Imaging of fluorescent small molecule transport into electropermeabilized cells reveals polarized patterns of entry, which must reflect in some way the mechanisms of the migration of these molecules across the compromised membrane barrier. In some reports, transport occurs primarily across the areas of the membrane nearest the positive electrode (anode), but in others cathode-facing entry dominates. Here we compare YO-PRO-1, propidium, and calcein uptake into U-937 cells after nanosecond (6 ns) and microsecond (220 µs) electric pulse exposures. Each of the three dyes exhibits a different pattern. Calcein shows no preference for anode- or cathode-facing entry that is detectable with our measurement system. Immediately after a microsecond pulse, YO-PRO-1 and propidium enter the cell roughly equally from the positive and negative poles, but transport through the cathode-facing side dominates in less than 1 s. After nanosecond pulse permeabilization, YO-PRO-1 and propidium enter primarily on the anode-facing side of the cell.
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http://dx.doi.org/10.1007/s00232-017-9962-1 | DOI Listing |
STAR Protoc
December 2024
Université Claude Bernard Lyon 1, CNRS UMR 5261, INSERM U1315, Institut NeuroMyoGène, Pathophysiology and Genetics of Neuron and Muscle Unit, 69008 Lyon, France.
Skeletal muscle is critically dependent on the function of muscle stem cells (MuSCs) for effective muscle repair following injury. Here, we detail a protocol for the isolation of primary muscle cells and subsequent analysis of proliferation capacity in vitro using EdU (5-ethynyl-2'-deoxyuridine) on fixed cells. We also describe a cell death analysis on living cells with the identification of early- and late-apoptotic cells, as well as necrotic cells, through the incorporation of propidium iodide and YO-PRO-1 staining.
View Article and Find Full Text PDFRadiol Oncol
September 2024
Faculty of Electrical Engineering, University of Ljubljana, Ljubljana, Slovenia.
Background: Administering gadolinium-based contrast agent before electroporation allows the contrast agent to enter the cells and enables MRI assessment of reversibly electroporated regions. The aim of this study was evaluation of contrast agent entrapment in Chinese hamster ovary (CHO) cells and comparison of these results with those determined by standard methods for assessing cell membrane permeability, cell membrane integrity and cell survival following electroporation.
Materials And Methods: Cell membrane permeabilization and cell membrane integrity experiments were performed using YO-PRO-1 dye and propidium iodide, respectively.
Hum Reprod
February 2024
Institute of Science and Technology, National Health Commission, Beijing, China.
Study Question: What is the mechanism behind cryoinjury in human sperm, particularly concerning the interplay between reactive oxygen species (ROS) and autophagy, and how does it subsequently affect sperm fate?
Summary Answer: The freeze-thaw operation induces oxidative stress by generating abundant ROS, which impairs sperm motility and activates autophagy, ultimately guiding the sperm toward programmed cell death such as apoptosis and necrosis, as well as triggering premature capacitation.
What Is Known Already: Both ROS-induced oxidative stress and autophagy are thought to exert an influence on the quality of frozen-thawed sperm.
Study Design, Size, Duration: Overall, 84 semen specimens were collected from young healthy fertile males, with careful quality evaluation.
Biomedicines
November 2023
Institute of Experimental Medicine, WCRC "Center for Personalized Medicine", Saint-Petersburg 197022, Russia.
Glioblastoma (GBM) is an aggressive and lethal malignancy of the central nervous system with a median survival rate of 15 months. We investigated the combined anticancer effects of nerve growth factor (NGF), cathelicidin (LL-37), and protegrin-1 (PG-1) with chemotherapy (temozolomide, doxorubicin, carboplatin, cisplatin, and etoposide) in the glioblastoma U251 cell line to overcome the limitations of conventional chemotherapy and to guarantee specific treatments to succeed. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was used to study cell viability and to determine the cytotoxic effects of NGF, LL-37, and PG-1 and their combination with chemotherapy in U251 cells.
View Article and Find Full Text PDFFolia Biol (Praha)
February 2024
Institute of Pathological Physiology, First Faculty of Medicine, Charles University, Prague, Czech Republic.
Propidium iodide (PI) and YO-PRO-1 (YPI) dyes are routinely used to determine sperm viability in many livestock species. It is commonly accepted that these dyes penetrate only sperm cells with damaged plasma membranes. Recently, however, the mechanism of dye uptake unrelated to damaged plasma membranes, but instead related to pannexin channels in dog and stallion sperm cells was demonstrated.
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