Abstract
Purpose: :
To characterize a spontaneous model of autoimmune uveoretinitis (EAU) in mice expressing hen egg lysozyme (HEL) under the control of the IRBP promoter and a transgenic HEL–specific T cell receptor (TCR).
Methods: :
Transgenic mice expressing membrane–bound HEL under the control of the IRBP–promoter were generated. HEL expression was detected with RT–PCR and confocal microscopy. The mice were crossed with HEL–specific TCR transgenic mice (3A9) and EAU scored histologically. The inflammatory infiltrates were characterized in frozen eye sections using confocal microscopy and the lymphocytes in the eye and the lymphoid tissues with flow cytometry. 3A9 T cells were isolated from spleens and lymph nodes using CD4 microbeads and activated in vitro with 10–6M HEL and non–transgenic splenocytes. Cytokines were measured in culture supernatants using the CBA assay. For in vivo proliferation assay the cells were labeled with 5 µM CFSE. Adoptive transfers were intraperitoneal.
Results: :
HEL was expressed at high levels in the photoreceptors of IRBP–HEL mice and at lower quantities in thymus. Crossed with the 3A9 mice, double–transgenics spontaneously developed EAU at 22–24 days of age despite severe thymic deletion of HEL–specific CD4+ cells. The disease quickly progressed to grade 4 EAU, which persisted into late adulthood. The pathology comprised patchy lesions with CD4+ T cells, MHC–II+ CD11c+dendritic cells, F4/80+ macrophages and a few CD8+ T cells. The CD4+ infiltrate consisted of Th1–polarised cells with medium to low avidity for HEL, which also had a higher activation status in the peripheral lymphoid tissues. EAU could be induced in the single–transgenic IRBP–HEL mice with a transfer of as few as 1x105 in vitro HEL–activated CD4+ 3A9 T cells, but not with anti–CD3/CD28 activated 3A9 T cells. Naïve 3A9 CD4+ cells did not transfer EAU to and showed no signs of division in HEL+ recipients, although present at frequencies comparable to those found in the double transgenic mice at the time of EAU onset.
Conclusions: :
The IRBP–HEL double–transgenic mice developed a chronic early–onset EAU, which progressed to total retinal destruction after several weeks. The physiology of the double transgenic mouse may be permissive to the development of EAU, as naïve T cells could not transfer EAU to single–tg IRBP–HEL mice. The predictable time of disease onset and its chronic duration makes this model suitable for dissecting the mechanism of EAU initiation and testing of therapeutics aimed at preventing and controlling ocular inflammation.
Keywords: autoimmune disease • uveitis-clinical/animal model • transgenics/knock-outs