Shubha Tiwari1, Rohini M Nair2, Praseeda Vamadevan2, Mohammad Javed Ali3, Milind N Naik3, Santosh G Honavar4, Geeta K Vemuganti5,6,7. 1. Sudhakar and Sreekant Ravi Stem Cell Biology Laboratory, L V Prasad Eye Institute, Hyderabad, India. 2. School of Medical Sciences, University of Hyderabad, Hyderabad, 500046, India. 3. Ophthalmic Plastic Surgery, Orbit &and Ocular Oncology, L V Prasad Eye Institute, Hyderabad, India. 4. Centre for Sight Superspeciality Eye Hospital, Hyderabad, India. 5. Sudhakar and Sreekant Ravi Stem Cell Biology Laboratory, L V Prasad Eye Institute, Hyderabad, India. gkvemuganti@gmail.com. 6. Ophthalmic Pathology Services, L V Prasad Eye Institute, Hyderabad, India. gkvemuganti@gmail.com. 7. School of Medical Sciences, University of Hyderabad, Hyderabad, 500046, India. gkvemuganti@gmail.com.
Abstract
PURPOSE: Lacrimal gland (LG) dysfunction leading to dry eye syndrome (DES) is an important cause of ocular morbidity. One of the potential and promising long-term management therapies for restoration of LG function could be transplantation of autologous ex vivo expanded stem cells. The present study was aimed at exploring the 2D and 3D cultures of human LG, identifying inherent stem cells and evaluating their secretory potential. METHODS: Fresh human lacrimal gland (HuLG) (n = 5) from patients undergoing therapeutic exenteration was harvested after ethical approval and informed consent. The gland was enzymatically digested and the isolated cells plated in Hepato-STIM media supplemented with l-glutamine, epidermal growth factor, fibroblast growth factor, and N-2 supplement. The native HuLG and the cultured spheres (DIV14-16) were evaluated for presence of stem cells (CD117 expression, quiescence, BrdU label retention, cell cycle, colony forming efficiency) and differentiation (secretion of tear proteins). RESULTS: Under the established culture conditions, suspension 3D cultures of human "lacrispheres" could be maintained and propagated for 3-4 weeks. The spheres consist of both acinar as well as ductal cells with evidence of stem cells (0.8 ± 0.05% CD117+ cells), BrdU label retention (9.31 ± 0.41%), G0/G1 profile similar to native lacrimal cells at isolation (76.9 versus 79.9%) and colony forming units (3.1%). The lacrispheres also secreted quantifiable levels of tear proteins (lysozyme, lactoferrin, scIgA) into the conditioned media. CONCLUSION: The study provides promising, first-of-its-kind evidence for the generation of lacrispheres from fresh HuLG, with enriched population of stem cells and secretory competent differentiated cells. The dual properties of these spheres make them a highly suitable source of transplantable cells for restoring the structure and function of damaged lacrimal gland.
PURPOSE: Lacrimal gland (LG) dysfunction leading to dry eye syndrome (DES) is an important cause of ocular morbidity. One of the potential and promising long-term management therapies for restoration of LG function could be transplantation of autologous ex vivo expanded stem cells. The present study was aimed at exploring the 2D and 3D cultures of human LG, identifying inherent stem cells and evaluating their secretory potential. METHODS: Fresh human lacrimal gland (HuLG) (n = 5) from patients undergoing therapeutic exenteration was harvested after ethical approval and informed consent. The gland was enzymatically digested and the isolated cells plated in Hepato-STIM media supplemented with l-glutamine, epidermal growth factor, fibroblast growth factor, and N-2 supplement. The native HuLG and the cultured spheres (DIV14-16) were evaluated for presence of stem cells (CD117 expression, quiescence, BrdU label retention, cell cycle, colony forming efficiency) and differentiation (secretion of tear proteins). RESULTS: Under the established culture conditions, suspension 3D cultures of human "lacrispheres" could be maintained and propagated for 3-4 weeks. The spheres consist of both acinar as well as ductal cells with evidence of stem cells (0.8 ± 0.05% CD117+ cells), BrdU label retention (9.31 ± 0.41%), G0/G1 profile similar to native lacrimal cells at isolation (76.9 versus 79.9%) and colony forming units (3.1%). The lacrispheres also secreted quantifiable levels of tear proteins (lysozyme, lactoferrin, scIgA) into the conditioned media. CONCLUSION: The study provides promising, first-of-its-kind evidence for the generation of lacrispheres from fresh HuLG, with enriched population of stem cells and secretory competent differentiated cells. The dual properties of these spheres make them a highly suitable source of transplantable cells for restoring the structure and function of damaged lacrimal gland.
Authors: Philipp Ackermann; Susann Hetz; Julia Dieckow; Martin Schicht; Anja Richter; Charli Kruse; Insa S Schroeder; Matthias Jung; Friedrich P Paulsen Journal: Invest Ophthalmol Vis Sci Date: 2015-07 Impact factor: 4.799
Authors: Shubha Tiwari; Mohammad Javed Ali; Murali M S Balla; Milind N Naik; Santosh G Honavar; Vijay Anand P Reddy; Geeta K Vemuganti Journal: PLoS One Date: 2012-01-13 Impact factor: 3.240
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