Intraocular T Cells of Patients with Herpes Simplex Virus (HSV)–Induced Acute Retinal Necrosis Recognize HSV Tegument Proteins VP11/12 and VP13/14

Abstract

It has previously been shown that T cells specific for the triggering virus infiltrate the eye of patients with herpes simplex virus type 1 (HSV-1)-induced acute retinal necrosis (ARN). The T cells were mainly directed against 0.67-0.73 HSV-1 map region encoded antigens. The fine specificities of genetically different T cell clones (TCC), obtained from affected eyes of 3 patients with HSV-induced ARN and reactive toward this genomic region of HSV-1, were analyzed with recombinant HSV viruses and synthetic peptides. For 1 patient, the HSV-1 UL46 gene encoded tegument protein VP11/12 was identified as the target antigen. Two separate CD4(+) T cell epitopes were defined in VP11/12. TCC from the other 2 patients recognized the HSV-1 UL47 gene encoded tegument protein VP13/14. Two separate CD4(+) VP13/14 T cell epitopes were identified in these patients. Analysis of the data indicates that HSV-1 VP11/12 and VP13/14 are major target antigens for T cells obtained from vitreous fluid samples of the HSV-induced ARN patients studied.

Functional Characterization of Ocular-Derived Human Alphaherpesvirus Cross-Reactive CD4 T Cells

“…21), which recognizes HSV-1, but not HSV-2 and VZV (Ref. 20; data not shown). Analogous to TCC 53, TCC 44 did not inhibit HSV-1 replication in RPE cells (Fig.…”

“…First, BLCL were infected overnight with HSV at a multiplicity of infection (MOI) of 10 or pulsed overnight with predefined optimal concentrations of protein lysates obtained from VZV-, HSV-, and mock-infected cells. Second, BLCL were pulsed overnight with the indicated synthetic peptides in RPMI 1640 medium (2 mM or as indicated) (20). Third, BLCL were nucleofected with pEGFP-N1 control plasmid and plasmids expressing HSV-1 ICP4 aa 945-1054 (pEGFP-N1/HSV-1: ICP4 945-1054 ) or HSV-2 ICP4 aa 972-1082 (pEGFP-N1/HSV-2:ICP4 972-1082 ) using Amaxa Cell Line Nucleofector kit V (Lonza).…”

“…IFN-g ELISPOT assays were performed in triplicate, according to the manufacturer's instructions (Mabtech AB). Briefly, T cells (5-20 3 10 3 cells/well) were preincubated with BLCL (2 3 10 4 cells/well) for 1.5 h in U-bottom 96-well plates (Greiner Bio-One) at 37˚C, followed by an additional 6 h incubation at 37˚C in anti-IFN-g precoated Multiscreen HTS-IP ELISPOT plates (Millipore) (20). T cells were washed away, and the secondary anti-IFN-g mAb, followed by the streptavidin-alkaline-phosphatase mAb were added to the wells.…”

“…The HLA-II genotype of the uveitis patients and RPE cell lines, and the HLA-II restriction of TCC were previously determined (17,18,20). The RPE 171 cell line expresses HLA-DRB1*0301;0701 compatible with the HLA-DRB1*0703 restriction element of TCC 53, whereas RPE 172 cells express HLA-DRB1*1201;1501 compatible with the HLA-DRB1*1501 restriction element of TCC 44 (17,18,20). Human RPE cells were infected with HSV-1.GFP-VP16 (5 3 10 3 PFU/well), resulting in a MOI of 0.5-1, or incubated with VZV.GFP-66-infected MeWo cells (ratio of approximately one infected cell to four uninfected cells) for 4 h at 37˚C.…”

“…Studies on the experimental HSV-1 ARN mouse model have demonstrated the pathogenic role of ocular infiltrating T cells (11,16). The intraocular virus-specific T cell response in aHHV uveitis patients, mediated by cytotoxic CD4 and CD8 Th0/Th1-like T cells, is preferentially directed to the causative herpesvirus (17)(18)(19)(20)(21). Herpesvirus particles and Ags have been detected in the neural retina and retinal pigment epithelial (RPE) cell layer of patients with HSV and VZV posterior uveitis (22)(23)(24).…”