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Establishing and characterizing lacrispheres from human lacrimal gland for potential clinical application

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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.

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Acknowledgments

The authors would like to thank Mr. Sreedhar Bowenpalli and Mr. G. Chenchu Naidu for the technical assistance provided in immunohistochemistry experiments.

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Authors and Affiliations

  1. Sudhakar and Sreekant Ravi Stem Cell Biology Laboratory, L V Prasad Eye Institute, Hyderabad, India

    Shubha Tiwari & Geeta K. Vemuganti

  2. Ophthalmic Plastic Surgery, Orbit &and Ocular Oncology, L V Prasad Eye Institute, Hyderabad, India

    Mohammad Javed Ali & Milind N. Naik

  3. Ophthalmic Pathology Services, L V Prasad Eye Institute, Hyderabad, India

    Geeta K. Vemuganti

  4. School of Medical Sciences, University of Hyderabad, Hyderabad, 500046, India

    Rohini M. Nair, Praseeda Vamadevan & Geeta K. Vemuganti

  5. Centre for Sight Superspeciality Eye Hospital, Hyderabad, India

    Santosh G. Honavar

Authors
  1. Shubha Tiwari
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  2. Rohini M. Nair
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  3. Praseeda Vamadevan
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  4. Mohammad Javed Ali
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  5. Milind N. Naik
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  6. Santosh G. Honavar
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  7. Geeta K. Vemuganti
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Corresponding author

Correspondence to Geeta K. Vemuganti.

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Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Informed consent

Informed consent was obtained from all individual participants included in the study.

Financial support

The authors thank International Atomic Energy Agency (IAEA), Hyderabad Eye Research Foundation (HERF), Champalimaud Translational Research Foundation (C-Tracer), Department of Biotechnology (DBT), and Council for Scientific and Industrial Research (CSIR) for their financial support.

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None.

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Tiwari, S., Nair, R.M., Vamadevan, P. et al. Establishing and characterizing lacrispheres from human lacrimal gland for potential clinical application. Graefes Arch Clin Exp Ophthalmol 256, 717–727 (2018). https://doi.org/10.1007/s00417-018-3926-8

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  • DOI: https://doi.org/10.1007/s00417-018-3926-8

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