scholarly journals Growth of Hepatitis A Virus in a Mouse Liver Cell Line

2005 ◽  
Vol 79 (5) ◽  
pp. 2950-2955 ◽  
Author(s):  
Dino A. Feigelstock ◽  
Peter Thompson ◽  
Gerardo G. Kaplan
Keyword(s):  
Growth Factors ◽  
Mouse Liver ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Growth Conditions ◽  
Liver Cells ◽  
Nucleotide Sequence Analysis ◽  
Mouse Liver Cell ◽  
A Cell ◽  
Cells Cultured

ABSTRACT Hepatitis A virus (HAV) has been adapted to grow efficiently in primate and some nonprimate cell lines but not in cells of murine origin. To understand the inability of the virus to grow in mouse cells, we studied the replication of HAV in immortalized and nontransformed MMH-D3 mouse liver cells, which require growth factors and collagen to maintain their phenotype. HAV grew in MMH-D3 cells transfected with virion RNA but not in those infected with viral particles, indicating a cell entry block for HAV. However, MMH-D3 cells cultured under suboptimal conditions in the absence of growth factors acquired susceptibility to HAV infection. Serial passages of the virus in MMH-D3 cells under suboptimal growth conditions resulted in the selection of HAV variants that grew efficiently in MMH-D3 cells cultured under both optimal and suboptimal conditions. Nucleotide sequence analysis of the MMH-D3 cell-adapted HAV revealed that N1237D and D2132G substitutions were present in the capsid regions of six viral clones. These two mutations are most likely located on the surface of the virion and may play a role in the entry of HAV into the mouse liver cells. Our results demonstrate that mouse hepatocyte-like cells code for all factors required for the efficient growth of HAV in cell culture.

Modulation of fetal mouse liver cells cultured on a pigskin substrate.

1983 ◽  
Vol 140 (1) ◽  
pp. 15-28 ◽  
Author(s):  
KOICHI HIRATA ◽  
KOJI SHIRAMATSU ◽  
TOMOAKI USUI ◽  
YUTAKA YOSHIDA ◽  
AARON E. FREEMAN ◽  
...  
Keyword(s):  
Mouse Liver ◽  
Liver Cells ◽  
Fetal Mouse ◽  
Fetal Mouse Liver ◽  
Cells Cultured

scholarly journals Determinants in 3Dpol Modulate the Rate of Growth of Hepatitis A Virus

2010 ◽  
Vol 84 (16) ◽  
pp. 8342-8347 ◽  
Author(s):  
Krishnamurthy Konduru ◽  
Gerardo G. Kaplan
Keyword(s):  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Rna Replication ◽  
Virus Genome ◽  
Nucleotide Sequence Analysis ◽  
Reverse Genetic ◽  
Genetic Studies ◽  
High Concentrations ◽  
Kinetics Of ◽  
Atypical Member

ABSTRACT Hepatitis A virus (HAV), an atypical member of the Picornaviridae, grows poorly in cell culture. To define determinants of HAV growth, we introduced a blasticidin (Bsd) resistance gene into the virus genome and selected variants that grew at high concentrations of Bsd. The mutants grew fast and had increased rates of RNA replication and translation but did not produce significantly higher virus yields. Nucleotide sequence analysis and reverse genetic studies revealed that a T6069G change resulting in a F42L amino acid substitution in the viral polymerase (3Dpol) was required for growth at high Bsd concentrations whereas a silent C7027T mutation enhanced the growth rate. Here, we identified a novel determinant(s) in 3Dpol that controls the kinetics of HAV growth.

Antiviral activity of recombinant interferon-α on hepatitis A virus replication in human liver cells

1995 ◽  
Vol 28 (1) ◽  
pp. 69-80 ◽  
Author(s):  
Jean-Marc Crance ◽  
Françoise Lévêque ◽  
Suzanne Chousterman ◽  
Alain Jouan ◽  
Christian Trépo ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Virus Replication ◽  
Human Liver ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Liver Cells ◽  
Interferon Α ◽  
Recombinant Interferon ◽  
Human Liver Cells

scholarly journals INTERACTION OF DIPHTHERIA TOXIN WITH CELL CULTURES FROM SUSCEPTIBLE AND RESISTANT ANIMALS

1966 ◽  
Vol 123 (4) ◽  
pp. 723-732 ◽  
Author(s):  
Janis Gabliks ◽  
Marcia Falconer
Keyword(s):  
Diphtheria Toxin ◽  
Mouse Liver ◽  
Diffusion Rate ◽  
Liver Cells ◽  
Cell Protein ◽  
Resistant Mouse ◽  
Mouse Liver Cell ◽  
The Individual ◽  
Chang Liver ◽  
Rna And Dna

The cytotoxicity (TD) level of diphtheria toxin for human Chang liver strain was 0.1 guinea pig MLD per ml; for mouse liver NCTC 1469 strain, the TD was 500 MLD per ml. The results of cell culture toxicity correlated well with relative susceptibility of both man and mouse. The initial effects of toxin on the susceptible Chang liver cells were studied at one half the TD level (0.05 MLD per ml). At this low concentration of toxin, the number of cells per culture was reduced slightly below the "0" hr values, whereas the amounts of cell protein, RNA, and DNA were increased. Analysis of the toxin-treated cells indicated an enlargement of the cells. The individual cells contained significantly more protein, RNA, and DNA than the control cells and the cell volume was increased 1.9 times. When purified diphtheria toxin was incubated with the susceptible Chang liver cells and then tested for its biological activity, the results showed an increased diffusion rate in parabiotic culture chambers and definite cytotoxicity to the normally resistant mouse liver cells. The cytotoxicity was neutralizable with antitoxin. The results suggest that the toxin-susceptible cells transform the toxin molecule to a more active derivative which affects the highly resistant mouse liver cell.

Sofosbuvir inhibits hepatitis A virus replication in vitro assessed by a cell-based fluorescent reporter system

2018 ◽  
Vol 154 ◽  
pp. 51-57 ◽  
Author(s):  
Wang Jiang ◽  
Fawad Muhammad ◽  
Pengjuan Ma ◽  
Xiyu Liu ◽  
Gang Long
Keyword(s):  
Virus Replication ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Reporter System ◽  
Fluorescent Reporter ◽  
A Cell

scholarly journals Utilization of Chimeras between Human (HM-175) and Simian (AGM-27) Strains of Hepatitis A Virus To Study the Molecular Basis of Virulence

1998 ◽  
Vol 72 (9) ◽  
pp. 7467-7475 ◽  
Author(s):  
Gopa Raychaudhuri ◽  
Sugantha Govindarajan ◽  
Max Shapiro ◽  
Robert H. Purcell ◽  
and Suzanne U. Emerson
Keyword(s):  
Cell Culture ◽  
Hepatitis A ◽  
Cultured Cells ◽  
Hepatitis A Virus ◽  
Kinetic Studies ◽  
Simian Virus ◽  
Saguinus Mystax ◽  
Amino Terminal ◽  
A Cell ◽  
Carboxy Terminal

ABSTRACT Chimeras between human (HM-175) and simian (AGM-27) strains of hepatitis A virus (HAV) were constructed to evaluate the effect of the 2C gene of AGM-27 on HAV replication in cell culture and virulence in tamarins (Saguinus mystax) and chimpanzees (Pan troglodytes). Kinetic studies and radioimmunofocus assays demonstrated that replacement of the 2C gene of HAV/7, a cell culture-adapted strain of HM-175, with that of AGM-27 drastically reduced the ability of the virus to replicate in cultured cells. Intragenic chimeras containing AGM-27 sequences in either the 5′ or 3′ half of the 2C gene replicated in cell culture at an intermediate level. Whereas HAV/7 is attenuated for tamarins, a chimera containing the simian virus 2C gene in the HAV/7 background was virulent in tamarins, demonstrating that the simian virus 2C gene alone can confer the phenotype of virulence to an otherwise attenuated virus. Clusters of AGM-27-specific residues near both ends of the 2C protein were required for virulence since a chimera containing AGM-27 sequences in the carboxy-terminal half of 2C was partially attenuated for tamarins while one containing AGM-27 sequences only in the amino-terminal half of 2C was even more attenuated. Chimeras containing either the entire or only the 3′ half of the simian virus 2C gene in the HAV/7 background were attenuated for chimpanzees.

scholarly journals Analysis of Deletion Mutants Indicates that the 2A Polypeptide of Hepatitis A Virus Participates in Virion Morphogenesis

2002 ◽  
Vol 76 (15) ◽  
pp. 7495-7505 ◽  
Author(s):  
Lisette Cohen ◽  
Danièle Bénichou ◽  
Annette Martin
Keyword(s):  
Capsid Protein ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Protein A ◽  
Growth Conditions ◽  
Protein Precursor ◽  
Synthetic Genome ◽  
Vaccinia Viruses ◽  
One Step ◽  
Capsid Protein Vp1

ABSTRACT Unlike all other picornaviruses, the primary cleavage of the hepatitis A virus (HAV) polyprotein occurs at the 2A/2B junction and is carried out by the only proteinase encoded by the virus, 3Cpro. The resulting P1-2A capsid protein precursor is subsequently cleaved by 3Cpro to generate VP0, VP3, and VP1-2A, which associate as pentamers. An unidentified cellular proteinase acting at the VP1/2A junction releases the mature capsid protein VP1 from VP1-2A later in the morphogenesis process. Although these aspects of polyprotein processing are well characterized, the function of 2A is unknown. To study its role in the viral life cycle, we assessed the infectivity of synthetic, genome-length RNAs containing 11 different in-frame deletions in the 2A region. Deletions in the N-terminal 40% of 2A abolished infectivity, whereas deletions in the C-terminal 60% resulted in viruses with a small-focus replication phenotype. C-terminal deletions in 2A had no effect on RNA replication kinetics under one-step growth conditions, nor did they have an effect on capsid protein synthesis and 3Cpro-mediated processing. However, C-terminal deletions in 2A altered the VP1/2A cleavage, resulting in accumulation of uncleaved VP1-2A precursor in virions and possibly accounting for a delay in the appearance of infectious particles with these mutants, as well as a fourfold decrease in specific infectivity of the virus particles. When the capsid proteins were expressed from recombinant vaccinia viruses, the N-terminal part of 2A was required for efficient cleavage of the P1-2A precursor by 3Cpro and assembly of structural precursors into pentamers. These data indicate that the N-terminal domain of 2A must be present as a C-terminal extension of P1 for folding of the capsid protein precursor to allow efficient 3Cpro-mediated cleavages and to promote pentamer assembly, after which cleavage at the VP1/2A junction releases the mature VP1 protein, a process that appears to be necessary to produce highly infectious particles.

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