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Journal of Histochemistry and Cytochemistry, Vol. 45, 1533-1546, Copyright © 1997 by The Histochemical Society, Inc.

ARTICLE

Retrovirus-mediated Gene Transfer into Rat Salivary Gland Cells In Vitro and In Vivo

Tibor Barkaa and Hendrika M. van der Noena
a Department of Cell Biology and Anatomy, Mount Sinai School of Medicine of The City University of New York, New York, New York

Correspondence to: Tibor Barka, Mount Sinai School of Medicine, Box 1007, New York, NY 10029.

A retroviral vector DAP that encodes the human placental alkaline phosphatase (PLAP) and the neomycin-resistant gene was used to transduce the salivary gland-derived cell line A5 in vitro and acinar cells in rat submandibular gland in vivo. Expression of the transduced PLAP gene was established by histochemical staining for heat-resistant AP and by determination of enzyme activity. From the in vitro experiments, we concluded that the salivary gland-derived cell line A5 can be infected by the retroviral vector DAP. In the transduced cells the viral long terminal repeat (LTR) promoter was effective, and the cells expressed heat-stable PLAP which was localized mostly in the plasma membrane and could be released by treatment with bromelain or phosphatidyinositol-specific phospholipase C. A5-DAP cells secreted PLAP into the medium. Clones of A5-DAP cells expressed various levels of the enzyme. The level of enzyme activity in different clones was unrelated to growth rate. Retrograde ductal injection of the viral vector into the duct of the submandibular gland of rats resulted in integration and long-term expression of PLAP gene in acinar cells. Expression of PLAP was seen up to 25 days, the limit of the observation period. To facilitate integration of the viral DNA, cell division of acinar cells was induced by administration of the ß-adrenergic agonist isoproterenol before administration of the virus. PLAP was secreted into submandibular saliva. The data support the notion that salivary glands are suitable targets for gene transfer in vivo by a retroviral vector. (J Histochem Cytochem 45:1533-1545, 1997)

Key Words: gene transfer, DAP retroviral vector, submandibular gland, placental alkaline phosphatase, tissue culture, isoproterenol, membrane enzyme


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T. Barka, E. W. Gresik, and H. van der Noen
Transduction of TAT-HA-{beta}-galactosidase Fusion Protein into Salivary Gland-derived Cells and Organ Cultures of the Developing Gland, and into Rat Submandibular Gland in Vivo
J. Histochem. Cytochem., November 1, 2000; 48(11): 1453 - 1460.
[Abstract] [Full Text]



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