scholarly journals A CYTOCHEMICAL DESCRIPTION OF THE MULTIPLICATION OF MENGOVIRUS IN L-929 CELLS

1962 ◽  
Vol 12 (1) ◽  
pp. 1-15 ◽  
Author(s):  
Richard M. Franklin
Keyword(s):  
Phase Contrast ◽  
Basic Protein ◽  
Virus Production ◽  
Cytological Change ◽  
Virus Particles ◽  
Contrast Microscopy ◽  
Infected Cells ◽  
Cytological Changes ◽  
Perinuclear Area

A correlation of cytochemical changes with virus production has been studied in L cells infected with Mengovirus. After a latent period of about 2 hours, virus was produced rapidly, reaching maximum titers of up to 12,000 particles per cell in 6 to 8 hours. The earliest cytological change was in the nucleus and consisted of a slight condensation of chromatin. There is no evidence, however, for the multiplication of either the viral RNA or protein in the nucleus. RNA, of high molecular weight, accumulated in the perinuclear area of the cytoplasm and was later found in inclusions. The perinuclear RNA was digestible with RNase and may be located in or on ribosomes. The inclusion RNA was resistant to RNase but could be removed by pepsin or potassium permanganate; it is probably in completed virus particles. Viral antigen was first observed in a perinuclear location and later in the above-mentioned inclusions. Although the viral protein contains appreciable amounts of arginine and lysine, it is not a basic protein of the histone type. Phase-contrast microscopy of living cells clearly demonstrated the role of the inclusions in release of virus from infected cells. A comparison is made between these cytological changes in Mengo-infected cells and those which have been found by other workers in polio-infected cells. There are many very similar changes.

scholarly journals Rough Dental Implant Surfaces and Peri-Implantitis: Role of Phase-Contrast Microscopy, Laser Protocols, and Modified Home Oral Hygiene in Maintenance. A 10-Year Retrospective Study

2021 ◽  
Vol 11 (11) ◽  
pp. 4985
Author(s):  
Gianluigi Caccianiga ◽  
Gérard Rey ◽  
Paolo Caccianiga ◽  
Alessandro Leonida ◽  
Marco Baldoni ◽  
...  
Keyword(s):  
Retrospective Study ◽  
Phase Contrast ◽  
Vertical Movement ◽  
Subgingival Plaque ◽  
Phase Contrast Microscopy ◽  
Implant Surface ◽  
Total Percentage ◽  
Contrast Microscopy

The aim of this study was to evaluate two different kinds of rough implant surface and to assess their tendency to peri-implantitis disease, with a follow-up of more than 10 years. Data were obtained from a cluster of 500 implants with Ti-Unite surface and 1000 implants with Ossean surface, with a minimum follow-up of 10 years. Implants had been inserted both in pristine bone and regenerated bone. We registered incidence of peri-implantitis and other causes of implant loss. All patients agreed with the following maintenance protocol: sonic brush with vertical movement (Broxo), interdental brushes, and oral irrigators (Broxo) at least two times every day. For all patients with implants, we evaluated subgingival plaque samples by phase-contrast microscopy every 4 months for a period of more than 10-years. Ti-Unite surface implants underwent peri-implantitis in 1.6% of the total number of implants inserted and Ossean surface implants showed peri-implantitis in 1.5% of the total number of implants. The total percentage of implant lost was 4% for Ti-Unite surfaces and 3.6% for Ossean surfaces. Strict control of implants leads to low percentage of peri-implantitis even for rough surfaces dental implants.

scholarly journals Functional analysis of the putative antiapoptotic genes, p49 and iap4, of Spodoptera litura nucleopolyhedrovirus with RNAi

2008 ◽  
Vol 89 (8) ◽  
pp. 1873-1880 ◽  
Author(s):  
Qian Yu ◽  
Tiehao Lin ◽  
Guozhong Feng ◽  
Kai Yang ◽  
Yi Pang
Keyword(s):  
Spodoptera Litura ◽  
Virus Production ◽  
Host Cells ◽  
Homology Search ◽  
Pcr Analysis ◽  
Viability Analysis ◽  
Infected Cells ◽  
Budded Virus ◽  
Almost All

A homology search of a public database revealed that Spodoptera litura nucleopolyhedrovirus (SpltNPV) possesses two putative, antiapoptotic genes, p49 and inhibitor of apoptosis 4 (iap4), but their function has not been investigated in its native host cells. In the present study, we used RNA interference (RNAi) to silence the expression of Splt-iap4 and Splt-p49, independently or together, to determine their roles during the SpltNPV life cycle. RT-PCR analysis and Western blot analysis showed the target gene expression had been knocked out in the SpltNPV-infected SpLi-221 cells after treatment with Splt-p49 or Splt-iap4 double-stranded RNA (dsRNA), respectively, confirming that the two genes were effectively silenced. In SpltNPV-infected cells treated with Splt-p49 dsRNA, apoptosis was observed beginning at 14 h, and almost all cells had undergone apoptosis by 48 h. In contrast, budded virus production and polyhedra formation progressed normally in infected cells treated with Splt-iap4 dsRNA. Cell viability analysis showed that Splt-IAP4 had no synergistic effect on the inhibition of apoptosis of SpLi-221 cells induced by SpltNPV infection. Interestingly, after Splt-iap4 dsRNA treatment, cells did not congregate like those infected with SpltNPV in the early infection phase, implying an unknown role of baculovirus iap4. Our results determine that Splt-p49 is necessary to prevent apoptosis; however, Splt-iap4 has no antiapoptotic function during SpltNPV infection.

scholarly journals What is in a Viral Stock: Perspectives and Current Limitations of Flow Virometry

2020 ◽  
Author(s):  
Timothy K. Soh ◽  
Jens B. Bosse
Keyword(s):  
Flow Cytometry ◽  
Full Potential ◽  
Multiple Objects ◽  
Virus Particles ◽  
Viral Particles ◽  
Viral Stock ◽  
Infected Cells ◽  
A Minor ◽  
Unique Capability

Herpesviruses produce a plethora of pleomorphic and heterogeneous particle populations. The composition and biological role of these is not understood. Detailed analysis has been challenging due to the lack of multidimensional identification and purification methodologies. Fluorescence-activated cell sorting (FACS), originally developed to sort objects with at least ten thousand-fold larger volumes, has recently been applied to cellular exosomes as well as viral particles and has been dubbed nanoscale flow cytometry or “flow virometry”. In comparison to other nanoparticles, herpesvirus concentrations can be measured with high precision using simple culturing methods. Here, we used this unique capability to evaluate a standard FACS sorter. We demonstrate that detection and separation capabilities were insufficient to distinguish infectious fluorescent viral populations from populations lacking fluorescence and infectivity. Moreover, fluorescent populations did not contain single virus particles but mostly aggregates. On top, analysis of viral samples by flow cytometry was confounded by swarm detection, as multiple objects are measured simultaneously and interpreted as a single object. Despite these technical difficulties, comparison of crude supernatant to gradient purified HCMV revealed that infectious virus is a minor proportion of the particles released from infected cells. Our data stress the need for a set of standardized controls and protocols when applying FACS to biological nanoparticles and highlights technical challenges that need to be solved before flow virometry can achieve its full potential.

Intracellular motility of mitochondria: role of the inner compartment in migration and shape changes of mitochondria in XTH-cells

1978 ◽  
Vol 30 (1) ◽  
pp. 99-115
Author(s):  
J. Bereiter-Hahn
Keyword(s):  
Phase Contrast ◽  
Heart Cells ◽  
Underlying Principle ◽  
Good Accord ◽  
Contrast Microscopy ◽  
Mitochondrial Motility ◽  
Time Required ◽  
Shape Changes ◽  
Intracellular Motility

Mitochondrial movements have been followed by phase-contrast microscopy in living XTH-cells (Xenopus laevis tadpole-heart cells) in tissue culture. The same organelles have been viewed subsequently in electron micrographs. Locomotion of mitochondria proceeds at velocities up to 100 micrometer/min. Formation of branches of mitochondria and other shape changes may occur with the same speed. Mitochondrial motility can be classified into 4 types: (I) Alternating extension and contraction at the two ends of rod-shaped mitochondria. (2) Lateral branching. (3) Alternate stretching and contraction of arbitrary parts of a mitochondrion amounting to a kind of peristaltic action. (4) Transverse wave propagation along the organelle. Types I to 3 can be reduced to the same underlying principle; they cause locomotion. Formation of mitochondrial extensions is due to elongation of cristae. The observations are discussed in terms of 4 specific proposals. (I) Intracellular locomotion of mitochondria is caused by local enlargements and contractions of the organelles. (2) The shape changes are correlated with alterations in the arrangement of the cristae. (3) Such arrangements are not associated with overall swelling or shrinkage of the mitochondrion; they are local features. (4) Estimates of the time required for rearrangement of the inner compartment amount to less than 0.3 s for single crista arrangements during the fastest shape changes, and less than 1–3 s during slower alterations. This high velocity is in good accord with the hypothesis of energy conservation by conformational events during oxidative phosphorylation.

scholarly journals Interaction of the Rabies Virus P Protein with the LC8 Dynein Light Chain

2000 ◽  
Vol 74 (21) ◽  
pp. 10212-10216 ◽  
Author(s):  
Hélène Raux ◽  
Anne Flamand ◽  
Danielle Blondel
Keyword(s):  
Rabies Virus ◽  
Light Chain ◽  
Cytoplasmic Dynein ◽  
Dynein Light Chain ◽  
Directed Movement ◽  
Virus Particles ◽  
P Protein ◽  
Infected Cells ◽  
Transcription And Replication

ABSTRACT The rabies virus P protein is involved in viral transcription and replication but its precise function is not clear. We investigated the role of P (CVS strain) by searching for cellular partners by using a two-hybrid screening of a PC12 cDNA library. We isolated a cDNA encoding a 10-kDa dynein light chain (LC8). LC8 is a component of cytoplasmic dynein involved in the minus end-directed movement of organelles along microtubules. We confirmed that this molecule interacts with P by coimmunoprecipitation in infected cells and in cells transfected with a plasmid encoding P protein. LC8 was also detected in virus particles. Series of deletions from the N- and C-terminal ends of P protein were used to map the LC8-binding domain to the central part of P (residues 138 to 172). These results are relevant to speculate that dynein may be involved in the axonal transport of rabies virus along microtubules through neuron cells.

scholarly journals Ubiquitination of the Cytoplasmic Domain of Influenza A Virus M2 Protein is Crucial for Production of Infectious Virus Particles

2017 ◽  
pp. JVI.01972-17 ◽  
Author(s):  
Wen-Chi Su ◽  
Wen-Ya Yu ◽  
Shih-Han Huang ◽  
Michael M.C. Lai
Keyword(s):  
Virus Replication ◽  
Influenza A ◽  
Functional Role ◽  
Infectious Virus ◽  
Virus Production ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Virus Particles

Virus replication is mediated by interactions between virus and host. Here, we demonstrate that influenza A virus membrane protein 2 (M2) can be ubiquitinated. The lysine residue at position 78, which is located in the cytoplasmic domain of M2, is essential for M2 ubiquitination. An M2-K78R (Lys78→Arg78) mutant, which produces ubiquitination-deficient M2, showed a severe defect in production of infectious virus particles. M2-K78R mutant progeny contained more HA proteins, less viral RNAs and less internal viral proteins, including M1 and NP, than the wild-type virus. Furthermore, most of the M2-K78R mutant viral particles lacked viral ribonucleoproteins upon examination under electron microscopy and exhibited slightly lower densities. We also found that mutant M2 colocalized with M1 protein to a lesser extent than for wild-type virus. These findings may account for the reduced incorporation of viral ribonucleoprotein into virions. By blocking the second round of virus infection, we showed that the M2 ubiquitination-defective mutant exhibited normal level of virus replication during the first round of infection, thereby proving that M2 ubiquitination is involved in the virus production step. Finally, we found that M2-K78R mutant virus induced autophagy and apoptosis earlier than wild-type virus. Collectively, these results suggest that M2 ubiquitination plays an important role in infectious virus production by coordinating efficient packaging of the viral genome into virus particles and timing of viral-induced cell death.IMPORTANCEAnnual epidemics and recurring pandemics of influenza viruses represent a very high global health and economic burden. Influenza virus M2 protein has been extensively studied for its important roles in virus replication, particularly in viral entry and release. Rimantadine, one of the most commonly used antiviral drugs, binds to the channel lumen near the N-terminus of M2 proteins. However, viruses resistant to Rimantadine have emerged. M2 undergoes several posttranslational modifications, such as phosphorylation and palmitoylation. Here, we reveal that ubiquitination mediates the functional role of M2. A ubiquitination-deficient M2 mutant predominately produced virus particles either lacking viral ribonucleoproteins or containing smaller amounts of internal viral components, resulting in lower infectivity. Our findings offer insights into the mechanism of influenza virus morphogenesis, particularly the functional role of M1-M2 interactions in viral particle assembly, and can be applied to the development of new influenza therapies.

scholarly journals Trypsin increases pseudorabies virus production through activation of the ERK signalling pathway

2006 ◽  
Vol 87 (5) ◽  
pp. 1109-1112 ◽  
Author(s):  
Béatrice Riteau ◽  
Christiane de Vaureix ◽  
François Lefèvre
Keyword(s):  
Pseudorabies Virus ◽  
Virus Production ◽  
Signalling Pathway ◽  
Herpesvirus Infection ◽  
Extracellular Proteases ◽  
Extracellular Signal ◽  
Inflammatory Processes ◽  
Infected Cells

Extracellular proteases that are expressed in primary and secondary foci of viral infection are potentially important mediators of infectious inflammatory processes. For some viruses, such as influenza virus and rotaviruses, proteases such as trypsin enhance infectivity by a direct proteolytic effect on some virion proteins. By using an in vitro model of herpesvirus infection, the possibility that proteases modulate the viral cycle through signalling delivered to the infected cell was investigated. It is reported that exposure of pseudorabies virus-infected cells to trypsin increased virus production. Moreover, this treatment induced synergistic and sustained activation of the extracellular signal-regulated kinase (ERK) 1/2 signalling pathway, which appeared to be necessary for this increased viral production. These results suggest that herpesviruses could take advantage of the inflammatory context and particularly of the presence of proteases to increase their replication. Thus, these data point to a potentially important role of extracellular proteases in herpesvirus infection.

Role Of Surface Negative Charge In Platelet Aggregation

1981 ◽  
Author(s):  
K Tanoue ◽  
S M Jung ◽  
I Isohisa ◽  
C Sakakibara ◽  
S Ariga ◽  
...  
Keyword(s):  
Phase Contrast ◽  
Negative Charge ◽  
Human Platelets ◽  
Platelet Surface ◽  
Rabbit Platelets ◽  
Contrast Microscopy ◽  
Physiologic Saline ◽  
And Control ◽  
The Relationship

The relationship between aggregability and electrophoretic mobility (EPM) was studied to clarify the role of surface negative charge in platelet function. 1. Human platelets were washed 3 times with Phillip’s buffer (pH 7.4), resuspended to 107 platelets/ml in modified Zeiller and Hannig’s buffer with 1 mM EDTA and incubated with various concentrations of ADP, adrenaline, thrombin and ristocetin at 37°C for 3 min. Without further washing, mean EPM of about 700 platelets was determined by the automatic Laser Zee System 3000 with good reproducibility with less than 1 % variation coefficient for 5 measurements. In all experiments, aggregating agents were dissolved in physiologic saline to obtain the same solution conductance both in test and control platelet suspensions to which saline alone was added. Neither 1-100 uM ADP nor 5-500 μM adrenaline had any effect on platelet EPM. Thrombin decreased EPM with dose response relation with -1.692±0.014 μm/sec/V/cm without thrombin, -1.580±0.084 with o725 , -1.578±0.001 with 1.0, -1.454±0.018 with 2.0 and -1.289±0.004 with 5.0 U/ml of thrombin. Ristocetin had decreasing effect on EPM at 0.2 mg/ml or less. After addition of above aggregating agents, the washed platelets did not aggregate under phase contrast microscopy. 2. EPM of washed platelets were compared with aggregability in the PRP collected from 33 clinical cases with various degrees of aggregation. There was no correlation between EPM and secondary and maximum aggregation induced by any of these four agents. However, intensity of primary aggregation by adrenaline correlated with EPM (r= 0.485, P<0.01). The same tendency was observed in primary aggregation by ADP. 3. Treatment of washed rabbit platelets with increasing doses of neuraminidase resulted in increasing aggregation by ADP and collagen associated with decrease in EPM. These findings suggested that platelet surface negative charge may be regulatory role in initiation of platelet activation.

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.

The Morphology and Origin of the Golgi Bodies and their Role in the Secretion of the Acrosome in the Spermatogenesis of Pulmonate Gastropods as Determined in the Living Material by Phase-contrast Microscopy

1956 ◽  
Vol s3-97 (39) ◽  
pp. 369-377
Author(s):  
VISHWA NATH ◽  
BRIJ L. GUPTA
Keyword(s):  
Phase Contrast ◽  
Duplex Structure ◽  
Golgi Bodies ◽  
Golgi Region ◽  
Contrast Microscope ◽  
Contrast Microscopy ◽  
Spermatid Nucleus ◽  
Living Material ◽  
Dark Contrast

In this paper are embodied our observations on the morphology, origin, and role of the Golgi bodies in the spermatogenesis of the slug, Anadenus altivagus, and the snail, Euaustenia cassida. Living cells have been studied under the phase-contrast microscope and photomicrographed. In the juxta-nuclear Golgi region in primary spermatocytes the Golgi bodies exist in the form of (1) granules and rods of dark contrast, (2) spheres of pale contrast, and (3) spheroids showing a duplex structure, each consisting of a sphere of pale contrast and an incomplete or complete cortex or sheath of dark contrast. The Golgi spheroids have been shown to arise three times in the course of spermatogenesis from the mitochondrial granules by a process of alignment. The acrosome is formed from a cap of tiny acrosomal granules, which are deposited in front of the spermatid nucleus by the Golgi bodies (acroblasts).

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