March 2012
Volume 53, Issue 14
ARVO Annual Meeting Abstract  |   March 2012
Role for Vitamin D3 at the Ocular Surface during Inflammation
Author Affiliations & Notes
  • Rose Y. Reins
    College of Optometry, Univ of Houston, Houston, Texas
  • Alison M. McDermott
    Optometry, University of Houston, Houston, Texas
  • Footnotes
    Commercial Relationships  Rose Y. Reins, None; Alison M. McDermott, None
  • Footnotes
    Support  Lions Foundation for Sight Grant
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 5241. doi:
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      Rose Y. Reins, Alison M. McDermott; Role for Vitamin D3 at the Ocular Surface during Inflammation. Invest. Ophthalmol. Vis. Sci. 2012;53(14):5241.

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

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Purpose: : In addition to a role in calcium homeostasis, vitamin D (D3) has many diverse functions, including an immunomodulatory role during infection and inflammation. However, relatively little is known about D3 function at the ocularsurface. The purpose of these studies was to determine corneal expression of D3 regulatory enzymes, modulation of these enzymes via Toll-like receptor (TLR) activation, cytokine treatment, and hyperosmolar stress (HOS), and the effect of D3 on antimicrobial peptide (AMP) production and cytokine modulation during inflammation.

Methods: : Primary human corneal epithelial cells (HCEC) or telomerase immortalized HCEC were stimulated with 10-7M D3, 25D3, or 1,25D3 for 24 hrs and expression of LL-37, CD14, and CYP24A1 analyzed by RT-PCR and immunoblot. Human corneas were incubated with 10-7M 1,25D3 for 24 hrs, then frozen for immunohistochemistry, or epithelial and endothelial RNA was collected to determine AMP (LL-37, hBD2) expression. For inflammatory stimuli, HCEC were treated with 1μg/ml agonist for TLR 2/1, 6/2, 3, or 5, IL-1β(10ng/ml), or HOS (400, 450, or 500 mOsm/kg) for 24 hrs and expression of D3 hydroxylases analyzed by RT-PCR. Cytokine production by HCEC treated with PolyI:C (20μg/ml) and 1,25D3 (10-7M) was determined by RT-PCR and ELISA. Supernatants from HCEC incubated with 1,25D3 (10-7M) were used in an antimicrobial assay against Pseudomonas aeruginosa (PA) 19660. All experiments were performed three times.

Results: : HCEC expressed hydroxylases CYP27B1 and CYP24A1 necessary for D3 activation and inactivation respectively, and the vitamin D receptor. Stimulating HCEC with all three D3 metabolites led to induction of LL-37, CD14, and CYP24A1, while both epithelium and endothelium from stimulated donor corneas had increased (5-11 fold) expression of hBD-2. All TLR agonists tested and IL-1β increased HCEC expression of CYP24A1 (3-30 fold) and CYP27B1 (3-15 fold), with TLR3 agonist PolyI:C having the greatest effect. HCECexposed to HOS upregulated CYP24A1 17±1.8 fold above control. 1,25D3downregulated IL-8 and TNFα induced by PolyI:C. In an in vitro antimicrobial assay, supernatants from D3 treated HCEC inhibited the growth of PA by 40 ± 2.5% compared to control supernatants.

Conclusions: : HCEC respond to D3 stimulation, as shown by enhanced expression of AMPs, and culture conditions mimicking inflammation induced the expression of hydroxylases important for regulating hormone activity. Further, D3 suppressed proinflammatory cytokine levels produced by TLR3 activation and enhanced HCEC antimicrobial activity. These results suggest an important role for D3 during ocular surface infection, inflammation, and stress.

Keywords: cornea: epithelium • cornea: basic science • inflammation 

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