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Journal of Virology, December 2001, p. 11747-11754, Vol. 75, No. 23
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.23.11747-11754.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Vesicular Stomatitis Virus G-Pseudotyped Lentivirus Vectors Mediate Efficient Apical Transduction of Polarized Quiescent Primary Alveolar Epithelial Cells

Zea Borok,1,* Jens Erik Harboe-Schmidt,2,3 Steven L. Brody,4 Yingjian You,4 Beiyun Zhou,1 Xian Li,1 Paula M. Cannon,2 Kwang-Jin Kim,1 Edward D. Crandall,1,5 and Noriyuki Kasahara2,3,5

Department of Medicine and Will Rogers Institute Pulmonary Research Center,1 Departments of Biochemistry and Molecular Biology2 and Pathology,5 and Institute for Genetic Medicine,3 Keck School of Medicine, University of Southern California, Los Angeles, California, and Department of Medicine, Washington University School of Medicine, St. Louis, Missouri4

Received 30 April 2001/Accepted 21 August 2001

We investigated the use of lentivirus vectors for gene transfer to quiescent alveolar epithelial cells. Primary rat alveolar epithelial cells (AEC) grown on plastic or as polarized monolayers on tissue culture-treated polycarbonate semipermeable supports were transduced with a replication-defective human immunodeficiency virus-based lentivirus vector pseudotyped with the vesicular stomatitis virus G (VSV-G) protein and encoding an enhanced green fluorescent protein reporter gene. Transduction efficiency, evaluated by confocal microscopy and quantified by fluorescence-activated cell sorting, was dependent on the dose of vector, ranging from 4% at a multiplicity of infection (MOI) of 0.1 to 99% at an MOI of 50 for AEC grown on plastic. At a comparable titer and MOI, transduction of these cells by a similarly pseudotyped murine leukemia virus vector was ~30-fold less than by the lentivirus vector. Importantly, comparison of lentivirus-mediated gene transfer from the apical or basolateral surface of confluent AEC monolayers (Rt > 2 kOmega · cm2; MOI = 10) revealed efficient transduction only when VSV-G-pseudotyped lentivirus was applied apically. Furthermore, treatment with EGTA to increase access to the basolateral surface did not increase transduction of apically applied virus, indicating that transduction was primarily via the apical membrane domain. In contrast, differentiated tracheal epithelial cells were transduced by apically applied lentivirus only in the presence of EGTA and at a much lower overall efficiency (~15-fold) than was observed for AEC. Efficient transduction of AEC from the apical cell surface supports the feasibility of using VSV-G-pseudotyped lentivirus vectors for gene transfer to the alveolar epithelium and suggests that differences exist between upper and lower airways in the polarity of available receptors for the VSV-G protein.

* Corresponding author. Mailing address: Division of Pulmonary and Critical Care Medicine, University of Southern California, GNH 11-900, 2025 Zonal Ave., Los Angeles, CA 90033. Phone: (323) 442-3329. Fax: (323) 442-2611. E-mail: zborok{at}hsc.usc.edu.

Journal of Virology, December 2001, p. 11747-11754, Vol. 75, No. 23
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.23.11747-11754.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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