July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Pigment Epithelium Derived Factor Secreted by Corneal Epithelial Cells Regulates Dendritic Cell Maturation in Dry Eye Disease.
Author Affiliations & Notes
  • Rohan Bir Singh
    Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States
  • Purushottam Jha
    Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States
  • Tomas Blanco
    Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States
  • Afsaneh Amouzegar
    Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States
  • Reza Dana
    Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States
  • Footnotes
    Commercial Relationships   Rohan Singh, None; Purushottam Jha, None; Tomas Blanco, None; Afsaneh Amouzegar, None; Reza Dana, None
  • Footnotes
    Support  NIH R01 EY20889
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4279. doi:
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      Rohan Bir Singh, Purushottam Jha, Tomas Blanco, Afsaneh Amouzegar, Reza Dana; Pigment Epithelium Derived Factor Secreted by Corneal Epithelial Cells Regulates Dendritic Cell Maturation in Dry Eye Disease.. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4279.

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

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Abstract

Purpose : Pigment Epithelium Derived Factor (PEDF)/SERPINf1 is a 50kD protein secreted by various tissues in the eye and has known neurotropic and anti-angiogenic functions in the cornea. In this study, we evaluated the expression of PEDF by corneal epithelial cells (CEpCs) and investigated its immunomodulatory effects on dendritic cell maturation using a validated murine model of dry eye disease (DED).

Methods : DED was induced in six to eight-week-old female C57BL/6 mice (N=20) using a controlled environment chamber for 7 days. The expression of PEDF in CEpCs from naïve and DED mice was confirmed using immunohistochemistry (IHC) staining (N=4) and quantified using real-time PCR (N=4) and ELISA (N=4). CEpCs from either naïve (N=8) or DED mice (N=8) were cultured with bone marrow-derived dendritic cell (BMDCs) in the presence of IFNγ (100ng/ml) for 48 hours. The frequencies of MHC IIhi and CD80+ BMDCs were determined using flow cytometry. Next, either recombinant PEDF (rPEDF, 200ng/ml) or PEDF receptor blocking antibody (anti-PEDF-R, 200ng/ml) was added to the co-culture, and BMDC expressions of above activation markers were evaluated.

Results : Our results demonstrated a 6.7-fold increase in SERPINf1 mRNA expression in CEpCs from DED mice compared to naïve mice (p=0.025). Our ELISA results showed a 1.7-fold increase in PEDF protein levels in CEpCs from DED mice as compared to naïve mice (3.4±0.343 ng/ml vs. 1.924±0.085 ng/ml, p=0.05). The frequencies of MHCIIhi and CD80+ BMDCs cultured in the presence of CEpCs from naïve mice were lower than BMDCs cultured with CEpCs from DED mice (MHCIIhi :42.8 ±1.45% vs.46.2±1.36%, p=0.134; CD80+:26.7±0.29% vs. 27.8±2.24%, p=0.783). Addition of rPEDF to the co-culture system resulted in a significant decrease in frequencies of MHC-IIhi (36.7±1.04% vs. 42.8±1.45%, p=0.017) and CD80+(23.8±0.38% vs. 26.7±0.29%, p=0.04) BMDCs. The addition of anti PEDF-R blocking antibody abrogated the suppressive effect on frequencies of MHC IIhi (45.2±0.87% vs. 42.8±1.45%, p<0.001) and CD80+ (30.8±1.56% vs. 23.8±0.38%, p=0.018) BMDCs.

Conclusions : Our results suggest that PEDF secreted by corneal epithelial cells inhibits dendritic cell maturation in response to desiccating stress. These findings indicate a possible immunoregulatory role for corneal epithelium-derived PEDF in DED.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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