IRF8 Transcription-Factor-Dependent Classical Dendritic Cells Are Essential for Intestinal T Cell Homeostasis.

The role of dendritic cells (DCs) in intestinal immune homeostasis remains incompletely defined. Here we show that mice lacking IRF8 transcription-factor-dependent DCs had reduced numbers of T cells in the small intestine (SI), but not large intestine (LI), including an almost complete absence of SI CD8αß(+) and CD4(+)CD8αα(+) T cells; the latter requiring ß8 integrin expression by migratory IRF8 dependent CD103(+)CD11b(-) DCs. SI homing receptor induction was impaired during T cell priming in mesenteric lymph nodes (MLN), which correlated with a reduction in aldehyde dehydrogenase activity by SI-derived MLN DCs, and inefficient T cell localization to the SI. These mice also lacked intestinal T helper 1 (Th1) cells, and failed to support Th1 cell differentiation in MLN and mount Th1 cell responses to Trichuris muris infection. Collectively these results highlight multiple non-redundant roles for IRF8 dependent DCs in the maintenance of intestinal T cell homeostasis.

The Quest for Orthologs orthology benchmark service in 2022.

The Orthology Benchmark Service (https://orthology.benchmarkservice.org) is the gold standard for orthology inference evaluation, supported and maintained by the Quest for Orthologs consortium. It is an essential resource to compare existing and new methods of orthology inference (the bedrock for many comparative genomics and phylogenetic analysis) over a standard dataset and through common procedures. The Quest for Orthologs Consortium is dedicated to maintaining the resource up to date, through regular updates of the Reference Proteomes and increasingly accessible data through the OpenEBench platform. For this update, we have added a new benchmark based on curated orthology assertion from the Vertebrate Gene Nomenclature Committee, and provided an example meta-analysis of the public predictions present on the platform.

InParanoid-DIAMOND: faster orthology analysis with the InParanoid algorithm.

SUMMARY: Predicting orthologs, genes in different species having shared ancestry, is an important task in bioinformatics. Orthology prediction tools are required to make accurate and fast predictions, in order to analyze large amounts of data within a feasible time frame. InParanoid is a well-known algorithm for orthology analysis, shown to perform well in benchmarks, but having the major limitation of long runtimes on large datasets. Here, we present an update to the InParanoid algorithm that can use the faster tool DIAMOND instead of BLAST for the homolog search step. We show that it reduces the runtime by 94%, while still obtaining similar performance in the Quest for Orthologs benchmark. AVAILABILITY AND IMPLEMENTATION: The source code is available at (https://bitbucket.org/sonnhammergroup/inparanoid). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Risk of cancer in young and middle-aged adults with childhood-onset type 1 diabetes in Sweden-A prospective cohort study.

AIMS/HYPOTHESIS: In persons with type 1 diabetes, the risk of cancer remains controversial. We wanted to examine the excess risk of cancer in a large population-based cohort diagnosed with type 1 diabetes before 15 years of age. STUDY POPULATION AND METHODS: From 1 July 1977 to 31 December 2013, we prospectively and on a national scale included 18,724 persons (53% men) with childhood-onset type 1 diabetes. For each person with type 1 diabetes, we selected four referents, matched for the date at birth and municipality of living at the time when the case developed diabetes. Cases and referents were linked to national registers of cancer and of the cause of death. RESULTS: A total of 125 persons (61% women) with diabetes had 135 different cancers, all diagnosed after the diabetes diagnosis. The median duration from diabetes diagnosis to first cancer diagnosis was 19 years (interquartile range 10-26). The median age at cancer diagnosis in the diabetes group was 28 years (interquartile range 20-35). The overall standardized incidence ratio (95%), using the Swedish general population as referents for women with diabetes was 1.28 (1.02, 1.58) and when comparing women with diabetes with matched referents, we found a hazard ratio of 1.42 (1.10, 1.85). No elevated risk was seen for men. Cancers of the breast and testis were the most common types in women and men respectively. CONCLUSIONS: Women with childhood-onset type 1 diabetes had a small but significantly elevated risk of cancer. No such tendency was seen for men. The reason behind this is unclear.

IRF4 transcription-factor-dependent CD103(+)CD11b(+) dendritic cells drive mucosal T helper 17 cell differentiation.

CD103(+)CD11b(+) dendritic cells (DCs) represent the major migratory DC population within the small intestinal lamina propria (SI-LP), but their in vivo function remains unclear. Here we demonstrate that intestinal CD103(+)CD11b(+) DC survival was dependent on interferon regulatory factor 4 (IRF4). Mice with a DC deletion in Irf4 displayed reduced numbers of intestinal interleukin 17 (IL-17)-secreting helper T 17 (Th17) cells and failed to support Th17 cell differentiation in draining mesenteric lymph nodes (MLN) following immunization. The latter was associated with a selective reduction in CD103(+)CD11b(+) MLN DCs and DC derived IL-6. Immunized Il6(-/-) mice failed to support Th17 cell differentiation in MLN in vivo and CD103(+)CD11b(+) MLN DCs supported IL-6-dependent Th17 cell differentiation in vitro. Together, our results suggest a central role for IRF4-dependent, IL-6 producing CD103(+)CD11b(+) DCs in intestinal Th17 cell differentiation.

Inhibition of STAT3 prevents bone metastatic progression of prostate cancer in vivo.

BACKGROUND: Prostate cancer (PC) metastasizes to the skeleton forming predominantly sclerotic lesions, and there is currently no cure for bone metastatic disease. The transcription factor signal transducer and activator of transcription 3 (STAT3) is implicated as a metastatic driver, but its potential as therapeutic target in bone metastasis has not been investigated. In this study, we evaluated for the first time a STAT3 inhibitor, Napabucasin, as a therapeutic option for bone metastatic PC. METHODS: Effects of STAT3 inhibitors, Stattic and Napabucasin, on metastatic potential in PC cells were studied in vitro by assessment of migration capacity, self-renewal potential, and tumorsphere formation. For evaluation of the role of STAT3 in initial skeletal establishment of PC cells as well as in progressed castration-resistant PC (CRPC) in bone, human VCaP prostate cancer cells were inoculated in the tibia of mice which subsequently were treated with the STAT3 inhibitor Napabucasin. Bone specimens were analyzed using computed tomography (CT), immunohistochemistry, and quantitative polymerase chain reaction. RESULTS: The small molecule STAT3 inhibitors Stattic and Napabucasin both effectively impaired metastatic potential of PC cells in vitro. Furthermore, treatment with Napabucasin prevented metastatic establishment in tibial bones in vivo and thereby also the tumor-induced sclerotic bone response seen in vehicle-treated VCaP xenografts. In addition, treatment with Napabucasin of established bone CRPC significantly decreased both tumor burden and tumor-induced trabecular bone volume compared with effects seen in vehicle-treated animals. Anti-mitotic effects were confirmed by decreased Ki67 staining in Napabucasin-treated xenografts compared with vehicle-treated xenografts. Alterations of gene expression in the femoral bone marrow (BM) niche toward the maintenance of hematopoietic stem cells and the myeloid lineage were demonstrated by quantitative real-time polymerase chain reaction and were further reflected by a substantial increase in the number of erythrocytes in BM of Napabucasin-treated mice. Furthermore, a unique pattern of STAT3 phosphorylation in osteoblasts/stromal cells surrounding the areas of tumor cells was demonstrated immunohistochemically in bone xenograft models using several different PC cell lines. CONCLUSION: Inhibition of STAT3 activity disrupts the bone metastatic niche and targets both the skeletal establishment of PC and advanced bone metastatic CRPC in mice, suggesting STAT3 as a candidate for molecular targeted therapies of skeletal metastatic disease.

Patient-derived scaffolds influence secretion profiles in cancer cells mirroring clinical features and breast cancer subtypes.

BACKGROUND: Breast cancer is a common malignancy with varying clinical behaviors and for the more aggressive subtypes, novel and more efficient therapeutic approaches are needed. Qualities of the tumor microenvironment as well as cancer cell secretion have independently been associated with malignant clinical behaviors and a better understanding of the interplay between these two features could potentially reveal novel targetable key events linked to cancer progression. METHODS: A newly developed human derived in vivo-like growth system, consisting of decellularized patient-derived scaffolds (PDSs) recellularized with standardized breast cancer cell lines (MCF7 and MDA-MB-231), were used to analyze how 63 individual patient specific microenvironments influenced secretion determined by proximity extension assays including 184 proteins and how these relate to clinical outcome. RESULTS: The secretome from cancer cells in PDS cultures varied distinctly from cells grown as standard monolayers and besides a general increase in secretion from PDS cultures, several secreted proteins were only detectable in PDSs. Monolayer cells treated with conditioned media from PDS cultures, further showed increased mammosphere formation demonstrating a cancer stem cell activating function of the PDS culture induced secretion. The detailed secretomic profiles from MCF7s growing on 57 individual PDSs differed markedly but unsupervised clustering generated three separate groups having similar secretion profiles that significantly correlated to different clinical behaviors. The secretomic profile that associated with cancer relapse and high grade breast cancer showed induced secretion of the proteins IL-6, CCL2 and PAI-1, all linked to cancer stem cell activation, metastasis and priming of the pre-metastatic niche. Cancer promoting pathways such as "Suppress tumor immunity" and "Vascular and tissue remodeling" was also linked to this more malignant secretion cluster. CONCLUSION: PDSs repopulated with cancer cells can be used to assess how cancer secretion is effected by specific and varying microenvironments. More malignant secretion patterns induced by specific patient based cancer microenvironments could further be identified pinpointing novel therapeutic opportunities targeting micro environmentally induced cancer progression via secretion of potent cytokines. Video abstract.

Detection of Unilateral Arm Paresis after Stroke by Wearable Accelerometers and Machine Learning.

Recent advances in stroke treatment have provided effective tools to successfully treat ischemic stroke, but still a majority of patients are not treated due to late arrival to hospital. With modern stroke treatment, earlier arrival would greatly improve the overall treatment results. This prospective study was performed to asses the capability of bilateral accelerometers worn in bracelets 24/7 to detect unilateral arm paralysis, a hallmark symptom of stroke, early enough to receive treatment. Classical machine learning algorithms as well as state-of-the-art deep neural networks were evaluated on detection times between 15 min and 120 min. Motion data were collected using triaxial accelerometer bracelets worn on both arms for 24 h. Eighty-four stroke patients with unilateral arm motor impairment and 101 healthy subjects participated in the study. Accelerometer data were divided into data windows of different lengths and analyzed using multiple machine learning algorithms. The results show that all algorithms performed well in separating the two groups early enough to be clinically relevant, based on wrist-worn accelerometers. The two evaluated deep learning models, fully convolutional network and InceptionTime, performed better than the classical machine learning models with an AUC score between 0.947-0.957 on 15 min data windows and up to 0.993-0.994 on 120 min data windows. Window lengths longer than 90 min only marginally improved performance. The difference in performance between the deep learning models and the classical models was statistically significant according to a non-parametric Friedman test followed by a post-hoc Nemenyi test. Introduction of wearable stroke detection devices may dramatically increase the portion of stroke patients eligible for revascularization and shorten the time to treatment. Since the treatment effect is highly time-dependent, early stroke detection may dramatically improve stroke outcomes.

Domainoid: domain-oriented orthology inference.

BACKGROUND: Orthology inference is normally based on full-length protein sequences. However, most proteins contain independently folding and recurring regions, domains. The domain architecture of a protein is vital for its function, and recombination events mean individual domains can have different evolutionary histories. It has previously been shown that orthologous proteins may differ in domain architecture, creating challenges for orthology inference methods operating on full-length sequences. We have developed Domainoid, a new tool aiming to overcome these challenges faced by full-length orthology methods by inferring orthology on the domain level. It employs the InParanoid algorithm on single domains separately, to infer groups of orthologous domains. RESULTS: This domain-oriented approach allows detection of discordant domain orthologs, cases where different domains on the same protein have different evolutionary histories. In addition to domain level analysis, protein level orthology based on the fraction of domains that are orthologous can be inferred. Domainoid orthology assignments were compared to those yielded by the conventional full-length approach InParanoid, and were validated in a standard benchmark. CONCLUSIONS: Our results show that domain-based orthology inference can reveal many orthologous relationships that are not found by full-length sequence approaches. AVAILABILITY: https://bitbucket.org/sonnhammergroup/domainoid/.

Osteoblast-derived factors promote metastatic potential in human prostate cancer cells, in part via non-canonical transforming growth factor ß (TGFß) signaling.

BACKGROUND: Transforming growth factor ß (TGFß) functions as a double-edged sword in prostate cancer tumorigenesis. In initial stages of the disease, TGFß acts as a growth inhibitor upon tumor cells, whereas it in later stages of disease rather promotes invasion and metastatic potential. One well-known cellular source of TGFß in the bone metastatic site is the bone-forming osteoblasts. Here we have studied the effects by osteoblast-derived factors on metastatic potential in several human prostate cancer cell lines. METHODS: Effects on metastatic potential in prostate cancer cells by osteoblast-derived factors were studied in vitro using several methods, including Transwell migration and evaluation of formation of pro-migratory protrusions. Confocal microscopy was used to evaluate possible changes in differentiation state in tumor cells by analysis of markers for epithelial-to-mesenchymal transition (EMT). The Matrigel-on-top 3D culture method was used for further assessment of metastatic characteristics in tumor cells by analysis of formation of filopodium-like protrusions (FLPs). RESULTS: Osteoblast-derived factors increased migration of PC-3U cells, an effect less prominent in cells overexpressing a mutated type I TGFß receptor (TßRI) preventing non-canonical TRAF6-dependent TGFß signaling. Osteoblast-derived factors also increased the formation of long protrusions and loss of cell-cell contacts in PC-3U cells, suggesting induction of a more aggressive phenotype. In addition, treatment with TGFß or osteoblast-derived factors of PC-3U cells in Matrigel-on-top 3D cultures promoted formation of FLPs, previously shown to be essential for metastatic establishment. CONCLUSIONS: These findings suggests that factors secreted from osteoblasts, including TGFß, can induce several cellular traits involved in metastatic potential of PC-3U cells, further strengthening the role for bone cells to promote metastatic tumor cell behavior.

Oncostatin M induces tumorigenic properties in non-transformed human prostate epithelial cells, in part through activation of signal transducer and activator of transcription 3 (STAT3).

Prostate cancer is one of the most common types of cancer in men in Western countries. Chronic inflammation in the prostate, regulated by a complex network of factors including inflammatory cytokines, is one of the established risk factors for development of prostate cancer. Interleukin-6 (IL-6) is a well-known promoter of inflammation-induced carcinogenesis and disease progression in prostate cancer. Presence in the prostate and possible roles in tumor development by other members of the IL-6 family of cytokines have, however, been less studied. Here we show that the IL-6-type cytokine oncostatin M (OSM) indeed induce cellular properties associated with tumorigenesis and disease progression in non-transformed human prostate epithelial cells, including morphological changes, epithelial-to-mesenchymal transition (EMT), enhanced migration and pro-invasive growth patterns. The effects by OSM were partly mediated by activation of signal transducer and activator of transcription 3 (STAT3), a transcription factor established as driver of cancer progression and treatment resistance in numerous types of cancer. The findings presented here further consolidate IL-6-type cytokines and STAT3 as promising future treatment targets for prostate cancer.

Secretion and Uptake of α-Synuclein Via Extracellular Vesicles in Cultured Cells.

In Parkinson's disease and other Lewy body disorders, the propagation of pathology has been accredited to the spreading of extracellular α-synuclein (α-syn). Although the pathogenic mechanisms are not fully understood, cell-to-cell transfer of α-syn via exosomes and other extracellular vesicles (EVs) has been reported. Here, we investigated whether altered molecular properties of α-syn can influence the distribution and secretion of α-syn in human neuroblastoma cells. Different α-syn variants, including α-syn:hemi-Venus and disease-causing mutants, were overexpressed and EVs were isolated from the conditioned medium. Of the secreted α-syn, 0.1-2% was associated with vesicles. The major part of EV α-syn was attached to the outer membrane of vesicles, whereas a smaller fraction was found in their lumen. For α-syn expressed with N-terminal hemi-Venus, the relative levels associated with EVs were higher than for WT α-syn. Moreover, such EV-associated α-syn:hemi-Venus species were internalized in recipient cells to a higher degree than the corresponding free-floating forms. Among the disease-causing mutants, A53T α-syn displayed an increased association with EVs. Taken together, our data suggest that α-syn species with presumably lost physiological functions or altered aggregation properties may shift the cellular processing towards vesicular secretion. Our findings thus lend further support to the tenet that EVs can mediate spreading of harmful α-syn species and thereby contribute to the pathology in α-synucleinopathies.

Intestinal dendritic cells in the regulation of mucosal immunity.

The intestine presents a huge surface area to the outside environment, a property that is of critical importance for its key functions in nutrient digestion, absorption, and waste disposal. As such, the intestine is constantly exposed to dietary and microbial-derived foreign antigens, to which immune cells within the mucosa must suitably respond to maintain intestinal integrity, while also providing the ability to mount effective immune responses to potential pathogens. Dendritic cells (DCs) are sentinel immune cells that play a central role in the initiation and differentiation of adaptive immune responses. In the intestinal mucosa, DCs are located diffusely throughout the intestinal lamina propria, within gut-associated lymphoid tissues, including Peyer's patches and smaller lymphoid aggregates, as well as in intestinal-draining lymph nodes, including mesenteric lymph nodes. The recognition that dietary nutrients and microbial communities in the intestine influence both mucosal and systemic immune cell development and function as well as immune-mediated disease has led to an explosion of literature in mucosal immunology in recent years and a growing interest in the functionality of intestinal DCs. In the current review, we discuss recent findings from our group and others that have provided important insights regarding murine and human intestinal lamina propria DCs and highlighted marked developmental and functional heterogeneity within this compartment. A thorough understanding of the role these subsets play in the regulation of intestinal immune homeostasis and inflammation will help to define novel strategies for the treatment of intestinal pathologies and contribute to improved rational design of mucosal vaccines.

Estimating marginal causal effects in a secondary analysis of case-control data.

When an initial case-control study is performed, data can be used in a secondary analysis to evaluate the effect of the case-defining event on later outcomes. In this paper, we study the example in which the role of the event is changed from a response variable to a treatment of interest. If the aim is to estimate marginal effects, such as average effects in the population, the sampling scheme needs to be adjusted for. We study estimators of the average effect of the treatment in a secondary analysis of matched and unmatched case-control data where the probability of being a case is known. For a general class of estimators, we show the components of the bias resulting from ignoring the sampling scheme and demonstrate a design-weighted matching estimator of the average causal effect. In simulations, the finite sample properties of the design-weighted matching estimator are studied. Using a Swedish diabetes incidence register with a matched case-control design, we study the effect of childhood onset diabetes on the use of antidepressant medication as an adult. Copyright © 2017 John Wiley & Sons, Ltd.

How vitamin A metabolizing dendritic cells are generated in the gut mucosa.

CD103(+) dendritic cells (DCs) represent the major migratory DC population in the intestinal lamina propria and are believed to play an essential role in the initiation and regulation of mucosal adaptive immune responses. Small intestine (SI) CD103(+) DCs have an enhanced capacity to generate the vitamin A metabolite, retinoic acid, a property that underlies their ability to induce the gut homing receptors CC chemokine receptor 9 and α4ß7 on responding T and B cells, and enhance forkhead box P3(+) T regulatory and IgA plasma cell differentiation in vitro. In this review, we discuss the environmental signals that appear to promote vitamin A metabolising activity in SI CD103(+) DCs in the steady state and thus which may contribute to driving the unique nature of SI immune responses.

Potential upstream regulators of cannabinoid receptor 1 signaling in prostate cancer: a Bayesian network analysis of data from a tissue microarray.

BACKGROUND: The endocannabinoid system regulates cancer cell proliferation, and in prostate cancer a high cannabinoid CB1 receptor expression is associated with a poor prognosis. Down-stream mediators of CB1 receptor signaling in prostate cancer are known, but information on potential upstream regulators is lacking. RESULTS: Data from a well-characterized tumor tissue microarray were used for a Bayesian network analysis using the max-min hill-climbing method. In non-malignant tissue samples, a directionality of pEGFR (the phosphorylated form of the epidermal growth factor receptor) → CB1 receptors were found regardless as to whether the endocannabinoid metabolizing enzyme fatty acid amide hydrolase (FAAH) was included as a parameter. A similar result was found in the tumor tissue, but only when FAAH was included in the analysis. A second regulatory pathway, from the growth factor receptor ErbB2 → FAAH was also identified in the tumor samples. Transfection of AT1 prostate cancer cells with CB1 receptors induced a sensitivity to the growth-inhibiting effects of the CB receptor agonist CP55,940. The sensitivity was not dependent upon the level of receptor expression. Thus a high CB1 receptor expression alone does not drive the cells towards a survival phenotype in the presence of a CB receptor agonist. CONCLUSIONS: The data identify two potential regulators of the endocannabinoid system in prostate cancer and allow the construction of a model of a dysregulated endocannabinoid signaling network in this tumor. Further studies should be designed to test the veracity of the predictions of the network analysis in prostate cancer and other solid tumors.

Dendritic cell subsets in the intestinal lamina propria: ontogeny and function.

The intestinal mucosa is exposed to large amounts of foreign antigen (Ag) derived from commensal bacteria, dietary Ags, and intestinal pathogens. Dendritic cells (DCs) are believed to be involved in the induction of tolerance to harmless Ags and in mounting protective immune responses to pathogens and, as such, to play key roles in regulating intestinal immune homeostasis. The characterization of classical DCs (cDCs) in the intestinal lamina propria has been under intense investigation in recent years but the use of markers (including CD11c, CD11b, MHC class II), which are also expressed by intestinal MΦs, has led to some controversy regarding their definition. Here we review recent studies that help to distinguish cDCs subsets from monocyte-derived cells in the intestinal mucosa. We address the phenotype and ontogeny of these cDC subsets and highlight recent findings indicating that these subsets play distinct roles in the regulation of mucosal immune responses in vivo.

Cysteine proteinase inhibitors regulate human and mouse osteoclastogenesis by interfering with RANK signaling.

The cysteine proteinase inhibitor cystatin C inhibited RANKL-stimulated osteoclast formation in mouse bone marrow macrophage cultures, an effect associated with decreased mRNA expression of Acp5, Calcr, Ctsk, Mmp9, Itgb3, and Atp6i, without effect on proliferation or apoptosis. The effects were concentration dependent with half-maximal inhibition at 0.3 µM. Cystatin C also inhibited osteoclast formation when RANKL-stimulated osteoclasts were cultured on bone, leading to decreased formation of resorption pits. RANKL-stimulated cells retained characteristics of phagocytotic macrophages when cotreated with cystatin C. Three other cysteine proteinase inhibitors, cystatin D, Z-RLVG-CHN2 (IC50 0.1 µM), and E-64 (IC50 3 µM), also inhibited osteoclast formation in RANKL-stimulated macrophages. In addition, cystatin C, Z-RLVG-CHN2, and E-64 inhibited osteoclastic differentiation of RANKL-stimulated CD14(+) human monocytes. The effect by cystatin C on differentiation of bone marrow macrophages was exerted at an early stage after RANKL stimulation and was associated with early (4 h) inhibition of c-Fos expression and decreased protein and nuclear translocation of c-Fos. Subsequently, p52, p65, IκBα, and Nfatc1 mRNA were decreased. Cystatin C was internalized in osteoclast progenitors, a process requiring RANKL stimulation. These data show that cystatin C inhibits osteoclast differentiation and formation by interfering intracellularly with signaling pathways downstream RANK.

STAT6 signaling pathway activated by the cytokines IL-4 and IL-13 induces expression of the Epstein-Barr virus-encoded protein LMP-1 in absence of EBNA-2: implications for the type II EBV latent gene expression in Hodgkin lymphoma.

In line with the B-lymphotropic nature of Epstein-Barr virus (EBV), the virus is present in several types of B-cell lymphomas. EBV expresses a different set of latent genes in the associated tumors, such as EBV nuclear antigen 1 (EBNA-1) and latent membrane proteins (LMPs; type II latency) in classical Hodgkin lymphomas (HLs). We previously reported that exposure of in vitro EBV-converted, HL-derived cell line KMH2-EBV to CD40-ligand and interleukin-4 (IL-4) induced the expression of LMP-1. Here, we show that exposure to IL-4 or IL-13 alone induced LMP-1 in the absence of EBNA-2. Induction of LMP-1 by IL-4 and IL-13 was mediated by the signal transducer signal transducer and activator of transcription 6 (STAT6) and a newly defined high-affinity STAT6-binding site in the LMP-1 promoter. IL-4 induced LMP-1 also in Burkitt lymphoma-derived lines and in tonsillar B cells infected with the EBNA-2-deficient EBV strain P3HR-1. Furthermore, coculture of EBV-carrying Burkitt lymphoma cells with activated CD4(+) T cells resulted in the induction of LMP-1 in the absence of EBNA-2. Because Hodgkin/Reed-Sternberg cells are known to secrete IL-13, to have constitutively activated STAT6, and to be closely surrounded by CD4(+) T cells, these mechanisms may be involved in the expression of LMP-1 in EBV-positive chronic HLs.