scholarly journals A New Model of SARS-CoV-2 Infection Based on (Hydroxy)Chloroquine Activity

2020 ◽  
Author(s):  
Robert J. Sheaff
Keyword(s):  
Oxidative Phosphorylation ◽  
Host Cell ◽  
Disease Progression ◽  
Therapeutic Benefit ◽  
Beta Oxidation ◽  
New Model ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Obesity And Diabetes ◽  
Infected Cells

Chloroquine and hydroxychloroquine [(H)CQ] are well known anti-malarial drugs, while their use against COVID-19 is more controversial. (H)CQ activity was examined in tissue culture cells to determine if their anti-viral benefits or adverse effects might be due to altering host cell pathways. Metabolic analysis revealed (H)CQ inhibit oxidative phosphorylation in mitochondria, likely by sequestering protons needed to drive ATP synthase. This activity could cause cardiotoxicity because heart muscle relies on beta oxidation of fatty acids. However, it might also explain their therapeutic benefit against COVID-19. A new model of SARS-CoV-2 infection postulates virus enters host cell mitochondria and uses its protons for genome release. Oxidative phosphorylation is eventually compromised, so glycolysis is upregulated to maintain ATP levels. (H)CQ could prevent viral infection and/or slow its replication by sequestering these protons. In support of this model other potential COVID-19 therapeutics also targeted mitochondria, as did tobacco smoke, which may underlie smokers protection. The mitochondria of young people are naturally more adaptable and resilient, providing a rationale for their resistance to disease progression. Conversely, obesity and diabetes could exacerbate disease severity by providing extra glucose to infected cells dependent on glycolysis. The description of (H)CQ function presented here, together with its implications for understanding SARS-CO-V2 infection, makes testable predictions about disease progression and identifies new approaches for treating COVID-19.

scholarly journals IMMUNOFLUORESCENT STUDIES OF HUMAN CELL CULTURES AND CHICK EMBRYOS INOCULATED WITH THE AMEBOID CELL-"LIPOVIRUS" COMPLEX

1966 ◽  
Vol 124 (6) ◽  
pp. 1167-1180 ◽  
Author(s):  
Chien Liu ◽  
Patricia Rodina
Keyword(s):  
Phase Contrast ◽  
Early Stage ◽  
Immunofluorescent Staining ◽  
Chick Embryos ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Human Cell Cultures ◽  
Contrast Microscopy ◽  
Infected Cells ◽  
Ameboid Cell

Tissue culture cells of human origin (HeLa, Lich, and AV3) were inoculated with the AL complex. By immunofluorescent staining, AL complex antigen was detectable in the cytoplasm of infected cells as punctate fluorescent granules during the early stage and as homogeneous fluorescence during the late stage of infection. By combining fluorescence and phase-contrast microscopy, many infected cells with cytoplasmic AL complex antigen were shown to have a normal nuclear morphology indistinguishable from uninfected cells. Initiation of AL complex infection was interpreted as occurring by transfer of transmissible factor(s) from cell to cell by contact and mutiplication of such factor(s) therein. Chick embryos were susceptible to AL complex infection following allantoic or amniotic inoculations. Antigen and infectious AL complex were demonstrable in the liver, brain, intestines, lungs, and embryonic membranes. Further investigations on AL complex and its relation to human disease are suggested.

scholarly journals Overexpression of the Rabies Virus Glycoprotein Results in Enhancement of Apoptosis and Antiviral Immune Response

2002 ◽  
Vol 76 (7) ◽  
pp. 3374-3381 ◽  
Author(s):  
Milosz Faber ◽  
Rojjanaporn Pulmanausahakul ◽  
Suchita S. Hodawadekar ◽  
Sergei Spitsin ◽  
James P. McGettigan ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Caspase 3 ◽  
Marked Decrease ◽  
Fluorescence Staining ◽  
Antiviral Immune Response ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Glycoprotein G ◽  
Antibody Titers ◽  
Infected Cells

ABSTRACT A recombinant rabies virus (RV) carrying two identical glycoprotein (G) genes (SPBNGA-GA) was constructed and used to determine the effect of RV G overexpression on cell viability and immunity. Immunoprecipitation analysis and flow cytometry showed that tissue culture cells infected with SPBNGA-GA produced, on average, twice as much RV G as cells infected with RV carrying only a single RV G gene (SPBNGA). The overexpression of RV G in SPBNGA-GA-infected NA cells was paralleled by a significant increase in caspase 3 activity followed by a marked decrease in mitochondrial respiration, neither of which was observed in SPBNGA-infected cells. Furthermore, fluorescence staining and confocal microscopy revealed an increased extent of apoptosis and markedly reduced neurofilament and F actin in SPBNGA-GA-infected primary neuron cultures compared with neuronal cells infected with SPBNGA, supporting the concept that RV G or motifs of the RV G gene trigger the apoptosis cascade. Mice immunized with SPBNGA-GA showed substantially higher antibody titers against the RV G and against the nucleoprotein than SPBNGA-immunized mice, suggesting that the speed or extent of apoptosis directly determines the magnitude of the antibody response.

scholarly journals Junctional Adhesion Molecule 1 Is a Functional Receptor for Feline Calicivirus

2006 ◽  
Vol 80 (9) ◽  
pp. 4482-4490 ◽  
Author(s):  
Akiko Makino ◽  
Masayuki Shimojima ◽  
Takayuki Miyazawa ◽  
Kentaro Kato ◽  
Yukinobu Tohya ◽  
...  
Keyword(s):  
Adhesion Molecule ◽  
Vero Cells ◽  
Prototype Strain ◽  
Feline Calicivirus ◽  
Progeny Production ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
The Family ◽  
Infected Cells ◽  
Functional Receptor

ABSTRACT The life cycle of calicivirus is not fully understood because most of the viruses cannot be propagated in tissue culture cells. We studied the mechanism of calicivirus entry into cells using feline calicivirus (FCV), a cultivable calicivirus. From the cDNA library of Crandell-Rees feline kidney (CRFK) cells, feline junctional adhesion molecule 1 (JAM-1), an immunoglobulin-like protein present in tight junctions, was identified as a cellular-binding molecule of the FCV F4 strain, a prototype strain in Japan. Feline JAM-1 expression in nonpermissive hamster lung cells led to binding and infection by F4 and all other strains tested. An anti-feline JAM-1 antibody reduced the binding of FCV to permissive CRFK cells and strongly suppressed the cytopathic effect (CPE) and FCV progeny production in infected cells. Some strains of FCV, such as F4 and F25, have the ability to replicate in Vero cells. We found that regardless of replication ability, FCV bound to Vero and 293T cells via simian and human JAM-1, respectively. In Vero cells, an anti-human JAM-1 antibody inhibited binding, CPE, and progeny production by F4 and F25. In addition, feline JAM-1 expression permitted FCV infection in 293T cells. Taken together, our results demonstrate that feline JAM-1 is a functional receptor for FCV, simian JAM-1 also functions as a receptor for some strains of FCV, and the interaction between FCV and JAM-1 molecules may be a determinant of viral tropism. This is the first report concerning a functional receptor for the viruses in the family Caliciviridae.

Respiration of L cells infected with Chlamydia psittaci

1970 ◽  
Vol 16 (11) ◽  
pp. 1033-1039 ◽  
Author(s):  
Shawn D'arcy Gill ◽  
Robert B. Stewart
Keyword(s):  
Tissue Culture ◽  
Oxidative Phosphorylation ◽  
Host Cell ◽  
Respiratory Quotient ◽  
Culture Medium ◽  
Mouse Fibroblast ◽  
Chlamydia Psittaci ◽  
Concomitant Increase ◽  
L Cells ◽  
Infected Cells

Strain 929 mouse fibroblast L cells demonstrated an increased rate of endogenous and exogenous respiration when infected by the 6BC strain of the psittacosis organism. The respiratory quotient (R.Q.) values for infected cultures were about 1.0. Studies involving the use of 2,4-DNPand reduced phosphate suggested that the accelerated respiration was a result of increased oxidative phosphorylation. Infection was accompanied by a decline in host cell ATP levels with a concomitant increase in the ADP levels. A reduction in tissue culture medium phosphate resulted in a reduced yield of psittacosis organisms from infected cells and lower cellular levels of ATP. These results are discussed in relation to an energy parasite hypothesis.

Freeze-Etching of Intranuclear Adenovirus

1971 ◽  
Vol 29 ◽  
pp. 384-385
Author(s):  
Dennis T. Brown ◽  
Byron T. Burlingham
Keyword(s):  
Osmium Tetroxide ◽  
Adenovirus Type ◽  
Etching Time ◽  
Human Carcinoma ◽  
Thin Sections ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Infected Cells ◽  
Freeze Etching

Adenoviruses are revealed by negative staining to be icosahedral in structure and composed of 252 subunits (insert Fig. 1). The vertices have a fiber-like projection which has been implicated in the adsorbtion of the virus to the host cell. Ultra thin sections of adenovirus-infected cells has revealed that virus morphogenesis occurs in the nucleus.Human carcinoma (KB) tissue culture cells were productively infected with adenovirus type 2 at a multiplicity of 10 and incubated for 40 hours at 37°C. The infected monolayers were fixed in place with 1%, gluteraldehyde buffered in Millonigs phosphate buffer. The fixed cells were suspended in 30%, glycerol and processed according to the procedure of Moor in a Balzers 360 M freeze etch unit with an etching time of 70 seconds at -100°C. A second portion of the fixed cells in glycerol was post-fixed with 1% osmium tetroxide and embeded in epon 812 for ultrathin sectioning.

scholarly journals An Alternative Efficient Procedure for Purification of the Obligate Intracellular Fish Bacterial Pathogen Piscirickettsia salmonis

2003 ◽  
Vol 69 (10) ◽  
pp. 6268-6271 ◽  
Author(s):  
Vitalia Henríquez ◽  
María Verónica Rojas ◽  
Sergio H. Marshall
Keyword(s):  
Tissue Culture ◽  
Host Cell ◽  
Dna Analysis ◽  
Bacterial Pathogen ◽  
Obligate Intracellular ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Piscirickettsia Salmonis ◽  
Chinook Salmon Embryo

ABSTRACT Piscirickettsia salmonis is an obligate intracellular bacterial pathogen of salmonid fish and the etiological agent of the aggressive disease salmonid rickettsial syndrome. Today, this disease, also known as piscirickettsiosis, is the cause of high mortality in net pen-reared salmonids in southern Chile. Although the bacteria can be grown in tissue culture cells, genetic analysis of the organism has been hindered because of the difficulty in obtaining P. salmonis DNA free from contaminating host cell DNA. In this report, we describe a novel procedure to purify in vitro-grown bacteria with iodixanol as the substrate to run differential centrifugation gradients which, combined with DNase I digestion, yield enough pure bacteria to do DNA analysis. The efficiency of the purification procedure relies on two main issues: semiquantitative synchrony of the P. salmonis-infected Chinook salmon embryo (CHSE-214) tissue culture cells and low osmolarity of iodixanol to better resolve bacteria from the membranous structures of the host cell. This method resulted in the isolation of intact piscirickettsia organisms and removed salmon and mitochondrial DNA effectively, with only 1.0% contamination with the latter.

scholarly journals Examination of Listeria monocytogenes Intracellular Gene Expression by Using the Green Fluorescent Protein ofAequorea victoria

1999 ◽  
Vol 67 (4) ◽  
pp. 1844-1852 ◽  
Author(s):  
Nancy E. Freitag ◽  
Kathleen E. Jacobs
Keyword(s):  
Gene Expression ◽  
Tissue Culture ◽  
Green Fluorescent Protein ◽  
Listeria Monocytogenes ◽  
Host Cell ◽  
Fluorescent Protein ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Green Fluorescent ◽  
Cell Spread

ABSTRACT The ActA protein of Listeria monocytogenes is an essential virulence factor and is required for intracellular bacterial motility and cell-to-cell spread. plcB, cotranscribed withactA, encodes a broad-specificity phospholipase C that contributes to lysis of host cell vacuoles and cell-to-cell spread. Construction of a transcriptional fusion between actA-plcBand the green fluorescent protein gene of Aequorea victoriahas facilitated the detailed examination of patterns ofactA/plcB expression within infected tissue culture cells.actA/plcB expression began approximately 30 min postinfection and was dependent upon entry of L. monocytogenes into the host cytosol. L. monocytogenes Δhly mutants, which are unable to escape from host cell vacuoles, did not express actA/plcB at detectable levels within infected tissue culture cells; however, complementation of thehly defect allowed entry of the bacteria into the host cytoplasm and subsequent actA/plcB expression. These results emphasize the ability of L. monocytogenes to sense the different host cell compartment environments encountered during the course of infection and to regulate virulence gene expression in response.

Cytoplasmic Tubules in Cells Infected with Laryngotracheitis Virus

1969 ◽  
Vol 27 ◽  
pp. 376-377
Author(s):  
A. M. Watrach
Keyword(s):  
Tissue Culture ◽  
Small Proportion ◽  
Chicken Embryo ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Morphologic Characteristics ◽  
Infectious Laryngotracheitis ◽  
Laryngotracheitis Virus ◽  
In Cells

During a study of the development of infectious laryngotracheitis (LT) virus in tissue culture cells, unusual tubular formations were found in the cytoplasm of a small proportion of the affected cells. It is the purpose of this report to describe the morphologic characteristics of the tubules and to discuss their possible association with the development of virus.The source and maintenance of the strain of LT virus have been described. Prior to this study, the virus was passed several times in chicken embryo kidney (CEK) tissue culture cells.

Studies of a Defective Measles Virus

1968 ◽  
Vol 26 ◽  
pp. 208-209
Author(s):  
R. M. McCombs ◽  
M. Benyesh-Melnick ◽  
J. P. Brunschwig
Keyword(s):  
Inclusion Bodies ◽  
Measles Virus ◽  
Giant Cells ◽  
Helical Structures ◽  
Thin Sections ◽  
Virus Particles ◽  
Extracellular Virus ◽  
Culture Cells ◽  
Infected Cells ◽  
Eosinophilic Inclusions

Measles virus is an agent that is capable of replicating in a number of different culture cells and generally causes the formation of multinucleated giant cells. As a result of infection, virus is released from the cells into the culture fluids and reinfection can be initiated by this cell-free virus. The extracellular virus has been examined by negative staining with phosphotungstic acid and has been shown to be a rather pleomorphic particle with a diameter of about 140 mμ. However, no such virus particles have been detected in thin sections of the infected cells. Rather, the only virus-induced structures present in the giant cells are eosinophilic inclusions (intracytoplasmic or intranuclear). These inclusion bodies have been shown to contain helical structures, resembling the nucleocapsid observed in negatively stained preparations.

Sulfhydryl bond formation is a prerequisite for proper cycling of centrosomes and chromosomes in mammalian tissue culture cells

1993 ◽  
Vol 51 ◽  
pp. 342-343
Author(s):  
Heide Schatten ◽  
Neidhard Paweletz ◽  
Ron Balczon
Keyword(s):  
Tissue Culture ◽  
Cell Cycle Progression ◽  
Group Formation ◽  
Sulfhydryl Group ◽  
Mammalian Tissue ◽  
Mitotic Chromosomes ◽  
Microtubule Network ◽  
Culture Cells ◽  
Tissue Culture Cells ◽  
Transmission Electron

To study the role of sulfhydryl group formation during cell cycle progression, mammalian tissue culture cells (PTK2) were exposed to 100¼M 2-mercaptoethanol for 2 to 6 h during their exponential phase of growth. The effects of 2-mercaptoethanol on centrosomes, chromosomes, microtubules, membranes and intermediate filaments were analyzed by transmission electron microscopy (TEM) and by immunofluorescence microscopy (IFM) methods using a human autoimmune antibody directed against centrosomes (SPJ), and a mouse monoclonal antibody directed against tubulin (E7). Chromosomes were affected most by this treatment: premature chromosome condensation was detected in interphase nuclei, and the structure in mitotic chromosomes was altered compared to control cells. This would support previous findings in dividing sea urchin cells in which chromosomes are arrested at metaphase while the centrosome splitting cycle continues. It might also support findings that certairt-sulfhydryl-blocking agents block cyclin destruction. The organization of the microtubule network was scattered probably due to a looser organization of centrosomal material at the interphase centers and at the mitotic poles.

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