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G. K. Vemuganti, S. Tiwari, S. G. Honavar, M. N. Naik, V. P. Reddy; Cultivation and Characterization of Human Lacrimal Gland Cells for Potential Clinical Application. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4171.
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© ARVO (1962-2015); The Authors (2016-present)
Xerophthalmia, xerostomia are some of the morbid complications following radiotherapy to the tumors of head and neck region. Due to the limited benefits of conventional medical treatment of radiation induced-dry eyes options like cell based therapy are being evaluated as promising alternatives. We herein present the data on in-vitro culture and characterization of lacrimal gland cells for potential clinical.
After IRB approval and informed consent from patients, fresh lacrimal gland (8) samples were obtained from patients undergoing exenteration. All the tissues were histologically confirmed as normal and free of tumor. Explant and suspended cultures were established using different enzyme cocktail, substrate and media composition. The cells were characterized by immunocytochemistry and flow cytometry for stem cell marker (ABCG2), epithelial markers (E-cadherin, CK3/12, and connexin 43) and mesenchymal markers (CD90, vimentin). In-vitro function of these acinar cells was evaluated by ELISA by testing for Ig A levels.
Successful cultures were established from all the samples of human lacrimal gland tissues- both as a monolayer as well as spheres. The epithelial cells were polygonal, with distinct cell borders and secretory granules and were immunoreactive for ABCG2, CK 3/12, connexin and E-cadherin. The 3 D spheres of epithelial cells were better formed in serum free medium, while monolayered epithelial cells were seen on denuded human amniotic membrane The conditioned media of the lacrimal tissues showed the presence of secretory IgA (Mean OD- Test: 0.32 Vs Mean OD-control: 0.24). In addition, the cultures also showed adherent spindle cells that were positive for CD 90 and vimentin. The epithelial cells were short lived (20-30 days) while the spindle cell survived for 3-4 months, especially in serum containing medium.
Successful 2 and 3 dimensional cultures were established from human lacrimal gland tissues with preserved secretory function. The presence of spindle cells is an intriguing finding which could represent a stromal origin and warrant further studies to determine the lineage and function
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