June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Lack of Goblet Cells in SPDEF KO Mice Increases Retention of IL-12 Producing Dendritic Cells in Conjunctiva
Author Affiliations & Notes
  • Byung Yi Ko
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
    Ophthamlology, Konyang University Hospital and College of Medicine, Daejeon, Korea (the Republic of)
  • Yangyan Xiao
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
    Second Xiangya Hospital, Central South University, Changsha, Hunan, China
  • Fang Bian
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
  • Jeffrey Whitsett
    Cincinnati Children’s Hospital, Cincinnati, Ohio, United States
  • Hans Clever
    Hubrecht Institute, UTRECHT, Netherlands
  • Flavia L Barbosa
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
  • Cintia S De Paiva
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
  • Stephen C Pflugfelder
    Department of Ophthalmology, Baylor College of Medicine , Houston, Texas, United States
  • Footnotes
    Commercial Relationships   Byung Yi Ko, None; Yangyan Xiao, None; Fang Bian, None; Jeffrey Whitsett, None; Hans Clever, None; Flavia Barbosa, None; Cintia De Paiva, None; Stephen Pflugfelder, None
  • Footnotes
    Support  NIH/NEI EY011915 (SCP), NIH Training Grant T32-AI053831 (FB), RPB Research to Prevent Blindness, The Oshman Foundation, William Stamps Farish Fund and The Hamill Foundation; (the Cytometry and Cell Sorting Core at Baylor College of Medicine - the NIH NIAID P30AI036211, NCI P30CA125123, and NCRR S10RR024574).
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 463. doi:
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    • Get Citation

      Byung Yi Ko, Yangyan Xiao, Fang Bian, Jeffrey Whitsett, Hans Clever, Flavia L Barbosa, Cintia S De Paiva, Stephen C Pflugfelder; Lack of Goblet Cells in SPDEF KO Mice Increases Retention of IL-12 Producing Dendritic Cells in Conjunctiva. Invest. Ophthalmol. Vis. Sci. 2017;58(8):463.

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

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Abstract

Purpose : Goblet cell (GC) mucins have been found to condition tolerogenic properties in intestinal dendritic cells (DCs). Aqueous tear deficiency is associated with GC loss and greater IFN-γ expression in the conjunctiva (Cj). We hypothesize that loss of Cj GCs is associated with greater dendritic cell T helper (Th)1 polarization in the SPDEF knock-out (KO) strain that lacks GCs.

Methods : Six to nine week-old female C57BL/6 wild type (WT) and SPDEF KO mice were used. CD11c+ and CD11b+ DCs were visualized in whole mount Cj by laser scanning confocal microscope using the Z-stack option. IL-12 expression in DC was quantified by flow cytometry and IL-12+ cells were also detected by immunostaining in Cj cryosections. Mixed lymphocyte reactions to evaluate T helper (Th) cell polarization and proliferation were performed with OVA 323-339 peptide pulsed cervical lymph node (CLN) cell suspensions and OT II CD4+ T cells. Th cytokine expression (IFN-γ and IL-17) was measured by flow cytometry and immunobead assay. The effects of WT conjunctival conditioned medium (CJCM) on lipopolysaccharide (LPS) -stimulated IL-12 expression in the Cj was evaluated after topical application in SPDEF KO mice.

Results : SPDEF KO had greater frequency of CD11b+CD11c- (p=0.01) and CD11b-CD11c+ conjunctival DCs and CD11b-CD11c+ DCs were more superficially located in WT mice. Immunohistochemistry demonstrated that SPDEF KO mice had a greater number of IL-12+ cells in Cj stroma than WT group. A greater percentage of CD11b+CD11c- and CD11b+CD11c+ cells (p ≤ 0.03) with higher IL-12 median fluorescent intensity (MFI) (both p ≤ 0.03) in the Cj compared to WT control was demonstrated by flow cytometry. Th cells primed by SPDEF KO CLN DCs showed a skewing towards Th1 and generated a lower frequency of CD4+Foxp3+ cells. Significantly increased IFN-g was measured in the supernatant of these cells (p=0.03). Following topical LPS stimulation, SPDEF KO mice had higher IL-12 expression in CD11b+CD11c+ Cj DCs than the control group (p=0.02) and reconstitution with topically applied WT CJCM reduced stimulated IL-12 production in CD11b+CD11c+ cells compared to vehicle control (p=0.04).

Conclusions : Our results support our hypothesis that CJ GCs have a tolerogenic effects on DCs by suppressing IL-12 production and Th1 polarization.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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