G. Seerp Baarsma

Detection of intraocular antibody production to herpesviruses in acute retinal necrosis syndrome

In order to improve the determination of the causative agent in acute retinal necrosis syndrome, we evaluated the detection of intraocular antibody production to herpesviruses in 28 patients with this disease. Intraocular antibody production was determined by calculation of the Goldmann-Witmer coefficient whereby specific antibody titers in the inflamed eye and circulation are related to the total IgG content in ocular fluid and serum. Specific antibody titers to herpesviruses and Toxoplasma were determined by the indirect immunofluorescence technique. Thirty-five patients with ocular toxoplasmosis, cataract, or proliferative vitreoretinal disorders were tested as controls. By this technique, intraocular antibody production to varicella zoster virus or herpes simplex virus could be established in 16 (57%) of the patients with the typical clinical features of acute retinal necrosis, compared to none of the controls. Of the 33 affected eyes, 21 (64%) had a visual outcome of less than 20/200. We concluded that detection of intraocular antibody production to herpesviruses may be a useful diagnostic tool in establishing the causative agents in acute retinal necrosis.

Vγ9Vδ2 T cells recovered from eyes of patients with Behçet's disease recognize non-peptide prenyl pyrophosphate antigens

The phenotype and antigen-specificity of T cells expanded by mitogenic stimulation from intra-ocular fluid (IOF) samples of affected eyes of six Behcets disease (BD) patients, and seven patients with other uveitis entities, were determined. High numbers of γδ T cells, predominantly Vγ9Vδ2 T cells, were only detected in the IOF-derived TCL of three BD patients. Whereas no TCL responded to heat shock protein (HSP) 65 kDa, reactivity to isopentyl pyrophosphate (IPP) and related non-peptide prenyl pyrophosphates (PPP) was restricted to the γδ T cell containing TCL. Upon IPP stimulation, these TCL secreted IFN-γ but no IL-4. By single-cell analysis of intracellular IFN-γ production and CD69 expression the IOF-derived IPP-specific T cells were identified as CD4−CD8− γδ T cells. The data presented suggest the infiltration of PPP-specific Vγ9Vδ2 Th1-like cells into the eye of BD patients with uveitis.

The neonatal Fc receptor is expressed by human retinal pigment epithelial cells and is downregulated by tumour necrosis factor-alpha

Background/aims The neonatal Fc receptor (FcRn) protects immunoglobulin G (IgG) from catabolism, controls its transport between cell layers and extends its serum half-life. In the human, vitreous IgG can be found, but how vitreous IgG is processed or transported is currently unknown. The FcRn is a candidate molecule to regulate these processes. The authors examined FcRn expression and regulation in human retinal pigment epithelium (RPE) cells. Methods In three primary RPE cell cultures (from three donor eyes) and in the human RPE cell line ARPE-19, FcRn and beta-2-microglobulin (β2M) mRNA levels were determined by real-time quantitative PCR. FcRn protein expression was analysed by western blot studies. Stimulation experiments were performed with recombinant human tumour necrosis factor (TNF)-α and interferon (IFN)-γ. HT-29, THP-1 and HeLa cell lines were used as FcRn positive and negative non-ocular controls, respectively. Results Expression of FcRn mRNA and protein was demonstrated in all three RPE cultures. After stimulation with TNF-α, FcRn expression is downregulated in RPE cells and upregulated in HT-29 and THP-1 cells. IFN-γ has no effect on FcRn expression in RPE cells. Conclusions Human RPE cells express FcRn. The proinflammatory cytokine TNF-α downregulates FcRn expression. The authors speculate that the FcRn may play a pivotal role in the immune privilege of the human eye.

Intraocular T cells of patients with herpes simplex virus (HSV)-induced acute retinal necrosis recognize HSV tegument proteins VP11/12 and VP13/14.

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.

T Cells Specific for the Triggering Virus Infiltrate the Eye in Patients with Herpes Simplex Virus-Mediated Acute Retinal Necrosis

Acute retinal necrosis (ARN) is a rare, potentially blinding retinal disease resulting from ocular infections with herpes simplex virus (HSV) or varicella-zoster virus (VZV). To determine the antigen specificity and functional characteristics of ocular infiltrating T cells in ARN, T cells were isolated and expanded nonspecifically from intraocular fluid (IOF) samples from 2 patients with HSV-1- and 3 with VZV-mediated ARN. HSV-specific T cell reactivity could be detected only in the IOF-derived T cell lines (TCLs) of the 2 patients with HSV-mediated ARN. These TCLs consisted of both HSV type-common and type-specific CD4+ and CD8+ T cell clones (TCCs) with differential T cell receptor usage. Irrespective of their phenotype, the TCCs were cytolytic and secreted interferon-gamma, tumor necrosis factor-alpha, interleukin-4, and interleukin-5. In both patients, the antigen specificity of a substantial number of HSV-1-specific TCCs could be mapped to approximately 0.67-0.73 HSV-1 map units. The data presented suggest the contribution of T cells, specific for the triggering virus, to the pathogenesis of ARN.

Analysis of Local Antibody Productionin the Vitreous Humor of Patients With Severe Uveitis

We analyzed the local antibody production in vitreous humor samples collected during vitrectomy in patients with severe vision-threatening uveitis. In 24 patients, paired serum and undiluted vitreous humor samples were collected and tested for antibodies against Toxoplasma gondii, herpes simplex virus, varicella-zoster virus, cytomegalovirus, Epstein-Barr virus, and Toxocara canis. Total IgG and the Goldmann-Witmer coefficient were determined. The initial diagnosis of ocular toxoplasmosis could be confirmed in six of the seven patients. The seventh patient showed a local antibody production against herpes simplex virus. One of the three patients with chronic panuveitis at initial diagnosis showed a local antibody production against T. gondii. These last two findings resulted in a change in medical treatment. Analysis of local antibody production in vitreous humor samples is a valuable diagnostic tool.

Multicolor flowcytometric immunophenotyping is a valuable tool for detection of intraocular lymphoma

OBJECTIVE Intraocular lymphoma (IOL) is a rare condition and frequently difficult to distinguish from uveitis or other uveitis-masquerading syndromes. The diagnosis is confirmed by cytologic examination of ocular fluid specimens and more recently by molecular-immunoglobulin heavy chain (IGH) translocation or cytokine analysis. However, some of these more recent methods have not been validated by follow-up studies. DESIGN Evaluation of a diagnostic test. PARTICIPANTS In a cohort of 51 consecutive patients with a clinical suspicion of IOL, vitreous analysis was performed via multicolor flowcytometric immunophenotyping. METHODS Multicolor flowcytometric immunophenotyping was performed with CD45, CD3, CD19, CD20, anti-SmIgκ, and anti-SmIgλ antibodies. The presence of a clear B-cell population showing a disequilibrium of Igκ versus Igλ expression was used to confirm the diagnosis of non-Hodgkin lymphoma (NHL). Patients were followed for a minimum of 2 years (mean, 5.9 ± 2.0 years) to validate the accuracy of the method. MAIN OUTCOME MEASURES The presence or absence of IOL during follow-up. RESULTS In 14 of 51 patients, a clinical diagnosis of IOL was confirmed using flowcytometric analysis. Of these 14 patients, 11 had primary IOL and 3 had metastasized secondary lymphomas. In 3 of 51 patients who were diagnosed with (central nervous system) NHL during follow-up, the test failed to confirm the presence of a clonal B-cell population. In 18 of the 34 other patients, an infectious or well-defined immunologic disorder was established during follow-up. The remaining 16 patients, with a minimal follow-up of 2 years, were diagnosed with idiopathic uveitis. CONCLUSIONS Multicolor flowcytometric analysis had 82.4% sensitivity and 100% specificity in patients with suspected IOL. This is comparable to the reported vitreous interleukin (IL)-6/IL-10 testing sensitivity of 0.8 and sensitivity of 0.65 to 0.95 by immunoglobulin heavy chain (IGH) gene arrangement testing in clinical cohorts. Because flowcytometric tests are readily performed in hematologic laboratories, this can be regarded as a useful method for confirming the clinical diagnosis of IOL. FINANCIAL DISCLOSURE(S) The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Acyclovir-resistant herpes simplex virus type 1 in intra-ocular fluid samples of herpetic uveitis patients

BACKGROUND Acyclovir (ACV) is the antiviral drug of choice to treat patients with herpes simplex virus type 1 (HSV-1) uveitis. The prevalence of intra-ocular ACV-resistant (ACV(R)) HSV-1 in herpetic uveitis is unknown and may have clinical consequences. In addition to its predictive value on ACV susceptibility, the polymorphic HSV-1 thymidine kinase (TK) gene facilitates differentiation between HSV-1 strains. OBJECTIVES The objective of this study was to determine the genetic composition and ACV susceptibility of the causative virus in intra-ocular fluid samples (IOF) of HSV-1 uveitis patients. STUDY DESIGN The intra-ocular HSV-1 pool from 11 HSV-1 uveitis patients was determined by sequencing IOF-derived viral TK genes. The ACV susceptibility profile of the cloned intra-ocular TK variants was defined by mass spectrometry. In addition, the ganciclovir (GCV) susceptibility of the ACV(R) HSV-1 TK variants was defined. RESULTS Intra-ocular fluid samples of HSV-1 uveitis patients contain HSV-1 quasispecies, principally consisting of one major and multiple genetically related minor patient-specific TK variants. Four of 10 patients analyzed had an intra-ocular ACV(R) HSV-1 of which 3 were cross-resistant to GCV. The ACV(R) profile of intra-ocular HSV-1 did not correlate with symptomatic ACV treatment. CONCLUSIONS Affected eyes of HSV-1 uveitis patients are commonly infected with a patient-specific HSV-1 quasispecies, including one major and multiple genetically related minor variants. A relatively high prevalence of intra-ocular ACV(R) HSV-1, mainly ACV/GCV cross-resistant viruses, was detected in HSV-1 uveitis patients.

Retinal vasculitis occurring with common variable immunodeficiency syndrome

PURPOSE To report severe retinal vasculitis causing decreased vision in three patients with the common variable immunodeficiency syndrome. METHOD Case report. Three patients with common variable immunodeficiency syndrome developed decreased vision secondary to retinal vasculitis. Fluorescein angiography was performed in all three patients. Peribulbar injections were given in one patient, and two patients were treated with oral steroids and cyclosporin. RESULTS All three patients were young and had classic common variable immunodeficiency syndrome. Bilateral retinal vasculitis and diffuse retinal edema were present in all three patients, and two patients had retinal neovascularization in the absence of ischemia. No evidence of intraocular infection was present, and none was detected systematically. Visual acuity decreased in five of the six eyes and was responsive to treatment in only one patient (both eyes). CONCLUSION Retinal vasculitis may be another autoimmune manifestation of common variable immunodeficiency syndrome.

HLA-A29 subtypes and birdshot chorioretinopathy

J Central Laboratory ofthe Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, The Netherlands 2 Department of Ophthalmology, Erasmus University, Rotterdam, The Netherlands 3 Department of Ophthalmo-Immunology of the Netherlands Ophthalmic Research Institute, Amsterdam, The Netherlands 4 Department of Haematology, University of Gent, Gent, Belgium 5 Department of Ophthalmology, University of Gent, Gent, Belgium