Human ocular-derived virus-specific CD4+ T cells control varicella zoster virus replication in human retinal pigment epithelial cells.

PURPOSE: Varicella zoster virus (VZV)-induced retinitis is characterized by the presence of virus-infected cells in the retinal layer and the ocular infiltration of VZV-specific T cells. Herein, the susceptibility of human retinal pigment epithelial (RPE) cells to VZV infection and the ability of virus-specific CD4(+) T cells to control VZV infection in RPE cells in vitro is addressed. METHODS: Human primary RPE cell cultures (n=2) were infected with a VZV strain expressing green fluorescent protein. The infection and viability of infected RPE cells was monitored by flow cytometry or by a fluorescent imager on RPE monolayers. RPE cells, pretreated with or without interferon-gamma (IFN-gamma), were infected with VZV and subsequently cultured with VZV-specific CD4(+) T-cell clones (TCCs; n=3) recognizing disparate VZV proteins presented by different HLA class II alleles. IFN-gamma production and cytotoxicity of the TCCs in response to VZV-infected RPE cells was determined by flow cytometry. RESULTS: Human RPE cells are permissive to a productive VZV infection. VZV-infected RPE cells presented the cognate antigen to the CD4(+) TCCs only if the RPE cells were pretreated with IFN-gamma and expressed the appropriate HLA class II allele. VZV-specific TCCs inhibited productive VZV infection in RPE cells, which was in part attributed to TCC-mediated killing of the VZV-infected RPE cells. CONCLUSIONS: The results presented suggest that RPE cells may play a role as retina-resident antigen-presenting cells in the intraocular, VZV-specific, T cell-mediated inflammatory response of VZV-induced uveitis.

Identification of viral antigens recognized by ocular infiltrating T cells from patients with varicella zoster virus-induced uveitis.

PURPOSE: Varicella zoster virus (VZV) is a common cause of infectious uveitis associated with an intraocular inflammatory response involving virus-specific T cells. In the current study, the functional characteristics and the antigen specificity of VZV-reactive T cells recovered from intraocular fluid (IOF) samples of five patients with VZV were determined. METHODS: B-cell lines were infected with a comprehensive panel of recombinant vaccinia viruses expressing 11 individual VZV open reading frames (ORFs), or alternatively pulsed with the corresponding peptides to generate antigen-presenting cells (APCs). T-cell responsiveness of the IOF-derived VZV-specific T cells toward APCs was monitored by interferon (IFN)-gamma enzyme-linked immunosorbent spot-forming assays on bulk T-cell cultures and subsequently T-cell clones (TCCs). The cytokine-secretion profile and cytotoxicity of the VZV-specific TCCs was determined by ELISA and flow cytometry, respectively. RESULTS: T-cell reactivity to VZV proteins encoded by ORF4, -10, -14, -18, -29, -31, -61, -62, -63, -67, and -68 was demonstrated, but specificity varied individually. T-cell epitopes on ORF62 and -68 were delineated. The TCCs secreted IFNgamma, but relatively low levels of interleukin-4 and -5, in response to VZV antigen-expressing APCs. The TCCs induced antigen-specific cytotoxic T-cell activity. CONCLUSIONS: The results suggest that the intraocular VZV-specific T-cell response in the patients with VZV analyzed is directed to a broad spectrum of VZV antigens, including the latency-associated VZV proteins from ORFs 4, 29, 63, and particularly ORF62. This local T-cell response was in part mediated by cytotoxic CD4(+) T cells with a Th1/0-like effector memory phenotype.

No evidence for circulating HuD-specific CD8+ T cells in patients with paraneoplastic neurological syndromes and Hu antibodies.

AIM: In paraneoplastic neurological syndromes (PNS) associated with small cell lung cancer (SCLC) and Hu antibodies (Hu-PNS), Hu antigens expressed by the tumour hypothetically trigger an immune response that also reacts with Hu antigens in the nervous system, resulting in tumour suppression and neuronal damage. To gain more insight into the hypothesized CD8(+ )T cell-mediated immune pathogenesis of these syndromes, we searched for circulating HuD-specific CD8(+) T cells in a large cohort of Hu-PNS patients and controls. PATIENTS AND METHODS: Blood was tested from 43 Hu-PNS patients, 31 Hu antibody negative SCLC patients without PNS and 54 healthy controls. Peripheral blood mononuclear cells (PBMC) were stimulated with HuD protein-spanning peptide pools (15-mers) and individual HuD-derived peptides (9-mers) and analysed by cytokine flow cytometry and interferon-gamma ELISPOT-assays. Additionally, HuD-based Class I HLA multimers were used to visualize HuD-specific CD8(+) T cells. RESULTS: No HuD-specific CD8(+ )T cells could be detected in the blood of Hu-PNS patients or controls. CONCLUSIONS: Our results do not support a role for HuD-specific CD8(+) T cells in Hu-PNS. Further studies should focus on the detection of circulating HuD-specific CD4(+ )T cells and examine the antigen specificity of T cells in affected tissues.

Selective retention of herpes simplex virus-specific T cells in latently infected human trigeminal ganglia.

Primary infection with herpes simplex virus 1 (HSV-1) and varicella zoster virus (VZV) results in lifelong latent infections of neurons in sensory ganglia such as the trigeminal ganglia (TG). It has been postulated that T cells retained in TG inhibit reactivation of latent virus. The acquisition of TG specimens of individuals within hours after death offered the unique opportunity to characterize the phenotype and specificity of TG-resident T cells in humans. High numbers of activated CD8(+) T cells expressing a late effector memory phenotype were found to reside in latently infected TG. The T cell infiltrate was oligoclonal, and T cells selectively clustered around HSV-1 but not VZV latently infected neurons. Neuronal damage was not observed despite granzyme B expression by the neuron-interacting CD8(+) T cells. The TG-resident T cells, mainly CD8(+) T cells, were directed against HSV-1 and not to VZV, despite neuronal expression of VZV proteins. The results implicate that herpesvirus latency in human TG is associated with a local, persistent T cell response, comprising activated late effector memory CD8(+) T cells that appear to control HSV-1 latency by noncytolytic pathways. In contrast, T cells do not seem to be directly involved in controlling VZV latency in human TG.

Characterization of the varicella zoster virus (VZV)-specific intra-ocular T-cell response in patients with VZV-induced uveitis.

Varicella zoster virus (VZV) is a well-known cause of infectious uveitis. The aim of this study was to characterize the VZV-specific T-cell response in eyes of patients with VZV-induced uveitis. T-cell lines (TCL) were generated by mitogenic stimulation of intra-ocular fluid (IOF) samples obtained from eight patients with VZV-induced uveitis. Two patients with herpes simplex virus (HSV)-induced uveitis were included as disease controls. Characterization of individual T-cells in the TCL was performed by stimulating the TCL with mock, HSV-1 and VZV antigen pulsed autologous B cells and subsequent flow cytometric analyses. Virus specificity and phenotype of the T-cells were identified by simultaneous detection of intracellular gamma interferon and cell surface markers CD4, CD8, CD3 or T-cell receptor (TCR) beta chain variable region (TCRBV) usage. The TCL obtained from patients with HSV-1-induced uveitis contained higher numbers of T-cells reactive to HSV-1 compared to VZV. VZV-specific T-cells were detected in all TCL of the patients diagnosed with VZV-induced uveitis. Four out of six TCL obtained from patients with VZV-induced uveitis that were assayed for both viruses, contained higher numbers of T-cells reactive to VZV compared to HSV-1. Detailed analyses of the TCL of two patients demonstrated that the VZV reactivity within the assayed TCL was dominated by T-cells expressing one specific TCRBV gene. The data implicate that VZV-reactive T-cells infiltrate and participate in the local inflammatory response in eyes of patients with VZV-induced uveitis.