Cell Cultures (Shanghai, China) between 2018 and 2021.

Techniques: Selection, Modification, Binding Assay, Derivative Assay, Sequencing, Control, Confocal Microscopy, Comparison, Generated, Incubation, Labeling, Flow Cytometry

Journal: Cancer research

Article Title: Development of novel aptamer-based targeted chemotherapy for bladder cancer

doi: 10.1158/0008-5472.CAN-21-2691

Figure Lengend Snippet: ( A ) Cells were incubated with Cy5.5-labeled aptamer B1, lysosomes and nuclei were stained with LysoTracker Green and Hoechst 33342, respectively. Scale bar: 40 μm. ( B and C ) Cells were first treated with an endocytosis inhibitor, and then incubated with Cy5.5-labeled aptamer B1, followed by flow cytometry analysis. Chlorpromazine inhibits clathrin-mediated endocytosis. Amiloride inhibits pinocytosis. (D) SDS-PAGE analysis of proteins recovered from pull-down experiments using biotin-labeled aptamer B1. Asterisk indicates a protein band specific to aptamer B1 treatment. (E) Knockdown of AP2B1 with siRNA reduced aptamer B1 internalization into T24 cells. Representative data of three independent experiments are shown.

Article Snippet: Human bladder cancer cell lines T24 (CLS Cat# 300352/p619_T24, RRID: CVCL_0554) ( 27 ), TCCSUP (ATCC Cat# HTB-5, RRID: CVCL_1738), J82 (KCLB Cat# 30001, RRID: CVCL_0359), SW780 (ATCC Cat# CRL-2169, RRID: CVCL_1728), UM-UC-3 (KCB Cat# KCB 2014053YJ, RRID: CVCL_1783), RT4 (CLS Cat# 300326/p10281_RT-4, RRID: CVCL_0036) and Biu87 (KCB Cat# KCB 2006103YJ, RRID: CVCL_6881) were purchased from the National Collection of Authenticated Cell Cultures (Shanghai, China) between 2018 and 2021.

Techniques: Incubation, Labeling, Staining, Flow Cytometry, SDS Page, Knockdown

Journal: Cancer research

Article Title: Development of novel aptamer-based targeted chemotherapy for bladder cancer

doi: 10.1158/0008-5472.CAN-21-2691

Figure Lengend Snippet: ( A ) Schematic of the self-assembled aptamer-tethered DNA nanotrain for cellular delivery of drugs. Aptamer directs drug-loaded nanotrain for selective internalization into bladder cancer cells. Chemotherapeutic drugs are unloaded inside the cell and induce cytotoxicity. Drug intrinsic fluorescence can be used to monitor its intercalation into DNA duplex and intracellular release. ( B ) Agarose gel analysis of nanotrain assembly. Lane1: M1+M2; Lane2: M1; Lane3: M2; Lane4: aptamer-tethered; Lane5: aptamer-tethered DNA nanotrain. ( C ) Flow cytometry analysis of nanotrain internalization into T24 and SV-huc-1 cells. ( D ) Confocal microscope images of nanotrain internalization into T24 and SV-huc-1 cells. Scale bars: 40 μm. Representative data of three independent experiments are shown.

Article Snippet: Human bladder cancer cell lines T24 (CLS Cat# 300352/p619_T24, RRID: CVCL_0554) ( 27 ), TCCSUP (ATCC Cat# HTB-5, RRID: CVCL_1738), J82 (KCLB Cat# 30001, RRID: CVCL_0359), SW780 (ATCC Cat# CRL-2169, RRID: CVCL_1728), UM-UC-3 (KCB Cat# KCB 2014053YJ, RRID: CVCL_1783), RT4 (CLS Cat# 300326/p10281_RT-4, RRID: CVCL_0036) and Biu87 (KCB Cat# KCB 2006103YJ, RRID: CVCL_6881) were purchased from the National Collection of Authenticated Cell Cultures (Shanghai, China) between 2018 and 2021.

Techniques: Fluorescence, Agarose Gel Electrophoresis, Flow Cytometry, Microscopy

Journal: Cancer research

Article Title: Development of novel aptamer-based targeted chemotherapy for bladder cancer

doi: 10.1158/0008-5472.CAN-21-2691

Figure Lengend Snippet: ( A ) Characterization of the loading of epirubicin into the nanotrain. The intrinsic fluorescence of EPI decreased with increasing equivalents of nanotrain boxcar component strands, as it was quenched upon intercalation into DNA duplex. ( B ) Stability of the EPI-loaded nanotrain in PBS was evaluated by dialysis. Free EPI molecules diffused across the dialysis membrane and resulted in a high fluorescence intensity in the exterior chamber. Interaction of EPI with nanotrain (NT-EPI) restricted its diffusion. (C) Release of EPI in the presence of DNase. Intercalation of EPI into boxcar DNA duplex of NT resulted in fluorescence quenching. Treatment with DNase Ⅰ degraded dsDNA and restored the intrinsic fluorescence of EPI. ( D ) Confocal microscope images showing targeted delivery of NT-EPI into T24 and KU-7 bladder cancer cells. Cells were treated with free EPI or NT-EPI for 1 h. The nuclei were stained with DAPI. Scale bar: 40 μm. ( E and F ) Aptamer B1 NT-EPI exhibits similar cytotoxicity to that of free EPI against T24 (E) and KU-7 (F) bladder cancer cells. ( G ) Aptamer B1 NT-EPI was much less toxic to SV-huc-1 cells compared with free EPI. Cells were treated for 48 h and cell viability was measured by MTT assay. Data are mean ± SD. Representative data of three independent experiments shown.

Article Snippet: Human bladder cancer cell lines T24 (CLS Cat# 300352/p619_T24, RRID: CVCL_0554) ( 27 ), TCCSUP (ATCC Cat# HTB-5, RRID: CVCL_1738), J82 (KCLB Cat# 30001, RRID: CVCL_0359), SW780 (ATCC Cat# CRL-2169, RRID: CVCL_1728), UM-UC-3 (KCB Cat# KCB 2014053YJ, RRID: CVCL_1783), RT4 (CLS Cat# 300326/p10281_RT-4, RRID: CVCL_0036) and Biu87 (KCB Cat# KCB 2006103YJ, RRID: CVCL_6881) were purchased from the National Collection of Authenticated Cell Cultures (Shanghai, China) between 2018 and 2021.

Techniques: Stable Transfection, Fluorescence, Membrane, Diffusion-based Assay, Microscopy, Staining, MTT Assay

Journal: Cancer research

Article Title: Development of novel aptamer-based targeted chemotherapy for bladder cancer

doi: 10.1158/0008-5472.CAN-21-2691

Figure Lengend Snippet: ( A ) Schematic of orthotopic xenograft model establishment and treatment regimen. Luciferase-labeled KU-7 human bladder cancer cells were implanted into the bladder of nude mice. Fourteen days later, mice with positive bladder fluorescence signal were randomly divided into three groups and treated with intravesical instillation of PBS, free EPI (0.4 mg/ml, 50 μl/instillation) or aptamer B1 NT-EPI (equal molar EPI molecules). ( B ) Luminescence imaging and ( C ) quantitative analysis of tumors in mouse bladder at the end of treatment. Data are mean ± SD (n = 9 mice/group). ** P < 0.01. ( D ) Representative H&E staining images of bladder tumor sections from each group. Sections of whole bladder was shown in the middle (scale bar: 500 μm), with magnification on both sides (scale bar: 100 μm). The NT-EPI group showed much less cystitis compared to free EPI, as indicated by reduced infiltration of inflammatory cells and interstitial edema, suggesting reduced damage to normal urothelium. Quantification of all nine mice in each group are summarized in Supplementary Fig. 6 .

Article Snippet: Human bladder cancer cell lines T24 (CLS Cat# 300352/p619_T24, RRID: CVCL_0554) ( 27 ), TCCSUP (ATCC Cat# HTB-5, RRID: CVCL_1738), J82 (KCLB Cat# 30001, RRID: CVCL_0359), SW780 (ATCC Cat# CRL-2169, RRID: CVCL_1728), UM-UC-3 (KCB Cat# KCB 2014053YJ, RRID: CVCL_1783), RT4 (CLS Cat# 300326/p10281_RT-4, RRID: CVCL_0036) and Biu87 (KCB Cat# KCB 2006103YJ, RRID: CVCL_6881) were purchased from the National Collection of Authenticated Cell Cultures (Shanghai, China) between 2018 and 2021.

Techniques: Inhibition, In Vivo, Luciferase, Labeling, Fluorescence, Imaging, Staining

Journal: Scientific Reports

Article Title: Breast cancer cells exhibits specific dielectric signature in vitro using the open-ended coaxial probe technique from 200 MHz to 13.6 GHz

doi: 10.1038/s41598-019-41124-1

Figure Lengend Snippet: Morphology of normal and breast cancer cells. Prior to measurement, monolayer grown normal mammary (MCF-10A) and breast cancer cells (MDA-MB-231, Hs578T, MCF-7 and T47-D) cells were observed under EVOS XL Core Cell Imaging System (Life Technologies) at 400x.

Article Snippet: Human breast cancer cells MCF-7 (300273) and T47D (300353) were obtained from Cell line service (CLS)-GmbH.

Techniques: Imaging

Journal: Nature methods

Article Title: Deep learning enables fast and dense single-molecule localization with high accuracy

doi: 10.1038/s41592-021-01236-x

Figure Lengend Snippet: a) DECODE can reduce acquisition times by one order of magnitude. The same sample of microtubules, labeled with anti-α-tubulin primary and AF647 secondary antibodies, imaged with different UV activation intensities to result in different emitter densities per frame, between 0.08 and 0.86μm −2 and acquisition times between 93 and 1120 s, while keeping the total number of localizations the same. For high-density activation, we show a comparison with CSpline. b) Fourier Ring Correlation curves for DECODE and CSpline for different emitter densities. c) Resolution estimates obtained using the Fourier Ring Correlation and 0.143 criterion across densities for both methods. d) Fast live-cell SMLM on the Golgi apparatus labeled with a -mannosidase II-mEos3.2. See Supplementary Movie 1. e) Fast live-cell SMLM on the endoplasmic reticulum labeled with calnexin-mEos3.2. See Supplementary Movie 2 and . f) Fast live-cell SMLM on the nuclear pore complex protein Nup96-mMaple acquired in 3 seconds. g) DECODE enables ultra-high labeling densities. Microtubules labeled with a high concentration of anti-α and anti-β-tubulin primary and Alexa Fluor 647 secondary antibodies. g1, g2) Magnified regions as indicated in g. Data acquired with high-density labeling shows continuous structures. As a comparison, the same sample was acquired after pre-bleaching of the fluorophores to reach the single-molecule blinking regime. Here, single labels are resolved in the superresolution reconstruction and lead to a sparse decoration of the microtubules. g3, g4) Side view reconstructions of regions as indicated in g1, g2 resolving the hollow, cylinder-like structure of immunolabeled microtubules. h) Representative raw camera frames for the high-density and single-emitter acquisitions, respectively. Scale bars: 10μm (f inset, h), 1 μm (a, d, e, f, g, g1, g2), 100nm (g3, g4).

Article Snippet: The pulse length of the 405nm laser was adjusted manually to maintain a high emitter density and to allow imaging of all fluorophores in the field of view in about 1 min. For the acquisition of live-cell data of Nup96-mMaple , coverslips containing Nup96-mMaple cells (catalog no. 300461; CLS Cell Line Service, Eppelheim, Germany) were rinsed twice with warm PBS before they were mounted in 1mL growth medium containing 20 mM HEPES buffer and imaged directly.

Techniques: Labeling, Activation Assay, Comparison, Concentration Assay, Immunolabeling

Journal: Cell systems

Article Title: Receptor-Driven ERK Pulses Reconfigure MAPK Signaling and Enable Persistence of Drug-Adapted BRAF-Mutant Melanoma Cells

doi: 10.1016/j.cels.2020.10.002

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Human: SKMEL28, Melanoma Cell Line , MGH Cancer Center, primary source ATCC , CLS Cat# 300337/p495_SK-MEL-28, RRID:CVCL_0526.

Techniques: Formalin-fixed Paraffin-Embedded, Recombinant, Gene Expression, RNA Sequencing, Quantitative Proteomics, Phospho-proteomics, Software, Mass Spectrometry, Targeted Proteomics, Over Expression, Knockdown, Stable Transfection, Expressing, CRISPR, High Throughput Screening Assay, Microscopy, Live Cell Imaging, Cytometry, Staining

Journal: Research

Article Title: Novel Combination of Irreversible Electroporation and Allogenic Chimeric Antigen Receptor T-Cell Therapy Synergizes Therapeutic Outcomes in a Preclinical Human Pancreatic Cancer Mouse Model

doi: 10.34133/research.1105

Figure Lengend Snippet: (A) Calculated irreversible electroporation (IRE) lethal electric field thresholds for Pan02 mouse pancreatic cancer using a 3-dimensional (3D) collagen hydrogel; mean ± SD; n = 5. (B) COMSOL Multiphysics finite element model (FEM) representing subcutaneous mouse tumors with a 5-mm spacing shown. (C) Simulated percent tumor coverage by the lethal electric field using the subcutaneous FEM with randomized tissue conductivities and tumor sizes (4 to 8 mm); mean ± SD; n = 30. (D) In Vivo Imaging System (IVIS) imaging of FLuc-eGFP + Pan02 cells in immunodeficient NOD/SCID/IL2gc-KO (NSG) mice (m) follows complete ablation and subsequent microtumor recurrence after IRE treatment at 2,500 V/cm; n = 6. (E) Tumor scabbing and flattening at day 3 post-treatment. (F) Total photon flux within the tumor region of interest from the IVIS images; mean and range presented; multiple 2-tailed t tests between the data compared to the initial day 0 total photon flux; n = 6. (G) Measured tumor size over time from tumor inoculation; mean and range presented; multiple 2-tailed t tests between the data compared to pre-treatment tumor size on day 30; n = 6. ** P < 0.01; **** P < 0.0001. FLuc-eGFP, firefly-luciferase-enhanced green fluorescent protein. SAT, subcutaneous fat.

Article Snippet: Pan02 mouse pancreatic cancer cells (Cytion, 300501), AsPC-1 human pancreatic cancer cells (American Type Culture Collection [ATCC], CRL-1682), and Jurkat immortalized human T lymphocytes (ATCC, TIB-152) were cultured in RPMI 1640 medium (Thermo Fisher, 11875093) supplemented with 10% (v/v) fetal bovine serum (Fisher Scientific, FB12999102) and 1% (v/v) 10,000 U/ml penicillin–streptomycin (Gibco, 16140122).

Techniques: Electroporation, In Vivo Imaging, Imaging, Luciferase

Journal: International Journal of Molecular Sciences

Article Title: Synthetic Retinoid Seletinoid G Improves Skin Barrier Function through Wound Healing and Collagen Realignment in Human Skin Equivalents

doi: 10.3390/ijms21093198

Figure Lengend Snippet: Cell viability test of seletinoid G on a human keratinocyte cell line (HaCaT) and normal human dermal fibroblasts (NHDF). The cell viability of HaCaT cells and NHDF treated with seletinoid G at different concentrations for 24 and 48 h was measured by CCK-8 assay. (* p < 0.05; ** p < 0.01; *** p < 0.001 vs. the untreated group).

Article Snippet: The human keratinocyte cell line, HaCaT, was purchased from CLS (#300493, Cell Lines Service, Eppelheim, Germany).

Techniques: CCK-8 Assay

Journal: International Journal of Molecular Sciences

Article Title: Synthetic Retinoid Seletinoid G Improves Skin Barrier Function through Wound Healing and Collagen Realignment in Human Skin Equivalents

doi: 10.3390/ijms21093198

Figure Lengend Snippet: In vitro wound-healing effect of seletinoid G on wounded HaCaT keratinocyte monolayers. ( A ) HaCaT cells were line-scratched and then treated with seletinoid G (SG) at concentrations of 6, 12, and 25 μM in Dulbecco’s modified Eagle’s medium (DMEM) containing 1% fetal bovine serum (FBS) for 48 h Scale bars indicate 500 μm. ( B ) Each line-scratched area was automatically measured every hour for 48 h using time-lapse imaging microscopy.

Article Snippet: The human keratinocyte cell line, HaCaT, was purchased from CLS (#300493, Cell Lines Service, Eppelheim, Germany).

Techniques: In Vitro, Modification, Imaging, Microscopy