Keratinocyte growth factor stimulates transduction of the respiratory epithelium by retroviral vectors.
Cell proliferation is required for transduction by standard retrovirus vectors derived from viruses in the murine leukemia virus (MuLV) group. Since proliferation rates are low in the mature pulmonary epithelium, we tested the hypothesis that the efficiency of retrovirus-mediated transduction of respiratory epithelial cells can be enhanced by stimulation of cell proliferation with recombinant human keratinocyte growth factor (rhKGF). A marked increase in proliferation of bronchiolar and alveolar epithelial cells was observed after intratracheal administration of rhKGF (30 mg/kg) to adult FVB/N mice. Two days after rhKGF or saline treatment, 10(7) AP+ FFU of LAPSN, a recombinant amphotropic retrovirus that expresses human placental alkaline phosphatase (AP), was instilled intratracheally into the mice. Transduction efficiency, measured 2 days after infection, was increased approximately 70-fold by rhKGF pretreatment. However, even after KGF treatment the total numbers of AP-expressing cells were few. Transduction efficiency was similar using either LAPSN packaged by amphotropic host range packaging cells or LAPSN pseudotyped with 10A1 MuLV envelope protein (0.091 +/- 0.006 versus 0.094 +/- 0.028 transduction events/mm2, respectively). Amphotropic vectors use Pit-2 for cell entry, while 10A1 MuLV vectors can use Pit-1 or Pit-2 for cell entry. By in situ hybridization the retroviral receptor Pit-2 (Ram-1) mRNA was expressed only in the pulmonary vasculature, and Pit-1 (Glvr-1) mRNA was expressed at low levels throughout the lung. In vitro studies demonstrated that retrovirus was inactivated by pulmonary surfactant. Stimulating proliferation of the respiratory epithelium increased retroviral transduction in vivo, but the paucity of retroviral receptors and inactivation by surfactant are additional barriers to high-level retroviral gene transfer in the lung.
Division of Pulmonary Biology, Children's Hospital Research Foundation, Cincinnati, OH 45229, USA.
Z K
ZK
Zsengellér
C
C
Halbert
A D
AD
Miller
S E
SE
Wert
J A
JA
Whitsett
C J
CJ
Bachurski
D000818:Animals; D001980:Bronchi / Q000187:drug effects*; D002352:Carrier Proteins / Q000032:analysis; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000032:analysis; D004305:Dose-Response Relationship, Drug; D004848:Epithelium / Q000033:anatomy & histology / Q000276:immunology / Q000378:metabolism; D005260:Female; D051526:Fibroblast Growth Factor 10; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006133:Growth Substances / Q000494:pharmacology*; D006801:Humans; D008565:Membrane Proteins; D051379:Mice; D008815:Mice, Inbred Strains; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids / Q000032:analysis; D011650:Pulmonary Alveoli / Q000187:drug effects*; D011994:Recombinant Proteins / Q000494:pharmacology; D012190:Retroviridae / Q000235:genetics; D013997:Time Factors; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000032:analysis; D014161:Transduction, Genetic
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D051526:Fibroblast Growth Factor 10; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D006133:Growth Substances; D008565:Membrane Proteins; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids; D011994:Recombinant Proteins; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; C496453:FGF7 protein, human; C496454:Fgf7 protein, mouse; C494187:POU1F1 protein, human; C494185:Pou1f1 protein, mouse