Purchase this article with an account.
Deborah Aviya Lipski, Remi Dewispelaere, Vincent Foucart, Laure E Caspers, Matthieu Defrance, Catherine Anne Bruyns, François Willermain; MHC class II Expression In The Retina During Experimental Autoimmune Uveitis. Invest. Ophthalmol. Vis. Sci. 2017;58(8):544.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Controversy exists regarding which cell types are responsible for auto-antigen presentation in the retina during experimental autoimmune uveitis (EAU) development. In this study, we use a combination of techniques to characterize retinal resident and infiltrating cells susceptible to express MHC Class II (MHCII).
EAU is induced by adoptive transfer of IRBP1-20-specific activated T cells in female C57BL/6 mice aged 6-8 weeks. At day 21, cryosections are prepared and MHCII, GFAP, endoglin or CD31 and IBA-1 expression analyzed by immunofluorescence (IF). For flow cytometry analysis (FC), retinas are processed into single cell suspensions by enzymatic digestion and tested for MHCII, CD11b, CD45, Ly6C, CD31, CD40, CD80 and CD86 membrane expression using specific antibodies coupled to different fluorochromes. RNA-Seq is performed on different retinal cell populations sorted by FC.
IF images demonstrate a strong induction of MHCII expression in EAU, especially in the inner retina at the level of vasculitis, extending to the outer nuclear layer in severely inflamed eyes, at the level of the retinal pigment epithelium and on cells infiltrating the vitreous. Most MHCII+ cells express the hematopoietic marker IBA-1. FC analysis demonstrates that retinal cell MHCII expression is significantly correlated with disease severity (p<0,001, N>20) and associated with upregulation of co-stimulatory molecules CD40, CD80 and CD86. Although FC studies pick up a CD45-CD11b- cell population with low MHCII expression, they confirm that most cells expressing high levels of MHCII are of hematopoietic origin. Both Ly6C+ and Ly6C- cells contribute to the increase in MHCII+CD45+CD11b+ cells that occurs during EAU, possibly corresponding to macrophage infiltration and resident microglial cell proliferation. RNA-Seq analysis of both FC-sorted cell populations leads to a clear sample clustering with some differential inflammatory gene expression. However, no clear difference in cell lineage is evidenced.
MHCII induction during EAU is correlated to disease severity and accompanied by upregulation of co-stimulatory molecule expression. Hematopoietic cells express higher levels of MHCII than non-hematopoietic cells and various levels of Ly6C. In these experimental conditions, Ly6C expression seems to be associated with different expression of some inflammatory genes but not to a single specific cell type.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
This PDF is available to Subscribers Only