Characterization of immortalized rabbit lacrimal gland epithelial cells

Abstract

To establish an immortalized lacrimal gland epithelial cell line, the orbital lacrimal glands of normal New Zealand White rabbits were multiply injected with an immortalizing amphotropic retroviral vector (LXSN16E6E7) containing the E6 and E7 genes of human papillomavirus type 16. Lacrimal glands were removed after 2 d and acinar epithelial cells were isolated and cultured on Matrigel-coated 60 mm2 plates containing DMEM-F12 supplemented with 5% Nu-serum V. Transformed cells were selected in G418 sulfate for 7 d and passaged. Morphology of the immortalized cells was similar to that described for normal acinar cells both in vivo and in vitro, with rough endoplasmic reticulum and secretory granules. These characteristics remained unchanged and the cells continued to exhibit typical polygonal epithelioid structure. The cells have been maintained in culture for 14 mo. and have gone through 58 passages without loss of proliferation or epithelial cell characteristics. Immunohistochemistry and Western blots showed positive reactivity to secretory component, transferrin, and transferrin receptor, which are typical proteins found in the lacrimal gland. Functional analysis by stimulation with a cholinergic agonist, carbachol (100 microM), resulted in a significant release of protein. This is the first report of an immortalized rabbit lacrimal epithelial cell. These cells will provide a valuable tool for the molecular analysis of lacrimal gland epithelial cell functions.

Down-regulation of Androgen Receptor Expression and Inhibition of Lacrimal Gland Cell Proliferation by Retinoic Acid

“…Therefore, we turned to an immortalized rabbit lacrimal acinar cell line for studies of effects of DHT and retinoic acid on cell proliferation. This cell line expresses transferrin and secretory component (Nguyen et al, 1999), as expected in lacrimal acinar cells. We have recently reported that cytokeratin and integrin expression in this cell line is similar to primary cells and that this cell line proliferates in culture (Saarloos et al, 1999).…”

“…Cells were cultured on 4 % Matrigel-coated six-well plates in a hormone supplemented DMEM/ Hams F-12 mixture, as previously described (Nguyen et al, 1999;Saarloos et al, 1999), or in HepatoStim medium without serum. Cells grew equally well in both media.…”

Sensory Denervation Modulates eIF-2 Alpha Kinase Expression in the Rabbit Lacrimal Gland

“…15 In brief, frozen lacrimal glands were homogenized in hot 10 mM Tris-Cl (pH 7.4) with 1% SDS, and total protein lysate was quantified using a bicinchoninic acid (BCA) assay kit (Pierce, Rockford, IL, USA). Equal amounts (50 μg) of protein were separated on 7.5% polyacrylamide gels and transferred overnight at 4°C to nitrocellulose membranes (Hybond-ECL; Amersham).…”

Scintigraphic Evaluation of Lacrimal Glands Using a Rabbit Experimental Model

“…Since the inferior lacrimal gland of the rabbit is a wellstudied gland [5,6] with a considerable volume and good surgical accessibility, scintigraphy could be used to study functional alteration of the glandular parenchyma.…”