July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
Immunomodulatory Activity of Tear Protein Lacritin
Author Affiliations & Notes
  • Afsaneh Amouzegar
    Department of Ophthalmology/Harvard Medical School, Schepens Eye Research Institute, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Sharad Mittal
    Department of Ophthalmology/Harvard Medical School, Schepens Eye Research Institute, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Mingshun Li
    Department of Ophthalmology/Harvard Medical School, Schepens Eye Research Institute, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Robert L McKown
    Department of Integrated Science and Technology, James Madison University, Harrisonburg, Virginia, United States
  • Gordon W Laurie
    Departments of Cell Biology, Ophthalmology and Biomedical Engineering, University of Virginia, Charlottesville, Virginia, United States
  • Sunil Chauhan
    Department of Ophthalmology/Harvard Medical School, Schepens Eye Research Institute, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Footnotes
    Commercial Relationships   Afsaneh Amouzegar, None; Sharad Mittal, None; Mingshun Li, None; Robert McKown, TearSolutions, Inc. (R); Gordon Laurie, TearSolutions, Inc. (F), UVa Licensing and Ventures Group (P); Sunil Chauhan, None
  • Footnotes
    Support  R01EY024602 (SC) and R01EY024327 (GWL)
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 3323. doi:
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      Afsaneh Amouzegar, Sharad Mittal, Mingshun Li, Robert L McKown, Gordon W Laurie, Sunil Chauhan; Immunomodulatory Activity of Tear Protein Lacritin. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3323.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Lacritin is a protein found in tear that has been shown to exert cytoprotective effects on human corneal epithelial cells and increase epithelial cell proliferation under stress. Despite the established role of Lacritin in corneal epithelial cell homeostasis, whether Lacritin is capable of acting directly on immune cells has not been studied. The purpose of this study was to investigate the effect of Lacritin on immune cell function, including conventional and regulatory T cell subsets and macrophages.

Methods : CD4+ T cells were isolated and pooled from the spleen and cervical draining lymph nodes of C57BL/6 mice using magnetic sorting. Isolated cells were stimulated with anti-CD3 and anti-CD28 antibodies and cultured in the presence or absence of 4μM recombinant Lacritin or control recombinant albumin protein for 72 hours. T cell proliferation was assessed using BrdU incorporation assay. The frequencies of CD4+Foxp3+ regulatory T cells (Tregs) among cultured cells were determined using flow cytometry. F4/80+ macrophages were isolated from mice by peritoneal lavage with ice-cold PBS. Isolated cells were stimulated with IFNγ and cultured in the presence or absence of Lacritin or control protein for 48 hours. Frequencies of MHC-IIhi macrophages were evaluated using flow cytometry.

Results : Stimulation with anti-CD3 and anti-CD28 led to an approximately 9-fold increase in CD4+ T cell proliferation (mean±SEM of optical density (OD) values: 8443±258 vs. 951±182; p=0.0001). There was a 13% reduction in CD4+ T cell proliferation in cultures treated with Lacritin compared to stimulated T cells cultured with control protein (OD: 7378±575 vs. 8443±258; p=0.16). In addition, Lacritin-treated cultures showed an 11% increase in frequencies of CD4+Foxp3+ Tregs compared to control-treated cultures (6.12% vs. 5.50%). Furthermore, Lacritin led to a significant (48%) decrease in the frequencies of MHC-IIhi macrophages compared to the control group (49.4±2.1% vs. 95.8±1.6%, p=0.002).

Conclusions : Lacritin significantly suppresses macrophage activation and exerts a moderate regulatory effect on CD4+ T cell proliferation and Treg expansion. These findings indicate a possible immunoregulatory role for Lacritin in inflammatory diseases of the ocular surface.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

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