Primary Intraocular B-cell Lymphoma Research Articles

Th17 Cells Are Involved in the Local Control of Tumor Progression in Primary Intraocular Lymphoma

BackgroundTh17 cells play an important role in the pathogenesis of many autoimmune diseases, but despite some reports of their antitumor properties, too little is known about their presence and role in cancers. Specifically, knowledge is sparse about the relation of Th17 to lymphoma microenvironments and, more particularly, to the microenvironment of primary intraocular B-cell lymphoma (PIOL), an aggressive lymphoma with a poor prognosis.Methods and Principal FindingsIn this work, we investigated the presence of Th17 cells and their related cytokines in a syngeneic model of PIOL, a subtype of non-Hodgkin lymphoma. The very small number of lymphocytes trafficking in normal eyes, which represent a low background as compared to tumor-bearing eyes, allows us to develop the present model to characterize the different lymphocyte subsets present when a tumor is developing. IL-21 mRNA was expressed concomitantly with IL-17 mRNA in tumor-bearing eyes and intracellular expression of IL-17A and IL-21 in infiltrating CD4+ T lymphocytes. Interestingly, IL-17A production by T cells was negatively correlated with tumor burden. We also showed that IL-21 but not IL-17 inhibits tumor cell proliferation in vitro.ConclusionsThese data demonstrate that IL-17A and IL-21-producing CD4+ T cells, referred as Th17 cells, infiltrate this tumor locally and suggest that Th17-related cytokines may counteract tumor progression via IL-21 production. Thus, Th17 cells or their related cytokines could be considered to be a new therapeutic approach for non-Hodgkin B-cell lymphomas, particularly those with an ocular localization.

Regulatory T‐cells induce an immune escape in a murine model of primary intraocular B‐cell lymphoma

Abstract Purpose The role of regulatory T cells (Tregs) in the immune evasion of tumors is usually prominent. The aim of this study was to assess the presence and role of Tregs in a murine model of primary intraocular B‐cell lymphoma (PIOL). Methods We used a syngeneic model of PIOL in immunocompetent BALB/c mice. Immunohistochemistry and flow cytometric analysis have been performed to study the tumor growth and the immune infiltrate. Depletion of Tregs was achieved using intraperitoneal injection of anti‐CD25 mAb (PC61). Splenectomy was performed to study the role of induced Tregs. Results Tregs were recruited in PIOL eyes (16.2%) compared to control eyes (1.2%, p<0.001). The number of infiltrating Tregs was correlated with tumor burden (r2=0.76), and inversely correlated with CD4+ T‐cells (r2=‐0.79). Recruitment of Tregs was also observed in the spleen of mice bearing tumor (2.9%) compared to controls (2.1%, p<0.001), but not in the draining lymph nodes. Depletion of Tregs resulted in a significant decrease of tumor burden (53.8% in control mice compared to 37.7% in PC61 mice, p=0.03), and an increase of the immune infiltrate. Splenectomy experiments comfirmed the role of iTregs in tumor escape. Eventhough the number of tumor cells decreased, tumor growth could not be abrogated. Conclusion We demonstrated that Tregs are recruited in the tumor microenvironment. Tregs limit the recruitement of CD4+ T‐cells in the eyes and are thought to inhibit their effector functions. The mechanisms by which Tregs are generated could be similar to ACAID, yet in the posterior chamber. Several escape mechanisms are responsible for tolerance toward the tumor in the eye.

Regulatory T-cells induce an immune escape in a murine model of primary intraocular B-cell lymphoma

Purpose The role of regulatory T cells (Tregs) in the immune evasion of tumors is usually prominent. The aim of this study was to assess the presence and role of Tregs in a murine model of primary intraocular B-cell lymphoma (PIOL). Methods We used a syngeneic model of PIOL in immunocompetent BALB/c mice. Immunohistochemistry and flow cytometric analysis have been performed to study the tumor growth and the immune infiltrate. Depletion of Tregs was achieved using intraperitoneal injection of anti-CD25 mAb (PC61). Splenectomy was performed to study the role of induced Tregs. Results Tregs were recruited in PIOL eyes (16.2%) compared to control eyes (1.2%, p<0.001). The number of infiltrating Tregs was correlated with tumor burden (r2=0.76), and inversely correlated with CD4+ T-cells (r2=-0.79). Recruitment of Tregs was also observed in the spleen of mice bearing tumor (2.9%) compared to controls (2.1%, p<0.001), but not in the draining lymph nodes. Depletion of Tregs resulted in a significant decrease of tumor burden (53.8% in control mice compared to 37.7% in PC61 mice, p=0.03), and an increase of the immune infiltrate. Splenectomy experiments comfirmed the role of iTregs in tumor escape. Eventhough the number of tumor cells decreased, tumor growth could not be abrogated. Conclusion We demonstrated that Tregs are recruited in the tumor microenvironment. Tregs limit the recruitement of CD4+ T-cells in the eyes and are thought to inhibit their effector functions. The mechanisms by which Tregs are generated could be similar to ACAID, yet in the posterior chamber. Several escape mechanisms are responsible for tolerance toward the tumor in the eye.

Impaired Th1/Tc1 Cytokine Production of Tumor-Infiltrating Lymphocytes in a Model of Primary Intraocular B-Cell Lymphoma

Primary intraocular lymphoma is a high-grade non-Hodgkin lymphoma with a pathogenesis that is still unclear. Microenvironment is known to be crucial in controlling tumor growth and maintenance. To study the immune microenvironment in intraocular lymphomas and to characterize the cytokine polarization of infiltrating T-lymphocytes, a new murine model of intraocular B-cell lymphoma was developed. Immunocompetent adult mice were injected intravitreally with a syngeneic lymphomatous B-cell line. Clinical, histologic, and flow cytometric analyses were performed to characterize the tumoral invasion and the immune infiltration. Cytokine production of ocular cells was investigated by RT-PCR and fluorescent immunoassay, with or without stimulation by anti-CD3(+) anti-CD28 antibodies. Intraocular lymphoma developed in eyes injected by lymphomatous B-cells. At day 19, the retina and the vitreous cavity were infiltrated by tumor cells. Up to 15% of living cells were T-lymphocytes. Cytokine profile analysis of the supernatant of ocular cells cultured ex vivo demonstrated the presence of IL10, IL6, IFNgamma, and TNFalpha. Stimulation of ocular cells with anti-CD3(+) anti-CD28 antibodies increased the IFNgamma level and led to the induction of IL2 production, completing the type 1 (Th1/Tc1-like) pattern of cytokine expression observed. IL12p70 and IL4, potent Th1 or Th2 differentiating factors, were undetectable, even after stimulation. The results suggest that T-cells from intraocular B-lymphomas are characterized by a Th1/Tc1-like profile that could be partially inhibited in vivo. These data raise the possibility of a T-cell immunostimulation to reactivate the Th1/Tc1-lymphocytes and improve intraocular antitumoral immunity.

Vogt-Koyanagi-Harada Disease

Vogt-Koyanagi-Harada disease (VKH) is a multisystem autoimmune disorder principally affecting pigmented tissues in the ocular, auditory, integumentary and central nervous systems. Patients are typically 20 to 50 years old and have no history of either surgical or accidental ocular trauma. Pigmented races are more commonly affected. Depending on revised diagnostic criteria, the disease is classified as complete, incomplete or probable based on the presence of extraocular findings (neurological, auditory and integumentary). The clinical course of VKH is divided into four phases: prodromal (mimics a viral infection), uveitic (bilateral diffuse uveitis with papillitis and exudative retinal detachment), convalescent (tissue depigmentation), and chronic recurrent (recurrent uveitis and ocular complications). The pathogenesis of VKH is thought to be related to an aberrant T cell-mediated immune response directed against self-antigens found on melanocytes. VKH has been linked to human leukocyte antigen DR4 (HLA-DR4) and HLA-Dw53, with strongest associated risk for HLA-DRB1*0405 haplotype. The diagnosis of VKH is clinical, and differential includes sympathetic ophthalmia, sarcoidosis, primary intraocular B-cell lymphoma, posterior scleritis, and uveal effusion syndrome. Treatment is typically initiated with high-dose oral corticosteroids, but other immunomondulatory agents (most oftentimes cyclosporine) may be needed for non-responsive patients or when corticosteroid side-effects are not tolerated. Visual prognosis is generally good with prompt diagnosis and aggressive immunomodulatory treatment.

Molecular pathology of primary intraocular lymphoma

To evaluate immunoglobulin heavy chain (IgH) gene rearrangements, cytokines and chemokines, and infectious agents in primary intraocular B-cell lymphoma (PIOL) cells, in order to better diagnose and understand PIOL.We studied ocular specimens from 57 patients with PIOL at the National Eye Institute from 1991 to 2001. Specimens were analyzed for IgH gene rearrangements using microdissection and polymerase chain reaction (PCR). We measured vitreal interleukin (IL)-10 and IL-6 levels by enzyme-linked immunosorbent assay. IL-10 mRNA was studied in PIOL cells using microdissection and reverse transcribed (RT)-PCR. Chemokine and chemokine receptor expression was examined by using immunohistochemistry. Infectious DNA of human herpetic virus-8 (HHV-8), Epstein-Bar virus (EBV), and Toxoplasma gondii was detected by using microdissection and PCR and was confirmed with Southern blot hybridization.IgH rearrangement(s) were demonstrated in all 50 tested cases. Cytokine levels were measured in the vitreous of 39 patients. Thirty-one had measurable cytokine levels: 24 of 31 had elevation of IL-10 relative to that of IL-6, and, in contrast, only 7 of 31 had elevation of IL-6 relative to IL-10. IL-10 mRNA was abundant in lymphoma cells of 6 examined cases. Lymphoma cells expressed chemokine receptors of CXCR4 and CXCR5 in three tested cases. HHV-8 DNA was found in 6 of 32 cases (18.8%), EBV DNA in 2 of 21 (9.5%), and T gondii DNA in 2 of 16 (12.5%).Molecular analyses detecting IgH rearrangements and vitreal levels of IL-10 and IL-6 are useful adjuncts for PIOL diagnosis. A role for specific infectious agents is hypothesized in the pathogenesis of some cases of PIOL. B-cell chemokine is likely involved in attracting PIOL cells into the eye.

Expression of chemokine receptors, CXCR4 and CXCR5, and chemokines, BLC and SDF-1, in the eyes of patients with primary intraocular lymphoma

Objective Chemokines have a range of biologic activities, including regulation of leukocyte trafficking, modulation of hematopoietic cell proliferation, and adhesion to extracellular matrix molecules. Specifically, B-lymphocyte chemoattractant (BLC); BCA-1; CXCL13, SCYB13) and stromal cell-derived factor-1 (SDF-1, CXCL12, SCYB12) are chemotactic for human B cells, and their ligands CXCR4 and CXCR5 are differentially expressed on B cells, including malignant B cells. We investigated the expression of these chemokine/chemokine receptors in eyes with primary intraocular B-cell lymphoma (PIOL). Design Observational case series (human tissue study). Methods Three freshly enucleated eyes with PIOL and a normal autopsied eye were frozen and sectioned. The sections were evaluated using immunohistochemistry (avidin-biotin-complex immunoperoxidase technique) for CXCR4, CXCR5, BLC, and SDF-1 to detect the expression and location. Reverse transcriptase-polymerase chain reaction was used to detect chemokine transcripts of CXCR4, CXCR5, BLC, and SDF-1 in PIOL and retinal pigment epithelium (RPE) cells after microdissection—either by laser capture (Arcturus) or by manual dissection—from frozen sections. Main outcome measures and results The three PIOL eyes showed similar pathology, with typical diffuse large B-lymphoma cells subjacent to the RPE. The eyes also demonstrated a similar chemokine profile. High expression levels of CXCR4 and CXCR5 were found limited to the lymphoma cells. In contrast, BLC protein was expressed in the RPE but not located in other ocular resident cells. SDF-1 was barely detected in a few RPE cells. CXCR4 and CXCR5 transcripts were detected abundantly in lymphoma cells, whereas BLC and SDF-1 transcripts were detected only in the RPE and not the malignant cells. No chemokine expression was detected on the RPE cells in the normal control eye. Conclusions Chemokines and chemokine receptors selective for B cells were identified in RPE and malignant B cells, respectively. BLC, and possibly SDF-1, attracts both normal and malignant B-cells while promoting migration of only small numbers of T cells and macrophages. We propose that B-cell chemokines may be involved in the pathogenesis of PIOL by selectively attracting lymphoma cells to the RPE from the choroidal circulation. Our data suggest that inhibition of B-cell chemoattractants could be a future strategy for the treatment of PIOL.

Detection of Toxoplasma Gondii DNA in Primary Intraocular B-Cell Lymphoma

Primary intraocular lymphoma, a variant of primary central nervous system lymphoma with ocular involvement, is a large B-cell non-Hodgkin's lymphoma. Some cases of primary intraocular lymphoma have been reported to be associated with microorganisms including Epstein-Barr virus (EBV) and human herpes virus-8 (HHV-8), but not parasites. We analyzed 10 cases of primary intraocular lymphoma using microdissection and PCR. Tumor and normal cells were microdissected from ocular tissue on slides and subjected to PCR for genes from Toxoplasma gondii, EBV, and HHV-8. We detected Toxoplasma gondii, not HHV-8 or EBV, DNA in the lymphoma but not in normal cells of two cases that resembled ocular toxoplasmosis clinically. We speculate that Toxoplasma gondii may play a role in some forms of primary intraocular B-cell lymphoma.