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Originally published In Press as doi:10.1167/iovs.08-2503 on December 20, 2008
 (Investigative Ophthalmology and Visual Science. 2009;50:1978-1987.)
© 2009 by The Association for Research in Vision and Ophthalmology, Inc.
doi:10.1167/iovs.08-2503
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Purification and Characterization of Mouse Lacrimal Gland Epithelial Cells and Reconstruction of an Acinarlike Structure in Three-Dimensional Culture

Yoshiaki Ueda,1 Yoko Karasawa,1 Yasushi Satoh,2 Shimpei Nishikawa,1 Junko Imaki,3 and Masataka Ito3

1From the Departments of Ophthalmology, 2Biochemistry, and 3Developmental Anatomy and Regenerative Biology, National Defense Medical College, Saitama, Japan.

PURPOSE. To clarify which factors are involved in postnatal lacrimal gland development, the expressions of several growth factors and their receptors were analyzed in purified lacrimal gland epithelial and mesenchymal cells. Reconstruction of acinarlike structures in three-dimensional culture conditions from this epithelial cell population was attempted.

METHODS. Lacrimal gland epithelial and mesenchymal cells were isolated from newborn mice and purified using nylon mesh and collagenase. By real-time reverse transcription–polymerase chain reaction, immunocytochemistry, and Western blotting, the expressions of several growth factors and their receptors were analyzed. Responses of epithelial cells to the growth factors were analyzed by the addition of these factors to the culture medium.

RESULTS. Fibroblast growth factor (FGF)10 and hepatocyte growth factor (HGF) were more intensely expressed in mesenchymal cells than in epithelial cells, and their receptors (FGFR2IIIb and c-Met, respectively) were less intensely expressed, whereas the expression of epidermal growth factor (EGF) and its receptor showed no significant difference between cell types. In the monolayer culture, cell viability was activated by the addition of EGF or HGF to epithelial cells, but no response was observed when FGF10 was added. The epithelial cells formed clusters with lumina when cultured on basement membrane matrix. These clusters contained secretion granules and showed positive immunostaining for aquaporin-5.

CONCLUSIONS. EGF and HGF were considered to act in an autocrine/paracrine manner in and around the postnatal lacrimal gland, whereas epithelial cells did not respond to FGF10. It was suggested that certain extracellular matrix conditions accommodate these epithelial cells to reconstruct functional acinarlike structures.






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