Regulation Of Chemokine Receptor Expression Research Articles

Delta-Like Ligand 4 (DLL4)/Notch axis regulates EAE development by modulating T cell trafficking across the blood brain barrier

Abstract The Notch receptor system has been suggested to play a critical role in the regulation of chemokine receptor expression on encephalitogenic T cells and subsequent T cell trafficking to the central nervous system (CNS). Here we tested whether Delta-Like Ligand (DLL) 4 (DLL4)/Notch axis regulates EAE development by modulating T effector cell trafficking mechanisms across the blood-brain barrier (BBB). Primary cultures of brain microvessel endothelial cells were established and migration of splenocytes was tested in the presence of DLL4 blocking antibody. Our data show that anti DLL4 blocks transendothelial migration of non-activated T cells, but does not influence the migration of anti CD28 and CD3-activated splenocytes. Additionally, activation of brain microvessel endothelial cells with TNFα abrogated the blocking effect of anti DLL4 in transendothelial migration of T cells. We also demonstrate that blockade of DLL4 function inhibits the development of a progressive model of clinical experimental autoimmune encephalomyelitis (EAE), in C57Bl6 mice. Based on these results, we propose that endothelial DLL4 engagement with Notch expressing T cells is critical in strengthening the neuroinflammation and blockage of DLL4 could provide novel therapies in early phases of CNS neuroinflammatory diseases.

Abstract 13813: Prior β-Blocker Treatment Dampens Immune Cell Responses Following Myocardial Infarction

Following myocardial infarction (MI), immune cell infiltration initiates and regulates cardiac remodeling and repair. Our previous work has shown the importance of β2-adrenergic receptors (β2AR) in modulating immune cell infiltration to the heart following injury through the regulation of chemokine receptor expression and splenic retention of bone marrow (BM)-derived leukocyte populations. Pharmacological inhibition of βAR is a common strategy for the treatment of many cardiovascular etiologies, however the impact of β-blocker therapy on immune cell responses following acute cardiac injury is unknown. We hypothesized that prior treatment with β-blockers with distinct isoform selectivity would differentially decrease leukocyte infiltration to the heart following MI. Mice were administered either vehicle, a β1AR-selective (Metoprolol; Met), a β2AR-selective (ICI 118,551; ICI) or a βAR non-selective (Carvedilol; Carv) antagonist for two weeks via minipump. Analysis of mice treated with ICI or Carv showed decreased BM expression of CCR2 and CXCR4, as well as elevated levels of leukocyte populations within the spleen. While these effects were not observed in mice infused with a β1AR-selective dose of Met, a higher concentration of Met (equivalent to moderate doses used in humans) also decreased CCR2 and CXCR4 expression in BM and increased splenic leukocyte retention. To determine the impact of β2AR-selective versus non-selective blockade on immune cell responses following acute cardiac injury, mice from each treatment group underwent sham or MI surgery. In addition to decreased CCR2 and CXCR4 expression in the BM, mice administered ICI, Carv and the non-selective dose of Met had impaired recruitment of leukocyte populations to the heart following MI. Toward clinically relevant ends, we assessed CCR2 and CXCR4 expression in leukocytes isolated from blood samples of healthy and heart failure patients and found that levels of each were decreased in patients having taken β-blockers. In all, these results demonstrate that prior β-blocker treatment negatively regulates immune cell responsiveness to acute cardiac injury.

Sphingosine kinase inhibitor suppresses dendritic cell migration by regulating chemokine receptor expression and impairing p38 mitogen‐activated protein kinase

The migration of dendritic cells (DCs) to secondary lymphoid organs plays a crucial role in the initiation of adaptive immune responses. Although lipopolysaccharide enhances chemokine receptor 7 (CCR7) expression on DCs, the second signal for the migration of DCs toward the chemokine CCL19 remains unknown. In this study, we show that sphingosine kinase inhibitor (SKI) inhibits the migration of DCs toward CCL19 through the down-regulation of CCR7. Inhibition of p38 mitogen-activated protein kinase (MAPK) activation by SKI may be responsible for the SKI-mediated effects on the regulation of chemokine receptor expression. Impairment of DC migration by the inhibition of p38 MAPK and down-regulation of CCR7 expression may contribute to the protective effects of SKI in DC-related disorders. These results suggest that sphingosine kinase-mediated signalling plays a role in the innate and adaptive immune responses by altering DC migration.

Regulation of Trafficking Receptor Expression in Human Forkhead Box P3+ Regulatory T Cells

Forkhead Box P3(+) (FOXP3(+)) T cells are regulatory cells important for maintaining immune tolerance. While chemokine- and other homing-receptors are important for T cell migration, it has been unclear how they are regulated in FOXP3(+) T cells. We thoroughly investigated, ex vivo and in vitro, the regulation of chemokine receptor expression on human FOXP3(+) T cells in neonatal cord blood, adult peripheral blood, and tonsils. We found that human FOXP3(+) T cells undergo changes in trafficking receptors according to their stages of activation and differentiation. FOXP3(+) T cells are divided into CD45RA(+) (naive type) and CD45RO(+) (memory type) FOXP3(+) T cells in neonatal blood, adult blood, and tonsils. CD45RA(+)FOXP3(+) T cells mainly express lymphoid tissue homing receptors (CD62L, CCR7, and CXCR4), while CD45RO(+)FOXP3(+) T cells highly express both Th1 and Th2-associated trafficking receptors along with the lymphoid tissue homing receptors at reduced frequencies. Up-regulation of Th1/Th2-associated trafficking receptors begins with activation of CD45RA(+)FOXP3(+) T cells and is completed after their differentiation to CD45RO(+) cells. Some chemokine receptors such as CXCR5 and CXCR6 are preferentially expressed by many FOXP3(+) cells at a specific stage (CD69(+)CD45RO(+)) in tonsils. Our in vitro differentiation study demonstrated that CD45RA(+)FOXP3(+) T cells indeed undergo chemokine receptor switch from CD45RA(+) (secondary lymphoid tissue homing) to CD45RO(+) type (lymphoid and nonlymphoid tissue homing). The orderly regulation of trafficking receptors in FOXP3(+) T cells according to stages of differentiation and activation is potentially important for their tissue-specific migration and regulation of immune responses in humans.

The effect of particulate material on the regulation of chemokine receptor expression in leukocytes

Cellular chemotaxis is one of the most significant components of the host response to implanted materials. However, the effect of materials and their particulates on the regulation of chemokine receptor expression is not well known. This study investigated the effects of alloy particulates on the regulation of CXCR1 and CXCR2 expression on leukocytes from whole blood and purified leukocyte cultures. The volumetric particle concentration and opsonisation dependent manner of CXCR1 and CXCR2 expression on neutrophils and CCR1 and CCR2 expression on monocytes/macrophages was analysed using both flow cytometry (FACS) and real time-PCR. Variation in volumetric concentrations of particulates demonstrated that cell:particulate ratios of 1:1 of non-opsonised NiCr alloy induced the down regulation of CXCR1, and 1:0.25 of opsonised NiCr induced the up regulation of CXCR2. CoCr alloy particulates had no effect on CXCR expression. This study demonstrated that the regulation of chemokine receptor expression could be effected by the properties of materials and the expression of these receptors may contribute to host cellular reactions such as chemotaxis. This study also demonstrated that the response of leukocytes can be demonstrated using a whole blood assay for the study of chemokine receptor expression after direct contact with materials.

The effect of TGF-β1 on adhesion molecule and chemokine receptor expression on naive T-cells

Rationale T-cell tissue-invasion is a complex interaction between adhesion molecules, cytokines, chemokines and their receptors. TGF-β1 is well known for its immunomodulation. In this study we investigated its effect on the expression of adhesion molecules and chemokine receptors on naive CD4+ vs. CD8+ T-cells. Methods Naive T-cells were isolated from human cord-blood by negative selection (n=5). Expression of the adhesion molecules L-selectin, CLA, integrin-α E and -β 7 and CXCR3 and CCR4 was evaluated by flowcytometry, before/after 96 hours of anti-CD3 stimulation (10μg/ml) with/without TGF-β1 (10ng/ml). Results TGF-β1 increased the expression of α E integrin on both CD4+ and CD8+ T-cells (p<0,02). Majority (81%) of naive T-cells were β 7+, whereas activation significantly decreased its expression (p<0,01). Since α E and β 7 integrins compose an active unit, surprisingly much higher proportion of T-cells were α E+ compared to β 7+. Activation through the TCR significantly decreased the expression of CCR4 on both CD4+ and CD8+ T-cells, (p<0,05). However, TGF-β1 induced the expression of CCR4+ on CD8+, but not CD4+ T-cells (p<0,05). Finally, evaluating the, TGF-β1 significantly downregulated the CCR4 and CXCR3 expression on α E+ T-cells (p<0,02). Conclusions These results indicate that TGF-β1 may differently regulate the expression of α E vs β 7 integrins on both CD4+ and CD8+ naive T-cells. In addition, T-cell sensitivity towards TGF-β1 induced regulation of chemokine receptor expression is driven in part by its phenotype and the strength of the TCR-stimulation. Such TGF-β1 induced immune-modulation may have important applications in the treatment of site-specific T-cell driven inflammatory diseases.

Cyclosporin A impairs dendritic cell migration by regulating chemokine receptor expression and inhibiting cyclooxygenase-2 expression

AbstractMigration of dendritic cells (DCs) into tissues and secondary lymphoid organs plays a crucial role in the initiation of innate and adaptive immunity. In this article, we show that cyclosporin A (CsA) impairs the migration of DCs both in vitro and in vivo. Exposure of DCs to clinical concentrations of CsA neither induces apoptosis nor alters development but does impair cytokine secretion, chemokine receptor expression, and migration. In vitro, CsA impairs the migration of mouse bone marrow–derived DCs toward macrophage inflammatory protein-3β (MIP-3β) and induces them to retain responsiveness to MIP-1α after lipopolysaccharide (LPS)–stimulated DC maturation, while in vivo administration of CsA inhibits the migration of DCs out of skin and into the secondary lymphoid organs. CsA impairs chemokine receptor and cyclooxygenase-2 (COX-2) expression normally triggered in LPS-stimulated DCs; administration of exogenous prostaglandin E2 (PGE2) reverses the effects of CsA on chemokine receptor expression and DC migration. Inhibition of nuclear factor–κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathway signaling by CsA may be responsible for the CsA-mediated effects on the regulation of chemokine receptor and cyclooxygenase-2 (COX-2) expression. Impairment of DC migration due to inhibition of PGE2 production and regulation of chemokine receptor expression may contribute, in part, to CsA-mediated immunosuppression.

Differential regulation of the expression of chemokine receptor 3 and 4 during plasma cell differentiation

The duration of specific antibody titers ranges from a few weeks up to several years. While the formation of antibody secreting plasma cells takes place in secondary lymphoid tissues, long-lasting antibody responses are provided by bone marrow plasma cells. The majority of plasma cells initially formed remains in secondary lymphoid tissues and die within a few days. Plasma cells that migrate into the bone marrow or into chronically inflamed tissues can survive for much longer periods of time. The capacity of plasma blasts to migrate into these tissues is regulated by chemokine receptors. Expression of CXCR4 is important for plasma cell migration into the bone marrow. CXCR3 ligands expressed in large quantities in chronically inflamed tissues (e.g. in effected kidneys in systemic lupus erythematosus) and can attract plasmablasts, thus mediating plasma cell accumulation at those sites. Here we analyzed the regulation of chemokine receptor expression during plasma cell differentiation in culture. Purified CXCR3-negative B cells were activated T dependently in a two-step culture system, for 3 days by CD40 ligand together with IL-2 and IL-10 and for 5 more days with IL-2 and IL-10. Alternatively, cells were stimulated T independently with CpG, IL-2 and IL-10. Addition of IFN-γ or monocyte/T-cell culture supernatant in the initial culture, but not at late stages, induced the expression of CXCR3 on plasma cells, thus suggesting that the regulation of this receptor is an early event during plasma cell differentiation. In contrast, CXCR4 was present on all plasma cells, formed under any stimulation conditions and even when CXCR4-negative B cells were activated to form plasma cells. These data indicate that expression of CXCR4 and, as a consequence, the potential to migrate into the bone marrow is generally associated with differentiation into plasma cells, whereas the expression of CXCR3 must be induced by the inflammatory cytokine IFN-γ.

Regulation of chemokine receptor expression in human microglia and astrocytes

It has been proposed that the positioning of mobile cells within a tissue is determined by their overall profile of chemokine receptors. This study examines the profiles of chemokine receptors expressed on resting and activated adult human microglial cells, astrocytes and a microglial cell line, CHME3. Microglia express highest levels of CXCR1, CXCR3 and CCR3. Astrocytes also have moderate levels of CXCR1 and CXCR3, and some CCR3, while both cell types also expressed CCR4, CCR5, CCR6, CXCR2, CXCR4 and CXCR5 at lower levels. Activation of the cells with the inflammatory cytokine tumour necrosis factor-α (TNFα) and interferon-γ (IFNγ) increased the expression of some but not all receptors over a period of 24 h. Microglia showed moderate enhancement of receptor expression, while astrocytes responded particularly strongly to TNFα with enhanced CXCR3, CCR3 and CXCR1. However, the migratory and proliferative responses of the microglia and astrocytes to the same chemokine were different, with microglia migrating and astrocytes proliferating in response to CXCL10. The data indicates a mechanism by which activated microglia and astrocytes become selectively more sensitive to inflammatory chemokines during CNS disease, and the paper discusses which of the many chemokines present in CNS would have priority of action on microglia and astrocytes.

Regulation of Chemokine Receptor Expression in Eosinophils

Signals via chemokine receptors play an important role in the accumulation of eosinophils at allergic inflammatory sites. Eosinophils constitutively express CC chemokine receptor 3 (CCR3) and, to a lesser extent, CCR1. CCR3 is mainly responsible for migration of resting eosinophils, and its specific ligand, eotaxin, represents the most potent chemoattractant for eosinophils. Some reports also suggest the expression of CXC chemokine receptor 1 (CXCR1) and/or CXCR2 in eosinophils. In addition, we recently reported the functional expression of CXCR4. The ligand of CXCR4, stromal cell-derived factor-1 (SDF-1), was able to induce a strong migratory response comparable to that by eotaxin. In contrast to the CCR3/eotaxin system which is mainly regulated at the level of ligand production, the CXCR4/SDF-1 system is regulated at the level of receptor expression. CXCR4 expression was completely attenuated by IL-4 and IL-5 and upregulated by IFN-γ and dexamethasone, while CCR3 expression was only marginally affected. The balance between the biological effects of these chemokine systems may affect the distribution and migration of eosinophils.

C-C chemokine receptor 4 expression defines a major subset of circulating nonintestinal memory T cells of both Th1 and Th2 potential.

CCR4, a chemokine receptor for macrophage-derived chemokine (MDC) and thymus and activation-regulated chemokine (TARC), has been implicated as a preferential marker for Th2 lymphocytes. Following in vitro polarization protocols, most Th2 lymphocytes express CCR4 and respond to its ligands TARC and MDC, whereas Th1 lymphocytes express CXC chemokine receptor 3 and CCR5 (but not CCR4). We show in this study that CCR4 is a major receptor for MDC and TARC on T lymphocytes, as anti-CCR4 mAbs significantly inhibit the migration of these cells to MDC and TARC. CCR4 is also highly expressed in most single-positive CD4(+) thymocytes and on a major fraction of blood nonintestinal (alpha(4)beta(7)(-)) memory CD4 lymphocytes, including almost all skin memory CD4(+) cells expressing the cutaneous lymphocyte Ag (CLA), but weakly or not expressed in other subsets in thymus and blood. Interestingly, major fractions of circulating CCR4(+) memory CD4 lymphocytes coexpress the Th1-associated receptors CXC chemokine receptor 3 and CCR5, suggesting a potential problem in using these markers for Th1 vs Th2 lymphocyte cells. Moreover, although production of Th2 cytokines in blood T cells is associated with CCR4(+) CD4 lymphocytes, significant numbers of freshly isolated circulating CCR4(+) memory CD4 lymphocytes (including both CLA(+) and CLA(-) fractions) readily express the Th1 cytokine IFN-gamma after short-term stimulation. Our results are consistent with a role for CCR4 as a major trafficking receptor for systemic memory T cells, and indicate that the patterns and regulation of chemokine receptor expression in vivo are more complex than indicated by current in vitro models of Th1 vs Th2 cell generation.

Glucocorticoids preferentially upregulate functional CXCR4 expression in eosinophils

Background: Chemokines play an important role in accumulation of eosinophils at allergic inflammatory sites. Systemic administration of glucocorticoids (GCCs) attenuates tissue eosinophilia. In vivo chemokine actions are regulated at levels of both ligand production and receptor expression. The inhibitory effects of GCCs on the production of eosinophil-active chemokines, such as eotaxin, have been well established. However, no data exist regarding the effects of GCCs on expression of chemokine receptors in eosinophils per se. Objective: The objective of this study was to investigate the regulation of chemokine receptor expression in eosinophils by GCCs. Methods: Chemokine receptor expression was analyzed by using flow cytometry and reverse transcriptase PCR. Intracellular Ca2+ influx and chemotaxis were also analyzed. Results: Eosinophil CCR3 expression was slightly downregulated by 24-hour treatment with dexamethasone (DEX). On the other hand, DEX-treated eosinophils showed markedly increased CXCR4 expression (~6 fold) in a time- and dose-dependent fashion. In contrast to eosinophils, CXCR4 expression in neutrophils was only marginally affected by DEX. In DEX-treated eosinophils, stromal cell–derived factor 1α, a natural ligand for CXCR4, induced a higher level of Ca2+ influx and chemotaxis compared with untreated cells. Conclusion: GCCs upregulate the expression of CXCR4 in eosinophils but not in neutrophils. Because stromal cell–derived factor 1α may play a role in baseline trafficking of eosinophils into extravascular tissues rather than recruiting them directly to inflammatory sites, upregulation of CXCR4 by GCCs may mediate the antiallergic property of these drugs by sequestering eosinophils from the circulation to extravascular tissues. (J Allergy Clin Immunol 2000;106:1132-9.)

Chemokine receptors and their role in inflammation and infectious diseases

Chemokines are small peptides that are potent activators and chemoattractants for leukocyte subpopulations and some nonhemopoietic cells. Their actions are mediated by a family of 7-transmembrane G-protein–coupled receptors, the size of which has grown considerably in recent years and now includes 18 members. Chemokine receptor expression on different cell types and their binding and response to specific chemokines are highly variable. Significant advances have been made in understanding the regulation of chemokine receptor expression and the intracellular signaling mechanisms used in bringing about cell activation. Chemokine receptors have also recently been implicated in several disease states including allergy, psoriasis, atherosclerosis, and malaria. However, most fascinating has been the observation that some of these receptors are used by human immunodeficiency virus type 1 in gaining entry into permissive cells. This review will discuss structural and functional aspects of chemokine receptor biology and will consider the roles these receptors play in inflammation and in infectious diseases.

The role of chemokine receptors in primary, effector, and memory immune responses.

The immune system is composed of single cells, and its function is entirely dependent on the capacity of these cells to traffic, localize within tissues, and interact with each other in a precisely coordinated fashion. There is growing evidence that the large families of chemokines and chemokine receptors provide a flexible code for regulating cell traffic and positioning in both homeostatic and inflammatory conditions. The regulation of chemokine receptor expression during development and following cell activation explains the complex migratory pathways taken by dendritic cells, T and B lymphocytes, providing new insights into the mechanisms that control priming, effector function, and memory responses.

Redox regulation of chemokine receptor expression.

Cytokines and reactive oxygen intermediates (ROI) are frequent companions at sites of acute inflammation. We have shown previously that in human monocytes, bacterial lipopolysaccharide, IL-1, and tumor necrosis factor-alpha induce a rapid down-regulation of the monocyte chemotactic protein-1 receptor CCR2 (CC chemokine receptor-2). These stimuli also induce production of ROI. In this paper, we investigate the influence of antioxidants and/or ROI on chemokine-receptor expression. In human monocytes, the antioxidant pyrrolidine dithiocarbamate (PDTC) rapidly inhibited CCR2 (95-100% of inhibition) and CCR5 (77-100% of inhibition) mRNA expression by strongly decreasing transcript stability. CCR2 half-life was decreased from 1.5 h to 45 min; CCR5 half-life was decreased from 2 h to 70 min. This inhibitory activity also included CXCR4 (CXC chemokine receptor-4) but not CXCR2 receptor and, although to a lesser extent, was shared by the antioxidants N-acetyl-l-cysteine and 2-mercaptoethanol. In contrast, the ROI-generating system xanthine/xanthine oxidase increased CCR5 and CXCR4 mRNA expression and counteracted the inhibitory effect of PDTC. Accordingly, H(2)O(2) and the glutathione-depleting drug buthionine sulfoximine increased to different extents CCR2, CCR5, and CXCR4 mRNA expression. The PDTC-mediated inhibition of CCR5 and CXCR4 mRNA expression was associated with decreased chemotactic responsiveness (>90% inhibition) and with a marked inhibition of surface-receptor expression. In contrast, xanthine/xanthine oxidase opposed the bacterial lipopolysaccharide- and tumor necrosis factor-alpha-mediated inhibition of CCR5 and CXCR4 mRNA expression and increased both the CCR5 surface expression and the cell migration (3-fold) in response to macrophage inflammatory protein-1beta. These results suggest that the redox status of cells is a crucial determinant in the regulation of the chemokine system.

TGF-beta 1 reciprocally controls chemotaxis of human peripheral blood monocyte-derived dendritic cells via chemokine receptors.

We examined the effect of TGF-beta 1 on the chemotactic migratory ability of human monocyte-derived dendritic cells (DCs). Treatment of immature DCs with TGF-beta 1 resulted in increased expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXC chemokine receptor-4 (CXCR-4), which were concomitant with enhanced chemotactic migratory responses to their ligands, RANTES (for CCR-1, CCR-3, and CCR-5), macrophage-inflammatory protein-3 alpha (MIP-3 alpha) (for CCR-6), or stromal cell-derived growth factor-1 alpha (for CXCR-4). Ligation by TNF-alpha resulted in down-modulation of cell surface expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXCR-4, and the chemotaxis for RANTES, MIP-3 alpha, and stromal cell-derived growth factor-1 alpha, whereas this stimulation up-regulated the expression of CCR-7 and the chemotactic ability for MIP-3beta. Stimulation of mature DCs with TGF-beta 1 also enhanced TNF-alpha-induced down-regulation of the expressions of CCR-1, CCR-3, CCR-5, CCR-6, and CXCR-4, and chemotaxis to their respective ligands, while this stimulation suppressed TNF-alpha-induced expression of CCR-7 and chemotactic migratory ability to MIP-3 beta. Our findings suggest that TGF-beta 1 reversibly regulates chemotaxis of DCs via regulation of chemokine receptor expression.

Coreceptor/Chemokine Receptor Expression on Human Hematopoietic Cells: Biological Implications for Human Immunodeficiency Virus–Type 1 Infection: Presented in part at the American Society of Hematology Meeting, San Diego, CA, 1997 and published in abstract form in Blood90:2144, 1997 (abstr, suppl 1).

AbstractThe recent discovery of chemokine receptors as coreceptors for human immunodeficiency virus–type 1 (HIV-1) entry offers new avenues for investigating the pathogenesis of acquired immunodeficiency syndrome (AIDS)-related cytopenias. To this end, we sought to (1) phenotype human hematopoietic cells for CD4 and the HIV-1 coreceptors CXCR4, CCR5, CCR3, and CCR2b; (2) correlate CD4 and chemokine receptor expression with their susceptibility to HIV-1 infection; and (3) examine any potential interplay between inflammatory cytokines released during HIV-1 infection and regulation of chemokine receptor expression. Fluorescence-activated cell sorting (FACS) analysis of bone marrow mononuclear cells (BMMNC), cells derived from serum-free expanded hematopoietic lineages (colony-forming unit–granulocyte-macrophage [CFU-GM], colony-forming unit-megakaryocyte [CFU-Meg], and burst-forming unit-erythroid [BFU-E]), and CD34+ cells showed differential expression of chemokine receptors and CD4 with some lineage specificity. Significantly, FACS-sorted CXCR4+/CD34+ cells had the same clonogeneic potential as CXCR4−/CD34+ cells. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of FACS-sorted human candidate stem cells (HSC; CD34+, c-kit+, Rho123low) showed the presence of CXCR4 mRNA but not CD4 mRNA. Infection studies with HIV-1 Env-pseudotyped luciferase reporter viruses indicated that X4 Env (CXCR4-using) pseudotypes infected megakaryocytic cells, whereas R5 Env (CCR5-using) pseudotypes did not. Similarly, R5 but not X4 Env-pseudotyped viruses infected granulocyte-macrophage cells in a CD4/CCR5-dependent manner. Erythroid cells were resistant to R5 or X4 viral infection. Finally, we found that γ-interferon treatment upregulated CXCR4 expression on primary hematopoietic cells. In summary, the delineation of chemokine receptor expression on primary hematopoietic cells is a first step towards dissecting the chemokine-chemokine receptor axes that may play a role in hematopoietic cell proliferation and homing. Furthermore, susceptibility of hematopoietic cells to HIV-1 infection is likely to be more complicated than the mere physical presence of CD4 and the cognate chemokine receptor. Lastly, our results suggest a potential interplay between γ-interferon secretion and CXCR4 expression.

Coreceptor/Chemokine Receptor Expression on Human Hematopoietic Cells: Biological Implications for Human Immunodeficiency Virus–Type 1 Infection

The recent discovery of chemokine receptors as coreceptors for human immunodeficiency virus-type 1 (HIV-1) entry offers new avenues for investigating the pathogenesis of acquired immunodeficiency syndrome (AIDS)-related cytopenias. To this end, we sought to (1) phenotype human hematopoietic cells for CD4 and the HIV-1 coreceptors CXCR4, CCR5, CCR3, and CCR2b; (2) correlate CD4 and chemokine receptor expression with their susceptibility to HIV-1 infection; and (3) examine any potential interplay between inflammatory cytokines released during HIV-1 infection and regulation of chemokine receptor expression. Fluorescence-activated cell sorting (FACS) analysis of bone marrow mononuclear cells (BMMNC), cells derived from serum-free expanded hematopoietic lineages (colony-forming unit-granulocyte-macrophage [CFU-GM], colony-forming unit-megakaryocyte [CFU-Meg], and burst-forming unit-erythroid [BFU-E]), and CD34(+) cells showed differential expression of chemokine receptors and CD4 with some lineage specificity. Significantly, FACS-sorted CXCR4(+)/CD34(+) cells had the same clonogeneic potential as CXCR4(-)/CD34(+) cells. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of FACS-sorted human candidate stem cells (HSC; CD34(+), c-kit+, Rho123(low)) showed the presence of CXCR4 mRNA but not CD4 mRNA. Infection studies with HIV-1 Env-pseudotyped luciferase reporter viruses indicated that X4 Env (CXCR4-using) pseudotypes infected megakaryocytic cells, whereas R5 Env (CCR5-using) pseudotypes did not. Similarly, R5 but not X4 Env-pseudotyped viruses infected granulocyte-macrophage cells in a CD4/CCR5-dependent manner. Erythroid cells were resistant to R5 or X4 viral infection. Finally, we found that gamma-interferon treatment upregulated CXCR4 expression on primary hematopoietic cells. In summary, the delineation of chemokine receptor expression on primary hematopoietic cells is a first step towards dissecting the chemokine-chemokine receptor axes that may play a role in hematopoietic cell proliferation and homing. Furthermore, susceptibility of hematopoietic cells to HIV-1 infection is likely to be more complicated than the mere physical presence of CD4 and the cognate chemokine receptor. Lastly, our results suggest a potential interplay between gamma-interferon secretion and CXCR4 expression.

Regulation of chemokine receptor expression in dendritic cells

Regulation of chemokine receptor expression in dendritic cells

The role of co-stimulation in regulation of chemokine receptor expression and HIV-1 infection in primary T lymphocytes

Fusion and entry of the human immunodeficiency virus (HIV) into CD4+T lymphocytes requires expression of CD4 and a coreceptor. At least eight chemokine receptors can serve as coreceptors for HIV. Accumulating evidence indicates that multiple factors, including the state of cellular differentia- tion and activation, regulate the expression of α- and β-chemokine receptors on lymphocytes. For example, binding of antibodies to the CD28 coreceptor can downregulate expression of β-chemokine receptors, and this appears to have important consequences on the susceptibility of CD4+T lymphocytes to infection by HIV-1. In contrast, binding of the natural CD28 ligand B7 or antibodies to the CD28 homologue CTLA-4 can upregulate CCR5 expression, sug- gesting a reciprocal interaction between CD28 and CTLA-4 and the regulation of β-chemokine receptor expression. Thus, the CD28/CTLA-4/B7 co-stimulation pathway is identi- fied as a potential novel target for the control of susceptibility to some strains of HIV-1 infection.