A5084552653- Extracellular vesicles in disease
- Cancer Cells and Metastasis
- MicroRNA in disease regulation
- Spectroscopy Techniques in Biomedical and Chemical Research
- RNA modifications and cancer
- Nanoplatforms for cancer theranostics
- Laser-Matter Interactions and Applications
- Neonatal Respiratory Health Research
- Pulmonary Hypertension Research and Treatments
- Photoacoustic and Ultrasonic Imaging
- Chemical Reactions and Isotopes
- Inhalation and Respiratory Drug Delivery
- Galectins and Cancer Biology
- Circular RNAs in diseases
- Spectroscopy and Quantum Chemical Studies
- Adipose Tissue and Metabolism
- Glycosylation and Glycoproteins Research
- Advanced Fluorescence Microscopy Techniques
University of California, San Diego
2022-2024
Abstract Extracellular vesicles (EV) have emerged as critical effectors in the cross-talk between cancer and normal cells by transferring intracellular materials adjacent or distant cells. Previous studies begun to elucidate how cells, secreting EVs, adapt at a metastatic site facilitate cell metastasis. In this study, we utilized high-content microscopic screening platform investigate mechanisms of EV uptake primary lung fibroblasts. A selected library containing 90 FDA-approved anticancer...
Photoluminescence (PL) imaging has broad applications in visualizing biological activities, detecting chemical species, and characterizing materials. However, the information encoded PL images is often limited by overlapping emission spectra of chromophores. Here, we report a microscopy based on nonlinear interactions between mid-infrared visible excitations matters, which termed MultiDimensional Widefield Infrared-encoded Spontaneous Emission (MD-WISE) microscopy. MD-WISE can distinguish...
<p>Pathway inhibitors Trametinib and Copanlisib inhibit the uptake effect of EVs from various breast cancer cells. <b>A,</b> Western blots lung fibroblasts treated with 200 nmol/L or 10 µg/mL EV for 24 hours. <b>B,</b> qRT-PCR–determined mRNA levels S100A4 FN1 without MDA-MB-231 treatment 48 were added together when indicated. Data normalized to GAPDH. <b>C,</b> Lung pretreated (200 mmol/L) hours before Lck-GFP–labeled derived 4T1 MDA-MB-468 cells...
<p>Human lung fibroblasts take up MDA-MB-231 EVs via dynamin- and caveolae-dependent endocytosis macropinocytosis. <b>A,</b> Fibroblasts were pretreated with 80 µmol/L Dynasore or an equal volume of DMSO (as a control) for 24 hours then incubated 10 µg/mL 6 in the continuous presence control (<i>n</i> = 3 wells per group; 5 images well). Data are presented as mean ± SD. ***, <i>P</i> < 0.001. <b>B,</b> Cells 50 EIPA, CPZ, 200 Genistein...
<div>Abstract<p>Extracellular vesicles (EV) have emerged as critical effectors in the cross-talk between cancer and normal cells by transferring intracellular materials adjacent or distant cells. Previous studies begun to elucidate how cells, secreting EVs, adapt at a metastatic site facilitate cell metastasis. In this study, we utilized high-content microscopic screening platform investigate mechanisms of EV uptake primary lung fibroblasts. A selected library containing 90...
<p>MDA-MB-231 EV uptake by lung fibroblasts requires MEK2 but not MEK1. <b>A,</b> Fibroblasts were transfected with siRNAs against MEK1 or (two independent used for each gene), a control siRNA, at 48 hours before EVs added an incubation of 6 (<i>n</i> = 3 wells per group). <b>B,</b> Western blots cells indicated showing the gene knockdown efficiency MEK1/2. <b>C,</b> Representative images untreated and siRNAs. Images captured 10 ×. Green,...
<p>Supplementary Table S1 shows the compounds tested in screen.</p>
<p>The high-content microscopic platform for quantitative measurement of EV uptake. <b>A,</b> A schematic graph showing how cellular uptake fluorescently-labeled EVs was detected and quantified. Briefly, cells were prestained with CFSE (green) DiI (red). Following the incubation, extracellular washed off cell nuclei stained Hoechst 33342 (blue). During fluorescent image analysis, signals used to delineate single-cell boundaries per thereafter ObjectID, ID single cells. MFI,...
<p>Supplementary Table S2 shows the antibodies used in this study.</p>
<p>Trametinib interferes with macropinocytosis in lung fibroblasts while Copanlisib both and clathrin-mediated endocytosis. <b>A,</b> Quantification of the cellular uptake indicated markers or without Trametinib treatment. Data are presented as mean ± SD (<i>n</i> = 3 wells per group; 5 images well). Faint symbols represent all 15 group. Dark value from each well. **, <i>P</i> < 0.01; ns, not significant. <b>B,</b> Representative...
<p>Pathway inhibitors Trametinib and Copanlisib inhibit the uptake effect of EVs from various breast cancer cells. <b>A,</b> Western blots lung fibroblasts treated with 200 nmol/L or 10 µg/mL EV for 24 hours. <b>B,</b> qRT-PCR–determined mRNA levels S100A4 FN1 without MDA-MB-231 treatment 48 were added together when indicated. Data normalized to GAPDH. <b>C,</b> Lung pretreated (200 mmol/L) hours before Lck-GFP–labeled derived 4T1 MDA-MB-468 cells...
<p>Supplementary Figure S1 shows characterization of EVs. (A) Western blots whole cell lysates (WCL) and EVs from MDA-MB-231 showing EV markers a Golgi marker (GM130, as negative control for EV-specific proteins). (B) Nanoparticle tracking analysis (NTA) size distribution (n=3 biological replicates). Data are presented mean ± standard error the (SEM).</p>
<p>Supplementary Figure S2 shows the effect of inhibitors different cell uptake pathways on marker uptake. Lung fibroblasts were pre-treated with 50 μM EIPA, 10 CPZ, or 200 Genistein for 24 h and then incubated FITC-transferrin, Alexa488-BSA FITC-dextran 6 in continuous presence inhibitor (n=3 wells per group; 5 images well). Data are presented as mean ± SD. *, p<0.05; **, p<0.01; ***, p<0.001; ns, not significant.</p>
<div>Abstract<p>Extracellular vesicles (EV) have emerged as critical effectors in the cross-talk between cancer and normal cells by transferring intracellular materials adjacent or distant cells. Previous studies begun to elucidate how cells, secreting EVs, adapt at a metastatic site facilitate cell metastasis. In this study, we utilized high-content microscopic screening platform investigate mechanisms of EV uptake primary lung fibroblasts. A selected library containing 90...
<p>Trametinib interferes with macropinocytosis in lung fibroblasts while Copanlisib both and clathrin-mediated endocytosis. <b>A,</b> Quantification of the cellular uptake indicated markers or without Trametinib treatment. Data are presented as mean ± SD (<i>n</i> = 3 wells per group; 5 images well). Faint symbols represent all 15 group. Dark value from each well. **, <i>P</i> < 0.01; ns, not significant. <b>B,</b> Representative...
<p>Supplementary Figure S2 shows the effect of inhibitors different cell uptake pathways on marker uptake. Lung fibroblasts were pre-treated with 50 μM EIPA, 10 CPZ, or 200 Genistein for 24 h and then incubated FITC-transferrin, Alexa488-BSA FITC-dextran 6 in continuous presence inhibitor (n=3 wells per group; 5 images well). Data are presented as mean ± SD. *, p<0.05; **, p<0.01; ***, p<0.001; ns, not significant.</p>
<p>Supplementary Table S1 shows the compounds tested in screen.</p>
<p>Supplementary Table S2 shows the antibodies used in this study.</p>
<p>Screening of a targeted library oncology drugs by high-content microscopy identifies compounds suppressing EV uptake. <b>A,</b> Cells were pretreated with (200 nmol/L final concentration) for 24 hours and then either assayed viability or incubated 10 µg/mL 6 Dynasore (80 µmol/L) was also included as control treatment. For both MTS assay uptake assay, <i>n</i> = 2 wells per drug tested. 5 images well captured analyzed. Each dot represents the mean value two...
<p>MDA-MB-231 EV uptake by lung fibroblasts requires MEK2 but not MEK1. <b>A,</b> Fibroblasts were transfected with siRNAs against MEK1 or (two independent used for each gene), a control siRNA, at 48 hours before EVs added an incubation of 6 (<i>n</i> = 3 wells per group). <b>B,</b> Western blots cells indicated showing the gene knockdown efficiency MEK1/2. <b>C,</b> Representative images untreated and siRNAs. Images captured 10 ×. Green,...
<p>Screening of a targeted library oncology drugs by high-content microscopy identifies compounds suppressing EV uptake. <b>A,</b> Cells were pretreated with (200 nmol/L final concentration) for 24 hours and then either assayed viability or incubated 10 µg/mL 6 Dynasore (80 µmol/L) was also included as control treatment. For both MTS assay uptake assay, <i>n</i> = 2 wells per drug tested. 5 images well captured analyzed. Each dot represents the mean value two...
<p>The high-content microscopic platform for quantitative measurement of EV uptake. <b>A,</b> A schematic graph showing how cellular uptake fluorescently-labeled EVs was detected and quantified. Briefly, cells were prestained with CFSE (green) DiI (red). Following the incubation, extracellular washed off cell nuclei stained Hoechst 33342 (blue). During fluorescent image analysis, signals used to delineate single-cell boundaries per thereafter ObjectID, ID single cells. MFI,...
<p>Human lung fibroblasts take up MDA-MB-231 EVs via dynamin- and caveolae-dependent endocytosis macropinocytosis. <b>A,</b> Fibroblasts were pretreated with 80 µmol/L Dynasore or an equal volume of DMSO (as a control) for 24 hours then incubated 10 µg/mL 6 in the continuous presence control (<i>n</i> = 3 wells per group; 5 images well). Data are presented as mean ± SD. ***, <i>P</i> < 0.001. <b>B,</b> Cells 50 EIPA, CPZ, 200 Genistein...