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A Obolensky, I Chowers, E Piontek, E Pikarsky, J Pe'er, A Panet, E Banin; Gene Delivery by Viral Vectors in Primary Cultures of Lacrimal Gland Tissue . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4623.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To test the feasibility of gene transfer into lacrimal gland tissue in primary culture, using different viral vectors. Methods:Lacrimal glands were obtained from adult SABRA rats and divided by pincers to 0.3-0.4 mm bits. Under primary organ culture conditions in DMEM medium (with Penicillin, Streptomycin and Glutamine), tissue could be maintained for up to three weeks. The ability of four different viral vectors to conduct ß -galactosidase (ß-gal) reporter gene delivery was examined: adenovirus Ad5CMVLacZ, vaccinia virus VSC9, herpesvirus tkLTRZ1, and lentivirus LentiLacZ. One of the viral vectors, or saline, was added to the growth medium and incubated at 37º for 30-60 min before transferring tissue bits to fresh medium. After 3 and 7 days, ß-gal expression was examined by X-gal staining in gross preparations and in histological sections. Results:After 3 days, ß-gal expression was observed in 33% of tissue bits exposed to the adenoviral vector, and in 62.5% of bits exposed to the vaccinia vector. With the herpes and lenti viruses no significant expression occurred at this time point. Following 7 days in culture, ß-gal expression was observed in 95% cases of adenoviral, 66.7% cases of vaccinia, and only in 14.3% of herpesviral and 5.5% of lentiviral applications. Histological examination revealed different tissue tropism of the viral vectors: with adenovirus, staining was especially prominent in the interacinar areas, mainly in the myoepithelial cells. With the vaccinia vector, ß-gal expression was clearly seen in lacrimal duct cells and in the acini. Conclusion:Adenovirus and vaccinia virus are efficient vectors for gene transfer into lacrimal gland tissue in primary culture. This may be the first step towards expressing genes whose products could be continuously delivered to the eye via the tears. The tissue tropism of the vaccinia vector suggests that it may be the preferred candidate for such gene delivery.
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