DOCSLIB.ORG

Valinomycin Induces Apoptosis of Ascites Hepatomacells (AH-130) in Relation to Mitochondrial Membranepotential

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Valinomycin Induces Apoptosis of Ascites Hepatomacells (AH-130) in Relation to Mitochondrial Membranepotential CELL STRUCTURE AND FUNCTION 22: 555-563 (1997) © 1997 by Japan Society for Cell Biology Valinomycin Induces Apoptosis of Ascites Hepatoma Cells (AH-130) in Relation to Mitochondrial MembranePotential Yoko Inai, Munehisa Yabuki*, Tomoko Kanno*2, Jitsuo Akiyama**, Tatsuji Yasuda, and Kozo Utsumi*1 Department of Cell Chemistry, Institute of Molecular and Cell Biology, Okayama University Medical School, Okayama 700, Japan, ^Institute of Medical Science, Kurashiki Medical Center, Kurashiki 710, Japan, and **Doonan Institute of Medical Science, Hakodate 041, Japan Key words: AH-130 cells/valinomycin/mitochondrial membrane potential/Bcl-2 family/caspase-3 ABSTRACT.Valinomycin is a potassium ionophore, and is well knownto cause the collapse of the mitochon- drial membranepotential. It has been reported that loss of mitochondrial membranepotential is observed in the early stages of apoptosis induced by various agents. Thus, the effects of valinomycin on tumor cells were exam- ined. Valinomycin induced uncoupling of respiration and depolarization of isolated mitochondria. Depolariza- tion of intact mitochondria in AH-130rat ascites hepatoma cells was also induced by valinomycin. Valinomycin induced apoptosis revealing the typical apoptotic characteristics such as fragmentation and ladder formation of DNA,shrinkage of cells, and formation of pycnotic nucleus. There was a correlation between the depolariza- tion of mitochondria and DNAfragmentation. After depolarization of mitochondria, the activity of caspase-3- like protease but not caspase-1-like protease increased markedly. In contrast, this apoptosis did not involve the release of reactive oxygen species from mitochondria, increase in intracellular calcium concentration, or protein synthesis. In addition, anti-apoptotic members of the Bcl-2 family (Bcl-xL and Bcl-2) were not correlated with apoptosis. These results indicate that valinomycin might induce apoptosis through degradation of the mitochon- drial membranepotential. Taken together, these observations suggest that there maybe a mechanismthat trans- mits the signal from mitochondrial depolarization to subsequent apoptosis execution steps. The removal of surplus cells plays an important role 26, 30). Recently, Shimizu et al. reported that Bcl-2 and in both cellular development and homeostasis. The elim- Bcl-xL protected mitochondria against the loss of mem- ination of such cells is accomplished by apoptosis, a brane potential during apoptosis and certain forms of well-defined mode of cell death with distinct biological necrotic cell death (25). Bcl-2 and Bcl-xL proteins have and morphological features. Misregulation of apopto- been to several intracellular membranesincluding those sis results in various diseases. Apoptosis is regulated by of mitochondria, the endoplasmic reticulum, and the a number of genes, with the bcl-2 gene family encoding nuclear membrane (2, 4, 9). In certain cells, however, several apoptosis-related proteins. Changes in mito- apoptosis was inhibited by Bcl-2 localized in the mito- chondrial function have been shownto be involved in chondria but not in the endoplasmic reticulum (31). apoptotic cell death. The mitochondrial membrane These observations indirectly implicate the involvement potential, which is the driving force of mitochondrial of mitochondrial function in the mechanismof apopto- ATP synthesis, decreases during apoptosis, while the sis. Onthe other hand, it has been reported that the loss maintenance of the membranepotential prevents the of mitochondrial membranepotential is not related to apoptosis induced by various stimuli (6, 17, 18, 22, 25, apoptosis induced by certain stimuli (8). Thus, the mo- lecular mechanismof the involvement of mitochondria 1 To whomcorrespondence should be addressed. in apoptotic cell death is not well understood. Tel & Fax: +81-86-426-8616 To determine whether mitochondrial dysfunction is a 2 On leave from the DoonanInstitute of Medical Science. Abbreviations used: AIF, apoptosis-inducing factor; AMC,7-ami- critical event in the apoptotic cascade, it is important to no-4-methyl-coumarin; CHX, cycloheximide; DEVD-CHO, acetyl- investigate the mechanisms of apoptosis induced by vari- Asp-Glu-Val-Asp-aldehyde; DMEM,Dulbecco's modified Eagle's me- ous agents that collapse the mitochondrial membrane dium; KRPbuffer, Krebs-Ringer phosphate buffer; Mit, mitochon- potential. Recently, Marchetti et al. reported that thym- dria; PBS, phosphate-buffered saline; PT, permeability transition; ocyte apoptosis was induced by addition of protopor- ROS,reactive oxygen species; Succ, sodium succinate; Val, valinomy- phyrin IX to the mediumwhich disrupted mitochondri- cin; YVAD-CHO, acetyl-Tyr-Val-Ala-Asp-aldehyde. al membranepotential via interaction with mitochondri- 555 Y. Inai et al. al benzodiazepin receptors and formation of mitochon- treated with 0.4mg/ml RNase A for 1 hour at 37°C and drial permeability transition (PT) pores. Furthermore, treated further by 0.4 mg/ml proteinase K under the same con- they reported that overexpression of Bcl-2 partially in- ditions. The solutions were precipitated at -20°C with 0.2 hibited the mitochondrial PT and apoptosis (17). volume of 5 MNaCl and 1 volume of isopropanol. DNAwas In this study, we investigated the mechanismof apop- pelleted by centrifugation and resuspended in TE (10 mM tosis induced by valinomycin, a potassium ionophore Tris-HCl, pH 7.6, and 1 mMEDTA) buffer. The DNAwas an- and an uncoupler of mitochondrial respiration. Valino- alyzed by electrophoresis in 2% agarose gels. mycin induced apoptosis of AH-130cells (rat ascites Quantification ofDNAfragmentation. The tumor cells (7 hepatoma) through the depolarization of mitochondria x 105 cells) were collected by centrifugation, and the pellets and activation of caspase-3-like protease. Although were dissolved with lysis buffer A and kept on ice for 15 min- Bcl-xL was up-regulated by valinomycin during apopto- utes. Samples were centrifuged at 15,000 rpm for 20 minutes, sis, cycloheximide, a protein synthesis inhibitor, had no and the DNAin supernatants and pellets was measured by the effect on DNAfragmentation. diphenylamine method as fragmented DNAand intact DNA, respectively (3). DNAfragmentation {%) was calculated by MATERIALS AND METHODS the following formula: Chemicals. Valinomycin and cycloheximide were pur- DNAfragmentation {%) = { fragmented DNA/(fragmented DNA chased from WakoCo. Ltd. (Japan) and Sigma Chemicals Co. (USA), respectively. Enhanced chemiluminescence de- +intact DNA)} x 100. tection system was obtained from AmershamCorp. (UK). Measurementof mitochondrial membranepotential in Antibodies were purchased from Santa Cruz Biotechnology cells. Rhodamine 123 was loaded into the cells by incubation (USA). Proteinase K, RNase A, and ATPbioluminescence in medium containing 10 ^M rhodamine 123 for 30 minutes. assay kit CLS II were purchased from Boehringer Mannheim Cells were washed three times with KRPbuffer by centrifuga- (Germany). Fluorogenic peptides and caspase family inhibi- tion. Content of rhodamine 123 was measured after dissolu- tors were obtained from Peptide Institute Inc. (Japan). All tion of cells by addition of 0. \% Triton X-100. Fluorospectro- other reagents were of analytical grade and purchased from metry (Hitachi 650-10LC) was performed at excitation and Nacalai Tesque Co. (Japan). emission wavelengths of 485 nmand 520 nm, respectively. Isolation of liver mitochondria. Male Donryu rats weigh- Western blotting. The tumor cells were washed with PBS ing 200 g were fasted overnight and their liver mitochondria twice, and dissolved in lysis buffer B (0.1% SDS, \% Nonidet were isolated by the method of Hogeboom(10) using medium P-40, 0.5% sodium deoxycholate, and 1 mMphenylmethylsul- containing 0.25 Msucrose, 10 mMTris-HCl, pH 7.4, and 0.1 fonyl fluoride). The cell lysate was separated by SDS-PAGE mMEDTA.The mitochondrial preparations were washed using 10 or 12.5% acrylamide gels and transferred to PVDF twice with respiratory medium (154 mMKC1 containing 5 membranes. After blocking with 5% skimmed milk over- mMK2HPO4, 5 mM KH2PO4, pH 7.4, 3 mM MgCl2, and 0.1 night, the membraneswere incubated with anti-Bcl-xL, or mMEDTA)and resuspended in the same medium. -Bcl-2 antibody. The blots were then, hybridized with second- Preparation ofascites hepatoma cells. About 0.5 ml of as- ary antibody conjugated with horseradish peroxidase. Specific cites containing exponentially growing AH-130cells was inoc- bands for Bcl-xL and Bcl-2 were visualized with an enhanced ulated into the peritoneal cavity of male Donryu rats. After 6- chemiluminescence detection system. 8 days, cells were harvested from the peritoneal cavity. The tu- Hydrogen peroxide measurement. Hydrogen peroxide re- mor cell suspension was hemolyzed with 0.2% NaCl and lease by mitochondria was measured fluorometricaHy by moni- washed by centrifugation in calcium-free Krebs-Ringer phos- toring the oxidation of /7-hydroxyphenylacetic acid coupled phate (KRP) buffer. Membrane integrity was measured by with the enzymatic reduction of hydrogen peroxide using trypan blue dye exclusion. horseradish peroxidase (1 1). The reaction mixture consisted Induction of apoptosis. AH-130cells were suspended in of 0. 15 mg protein/ml rat liver mitochondria, 0.5 mg/mlp-hy- Dulbecco's modified Eagle's medium (DMEM) containing droxyphenylacetic acid, 4 U/ml horseradish peroxidase, and 20% heat-inactivated fetal bovine serum at 1 x 106 cells/ml 7.5 mMsodium succinate in respiratory medium. The oxida- and precultured
Load more

Recommended publications
  • Dietary Supplements Compendium Volume 1
    2015 Dietary Supplements Compendium DSC Volume 1 General Notices and Requirements USP–NF General Chapters USP–NF Dietary Supplement Monographs USP–NF Excipient Monographs FCC General Provisions FCC Monographs FCC Identity Standards FCC Appendices Reagents, Indicators, and Solutions Reference Tables DSC217M_DSCVol1_Title_2015-01_V3.indd 1 2/2/15 12:18 PM 2 Notice and Warning Concerning U.S. Patent or Trademark Rights The inclusion in the USP Dietary Supplements Compendium of a monograph on any dietary supplement in respect to which patent or trademark rights may exist shall not be deemed, and is not intended as, a grant of, or authority to exercise, any right or privilege protected by such patent or trademark. All such rights and privileges are vested in the patent or trademark owner, and no other person may exercise the same without express permission, authority, or license secured from such patent or trademark owner. Concerning Use of the USP Dietary Supplements Compendium Attention is called to the fact that USP Dietary Supplements Compendium text is fully copyrighted. Authors and others wishing to use portions of the text should request permission to do so from the Legal Department of the United States Pharmacopeial Convention. Copyright © 2015 The United States Pharmacopeial Convention ISBN: 978-1-936424-41-2 12601 Twinbrook Parkway, Rockville, MD 20852 All rights reserved. DSC Contents iii Contents USP Dietary Supplements Compendium Volume 1 Volume 2 Members . v. Preface . v Mission and Preface . 1 Dietary Supplements Admission Evaluations . 1. General Notices and Requirements . 9 USP Dietary Supplement Verification Program . .205 USP–NF General Chapters . 25 Dietary Supplements Regulatory USP–NF Dietary Supplement Monographs .
    [Show full text]
  • L-Cycloserine Amplifies Anti-Tumor Activity of Glutamine Antagonist
    J. Clin. Biochem. Nutr., 14, 53-60, 1993 L-Cycloserine Amplifies Anti-Tumor Activity of Glutamine Antagonist Akira OKADA, Hiroo TAKEHARA, Kanehiro YOSHIDA, Masaharu NISHI, Hidenori MIYAKE, and Nobuhiko KOMI* The First Department of Surgery, School of Medicine, The University of Tokushima, Tokushima 770, Japan (Received September 22, 1992) Summary We examined the effect of simultaneous treatment with 6-diazo-5-oxo-L-norleucine (DON), a glutamine antagonist, and L- cycloserine, a transaminase inhibitor, on Yoshida sarcoma-bearing rats, with the aim of achieving inhibition of both glutamine catabolism and glutamate synthesis. L-Cycloserine, in a dosage of 1-30 mg/kg, was intraperitoneally administered to Yoshida sarcoma-bearing rats, but the administration of L-cycloserine alone did not show any significant effect on the tumor weight. However, combined treatment with DON (1 mg/kg) caused a dose-dependent reduction in the tumor weight. Amino acid analysis of the tumor tissue, performed 24 h after these treatments in order to confirm their influences on amino acid metabolism in the tumor, showed that the increased glutamate level seen after treatment with DON in a previous study was suppressed by simultaneous administration of DON and 10 mg/kg of L-cycloserine. These results suggest that pooled glutamate in the tumor tissue plays some role in resistance to the tumor- reducing activity of this glutamine antagonist. This, in turn, suggests that concurrent inhibition of glutamate production and blockage of glutamine utilization might result in enhanced tumor reduction in a clinical study. Key Words: glutamine antagonist, transaminase inhibitor, glutamate, Yoshida sarcoma, tumor reduction The glutamine antagonist 6-diazo-5-oxo-L-norleucine (DON) evoked tumor reduction in experimental tumors [1-3], but it did not show adequate efficacy in clinical studies [4, 5].
    [Show full text]
  • Selective Toxicity of Rhodamine 123 in Carcinoma Cells in Vitro
    [CANCER RESEARCH 43, 716-720, February 1983] 0008-5472/83/0043-0000502.00 Selective Toxicity of Rhodamine 123 in Carcinoma Cells in Vitro Theodore J. Lampidis, 1 Samuel D. Bernal, 2 lan C. Summerhayes, and Lan Bo Chen a Sidney Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115 ABSTRACT ing appears to be unique, since chemotherapeutic agents currently in clinical use have not been shown to be particularly The study of mitochondria in situ has recently been facilitated selective between tumorigenic and normal cells in vitro. through the use of rhodamine 123, a mitochondrial-specific fluorescent dye. It has been found to be nontoxic when applied MATERIALS AND METHODS for short periods to a variety of cell types and has thus become an invaluable tool for examining mitochondrial morphology and Cell Cultures. All cell types and cell lines were grown in Dulbecco's function in the intact living cell. In this report, however, we modified Eagle's medium supplemented with 10% calf serum (M. A. demonstrate that with continuous exposure, rhodamine 123 Bioproducts, Walkersville, Md.) at 37 ~ with 5% CO2. The following cell selectively kills carcinoma as compared to normal epithelial lines were obtained from the American Type Culture Collection: cells grown in vitro. At doses of rhodamine 123 which were CCL 105, CCL 15, CRL 1550, CRL 1420, CCL51, CCL77, CCL185, toxic to carcinoma cells, the conversion of mitochondrial-spe- CCL 34, PtK-1, CV-1, and BSC-I. The human bladder carcinoma line cific to cytoplasmic-nonspecific localization of the drug was (E J) was provided by Dr.
    [Show full text]
  • Biological Stains & Dyes
    BIOLOGICAL STAINS & DYES Developed for Biology, microbiology & industrial applications ACRIFLAVIN ALCIAN BLUE 8GX ACRIDINE ORANGE ALIZARINE CYANINE GREEN ANILINE BLUE (SPIRIT SOLUBLE) www.lobachemie.com BIOLOGICAL STAINS & DYES Staining is an important technique used in microscopy to enhance contrast in the microscopic image. Stains and dyes are frequently used in biology and medicine to highlight structures in biological tissues. Loba Chemie offers comprehensive range of Stains and dyes, which are frequently used in Microbiology, Hematology, Histology, Cytology, Protein and DNA Staining after Electrophoresis and Fluorescence Microscopy etc. Many of our stains and dyes have specifications complying certified grade of Biological Stain Commission, and suitable for biological research. Stringent testing on all batches is performed to ensure consistency and satisfy necessary specification particularly in challenging work such as histology and molecular biology. Stains and dyes offer by Loba chemie includes Dry – powder form Stains and dyes as well as wet - ready to use solutions. Features: • Ideally suited to molecular biology or microbiology applications • Available in a wide range of innovative chemical packaging options. Range of Biological Stains & Dyes Product Code Product Name C.I. No CAS No 00590 ACRIDINE ORANGE 46005 10127-02-3 00600 ACRIFLAVIN 46000 8063-24-9 00830 ALCIAN BLUE 8GX 74240 33864-99-2 00840 ALIZARINE AR 58000 72-48-0 00852 ALIZARINE CYANINE GREEN 61570 4403-90-1 00980 AMARANTH 16185 915-67-3 01010 AMIDO BLACK 10B 20470
    [Show full text]
  • Cycloserine and Threo-Dihydrosphingosine Inhibit TNF-A-Induced Cytotoxicity: Evidence for the Importance of De Novo Ceramide Synthesis in TNF-A Signaling
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Biochimica et Biophysica Acta 1643 (2003) 1–4 www.bba-direct.com Rapid report Cycloserine and threo-dihydrosphingosine inhibit TNF-a-induced cytotoxicity: evidence for the importance of de novo ceramide synthesis in TNF-a signaling Sybille G. E. Meyer*, Herbert de Groot Institut fu¨r Physiologische Chemie, Universita¨tsklinikum Essen, Hufelandstrasse 55, D-45147 Essen, Germany Received 2 July 2003; accepted 9 October 2003 Abstract Measuring the cell death induced by tumor necrosis factor (TNF-a) in L929 cells, we discovered for the first time that L-cycloserine, an established inhibitor of serine palmitoyltransferase, as well as DL-threo-dihydrosphingosine (threo-DHS, threo-sphinganine) significantly protected against TNF-a-induced cytotoxicity. Under the same conditions sphingosine and DL-erythro-dihydrosphingosine (erythro-DHS) did not change TNF-a-induced cytotoxicity, thus underlining the specificity of threo-DHS. In serine-labeled cells, newly (de novo) synthetized labeled ceramide was significantly diminished by threo-DHS alone or together with TNF-a, which makes the (dihydro) ceramide synthase the likely target of threo-DHS. These results suggest the decisive role of ceramide de novo synthesis in TNF signaling. D 2003 Elsevier B.V. All rights reserved. Keywords: Ceramide de novo synthesis; Safingol; Sphinganine; Sphingosine; Threo-dihydrosphingosine; Tumor necrosis factor-a (TNF-a) Tumor necrosis factor-a (TNF-a) is a pleiotropic cyto- ceramide was only shown in TNF/cycloheximide-induced kine produced mainly by macrophages. It mediates inflam- cell death in cerebral endothelial cells using Fumonisin B1 mation and regulates immune function by triggering the [9].
    [Show full text]
  • The Antiviral Effect of Epigallocatechin Gallate-Stearate on Enterovirus-69" (2017)
    Montclair State University Montclair State University Digital Commons Theses, Dissertations and Culminating Projects 8-2017 The Antiviral Effect of Epigallocatechin Gallate- Stearate on Enterovirus-69 Hager Mohamed Montclair State University Follow this and additional works at: https://digitalcommons.montclair.edu/etd Part of the Biology Commons Recommended Citation Mohamed, Hager, "The Antiviral Effect of Epigallocatechin Gallate-Stearate on Enterovirus-69" (2017). Theses, Dissertations and Culminating Projects. 16. https://digitalcommons.montclair.edu/etd/16 This Thesis is brought to you for free and open access by Montclair State University Digital Commons. It has been accepted for inclusion in Theses, Dissertations and Culminating Projects by an authorized administrator of Montclair State University Digital Commons. For more information, please contact [email protected]. Abstract Enteroviruses are single stranded RNA viruses responsible for diseases from respiratory illness to hand-foot-and-mouth disease. As a rapidly evolving group, enteroviruses often gain genetic variability which leads to emergence of additional types, one of which is Enterovirus 69 (EV-69). EV-69 has been associated with respiratory illness, possibly causing symptoms similar to those caused by EV-68. To challenge the ability of the virus to infect cells, this study examined the result of infection in lung cell lines A549 and MRC-5 in response to Epigallocatechin gallate-stearate (EGCG-S) treatment of the virus before infection. EGCG-S, a modified green tea polyphenol, was assessed for its efficacy against EV-69 through examination of changes in cell viability, cytopathic effects, cell proliferation, and reactive oxygen species levels. Apoptosis in treated versus untreated virus infected controls was also assessed.
    [Show full text]
  • PINK1-Dependent Mitophagy Is Driven by the UPS and Can Occur Independently of LC3 Conversion
    Cell Death & Differentiation https://doi.org/10.1038/s41418-018-0219-z ARTICLE PINK1-dependent mitophagy is driven by the UPS and can occur independently of LC3 conversion 1 1 1,2 1 1 Aleksandar Rakovic ● Jonathan Ziegler ● Christoph U. Mårtensson ● Jannik Prasuhn ● Katharina Shurkewitsch ● 3 4 1 Peter König ● Henry L. Paulson ● Christine Klein Received: 1 February 2017 / Revised: 17 September 2018 / Accepted: 2 October 2018 © The Author(s) 2018 Abstract The Parkinson’s disease (PD)-related ubiquitin ligase Parkin and mitochondrial kinase PINK1 function together in the clearance of damaged mitochondria. Upon mitochondrial depolarization, Parkin translocates to mitochondria in a PINK1-dependent manner to ubiquitinate outer mitochondrial membrane proteins. According to the current model, the ubiquitin- and LC3-binding adaptor protein SQSTM1 is recruited to mitochondria, followed by their selective degradation through autophagy (mitophagy). However, the role of the ubiquitin proteasome system (UPS), although essential for this process, still remains largely elusive. Here, we investigated the role of the UPS and autophagy by applying the potassium fi fi 1234567890();,: 1234567890();,: ionophore Valinomycin in PINK1-de cient human broblasts and isogenic neuroblastoma cell lines generated by CRISPR/ Cas9. Although identical to the commonly used CCCP/FCCP in terms of dissipating the mitochondrial membrane potential and triggering complete removal of mitochondria, Valinomycin did not induce conversion of LC3 to its autophagy-related form. Moreover, FCCP-induced conversion of LC3 occurred even in mitophagy-incompetent, PINK1-deficient cell lines. While both stressors required a functional UPS, the removal of depolarized mitochondria persisted in cells depleted of LC3A and LC3B. Our study highlights the importance of the UPS in PINK1-/Parkin-mediated mitochondrial quality control.
    [Show full text]
  • Bulk Drug Substances Nominated for Use in Compounding Under Section 503B of the Federal Food, Drug, and Cosmetic Act
    Updated June 07, 2021 Bulk Drug Substances Nominated for Use in Compounding Under Section 503B of the Federal Food, Drug, and Cosmetic Act Three categories of bulk drug substances: • Category 1: Bulk Drug Substances Under Evaluation • Category 2: Bulk Drug Substances that Raise Significant Safety Risks • Category 3: Bulk Drug Substances Nominated Without Adequate Support Updates to Categories of Substances Nominated for the 503B Bulk Drug Substances List1 • Add the following entry to category 2 due to serious safety concerns of mutagenicity, cytotoxicity, and possible carcinogenicity when quinacrine hydrochloride is used for intrauterine administration for non- surgical female sterilization: 2,3 o Quinacrine Hydrochloride for intrauterine administration • Revision to category 1 for clarity: o Modify the entry for “Quinacrine Hydrochloride” to “Quinacrine Hydrochloride (except for intrauterine administration).” • Revision to category 1 to correct a substance name error: o Correct the error in the substance name “DHEA (dehydroepiandosterone)” to “DHEA (dehydroepiandrosterone).” 1 For the purposes of the substance names in the categories, hydrated forms of the substance are included in the scope of the substance name. 2 Quinacrine HCl was previously reviewed in 2016 as part of FDA’s consideration of this bulk drug substance for inclusion on the 503A Bulks List. As part of this review, the Division of Bone, Reproductive and Urologic Products (DBRUP), now the Division of Urology, Obstetrics and Gynecology (DUOG), evaluated the nomination of quinacrine for intrauterine administration for non-surgical female sterilization and recommended that quinacrine should not be included on the 503A Bulks List for this use. This recommendation was based on the lack of information on efficacy comparable to other available methods of female sterilization and serious safety concerns of mutagenicity, cytotoxicity and possible carcinogenicity in use of quinacrine for this indication and route of administration.
    [Show full text]
  • "Assessment of Cell Viability". In: Current Protocols in Cytometry
    Assessment of Cell Viability UNIT 9.2 Simon Johnson,1 Vy Nguyen,1 and David Coder2 1University of Washington School of Medicine, Seattle, Washington 2Baybioscience, Irvine, California ABSTRACT Cell viability may be judged by morphological changes or by changes in membrane per- meability and/or physiological state inferred from the exclusion of certain dyes or the up- take and retention of others. This unit presents methods based on dye exclusion, esterase activity, and mitochondrial membrane potential, as well as protocols for determining the pre-fixation viability of fixed cells either before or after fixation with amine-reactive dyes suitable for a range of excitation wavelengths. Membrane-impermeable dead cell and live cell dyes as well as dye-exclusion procedures for microscopy are also included. Curr. Protoc. Cytom. 64:9.2.1-9.2.26. C 2013 by John Wiley & Sons, Inc. Keywords: cytometry r flow cytometry r imaging r probes r toxicology r assessment of cell toxicity INTRODUCTION The method used to determine cell viability (and to a degree, the definition of viability) is often related to the phenomenon studied. Frequently, cell viability is thought of in somewhat negative terms. That is, one needs to exclude dead cells because they generate artifacts as a result of nonspecific binding and/or uptake of fluorescent probes. In addition to simple enumeration of live or dead cells present, there is a broad range of biologically relevant cytometric procedures that are related to the physiological state of the cells measured. For example, one may wish to measure cell morbidity in apoptosis, cell survival as a result of cytotoxicity, the potential of bacteria or microalgae to survive environmental insult and grow once normal conditions have been restored, or the potential of intracellular protozoa to undergo division cycles as part of the infective process.
    [Show full text]
  • Using Medications, Cosmetics, Or Eating Certain Foods Can Increase Sensitivity to Ultraviolet Radiation
    USING MEDICATIONS, COSMETICS, OR EATING CERTAIN FOODS CAN INCREASE SENSITIVITY TO ULTRAVIOLET RADIATION. INDIVIDUALS SHOULD CONSULT A PHYSICIAN BEFORE USING A SUNLAMP, IF THEY ARE TAKING MEDICATIONS. THE ITEMS LISTED ARE POTENTIAL PHOTOSENSITIZING AGENTS THAT MAY INCREASE SENSITIVITY TO ULTRAVIOLET LIGHT THAT MAY RESULT IN A PHOTOTOXIC OR PHOTOALLERGIC RESPONSES. PHOTOSENSITIZING MEDICATIONS Acetazolamide Amiloride+Hydrochlorothizide Amiodarone Amitriptyline Amoxapine Astemizole Atenolol+Chlorthalidone Auranofin Azatadine (Optimine) Azatidine+Pseudoephedrine Bendroflumethiazide Benzthiazide Bromodiphenhydramine Bromopheniramine Captopril Captopril+Hydrochlorothiazide Carbaamazepine Chlordiazepoxide+Amitriptyline Chlorothiazide Chlorpheniramine Chlorpheniramin+DPseudoephedrine Chlorpromazine Chlorpheniramine+Phenylopropanolamine Chlorpropamide Chlorprothixene Chlorthalidone Chlorthalidone+Reserpine Ciprofloxacin Clemastine Clofazime ClonidineChlorthalisone+Coal Tar Coal Tar Contraceptive (oral) Cyclobenzaprine Cyproheptadine Dacarcazine Danazol Demeclocycline Desipramine Dexchlorpheniramine Diclofenac Diflunisal Ditiazem Diphenhydramine Diphenylpyraline Doxepin Doxycycline Doxycycline Hyclate Enalapril Enalapril+Hydrochlorothiazide Erythromycin Ethylsuccinate+Sulfisoxazole Estrogens Estrogens Ethionamide Etretinate Floxuridine Flucytosine Fluorouracil Fluphenazine Flubiprofen Flutamide Gentamicin Glipizide Glyburide Gold Salts (compounds) Gold Sodium Thiomalate Griseofulvin Griseofulvin Ultramicrosize Griseofulvin+Hydrochlorothiazide Haloperidol
    [Show full text]
  • Vitreoretinal Staining Solutions Part Two of a Three-Part Series in Which Surgeons Share Pointers on the Optimal Use of Vital Dyes in Retina Surgery
    RETINA SURGERY FEATURE STORY Vitreoretinal Staining Solutions Part two of a three-part series in which surgeons share pointers on the optimal use of vital dyes in retina surgery. Considerations in Vital Dye orange-red color of the fundus background, a blue or green dye provides the best color contrast on the Staining During Retinal Surgery retinal surface. By Timothy L. Jackson, PhD, FRCOphth The relationship between dye concentration and tissue staining varies among dyes. Image analysis from he use of intraocular vital dyes to enhance one ex vivo study showed that staining increased lin- tissue visibility during retinal surgery has the early with dye concentration for some dyes, whereas potential to improve surgical performance others displayed an exponential relationship.1 Given and reduce risk. The ideal retinal vital dye that dyes are variably diluted during application in a forT selective membrane staining during vitreoretinal saline-filled eye, a linear relationship has advantages, surgery should be water soluble, reasonably priced, making staining more predictable. However, some and provide good color contrast, effective staining, dyes provided uneven tissue staining despite a con- and safety. Vital dyes should also offer repeatable, avid stant concentration. Agents shown to have favorable staining, yet be easily removable. Internationally, several staining characteristics were Evans blue, rose bengal, ophthalmic vital dyes are now available, although only naphthol green, natural red, and trypan blue. Some of one formulation is currently approved for ophthalmic these dyes (eg, rose bengal) may not have a suitable use by the US Food and Drug Administration (FDA). retinal safety profile.1 Characteristics of several vital dyes are discussed below.
    [Show full text]
  • Evans Blue Dye : a Revisit of Its Applications in Biomedicine
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Lirias Evans Blue Dye: A Revisit of Its Applications in Biomedicine Linpeng Yao1, Xing Xue1, Peipei Yu1,2, Yicheng Ni3, Feng Chen1* 1: Department of Radiology, The first affiliated hospital, Zhejiang University School of Medicine, Hangzhou, 79 Qingchun Road, 310003, Zhejiang Province, China 2: Department of Radiology, Sanmen County People’s Hospital, Sanmen, 317100, Zhejiang Province, China 3: Radiology Section, University Hospitals, University of Leuven, Leuven, 3000, Belgium *Correspondence to: Feng Chen, MD, PhD Director, Department of Radiology, The first affiliated hospital, Zhejiang University School of Medicine, Hangzhou, 79 Qingchun Road, 310003, Zhejiang Province, China Email: [email protected] Tel: +86 137 3558 9621 Keywords: Evans blue, necrosis, liposomal nanocarrier, theranostics, derivatives, enhanced permeability and retention (EPR) effect 1 Abstract Evans blue (EB) dye has owned a long history as a biological dye and diagnostic agent since its first staining application by Herbert McLean Evans in 1914. Due to its high water solubility and slow excretion, as well as its tightly binding to serum albumin, EB has been widely used in biomedicine, including its use in estimating blood volume and vascular permeability, detecting lymph nodes, and localizing the tumor lesions. Recently, a series of EB derivatives have been labeled with PET isotopes and can be used as theranostics with a broad potential due to their improved half-life in the blood and reduced release. Some of EB derivatives have even been used in translational applications in clinics. In addition, a novel necrosis-avid feature of EB has recently been reported in some preclinical animal studies.
    [Show full text]