scholarly journals Electrodelivery of Drugs into Cancer Cells in the Presence of Poloxamer 188

2010 ◽  
Vol 2010 ◽  
pp. 1-11 ◽  
Author(s):  
Iana Tsoneva ◽  
Iordan Iordanov ◽  
Annette J. Berger ◽  
Toma Tomov ◽  
Biliana Nikolova ◽  
...  
Keyword(s):  
Small Molecules ◽  
Cancer Cells ◽  
Nude Mice ◽  
Propidium Iodide ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Electrical Pulse ◽  
Red Fluorescent Protein ◽  
Poloxamer 188 ◽  
Pore Structures

In the present study it is shown that poloxamer 188, added before or immediately after an electrical pulse used for electroporation, decreases the number of dead cells and at the same time does not reduce the number of reversible electropores through which small molecules (cisplatin, bleomycin, or propidium iodide) can pass/diffuse. It was suggested that hydrophobic sections of poloxamer 188 molecules are incorporated into the edges of pores and that their hydrophilic parts act as brushy pore structures. The formation of brushy pores may reduce the expansion of pores and delay the irreversible electropermeability. Tumors were implanted subcutaneously in both flanks of nude mice using HeLa cells, transfected with genes for red fluorescent protein and luciferase. The volume of tumors stopped to grow after electrochemotherapy and the use of poloxamer 188 reduced the edema near the electrode and around the subcutaneously growing tumors.

Identification of microRNAs that stabilize p53 in HPV-positive cancer cells

2021 ◽  
Author(s):  
Gustavo Martínez-Noël ◽  
Patricia Szajner ◽  
Rebecca E. Kramer ◽  
Kathleen A. Boyland ◽  
Asma Sheikh ◽  
...  
Keyword(s):  
Cancer Cells ◽  
High Throughput ◽  
Ubiquitin Ligase ◽  
Hela Cells ◽  
Fluorescent Protein ◽  
Human Papillomaviruses ◽  
Live Cell ◽  
Viral Oncoprotein ◽  
Protein Levels ◽  
P53 Stability

Etiologically, 5% of all cancers worldwide are caused by the high-risk human papillomaviruses (hrHPVs). These viruses encode two oncoproteins (E6 and E7) whose expression is required for cancer initiation and maintenance. Among their cellular targets are the p53 and the retinoblastoma tumor suppressor proteins. Inhibition of the hrHPV E6-mediated ubiquitylation of p53 through the E6AP ubiquitin ligase results in the stabilization of p53, leading to cellular apoptosis. We utilized a live cell high throughput screen to determine whether exogenous microRNA (miRNA) transfection had the ability to stabilize p53 in hrHPV-positive cervical cancer cells expressing a p53-fluorescent protein as an in vivo reporter of p53 stability. Among the miRNAs whose transfection resulted in the greatest p53 stabilization was 375-3p that has previously been reported to stabilize p53 in HeLa cells, providing validation of the screen. The top 32 miRNAs in addition to 375-3p were further assessed using a second cell-based p53 stability reporter system as well as in non-reporter HeLa cells to examine their effects on endogenous p53 protein levels, resulting in the identification of 23 miRNAs whose transfection increased p53 levels in HeLa cells. While a few miRNAs that stabilized p53 led to decreases in E6AP protein levels, all targeted HPV oncoprotein expression. We further examined subsets of these miRNAs for their abilities to induce apoptosis and determined whether it was p53-mediated. The introduction of specific miRNAs revealed surprisingly heterogeneous responses in different cell lines. Nonetheless, some of the miRNAs described here have potential as therapeutics for treating HPV-positive cancers. Importance Human papillomaviruses cause approximately 5% of all cancers worldwide and encode genes that contribute to both the initiation and maintenance of these cancers. The viral oncoprotein E6 is expressed in all HPV-positive cancers and functions by targeting the degradation of p53 through the engagement of the cellular ubiquitin ligase E6AP. Inhibiting the degradation of p53 leads to apoptosis in HPV-positive cancer cells. Using a high throughput live cell assay we identified several miRNAs whose transfection stabilize p53 in HPV-positive cells. These miRNAs have the potential to be used in the treatment of HPV-positive cancers.

Effect of salmonella typhimurium A1-R on quiescent cancer cells that do not respond to standard chemotherapy.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e13576-e13576
Author(s):  
Shuya Yano ◽  
Yong Zhang ◽  
Fuminari Uehara ◽  
Yukihiko Hiroshima ◽  
Shinji Miwa ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Salmonella Typhimurium ◽  
Cancer Cells ◽  
Nude Mice ◽  
Fluorescent Protein ◽  
Confocal Imaging ◽  
Cytotoxic Agents ◽  
Cycle Phase ◽  
Solid Cancer ◽  
Proliferating Cells

e13576 Background: Quiescent cancer cells are a major impediment to treating solid cancer with chemotherapy, since in most tumors the majority of the cells are quiescent. Methods: The cell-cycle phase of each human MKN45 gastric cancer cell was imaged using a fluorescence ubiquitination cell cycle indicator (FUCCI). With FUCCI, quiescent cancer cells express mKusabira-Orange fluorescent protein (red) and proliferating cells express mAzami-Green fluorescent protein (green). FUCCI-labeled cancer cells in tumor spheres and subcutaneous tumor in nude mice were treated with Salmonella typhimurium A1-R. Results: Time-lapse confocal imaging showed that cancer cells in tumor spheres in serum-free culture become and remained quiescent. S. typhimurium A1-R infected and killed quiescent cancer cells in tumor spheres. In contrast, cytotoxic agents did not kill the quiescent cancer cells in the tumor spheres. S. typhimurium A1-R targeting of FUCCI-expressing subcutaneous tumors growing in nude mice resulted in the killing of quiescent cancer cells resistant to cytotoxic agents. Conclusions: S. typhimurium A1-R can kill quiescent cancer cells which suggests a new therapeutic paradigm potentially more effective than current therapeutics which are ineffective against quiescent cancer cells.

scholarly journals Fluorescently Tagged CCL19 and CCL21 to Monitor CCR7 and ACKR4 Functions

2018 ◽  
Vol 19 (12) ◽  
pp. 3876 ◽  
Author(s):  
Vladimir Purvanov ◽  
Christoph Matti ◽  
Guerric P. B. Samson ◽  
Ilona Kindinger ◽  
Daniel F. Legler
Keyword(s):  
Cancer Cells ◽  
Chemokine Receptors ◽  
Fluorescent Protein ◽  
Adaptive Immune Response ◽  
Cell Types ◽  
Red Fluorescent Protein ◽  
Adaptive Immune ◽  
Guidance Cues ◽  
Cell Locomotion ◽  
Health And Disease

Chemokines are essential guidance cues orchestrating cell migration in health and disease. Cognate chemokine receptors sense chemokine gradients over short distances to coordinate directional cell locomotion. The chemokines CCL19 and CCL21 are essential for recruiting CCR7-expressing dendritic cells bearing pathogen-derived antigens and lymphocytes to lymph nodes, where the two cell types meet to launch an adaptive immune response against the invading pathogen. CCR7-expressing cancer cells are also recruited by CCL19 and CCL21 to metastasize in lymphoid organs. In contrast, atypical chemokine receptors (ACKRs) do not transmit signals required for cell locomotion but scavenge chemokines. ACKR4 is crucial for internalizing and degrading CCL19 and CCL21 to establish local gradients, which are sensed by CCR7-expressing cells. Here, we describe the production of fluorescently tagged chemokines by fusing CCL19 and CCL21 to monomeric red fluorescent protein (mRFP). We show that purified CCL19-mRFP and CCL21-mRFP are versatile and powerful tools to study CCR7 and ACKR4 functions, such as receptor trafficking and chemokine scavenging, in a spatiotemporal fashion. We demonstrate that fluorescently tagged CCL19 and CCL21 permit the visualization and quantification of chemokine gradients in real time, while CCR7-expressing leukocytes and cancer cells sense the guidance cues and migrate along the chemokine gradients.

scholarly journals Analysis of Stroma Labeling During Multiple Passage of a Sarcoma Imageable Patient-Derived Orthotopic Xenograft (iPDOX) in Red Fluorescent Protein Transgenic Nude Mice

2017 ◽  
Vol 118 (10) ◽  
pp. 3367-3371 ◽  
Author(s):  
Tasuku Kiyuna ◽  
Takashi Murakami ◽  
Yasunori Tome ◽  
Kei Kawaguchi ◽  
Kentaro Igarashi ◽  
...  
Keyword(s):  
Nude Mice ◽  
Fluorescent Protein ◽  
Red Fluorescent Protein ◽  
Orthotopic Xenograft ◽  
Multiple Passage

Fluorescent proteins for in vivo imaging, where's the biliverdin?

2020 ◽  
Vol 48 (6) ◽  
pp. 2657-2667
Author(s):  
Felipe Montecinos-Franjola ◽  
John Y. Lin ◽  
Erik A. Rodriguez
Keyword(s):  
Hydrogen Peroxide ◽  
In Vivo Imaging ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Fluorescent Imaging ◽  
Infrared Light ◽  
Red Fluorescent Protein ◽  
Color Palette

Noninvasive fluorescent imaging requires far-red and near-infrared fluorescent proteins for deeper imaging. Near-infrared light penetrates biological tissue with blood vessels due to low absorbance, scattering, and reflection of light and has a greater signal-to-noise due to less autofluorescence. Far-red and near-infrared fluorescent proteins absorb light >600 nm to expand the color palette for imaging multiple biosensors and noninvasive in vivo imaging. The ideal fluorescent proteins are bright, photobleach minimally, express well in the desired cells, do not oligomerize, and generate or incorporate exogenous fluorophores efficiently. Coral-derived red fluorescent proteins require oxygen for fluorophore formation and release two hydrogen peroxide molecules. New fluorescent proteins based on phytochrome and phycobiliproteins use biliverdin IXα as fluorophores, do not require oxygen for maturation to image anaerobic organisms and tumor core, and do not generate hydrogen peroxide. The small Ultra-Red Fluorescent Protein (smURFP) was evolved from a cyanobacterial phycobiliprotein to covalently attach biliverdin as an exogenous fluorophore. The small Ultra-Red Fluorescent Protein is biophysically as bright as the enhanced green fluorescent protein, is exceptionally photostable, used for biosensor development, and visible in living mice. Novel applications of smURFP include in vitro protein diagnostics with attomolar (10−18 M) sensitivity, encapsulation in viral particles, and fluorescent protein nanoparticles. However, the availability of biliverdin limits the fluorescence of biliverdin-attaching fluorescent proteins; hence, extra biliverdin is needed to enhance brightness. New methods for improved biliverdin bioavailability are necessary to develop improved bright far-red and near-infrared fluorescent proteins for noninvasive imaging in vivo.

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