June 2020
Volume 61, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2020
Plasmacytoid Dendritic Cells Prevent T Regulatory Cell (Tregs) Re-programming to Effector Ex-Tregs in Herpes Simplex Keratitis
Author Affiliations & Notes
  • Deshea L Harris
    Center for Translational Ocular Immunology and Department of Ophthalmology, Tufts Medical Center and Tufts University School of Medicine, Boston, Massachusetts, United States
  • Arsia Jamali
    Center for Translational Ocular Immunology and Department of Ophthalmology, Tufts Medical Center and Tufts University School of Medicine, Boston, Massachusetts, United States
  • Maria J. Lopez
    Center for Translational Ocular Immunology and Department of Ophthalmology, Tufts Medical Center and Tufts University School of Medicine, Boston, Massachusetts, United States
  • Gustavo Ortiz
    Center for Translational Ocular Immunology and Department of Ophthalmology, Tufts Medical Center and Tufts University School of Medicine, Boston, Massachusetts, United States
  • Pedram Hamrah
    Center for Translational Ocular Immunology and Department of Ophthalmology, Tufts Medical Center and Tufts University School of Medicine, Boston, Massachusetts, United States
    Cornea Service, New England Eye Center, Boston, Massachusetts, United States
  • Footnotes
    Commercial Relationships   Deshea Harris, None; Arsia Jamali, None; Maria Lopez, None; Gustavo Ortiz, None; Pedram Hamrah, None
  • Footnotes
    Support  NIH-R01- EY022695 and NIH-R01- EY022695-S1, NIH-R01- EY026963 , NIH-R21- EY025393 , Massachusetts Lions Eye Research Fund, Inc., Research to Prevent Blindness Challenge Grant and Tufts Medical Center Institutional Support (PH)
Investigative Ophthalmology & Visual Science June 2020, Vol.61, 2324. doi:
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    • Get Citation

      Deshea L Harris, Arsia Jamali, Maria J. Lopez, Gustavo Ortiz, Pedram Hamrah; Plasmacytoid Dendritic Cells Prevent T Regulatory Cell (Tregs) Re-programming to Effector Ex-Tregs in Herpes Simplex Keratitis. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2324.

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

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Abstract

Purpose : Despite our recent understating on the role of pDCs in limiting herpes simplex virus (HSV)-1 keratitis, their significance in regulating Treg survival remains to be elucidated.

Methods : Bone marrow chimeric mice were generated by irradiating and reconstituting wildtype C57BL/6 mice by adoptive transfer of bone marrow cells from BDCA2-DTR (pDCs can be ablated by administration of diphtheria toxin [DT]) and Foxp3-eGFP/cre×Rosa-tdTomato mice (Foxp3+ Tregs are GFP+ tdTomato+ and Foxp3neg ex-Tregs are converted to eGFPneg tdTomato+) enabling concurrent depletion of pDCs and assessing Treg fate. pDC depletion was performed by subconjunctival (sconj.) injection of 30ng DT prior to HSV-1 inoculation and was repeated every 48 hours. Sconj. injection of PBS served as control. HSV-1 keratitis was induced by inoculation of 2×106 PFU of HSV-1 strain McKrae on scarified corneas, 4 weeks after generation of chimeric mice. Corneas and draining lymph nodes (dLNs) were evaluated by confocal microscopy and flow cytometry, respectively. Co-cultures of pDCs and Tregs under treatment with 105 PFU UV-inactivated HSV-1 were performed. T test was used to compare the groups. P<0.05 was considered significant.

Results : Confocal microscopy of whole-mounted corneas showed that pDC depletion resulted in a higher density of infiltrating eGFPneg Tdtomato+ ex-Tregs compared to sham depletion in both the peripheral (89.6±20.4 cells/mm2 vs 17.3±6.6; p<0.001) and central (20.8±9.6 vs. 3.3±1.9; p<0.001) corneas on day 7 following HSV-1 inoculation. Similarly, pDC depletion lead to 2.5-fold higher density of eGFPnegTdtomato+ ex-Tregs in the dLNs, suggesting that pDC depletion facilitates re-programming of Foxp3+Tregs to Foxp3neg ex-Tregs. Further phenotyping of Foxp3neg ex-Tregs in the dLNs showed that in both control and pDC-depleted mice, the majority of Foxp3neg ex-Tregs expressed IFN-γ, suggesting their re-programming to effector T cells. Co-culture of splenic Tregs with different densities of splenic pDCs under stimulation of UV-irradiated HSV-1 showed that while in the presence of UV-irradiated HSV-1, Tregs converted to ex-Tregs, the presence of 1:10 and 1:5 densities of pDCs prevented their re-programming to effector ex-Tregs by 2.6-folds and 3.4-folds, respectively.

Conclusions : pDCs prevent re-programing of Tregs to effector ex-Tregs in the dLNs and promote Treg survival during HSV keratitis.

This is a 2020 ARVO Annual Meeting abstract.

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