scholarly journals Visual Recognition and Efficient Isolation of Apoptotic Cells with Fluorescent-Magnetic-Biotargeting Multifunctional Nanospheres

2007 ◽  
Vol 53 (12) ◽  
pp. 2177-2185 ◽  
Author(s):  
Er-Qun Song ◽  
Guo-Ping Wang ◽  
Hai-Yan Xie ◽  
Zhi-Ling Zhang ◽  
Jun Hu ◽  
...  
Keyword(s):  
Ultraviolet Light ◽  
Clinical Diagnosis ◽  
Propidium Iodide ◽  
Biomedical Applications ◽  
Visual Recognition ◽  
Nuclear Dna ◽  
Annexin V ◽  
Living Cells ◽  
Apoptotic Cells ◽  
Propidium Iodide Staining

Abstract Background: Fluorescent-magnetic-biotargeting multifunctional nanospheres are likely to find important applications in bioanalysis, biomedicine, and clinical diagnosis. We have been developing such multifunctional nanospheres for biomedical applications. Methods: We covalently coupled avidin onto the surfaces of fluorescent-magnetic bifunctional nanospheres to construct fluorescent-magnetic-biotargeting trifunctional nanospheres and analyzed the functionality and specificity of these trifunctional nanospheres for their ability to recognize and isolate apoptotic cells labeled with biotinylated annexin V, which recognizes phosphatidylserine exposed on the surfaces of apoptotic cells. Results: The multifunctional nanospheres can be used in combination with propidium iodide staining of nuclear DNA to identify cells at different phases of the apoptotic process. Furthermore, we demonstrate that apoptotic cells induced by exposure to ultraviolet light can be isolated simply with a magnet from living cells at an efficiency of at least 80%; these cells can then be easily visualized with a fluorescence microscope. Conclusions: Our results show that fluorescent-magnetic-biotargeting trifunctional nanospheres can be a powerful tool for rapidly recognizing, magnetically enriching and sorting, and simultaneously identifying different kinds of cells.

Annexin V staining during reperfusion detects cardiomyocytes with unique properties

2001 ◽  
Vol 281 (5) ◽  
pp. H1931-H1937 ◽  
Author(s):  
Prakash Narayan ◽  
Robert M. Mentzer ◽  
Robert D. Lasley
Keyword(s):  
Propidium Iodide ◽  
Ventricular Myocytes ◽  
Contractile Function ◽  
Ischemia Reperfusion ◽  
Annexin V ◽  
Cell Width ◽  
Simulated Ischemia ◽  
Propidium Iodide Staining ◽  
Membrane Blebs ◽  
Apoptosis And Necrosis

With the use of markers of sarcolemmal membrane permeability, cardiomyocyte models of ischemic injury have primarily addressed necrotic death during ischemia. In the present study, we used annexin V-propidium iodide staining to examine apoptosis and necrosis after simulated ischemia and simulated reperfusion in rat ventricular myocytes. Annexin V binds phosphatidylserine, a phosphoaminolipid thought to be externalized during apoptosis or programmed cell death. Propidium iodide is a marker of cell necrosis. Under baseline conditions, <1% of cardiomyocytes stained positive for annexin V. After 20 or 60 min of simulated ischemia, there was no increase in annexin V staining, although 60-min simulated ischemia resulted in significant propidium iodide staining. Twenty minutes of simulated ischemia, followed by 20 or 60 min of simulated reperfusion, resulted in 8–10% of myocytes staining positive for annexin V. Annexin V-positive cells retained both rod-shaped morphology and contractile function but exhibited the decreased cell width indicative of cell shrinkage. Baseline mitochondrial free Ca2+(111 ± 14 nM) was elevated in reperfused annexin V-negative cells (214 ± 22 nM), and further elevated in annexin V-positive myocytes (382 ± 9 nM). After 60 min of simulated reperfusion, caspase-3-like activity was observed in ∼3% of myocytes, which had a rounded appearance and membrane blebs. These results suggest that the use of annexin V after simulated ischemia-reperfusion uncovers a population of cardiomyocytes whose characteristics appear to be consistent with cells undergoing apoptosis.

Detection of Apoptotic Cells Using Propidium Iodide Staining

2014 ◽  
Vol 2014 (11) ◽  
pp. pdb.prot082545 ◽  
Author(s):  
Andrea Newbold ◽  
Ben P. Martin ◽  
Carleen Cullinane ◽  
Michael Bots
Keyword(s):  
Propidium Iodide ◽  
Apoptotic Cells ◽  
Propidium Iodide Staining

scholarly journals Effect and Mechanism of Survivin on Hypoxia-Induced Multidrug Resistance of Human Laryngeal Carcinoma Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-6
Author(s):  
Dan Xu ◽  
Da Wei Li ◽  
Jin Xie ◽  
Xin Wei Chen
Keyword(s):  
Multidrug Resistance ◽  
Rna Interference ◽  
Cancer Cells ◽  
Propidium Iodide ◽  
Laryngeal Carcinoma ◽  
Survivin Expression ◽  
Annexin V ◽  
Carcinoma Cells ◽  
Propidium Iodide Staining ◽  
Human Laryngeal Carcinoma

This study aimed at clarifying the mechanism and role of survivin in hypoxia-induced multidrug resistance (MDR) of laryngeal carcinoma cells. Human laryngeal cancer cells were incubated under hypoxia or normoxia. The expression of survivin was silenced by performing RNA interference. Additionally, by Western blot and real-time quantitative RT-PCR, survivin expression was detected. The sensitivity of human laryngeal carcinoma cells to multiple drugs was measured by CCK-8 assay. Meanwhile, the apoptosis of cells induced by cisplatin or paclitaxel was assessed by Annexin-V/propidium iodide staining analysis. Under hypoxic conditions, the upregulation of survivin was abolished by RNA interference. Then, CCK-8 analysis demonstrated that the sensitivity to multiple agents of laryngeal carcinoma cells could be increased by inhibiting survivin expression (P<0.05). Moreover, Annexin-V/propidium iodide staining analysis revealed that decreased expression of survivin could evidently increase the apoptosis rate of laryngeal carcinoma cells that were induced by cisplatin or paclitaxel evidently (P<0.05). Our data suggests that hypoxia-elicited survivin may exert a pivotal role in regulating hypoxia-induced MDR of laryngeal cancer cells by preventing the apoptosis of cells induced by chemotherapeutic drug. Thus, blocking survivin expression in human laryngeal carcinoma cells may provide an avenue for gene therapy.

scholarly journals Lithium Sensitivity of Store Operated Ca2+ Entry and Survival of Fibroblasts Isolated from Chorea-Acanthocytosis Patients

2017 ◽  
Vol 42 (5) ◽  
pp. 2066-2077 ◽  
Author(s):  
Lisann Pelzl ◽  
Bhaeldin Elsir ◽  
Itishri Sahu ◽  
Rosi Bissinger ◽  
Yogesh Singh ◽  
...  
Keyword(s):  
Propidium Iodide ◽  
Neurodegenerative Disorder ◽  
Lithium Treatment ◽  
Annexin V ◽  
Supporting Cell ◽  
Pi3k Pathway ◽  
Loss Of Function ◽  
Propidium Iodide Staining ◽  
Phosphoinositide 3 Kinase ◽  
Vacuolar Protein

Background: The widely expressed protein chorein fosters activation of the phosphoinositide 3 kinase (PI3K) pathway thus supporting cell survival. Loss of function mutations of the chorein encoding gene VPS13A (vacuolar protein sorting-associated protein 13A) causes chorea-acanthocytosis (ChAc), a neurodegenerative disorder paralleled by deformations of erythrocytes. In mice, genetic knockout of chorein leads to enhanced neuronal apoptosis. PI3K dependent signalling upregulates Orai1, a pore forming channel protein accomplishing store operated Ca2+ entry (SOCE). Increased Orai1 expression and SOCE have been shown to confer survival of tumor cells. SOCE could be up-regulated by lithium. The present study explored, whether SOCE and/or apoptosis are altered in ChAc fibroblasts and could be modified by lithium treatment. Methods: Fibroblasts were isolated from ChAc patients and age-matched healthy volunteers. Cytosolic Ca2+ activity ([Ca2+]i) was estimated from Fura-2-fluorescence, SOCE from increase of [Ca2+]i following Ca2+ re-addition after Ca2+-store depletion with sarcoendoplasmatic Ca2+-ATPase (SERCA) inhibitor thapsigargin (1 µM), and apoptosis from annexin-V/propidium iodide staining quantified in flow cytometry. Results: SOCE was significantly smaller in ChAc fibroblasts than in control fibroblasts. Lithium (2 mM, 24 hours) significantly increased and Orai1 blocker 2-Aminoethoxydiphenyl Borate (2-APB, 50 µM, 24 hours) significantly decreased SOCE. Annexin-V-binding and propidium iodide staining were significantly higher in ChAc fibroblasts than in control fibroblasts. In ChAc fibroblasts annexin-V-binding and propidium iodide staining were significantly decreased by lithium treatment, significantly increased by 2-APB and virtually lithium insensitive in the presence of 2-APB. Conclusions: In ChAc fibroblasts, downregulation of SOCE contributes to enhanced susceptibility to apoptosis. Both, decreased SOCE and enhanced apoptosis of ChAc fibroblasts can be reversed by lithium treatment.

scholarly journals Corrigendum: Detection of Apoptotic Cells Using Propidium Iodide Staining

2018 ◽  
Vol 2018 (6) ◽  
pp. Corr104943
Author(s):  
Andrea Newbold ◽  
Ben P. Martin ◽  
Carleen Cullinane ◽  
Michael Bots
Keyword(s):  
Propidium Iodide ◽  
Apoptotic Cells ◽  
Propidium Iodide Staining

scholarly journals Modular Marx Generator Based on SiC-MOSFET Generating Adjustable Rectangular Pulses

Energies ◽  
2021 ◽  
Vol 14 (12) ◽  
pp. 3492
Author(s):  
Yahia Achour ◽  
Jacek Starzyński ◽  
Jacek Rąbkowski
Keyword(s):  
Silicon Carbide ◽  
Repetition Rate ◽  
Pulse Width ◽  
Biomedical Applications ◽  
Living Cells ◽  
Marx Generator ◽  
Electrical Field ◽  
Rectangular Shape ◽  
Overall Performance ◽  
Pulsed Electrical Field

The paper introduces a new design of Marx generator based on modular stages using Silicon Carbide MOSFETs (SiC-MOSFET) aimed to be used in biomedical applications. In this process, living cells are treated with intense nanosecond Pulsed Electrical Field (nsPEF). The electric field dose should be controlled by adjusting the pulse parameters such as amplitude, repetition rate and pulse-width. For this purpose, the structure of the proposed generator enables negative pulses with a quasi-rectangular shape, controllable amplitude, pulse-width and repetition-rate. A complete simulation study was conducted in ANSYS-Simplorer to verify the overall performance. A compact, modular prototype of Marx generator was designed with 1.7 kV rated SiC-MOSFETs and, finally, a set of experiments confirmed all expected features.

Type II secretory phospholipase A2 binds to ischemic flip-flopped cardiomyocytes and subsequently induces cell death

2003 ◽  
Vol 285 (5) ◽  
pp. H2218-H2224 ◽  
Author(s):  
R. Nijmeijer ◽  
M. Willemsen ◽  
C. J. L. M. Meijer ◽  
C. A. Visser ◽  
R. H. Verheijen ◽  
...  
Keyword(s):  
Cell Death ◽  
Propidium Iodide ◽  
Caspase 3 ◽  
Ischemia Reperfusion ◽  
Annexin V ◽  
Border Zone ◽  
Metabolic Inhibitors ◽  
Type Ii ◽  
Secretory Phospholipase

Type II secretory phospholipase A2 (sPLA2) is a cardiovascular risk factor. We recently found depositions of sPLA2 in the necrotic center of infarcted human myocardium and normally appearing cardiomyocytes adjacent to the border zone. The consequences of binding of sPLA2 to ischemic cardiomyocytes are not known. To explore a potential effect of sPLA2 on ischemic cardiomyocytes at a cellular level we used an in vitro model. The cardiomyocyte cell line H9c2 or adult cardiomyocytes were isolated from rabbits that were incubated with sPLA2 in the presence of metabolic inhibitors to mimic ischemia-reperfusion conditions. Cell viability was established with the use of annexin V and propidium iodide or 7-aminoactinomycin D. Metabolic inhibition induced an increase of the number of flip-flopped cells, including a population that did not stain with propidium iodide and that was caspase-3 negative. sPLA2 bound to the flip-flopped cells, including those negative for caspase-3. sPLA2 binding induced cell death in these latter cells. In addition, sPLA2 potentiated the binding of C-reactive protein (CRP) to these cells. We conclude that by binding to flip-flopped cardiomyocytes, including those that are caspase-3 negative and presumably reversibly injured, sPLA2 may induce cell death and tag these cells with CRP.

scholarly journals Total Phenols, Identification of Active Compounds and Anticancer Activity of Salvia judaica Boiss against the breast Cancer Cell MDA-231

2021 ◽  
Keyword(s):  
Breast Cancer ◽  
Cell Cycle ◽  
Essential Oil ◽  
Anticancer Activity ◽  
Breast Cancer Cell ◽  
Annexin V ◽  
Ethanol Extract ◽  
G1 Phase ◽  
Apoptotic Cells ◽  
Total Phenols

Salvia judaica is an annual herb from genus Salvia L.; the largest genera of Lamiaceae. It’s a medicinal plant prominent in pharmaceutical applications in many countries around the world. This study aimed to explore bioactive compounds likely to be responsible for the plant anticancer activity, and evaluate anticancer effects, after determining the total content of phenols in the ethanol extract and essential oil in this species. Ethanol extract (EE) and essential oil (EO) were prepared from dried aerial parts (leaves and the flower). GC-MS analysis of EO showed the presence of/43/ effective compounds in varying proportions, the major compounds were sesquiterpenes like delta-cadinene, alpha-Gurjunene, beta-humulene, and alpha-caryophyllene. This is the first study revealed that S.judaica is so rich in phenols which proceeded S.officinalis, noting the superiority of the EE over the EO samples in the total phenols. Anticancer properties of EE and EO of S. judaica against MDA-231 breast cancer cell line were studied -for the first time - by cell cycle analysis and Annexin V/PI apoptosis assay using Flow cytometry technique. Cells were treated with EE (0.001, 0.01, 0.02, 0.1mg/ml) and EO (0.005, 0.01, 0.02, 0.03, 0.04 mg/ml) at various concentrations for48 h. The results revealed that both EE and EO induced cell cycle arrest at G1-phase. Cells treated with EE and EO for 48h showed increasing the percentage of cells in G1-phase and decreasing the percentage of cells in S-phase with increasing concentration compared with untreated cells (control). Annexin V-FITC/PI assay confirmed that EO and EE were able to induce apoptosis. Cells treated with EOat (0.04 mg/ml) for 48h resulted in apoptotic cells at 96.68%, and necrotic cells at 0.12%, compared with untreated cells. On the other hand, Cells treated with EE at (0.1 mg/ml) for 48h resulted in apoptotic cells at 94.43%, and necrotic cells at 0.47%, compared with control. Results revealed that EO is better than EE as anticancer; treatment with EO resulted in more apoptotic cells and less necrotic cells, and there were significant differences between them. This confirmed that EO contains specific anticancer compounds as showed by GC-MS analysis. However, more studies should be performed to explore antioxidants present in S.judaica and determine the underlying mechanism of their anti-breast cancer properties.

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