The Experts below are selected from a list of 213 Experts worldwide ranked by ideXlab platform
Marc Feldmann - One of the best experts on this subject based on the ideXlab platform.
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RelB/p50 regulates CCL19 production, but fails to promote human DC maturation.
European journal of immunology, 2009Co-Authors: Chiara Gasparini, Brian M. J. Foxwell, Marc FeldmannAbstract:DC, when fully matured are the APC best able to activate naive T cells. Recently, we demonstrated using adenoviruses overexpressing IkappaBalpha and proteosome inhibitors that NF-kappaB is involved in DC activation, but the role of the individual subunits is still not clear. We investigated the role of the NF-kappaB subunits RelB and p50 in human DC activation using adenoviral vectors expressing RelB or p50. Nuclear RelB, in the form of RelB/p50, was active only in DC infected with both viruses, this induced the production of the soluble homeostatic Chemokine CCL19, but not other homeostatic Chemokines, particularly in LPS-matured DC. However, RelB/p50 did not affect the expression of costimulatory and antigen-presenting molecules, and increased the allogeneic mixed lymphocyte reaction only in LPS-matured DC. This enhanced mixed lymphocyte reaction is most likely due to enhanced CCL19 production, which sustains the interaction between mature DC and naive T cells. In conclusion, we demonstrated that RelB/p50 was active only in DC expressing both RelB and p50, and induced CCL19 production, but not DC maturation.
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relb p50 regulates CCL19 production but fails to promote human dc maturation
European Journal of Immunology, 2009Co-Authors: Chiara Gasparini, Brian M. J. Foxwell, Marc FeldmannAbstract:DC, when fully matured are the APC best able to activate naive T cells. Recently, we demonstrated using adenoviruses overexpressing IkappaBalpha and proteosome inhibitors that NF-kappaB is involved in DC activation, but the role of the individual subunits is still not clear. We investigated the role of the NF-kappaB subunits RelB and p50 in human DC activation using adenoviral vectors expressing RelB or p50. Nuclear RelB, in the form of RelB/p50, was active only in DC infected with both viruses, this induced the production of the soluble homeostatic Chemokine CCL19, but not other homeostatic Chemokines, particularly in LPS-matured DC. However, RelB/p50 did not affect the expression of costimulatory and antigen-presenting molecules, and increased the allogeneic mixed lymphocyte reaction only in LPS-matured DC. This enhanced mixed lymphocyte reaction is most likely due to enhanced CCL19 production, which sustains the interaction between mature DC and naive T cells. In conclusion, we demonstrated that RelB/p50 was active only in DC expressing both RelB and p50, and induced CCL19 production, but not DC maturation.
Suha Saleh - One of the best experts on this subject based on the ideXlab platform.
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HIV integration and the establishment of latency in CCL19-treated resting CD4(+) T cells require activation of NF-κB.
Retrovirology, 2016Co-Authors: Suha Saleh, Vanessa A. Evans, David Harisson, Jingling Zhou, Anthony Jaworowski, Georgina Sallmann, Karey Y. Cheong, Talia M. Mota, Surekha TennakoonAbstract:Background Eradication of HIV cannot be achieved with combination antiretroviral therapy (cART) because of the persistence of long-lived latently infected resting memory CD4+ T cells. We previously reported that HIV latency could be established in resting CD4+ T cells in the presence of the Chemokine CCL19. To define how CCL19 facilitated the establishment of latent HIV infection, the role of Chemokine receptor signalling was explored.
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hiv integration and the establishment of latency in CCL19 treated resting cd4 t cells require activation of nf κb
Retrovirology, 2016Co-Authors: Suha Saleh, Vanessa A. Evans, David Harisson, Jingling Zhou, Anthony Jaworowski, Georgina SallmannAbstract:Background Eradication of HIV cannot be achieved with combination antiretroviral therapy (cART) because of the persistence of long-lived latently infected resting memory CD4+ T cells. We previously reported that HIV latency could be established in resting CD4+ T cells in the presence of the Chemokine CCL19. To define how CCL19 facilitated the establishment of latent HIV infection, the role of Chemokine receptor signalling was explored.
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HIV integration and the establishment of latency in CCL19-treated resting CD4^+ T cells require activation of NF-κB
Retrovirology, 2016Co-Authors: Suha Saleh, David Harisson, Jingling Zhou, Anthony Jaworowski, Georgina Sallmann, Karey Y. Cheong, Talia M. Mota, Surekha Tennakoon, Vanessa Evans, Thomas A. AngelovichAbstract:Background Eradication of HIV cannot be achieved with combination antiretroviral therapy (cART) because of the persistence of long-lived latently infected resting memory CD4^+ T cells. We previously reported that HIV latency could be established in resting CD4^+ T cells in the presence of the Chemokine CCL19. To define how CCL19 facilitated the establishment of latent HIV infection, the role of Chemokine receptor signalling was explored. Results In resting CD4^+ T cells, CCL19 induced phosphorylation of RAC-alpha serine/threonine-protein kinase (Akt), nuclear factor kappa B (NF-κB), extracellular-signal-regulated kinase (ERK) and p38. Inhibition of the phosphoinositol-3-kinase (PI3K) and Ras/Raf/Mitogen-activated protein kinase/ERK kinase (MEK)/ERK signalling pathways inhibited HIV integration, without significant reduction in HIV nuclear entry (measured by Alu-LTR and 2-LTR circle qPCR respectively). Inhibiting activation of MEK1/ERK1/2, c-Jun N-terminal kinase (JNK), activating protein-1 (AP-1) and NF-κB, but not p38, also inhibited HIV integration. We also show that HIV integrases interact with Pin1 in CCL19-treated CD4^+ T cells and inhibition of JNK markedly reduced this interaction, suggesting that CCL19 treatment provided sufficient signals to protect HIV integrase from degradation via the proteasome pathway. Infection of CCL19-treated resting CD4^+ T cells with mutant strains of HIV, lacking NF-κB binding sites in the HIV long terminal repeat (LTR) compared to infection with wild type virus, led to a significant reduction in integration by up to 40-fold (range 1–115.4, p = 0.03). This was in contrast to only a modest reduction of 5-fold (range 1.7–11, p > 0.05) in fully activated CD4^+ T cells infected with the same mutants. Finally, we demonstrated significant differences in integration sites following HIV infection of unactivated, CCL19-treated, and fully activated CD4^+ T cells. Conclusions HIV integration in CCL19-treated resting CD4^+ T cells depends on NF-κB signalling and increases the stability of HIV integrase, which allow subsequent integration and establishment of latency. These findings have implications for strategies needed to prevent the establishment, and potentially reverse, latent infection.
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entry of hiv in primary human resting cd4 t cells pretreated with the Chemokine CCL19
AIDS Research and Human Retroviruses, 2014Co-Authors: Jenny L. Anderson, Karey Cheong, Amas K H Lee, Suha Saleh, Candida Da Fonseca Pereira, Paul U. Cameron, Sharon R LewinAbstract:Resting CD4+ T cells latently infected with HIV are a major barrier to a cure and therefore antiretroviral therapy is required life-long. There are two major potential pathways leading to the establishment of latency. One pathway, which we refer to as postactivation latency, occurs with the reversion of an activated infected CD4+ T cell to the resting memory state with an integrated silenced provirus. The other pathway, preactivation latency, occurs via direct infection of resting CD4+ T cells. The relative contribution in vivo of preactivation and postactivation latency is unclear, but latency can be established in vitro via both pathways.1 We have previously demonstrated that although unstimulated resting CD4+ T cells are relatively resistant to HIV infection, incubation of resting CD4+ T cells with Chemokines that bind the Chemokine receptors, CCR7, CXCR3, and CCR6, which are all highly expressed on resting CD4+T cells, facilitates latent infection in vitro.2,3 Pretreatment with the Chemokine CCL19, the ligand for CCR7, facilitated rapid nuclear localization and efficient integration. This was in part mediated via CCL19-induced changes in the actin cytoskeleton.3 Our aim here was to visualize the early events following HIV entry into a resting CD4+ T cell. We used fluorescent HIV to infect CCL19-treated resting CD4+ T cells to track the migration of virus to the nucleus. Resting CD4+ T cells were incubated with 30 nM CCL19 and infected at an MOI of 0.1 with HIVNL4.3 that was dual labeled with green fluorescent protein (GFP) fused to HIV Vpr and S15tomato fused to the viral membrane. GFPVpr associates with HIV cores and tracks HIV complexes in the cytoplasm of target cells. S15tomato is a fluorescent variant of S15mCherry that is no longer visible when virus particles fuse to target cells.4 Therefore, HIV that has fused and entered a cell can be identified as GFPVpr+ S15tomato−.4 Two hours after the addition of virus, cells were fixed and filamentous actin (F-actin) was stained using Alexa Fluor 350.phalloidin (Molecular Probes). The cells were placed in a Labtek II chamber slide and images were captured in a z series using a CoolSNAP camera (Photometrics) on a DeltaVision microscope (100×1.4 numerical aperture oil immersion lens, Applied Precision). Images were deconvolved using softWoRx deconvolution software (Applied Precision). A single z section of a deconvolved image is shown. Cells were also cultured for 4 days and quantification of integrated DNA was performed using Alu-LTR real time PCR.3 Reverse transcriptase (RT) was quantified in supernatant. The frequency of integrated HIV was 92,000 copies/106 CD4+ T cells and there was minimal RT production consistent with latent infection. In the image shown in Fig. 1, fused HIV complexes (GFPVpr+ S15tomato−) that have penetrated cortical F-actin at the cell periphery and are located in the resting CD4+ T cell cytoplasm can be seen. We are now examining how HIV interacts with other components of the cell cytoskeleton and defining key cellular proteins required for rapid transit of HIV to the nucleus in CCL19-treated resting CD4+ T cells. FIG. 1. Fused HIV in the cytoplasm of CCL19-treated resting CD4+ T cells. Resting CD4+ T cells pretreated with 30 nM CCL19 were infected for 2 h with HIVNL4.3 dual labeled with GFPVpr (yellow) and S15tomato (majenta). Fixed cells were stained ...
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Entry of HIV in primary human resting CD4(+) T cells pretreated with the Chemokine CCL19.
AIDS research and human retroviruses, 2014Co-Authors: Jenny L. Anderson, Karey Cheong, Amas K H Lee, Suha Saleh, Candida Da Fonseca Pereira, Paul U. Cameron, Sharon R LewinAbstract:Resting CD4+ T cells latently infected with HIV are a major barrier to a cure and therefore antiretroviral therapy is required life-long. There are two major potential pathways leading to the establishment of latency. One pathway, which we refer to as postactivation latency, occurs with the reversion of an activated infected CD4+ T cell to the resting memory state with an integrated silenced provirus. The other pathway, preactivation latency, occurs via direct infection of resting CD4+ T cells. The relative contribution in vivo of preactivation and postactivation latency is unclear, but latency can be established in vitro via both pathways.1 We have previously demonstrated that although unstimulated resting CD4+ T cells are relatively resistant to HIV infection, incubation of resting CD4+ T cells with Chemokines that bind the Chemokine receptors, CCR7, CXCR3, and CCR6, which are all highly expressed on resting CD4+T cells, facilitates latent infection in vitro.2,3 Pretreatment with the Chemokine CCL19, the ligand for CCR7, facilitated rapid nuclear localization and efficient integration. This was in part mediated via CCL19-induced changes in the actin cytoskeleton.3 Our aim here was to visualize the early events following HIV entry into a resting CD4+ T cell. We used fluorescent HIV to infect CCL19-treated resting CD4+ T cells to track the migration of virus to the nucleus. Resting CD4+ T cells were incubated with 30 nM CCL19 and infected at an MOI of 0.1 with HIVNL4.3 that was dual labeled with green fluorescent protein (GFP) fused to HIV Vpr and S15tomato fused to the viral membrane. GFPVpr associates with HIV cores and tracks HIV complexes in the cytoplasm of target cells. S15tomato is a fluorescent variant of S15mCherry that is no longer visible when virus particles fuse to target cells.4 Therefore, HIV that has fused and entered a cell can be identified as GFPVpr+ S15tomato−.4 Two hours after the addition of virus, cells were fixed and filamentous actin (F-actin) was stained using Alexa Fluor 350.phalloidin (Molecular Probes). The cells were placed in a Labtek II chamber slide and images were captured in a z series using a CoolSNAP camera (Photometrics) on a DeltaVision microscope (100×1.4 numerical aperture oil immersion lens, Applied Precision). Images were deconvolved using softWoRx deconvolution software (Applied Precision). A single z section of a deconvolved image is shown. Cells were also cultured for 4 days and quantification of integrated DNA was performed using Alu-LTR real time PCR.3 Reverse transcriptase (RT) was quantified in supernatant. The frequency of integrated HIV was 92,000 copies/106 CD4+ T cells and there was minimal RT production consistent with latent infection. In the image shown in Fig. 1, fused HIV complexes (GFPVpr+ S15tomato−) that have penetrated cortical F-actin at the cell periphery and are located in the resting CD4+ T cell cytoplasm can be seen. We are now examining how HIV interacts with other components of the cell cytoskeleton and defining key cellular proteins required for rapid transit of HIV to the nucleus in CCL19-treated resting CD4+ T cells. FIG. 1. Fused HIV in the cytoplasm of CCL19-treated resting CD4+ T cells. Resting CD4+ T cells pretreated with 30 nM CCL19 were infected for 2 h with HIVNL4.3 dual labeled with GFPVpr (yellow) and S15tomato (majenta). Fixed cells were stained ...
Kateryna O Cybulsky - One of the best experts on this subject based on the ideXlab platform.
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CCL19-CCR7–dependent reverse transendothelial migration of myeloid cells clears Chlamydia muridarum from the arterial intima
Nature Immunology, 2016Co-Authors: Mark Roufaiel, Eric Gracey, Hisham Ibrahim, Marwan G. Althagafi, Sharon J. Hyduk, Allan Siu, Su-ning Zhu, Andrew Lau, Kelly Tai, Kateryna O CybulskyAbstract:Cybulsky and colleagues show that myeloid cells in the arterial intima undergo reverse transendothelial migration into the arterial circulation that is dependent on the Chemokine CCL19 and its receptor CCR7. Regions of the normal arterial intima predisposed to atherosclerosis are sites of ongoing monocyte trafficking and also contain resident myeloid cells with features of dendritic cells. However, the pathophysiological roles of these cells are poorly understood. Here we found that intimal myeloid cells underwent reverse transendothelial migration (RTM) into the arterial circulation after systemic stimulation of pattern-recognition receptors (PRRs). This process was dependent on expression of the Chemokine receptor CCR7 and its ligand CCL19 by intimal myeloid cells. In mice infected with the intracellular pathogen Chlamydia muridarum , blood monocytes disseminated infection to the intima. Subsequent CCL19-CCR7–dependent RTM was critical for the clearance of intimal C. muridarum . This process was inhibited by hypercholesterolemia. Thus, RTM protects the normal arterial intima, and compromised RTM during atherogenesis might contribute to the intracellular retention of pathogens in atherosclerotic lesions.
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CCL19-CCR7-dependent reverse transendothelial migration of myeloid cells clears Chlamydia muridarum from the arterial intima
Nature Immunology, 2016Co-Authors: Mark Roufaiel, Eric Gracey, Hisham Ibrahim, Marwan G. Althagafi, Sharon J. Hyduk, Kateryna O CybulskyAbstract:Cybulsky and colleagues show that myeloid cells in the arterial intima undergo reverse transendothelial migration into the arterial circulation that is dependent on the Chemokine CCL19 and its receptor CCR7.
Chiara Gasparini - One of the best experts on this subject based on the ideXlab platform.
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RelB/p50 regulates CCL19 production, but fails to promote human DC maturation.
European journal of immunology, 2009Co-Authors: Chiara Gasparini, Brian M. J. Foxwell, Marc FeldmannAbstract:DC, when fully matured are the APC best able to activate naive T cells. Recently, we demonstrated using adenoviruses overexpressing IkappaBalpha and proteosome inhibitors that NF-kappaB is involved in DC activation, but the role of the individual subunits is still not clear. We investigated the role of the NF-kappaB subunits RelB and p50 in human DC activation using adenoviral vectors expressing RelB or p50. Nuclear RelB, in the form of RelB/p50, was active only in DC infected with both viruses, this induced the production of the soluble homeostatic Chemokine CCL19, but not other homeostatic Chemokines, particularly in LPS-matured DC. However, RelB/p50 did not affect the expression of costimulatory and antigen-presenting molecules, and increased the allogeneic mixed lymphocyte reaction only in LPS-matured DC. This enhanced mixed lymphocyte reaction is most likely due to enhanced CCL19 production, which sustains the interaction between mature DC and naive T cells. In conclusion, we demonstrated that RelB/p50 was active only in DC expressing both RelB and p50, and induced CCL19 production, but not DC maturation.
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relb p50 regulates CCL19 production but fails to promote human dc maturation
European Journal of Immunology, 2009Co-Authors: Chiara Gasparini, Brian M. J. Foxwell, Marc FeldmannAbstract:DC, when fully matured are the APC best able to activate naive T cells. Recently, we demonstrated using adenoviruses overexpressing IkappaBalpha and proteosome inhibitors that NF-kappaB is involved in DC activation, but the role of the individual subunits is still not clear. We investigated the role of the NF-kappaB subunits RelB and p50 in human DC activation using adenoviral vectors expressing RelB or p50. Nuclear RelB, in the form of RelB/p50, was active only in DC infected with both viruses, this induced the production of the soluble homeostatic Chemokine CCL19, but not other homeostatic Chemokines, particularly in LPS-matured DC. However, RelB/p50 did not affect the expression of costimulatory and antigen-presenting molecules, and increased the allogeneic mixed lymphocyte reaction only in LPS-matured DC. This enhanced mixed lymphocyte reaction is most likely due to enhanced CCL19 production, which sustains the interaction between mature DC and naive T cells. In conclusion, we demonstrated that RelB/p50 was active only in DC expressing both RelB and p50, and induced CCL19 production, but not DC maturation.
Meike Burger - One of the best experts on this subject based on the ideXlab platform.
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CCL19 is a specific ligand of the constitutively recycling atypical human Chemokine receptor cram b
Immunology, 2010Co-Authors: Marion Leick, Julie Catusse, Marie Follo, Robert J. B. Nibbs, Tanja Nicole Hartmann, Hendrik Veelken, Meike BurgerAbstract:The human Chemokine receptor CRAM (Chemokine receptor on activated macrophages), encoded by the gene CCRL2, is a new candidate for the atypical Chemokine receptor family that includes the receptors DARC, D6 and chemocentryx Chemokine receptor (CCX-CKR). CRAM is maturation-stage-dependently expressed on human B lymphocytes and its surface expression is up-regulated upon short-term CCL5 exposure. Here, we demonstrate that the homeostatic Chemokine CCL19 is a specific ligand for CRAM. In radioactive labelling studies CCL19 bound to CRAM-expressing cells with an affinity similar to the described binding of its other receptor CCR7. In contrast to the known CCL19/CCR7 ligand/receptor pair, CRAM stimulation by CCL19 did not result in typical Chemokine-receptor-dependent cellular activation like calcium mobilization or migration. Instead, we demonstrate that CRAM is constitutively recycling via clathrin-coated pits and able to internalize CCL19 as well as anti-CRAM antibodies. As this absence of classical Chemokine receptor responses and the recycling and internalization features are characteristic for non-classical Chemokine receptors, we suggest that CRAM is the newest member of this group. As CCL19 is known to be critically involved in lymphocyte and dendritic cell trafficking, CCL19-binding competition by CRAM might be involved in modulating these processes.
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CCL19 is a specific ligand of the constitutively recycling atypical human Chemokine receptor CRAM‐B
Immunology, 2009Co-Authors: Marion Leick, Julie Catusse, Marie Follo, Robert J. B. Nibbs, Tanja Nicole Hartmann, Hendrik Veelken, Meike BurgerAbstract:The human Chemokine receptor CRAM (Chemokine receptor on activated macrophages), encoded by the gene CCRL2, is a new candidate for the atypical Chemokine receptor family that includes the receptors DARC, D6 and chemocentryx Chemokine receptor (CCX-CKR). CRAM is maturation-stage-dependently expressed on human B lymphocytes and its surface expression is up-regulated upon short-term CCL5 exposure. Here, we demonstrate that the homeostatic Chemokine CCL19 is a specific ligand for CRAM. In radioactive labelling studies CCL19 bound to CRAM-expressing cells with an affinity similar to the described binding of its other receptor CCR7. In contrast to the known CCL19/CCR7 ligand/receptor pair, CRAM stimulation by CCL19 did not result in typical Chemokine-receptor-dependent cellular activation like calcium mobilization or migration. Instead, we demonstrate that CRAM is constitutively recycling via clathrin-coated pits and able to internalize CCL19 as well as anti-CRAM antibodies. As this absence of classical Chemokine receptor responses and the recycling and internalization features are characteristic for non-classical Chemokine receptors, we suggest that CRAM is the newest member of this group. As CCL19 is known to be critically involved in lymphocyte and dendritic cell trafficking, CCL19-binding competition by CRAM might be involved in modulating these processes.
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The Atypical Chemokine Receptor CRAM Mediates CCL19 Transcytosis through Endothelial Cells and Modulates CCL19 Activation of Non-Hodgkin Lymphoma B Cells.
Blood, 2009Co-Authors: Marion Leick, Julie Catusse, Meike BurgerAbstract:Abstract 2672 Poster Board II-648 Introduction: Chemokines work as cellular recruitment molecules. Specific combinations of Chemokines, receptors, and adhesion molecules determine which subgroups of leukocytes migrate and what their destinations are. Chemokine receptor expression and activation on malignant cells may be involved in the growth, survival and migration of cancer cells as well as in the tumor vascularisation. CCR7, by binding the Chemokines CCL19 and CCL21, is centrally involved in B cell localisation to the secondary lymphoid organs and therefore implicated in lymphadenopathy of various non-hodgkin lymphomas (NHL). In addition to Chemokine receptors that have been cloned and described, various orphan receptors with a Chemokine receptor-like structure are still not characterized. Atypical, non-signaling Chemokine receptors are members of a newly described class of receptors and have been implicated with Chemokine clearance and influencing of other signalling receptors. They are consequently considered as potent immuno-modulators and as anti-inflammatory factors and are implicated in progression of cancer. Among these receptors, we are investigating the role of the orphan Chemokine (C-C motif) receptor-like 2 (CCRL2), also known as CRAM, a receptor expressed on endothelial cells and B cells in a maturation stage dependent manner, but for which functions and ligands are poorly characterized so far. In an effort to elucidate the role of CRAM and its implication in neoplasias, we have focussed research on identification of ligands and the implication of CRAM in regulating B cell migration in samples from healthy donors and from non-Hodgkin lymphomas. Methods: We characterised the receptor's expression profile by flow cytometry in peripheral blood, bone marrow and lymph node sections of different B cell NHL and correlated it to expression levels of CCR7 and CXCR4. In addition, a screening for ligands was performed using radiolabelled binding assays. The role of CRAM was elucidated using various functional assays, internalisation and transcytosis experiments. Results: We show that CRAM is an alternative, but non-signaling receptor for the CCR7-activating Chemokine CCL19. CRAM is constitutively recycling to and from the cell surface and internalizing the Chemokine without degrading it. We found that the receptor is responsible for transcytosis of CCL19 through endothelial cell layers and subsequent presentation, a crucial step in homing of leukocytes to the lymph nodes. On the other hand, when expressed on B cells, CRAM interferes in CCL19 binding to CCR7. We thereby show that CRAM can act as an integrator of different signals, by binding different Chemokines and controlling their activity toward surrounding ligands. Chemotaxis experiments demonstrate that CRAM is a negative modulator of CCL19 B cell recruitment. In addition, we have found increased expression in activated B cells, dendritic cells, and also in the B cell malignancies chronic lymphocytic leukemia (B-CLL) and pre-B cell acute lymphoblastic leukemia (pre-B ALL), and are currently evaluating CRAM as a possible prognostic marker in various B-NHLs. Conclusions: CRAM is a newly identified member of the silent or atypical Chemokine receptor group, already known for modulating Chemokine availability, together with D6, DARC and CCX-CKR. We have shown here that it contributes to lymphocyte recruitment into peripheral lymphoid tissue by presenting CCL19 on endothelium. It is also involved in CCR7 driven recruitment of B cells by regulating CCL19 availability. Expression of CRAM differs in B cell malignancies for which both CCR7 ligands, CCL19 and CCL21, have already been shown to be implicated in the development of lymphadenopathies. We therefore suggest that CRAM is an additional player and potential biomarker in determining outcome and development of disease. Disclosures: No relevant conflicts of interest to declare.