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Journal of Virology, October 2001, p. 9857-9871, Vol. 75, No. 20
Department of Microbiology and
Immunology,1 Cell and Molecular Biology
Graduate Program,2 and Department of
Pathology,4 Milton S. Hershey Medical Center,
The Pennsylvania State University, College of Medicine, Hershey,
Pennsylvania 17033, and Department of Neurology,
Massachusetts General Hospital, Boston, Massachusetts
021293
Received 3 May 2001/Accepted 9 July 2001
Baculovirus infection has extended the capabilities for
transfection of exogenous genes into a variety of mammalian cell types. Because rat hepatocytes plated on collagen-coated dishes and maintained in dimethyl sulfoxide (DMSO)-supplemented chemically defined medium are
an excellent model system for studying liver function in vitro, we
investigated the ability of baculoviruses to infect and deliver exogenous genes to cells in this culture system. Efficient delivery to
hepatocytes in short-term culture becomes restricted to peripheral cells, or "edge" cells, as the hepatocytes acquire intercellular junctions and form islands with time in culture. This barrier to
baculovirus entry can be overcome, and the percentage of internal cells
within the hepatocyte islands that are infected with the baculovirus
can be increased more than 100-fold, when cells are subjected to
transient calcium depletion before and during infection. These findings
suggest that at least in some cell types, such as hepatocytes,
baculovirus entry may require contact with the basolateral surface. We
conclude from this study that recombinant baculovirus infection
following transient depletion of extracellular calcium results in
delivery of exogenous genes to at least 75% of hepatocytes in
long-term DMSO culture, thereby making it possible for the first time
to carry out gain-of-function and loss-of-function studies in this cell system.
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.20.9857-9871.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Transient Disruption of Intercellular Junctions
Enables Baculovirus Entry into Nondividing Hepatocytes
*
Corresponding author. Mailing address: Department of
Microbiology and Immunology, Milton S. Hershey Medical Center, The
Pennsylvania State University, College of Medicine, 500 University Dr.,
P.O. Box 850, Hershey, PA 17033. Phone: (717) 531-8609. Fax: (717) 531-4133. E-mail: hisom{at}psu.edu.
Present address: Gilead Sciences, Foster City, CA 94404.
Journal of Virology, October 2001, p. 9857-9871, Vol. 75, No. 20
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.20.9857-9871.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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