Macrophage inflammatory protein 3alpha is involved in the constitutive trafficking of epidermal langerhans cells

Certain types of dendritic cells (DCs) appear in inflammatory lesions of various etiologies, whereas other DCs, e.g., Langerhans cells (LCs), populate peripheral organs constitutively. Until now, the molecular mechanism behind such differential behavior has not been elucidated. Here, we show that CD1a(+) LC precursors respond selectively and specifically to the CC chemokine macrophage inflammatory protein (MIP)-3alpha. In contrast, CD14(+) precursors of DC and monocytes are not attracted by MIP-3alpha. LCs lose the migratory responsiveness to MIP-3alpha during their maturation, and non-LC DCs do not acquire MIP-3alpha sensitivity. The notion that MIP-3alpha may be responsible for selective LC recruitment into the epidermis is further supported by the following observations: (a) MIP-3alpha is expressed by keratinocytes and venular endothelial cells in clinically normal appearing human skin; (b) LCs express CC chemokine receptor (CCR)6, the sole MIP-3alpha receptor both in situ and in vitro; and (c) non-LC DCs that are not found in normal epidermis lack CCR6. The mature forms of LCs and non-LC DCs display comparable sensitivity for MIP-3beta, a CCR7 ligand, suggesting that DC subtype-specific chemokine responses are restricted to the committed precursor stage. Although LC precursors express primarily CCR6, non-LC DC precursors display a broad chemokine receptor repertoire. These findings reflect a scenario where the differential expression of chemokine receptors by two different subpopulations of DCs determines their functional behavior. One type, the LC, responds to MIP-3alpha and enters skin to screen the epidermis constitutively, whereas the other type, the "inflammatory" DC, migrates in response to a wide array of different chemokines and is involved in the amplification and modulation of the inflammatory tissue response.     

Macrophage inflammatory protein 3alpha is expressed at inflamed epithelial surfaces and is the most potent chemokine known in attracting Langerhans cell precursors

Dendritic cells (DCs) form a network comprising different populations that initiate and differentially regulate immune responses. Langerhans cells (LCs) represent a unique population of DCs colonizing epithelium, and we present here observations suggesting that macrophage inflammatory protein (MIP)-3alpha plays a central role in LC precursor recruitment into the epithelium during inflammation. (a) Among DC populations, MIP-3alpha was the most potent chemokine inducing the selective migration of in vitro-generated CD34(+) hematopoietic progenitor cell-derived LC precursors and skin LCs in accordance with the restricted MIP-3alpha receptor (CC chemokine receptor 6) expression to these cells. (b) MIP-3alpha was mainly produced by epithelial cells, and the migration of LC precursors induced by the supernatant of activated skin keratinocytes was completely blocked with an antibody against MIP-3alpha. (c) In vivo, MIP-3alpha was selectively produced at sites of inflammation as illustrated in tonsils and lesional psoriatic skin where MIP-3alpha upregulation appeared associated with an increase in LC turnover. (d) Finally, the secretion of MIP-3alpha was strongly upregulated by cells of epithelial origin after inflammatory stimuli (interleukin 1beta plus tumor necrosis factor alpha) or T cell signals. Results of this study suggest a major role of MIP-3alpha in epithelial colonization by LCs under inflammatory conditions and immune disorders, and might open new ways to control epithelial immunity.     

Accumulation of immature Langerhans cells in human lymph nodes draining chronically inflamed skin

The coordinated migration and maturation of dendritic cells (DCs) such as intraepithelial Langerhans cells (LCs) is considered critical for T cell priming in response to inflammation in the periphery. However, little is known about the role of inflammatory mediators for LC maturation and recruitment to lymph nodes in vivo. Here we show in human dermatopathic lymphadenitis (DL), which features an expanded population of LCs in one draining lymph node associated with inflammatory lesions in its tributary skin area, that the Langerin/CD207(+) LCs constitute a predominant population of immature DCs, which express CD1a, and CD68, but not CD83, CD86, and DC-lysosomal-associated membrane protein (LAMP)/CD208. Using LC-type cells generated in vitro in the presence of transforming growth factor (TGF)-beta1, we further found that tumor necrosis factor (TNF)-alpha, as a prototype proinflammatory factor, and a variety of inflammatory stimuli and bacterial products, increase Langerin expression and Langerin dependent Birbeck granules formation in cell which nevertheless lack costimulatory molecules, DC-LAMP/CD208 and potent T cell stimulatory activity but express CCR7 and respond to the lymph node homing chemokines CCL19 and CCL21. This indicates that LC migration and maturation can be independently regulated events. We suggest that during DL, inflammatory stimuli in the skin increase the migration of LCs to the lymph node but without associated maturation. Immature LCs might regulate immune responses during chronic inflammation.     

Redirecting migration of T cells to chemokine secreted from tumors by genetic modification with CXCR2

T-cell-based immunotherapies provide a promising means of cancer treatment although durable antitumor responses are infrequent. A potential reason for these shortcomings may lie in the observed lack of trafficking of specific T cells to tumor. Our increasing knowledge of the process of trafficking involving adhesion molecules and chemokines affords us the opportunity to intervene and correct deficiencies in this process. Chemokines can be expressed by a range of tumors and may serve as suitable targets for directing specific T cells toward tumor. We initially sought to identify which chemokines were produced by a range of human tumor cell lines, and which chemokines and chemokine receptors were expressed by cultured T cells. We identified two chemokines: Growth-Regulated Oncogene-alpha (Gro-alpha; CXCL1) and Regulated on Activation Normal T Cell-Expressed and Secreted (RANTES; CCL5), to be secreted by several human tumor cell lines. Expression was also detected in fine-needle aspirates of melanoma from patients. In addition, we determined the expression of several chemokine receptors on cultured human T cells including CCR1, CCR2, CCR4, CCR5, CXCR3, and CXCR4. Cultured, activated human T cells expressed the chemokines lymphotactin (XCL1), RANTES, macrophage inflammatory protein-1 alpha (MIP-1 alpha; CCL3) and MIP-1 beta (CCL4), but no appreciable Gro-alpha. In a strategy to direct T cells toward chemokines expressed by tumors we chose Gro-alpha as the target chemokine because it was produced by tumor and not by T cells themselves. However, T cells did not express the receptor for Gro-alpha, CXCR2, and therefore, T cells were transduced with a retroviral vector encoding CXCR2. Calcium ion mobilization, an important first step in chemokine receptor signaling, was subsequently demonstrated in transduced T cells in response to Gro-alpha. In addition, Gro-alpha was chemotactic for T cells expressing CXCR2 in vitro toward both recombinant protein and tumor-derived chemokine. Interestingly we demonstrate, for the first time, that Gro-alpha was able to induce interferon-gamma (IFN-gamma) secretion from transduced T cells, thereby extending our knowledge of other potential functions of CXCR2. This study demonstrates the feasibility of redirecting the migration properties of T cells toward chemokines secreted by tumors.     

Lack of chemokine receptor CCR1 enhances Th1 responses and glomerular injury during nephrotoxic nephritis

During the development of nephrotoxic nephritis (NTN) in the mouse, we find that a variety of chemokines and chemokine receptors are induced: CCR1 (RANTES, MIP-1alpha), CCR2 (MCP-1), CCR5 (RANTES, MIP-1alpha, MIP-1beta), CXCR2 (MIP-2), and CXCR3 (IP-10). Their timing of expression indicated that CXCR2 and CCR1 are probably important in the neutrophil-dependent heterologous phase of the disease, whereas CCR1, CCR2, CCR5, and CXCR3 accompany the subsequent mononuclear cell infiltration characteristic of autologous disease. We therefore assessed the role of CCR1 in NTN using CCR1(-/-) mice. We found that neutrophil accumulation in CCR1(-/-) mice was comparable to that in wild-type animals but that renal recruitment of CD4(+) and CD8(+) T cells and macrophages increased significantly. Moreover, CCR1(-/-) mice developed more severe glomerulonephritis than did controls, with greater proteinuria and blood urea nitrogen, as well as a higher frequency of crescent formation. In addition, CCR1(-/-) mice showed enhanced Th1 immune responses, including titers of antigen-specific IgG2a antibody, delayed-type hypersensitivity responses, and production of IFN-gamma and TNF-alpha. Lastly, using recombinant proteins and transfected cells that overexpressed CCR1, we demonstrated that MIP-1alpha, but not RANTES, bound CCR1 and induced cell chemotaxis. Thus, rather than simply promoting leukocyte recruitment during NTN, CCR1 expression profoundly alters the effector phase of glomerulonephritis. Therapeutic targeting of chemokine receptors may, on occasion, exacerbate underlying disease.     

CC chemokine receptor (CCR)2 is required for langerhans cell migration and localization of T helper cell type 1 (Th1)-inducing dendritic cells. Absence of CCR2 shifts the Leishmania major-resistant phenotype to a susceptible state dominated by Th2 cytokines, b cell outgrowth, and sustained neutrophilic inflammation

There is growing evidence that chemokines and their receptors regulate the movement and interaction of antigen-presenting cells such as dendritic cells (DCs) and T cells. We tested the hypothesis that the CC chemokine receptor (CCR)2 and CCR5 and the chemokine macrophage inflammatory protein (MIP)-1alpha, a ligand for CCR5, influence DC migration and localization. We found that deficiency of CCR2 but not CCR5 or MIP-1alpha led to distinct defects in DC biology. Langerhans cell (skin DC) density in CCR2-null mice was normal, and their ability to migrate into the dermis was intact; however, their migration to the draining lymph nodes was markedly impaired. CCR2-null mice had lower numbers of DCs in the spleen, and this was primarily due to a reduction in the CD8alpha(1) T helper cell type 1 (Th1)-inducing subset of DCs. Additionally, there was a block in the Leishmania major infection-induced relocalization of splenic DCs from the marginal zone to the T cell areas. We propose that these DC defects, in conjunction with increased expression of B lymphocyte chemoattractant, a B cell-specific chemokine, may collectively contribute to the striking B cell outgrowth and Th2 cytokine-biased nonhealing phenotype that we observed in CCR2-deficient mice infected with L. major. This disease phenotype in mice with an L. major-resistant genetic background but lacking CCR2 is strikingly reminiscent of that observed typically in mice with an L. major-susceptible genetic background. Thus, CCR2 is an important determinant of not only DC migration and localization but also the development of protective cell-mediated immune responses to L. major.     

Dominant myelopoietic effector functions mediated by chemokine receptor CCR1.

Macrophage inflammatory protein (MIP)-1alpha, a CC chemokine, enhances proliferation of mature subsets of myeloid progenitor cells (MPCs), suppresses proliferation of immature MPCs, and mobilizes mature and immature MPCs to the blood. MIP-1alpha binds at least three chemokine receptors. To determine if CCR1 was dominantly mediating the above activities of MIP-1alpha, CCR1-deficient (-/-) mice, produced by targeted gene disruption, were used. MIP-1alpha enhanced colony formation of marrow granulocyte/macrophage colony-forming units (CFU-GM), responsive to stimulation by granulocyte/macrophage colony-stimulating factor (GM-CSF), and CFU-M, responsive to stimulation by M-CSF, from littermate control CCR1(+/+) but not CCR1(-/-) mice. Moreover, MIP-1alpha did not mobilize MPCs to the blood or synergize with G-CSF in this effect in CCR1(-/-) mice. However, CCR1(-/-) mice were increased in sensitivity to MPC mobilizing effects of G-CSF. Multi-growth factor-stimulated MPCs in CCR1(-/-) and CCR1(+/+) marrow were equally sensitive to inhibition by MIP-1alpha. These results implicate CCR1 as a dominant receptor for MIP-1alpha enhancement of proliferation of lineage-committed MPCs and for mobilization of MPCs to the blood. CCR1 is not a dominant receptor for MIP-1alpha suppression of MPC proliferation, but it does negatively impact G-CSF-induced MPC mobilization.     

Interleukin 15 skews monocyte differentiation into dendritic cells with features of Langerhans cells

Langerhans cells (LCs) represent a subset of immature dendritic cells (DCs) specifically localized in the epidermis and other mucosal epithelia. As surrounding keratinocytes can produce interleukin (IL)-15, a cytokine that utilizes IL-2Rgamma chain, we analyzed whether IL-15 could skew monocyte differentiation into LCs. Monocytes cultured for 6 d with granulocyte/macrophage colony-stimulating factor (GM-CSF) and IL-15 differentiate into CD1a(+)HLA-DR(+)CD14(-)DCs (IL15-DCs). Agents such as lipopolysaccharide (LPS), tumor necrosis factor (TNF)alpha, and CD40L induce maturation of IL15-DCs to CD83(+), DC-LAMP(+) cells. IL15-DCs are potent antigen-presenting cells able to induce the primary (mixed lymphocyte reaction [MLR]) and secondary (recall responses to flu-matrix peptide) immune responses. As opposed to cultures made with GM-CSF/IL-4 (IL4-DCs), a proportion of IL15-DCs expresses LC markers: E-Cadherin, Langerin, and CC chemokine receptor (CCR)6. Accordingly, IL15-DCs, but not IL4-DCs, migrate in response to macrophage inflammatory protein (MIP)-3alpha/CCL20. However, IL15-DCs cannot be qualified as "genuine" Langerhans cells because, despite the presence of the 43-kD Langerin, they do not express bona fide Birbeck granules. Thus, our results demonstrate a novel pathway in monocyte differentiation into dendritic cells.     

Blockade of CC chemokine receptor 5 (CCR5)-tropic human immunodeficiency virus-1 replication in human lymphoid tissue by CC chemokines.

The CC chemokines MIP-1alpha, MIP-1beta, and RANTES suppress replication of certain HIV-1 strains in cultured PBMC and T cell lines by blocking interaction of gp120 with CC chemokine receptor 5 (CCR5). However, the same chemokines can enhance HIV-1 replication in cultured macrophages. The net effect of chemokines on HIV-1 infection in intact lymphoid tissue, the major reservoir of HIV-1 in vivo, is unknown and unpredictable since the tissue contains both T lymphocytes and macrophages. Here we show that exogenous MIP-1alpha, MIP-1beta, and RANTES markedly suppressed replication of CCR5-tropic HIV-1 strains in blocks of human lymphoid tissue infected ex vivo. Moreover, endogenous MIP-1alpha, MIP-1beta, and RANTES were upregulated in tissues infected ex vivo with CXC chemokine receptor 4-tropic but not CCR5-tropic HIV-1. Such an upregulation may contribute to the virus phenotype shift in the course of HIV disease in vivo.     

Unique chemotactic response profile and specific expression of chemokine receptors CCR4 and CCR8 by CD4(+)CD25(+) regulatory T cells

Chemokines dictate regional trafficking of functionally distinct T cell subsets. In rodents and humans, a unique subset of CD4(+)CD25(+) cytotoxic T lymphocyte antigen (CTLA)-4(+) regulatory T cells (Treg) has been proposed to control peripheral tolerance. However, the molecular basis of immune suppression and the trafficking properties of Treg cells are still unknown. Here, we determined the chemotactic response profile and chemokine receptor expression of human blood-borne CD4(+)CD25(+) Treg cells. These Treg cells were found to vigorously respond to several inflammatory and lymphoid chemokines. Treg cells specifically express the chemokine receptors CCR4 and CCR8 and represent a major subset of circulating CD4(+) T cells responding to the chemokines macrophage-derived chemokine (MDC)/CCL22, thymus and activation-regulated chemokine (TARC)/CCL17, I-309/CCL1, and to the virokine vMIP-I (ligands of CCR4 and CCR8). Blood-borne CD4(+) T cells that migrate in response to CCL1 and CCL22 exhibit a reduced alloproliferative response, dependent on the increased frequency of Treg cells in the migrated population. Importantly, mature dendritic cells preferentially attract Treg cells among circulating CD4(+) T cells, by secretion of CCR4 ligands CCL17 and CCL22. Overall, these results suggest that CCR4 and/or CCR8 may guide Treg cells to sites of antigen presentation in secondary lymphoid tissues and inflamed areas to attenuate T cell activation.     

A key role for CC chemokine receptor 4 in lipopolysaccharide-induced endotoxic shock

CC chemokine receptor (CCR)4, a high affinity receptor for the CC chemokines thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC), is expressed in the thymus and spleen, and also by peripheral blood T cells, macrophages, platelets, and basophils. Recent studies have shown that CCR4 is the major chemokine receptor expressed by T helper type 2 (Th2) polarized cells. To study the in vivo role of CCR4, we have generated CCR4-deficient (CCR4(-/-)) mice by gene targeting. CCR4(-/-) mice developed normally. Splenocytes and thymocytes isolated from the CCR4(-/-) mice failed to respond to the CCR4 ligands TARC and MDC, as expected, but also surprisingly did not undergo chemotaxis in vitro in response to macrophage inflammatory protein (MIP)-1alpha. The CCR4 deletion had no effect on Th2 differentiation in vitro or in a Th2-dependent model of allergic airway inflammation. However, CCR4(-/-) mice exhibited significantly decreased mortality on administration of high or low dose bacterial lipopolysaccharide (LPS) compared with CCR4(+/+) mice. After high dose LPS treatment, serum levels of tumor necrosis factor alpha, interleukin 1beta, and MIP-1alpha were reduced in CCR4(-/-) mice, and decreased expression of MDC and MIP-2 mRNA was detected in peritoneal exudate cells. Analysis of peritoneal lavage cells from CCR4(-/)- mice by flow cytometry also revealed a significant decrease in the F4/80(+) cell population. This may reflect a defect in the ability of the CCR4(-/-) macrophages to be retained in the peritoneal cavity. Taken together, our data reveal an unexpected role for CCR4 in the inflammatory response leading to LPS-induced lethality.     

CC chemokine receptor (CCR)4 and the CCR10 ligand cutaneous T cell-attracting chemokine (CTACK) in lymphocyte trafficking to inflamed skin.

The chemokine thymus and activation-regulated chemokine (TARC; CCL17) is displayed by cutaneous (but not intestinal) venules, and is thought to trigger vascular arrest of circulating skin homing memory T cells, which uniformly express the TARC receptor CC chemokine receptor (CCR)4. Cutaneous T cell-attracting chemokine (CTACK; CCL27), expressed by skin keratinocytes, also attracts cutaneous memory T cells, and is hypothesized to assist in lymphocyte recruitment to skin as well. Here we show that chronic cutaneous inflammation induces CD4 T cells expressing E-selectin binding activity (a marker of skin homing memory cells) in draining lymph node, and that these E-selectin ligand+ T cells migrate efficiently to TARC and to CTACK. In 24 h in vivo homing assays, stimulated lymph node T cells from wild-type mice or, surprisingly, from CCR4-deficient donors migrate efficiently to inflamed skin; and an inhibitory anti-CTACK antibody has no effect on wild-type lymphocyte recruitment. However, inhibition with anti-CTACK monoclonal antibody abrogates skin recruitment of CCR4-deficient T cells. We conclude that CTACK and CCR4 can both support homing of T cells to skin, and that either one or the other is required for lymphocyte recruitment in cutaneous delayed type hypersensitivity.     

Selective Recruitment of Immature and Mature Dendritic Cells by Distinct Chemokines Expressed in Different Anatomic Sites

DCs (dendritic cells) function as sentinels of the immune system. They traffic from the blood to the tissues where, while immature, they capture antigens. They then leave the tissues and move to the draining lymphoid organs where, converted into mature DC, they prime naive T cells. This suggestive link between DC traffic pattern and functions led us to investigate the chemokine responsiveness of DCs during their development and maturation. DCs were differentiated either from CD34⁺ hematopoietic progenitor cells (HPCs) cultured with granulocyte/macrophage colony–stimulating factor (GM-CSF) plus tumor necrosis factor (TNF)-α or from monocytes cultured with GM-CSF plus interleukin 4. Immature DCs derived from CD34⁺ HPCs migrate most vigorously in response to macrophage inflammatory protein (MIP)-3α, but also to MIP-1α and RANTES (regulated on activation, normal T cell expressed and secreted). Upon maturation, induced by either TNF-α, lipopolysaccharide, or CD40L, DCs lose their response to these three chemokines when they acquire a sustained responsiveness to a single other chemokine, MIP-3β. CC chemokine receptor (CCR)6 and CCR7 are the only known receptors for MIP-3α and MIP-3β, respectively. The observation that CCR6 mRNA expression decreases progressively as DCs mature, whereas CCR7 mRNA expression is sharply upregulated, provides a likely explanation for the changes in chemokine responsiveness. Similarly, MIP-3β responsiveness and CCR7 expression are induced upon maturation of monocyte- derived DCs. Furthermore, the chemotactic response to MIP-3β is also acquired by CD11c⁺ DCs isolated from blood after spontaneous maturation. Finally, detection by in situ hybridization of MIP-3α mRNA only within inflamed epithelial crypts of tonsils, and of MIP-3β mRNA specifically in T cell–rich areas, suggests a role for MIP-3α/CCR6 in recruitment of immature DCs at site of injury and for MIP-3β/CCR7 in accumulation of antigen-loaded mature DCs in T cell–rich areas.     

Macrophage-tropic HIV induces and exploits dendritic cell chemotaxis

Immature dendritic cells (iDCs) express the CC chemokine receptor (CCR)5, which promotes chemotaxis toward the CC chemokines regulated on activation, normal T cell expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta. By contrast, mature DCs downregulate CCR5 but upregulate CXC chemokine receptor (CXCR)4, and as a result exhibit enhanced chemotaxis toward stromal cell-derived factor (SDF)-1alpha. CCR5 and CXCR4 also function as coreceptors for macrophage-tropic (M-tropic) and T cell-tropic (T-tropic) human immunodeficiency virus (HIV)-1, respectively. Here, we demonstrate chemotaxis of iDCs toward M-tropic (R5) but not T-tropic (X4) HIV-1. Furthermore, preexposure to M-tropic HIV-1 or its recombinant envelope protein prevents migration toward CCR5 ligands. The migration of iDCs toward M-tropic HIV-1 may enhance formation of DC-T cell syncytia, thus promoting viral production and destruction of both DC and T helper lymphocytes. Therefore, disturbance of DC chemotaxis by HIV-1 is likely to contribute to immunosuppression in primary infection and AIDS. In addition, migration of iDCs toward HIV-1 may aid the capture of R5 HIV-1 virions by the abundant DC cell surface protein DC-specific intercellular adhesion molecule (ICAM)3-grabbing nonintegrin (DC-SIGN). HIV-1 bound to DC cell-specific DC-SIGN retains the ability to infect replication-permissive T cells in trans for several days. Consequently, recruitment of DC by HIV-1 could combine with the ability of DC-SIGN to capture and transmit the virus to T cells, and so facilitate dissemination of virus within an infected individual.     

CC-chemokine receptor 5 (CCR5) in hepatitis C--at the crossroads of the antiviral immune response?

An effective immune response to hepatitis C virus (HCV) infection requires efficient recruitment and activation of inflammatory cells to the liver, the site of infection. Chemokines are critically involved in this process, since they exert both chemotactic and immunoregulatory actions. In particular, the interaction between chemokines CCL3 (MIP-1alpha), CCL4 (MIP-1beta) and CCL5 (RANTES) and their receptor, CC-chemokine receptor 5 (CCR5), may be critical in regulating T cell functions by mediating recruitment, polarization, activation and differentiation of antiviral type 1 cytokine secreting T helper and cytotoxic T cells. A 32 bp deletion in the encoding region of CCR5 leads to complete loss of the functional CCR5 receptor in subjects homozygous for this mutation and decreased expression in heterozygous patients. This fact provides the unique opportunity to study the role of the CCR5 receptor in chronic hepatitis C infection by comparing immune responses between HCV infected CCR5-Delta32 carriers and CCR5 wild-type patients. This article will summarize and discuss the available data with respect to possibly altered disease susceptibility, clinical course and treatment outcomes associated with the CCR5-Delta32 mutation in hepatitis C.     

Developmental switches in chemokine response profiles during B cell differentiation and maturation

Developing B cells undergo dramatic changes in their responses to chemoattractant cytokines (chemokines) and in expression of chemokine receptors. Bone marrow pre-pro-B cells (AA4.1(+)/natural killer 1.1(-) Fraction A cells) and cells capable of generating pro-B colonies in the presence of interleukin 7 and flt3 ligand migrate to thymus-expressed chemokine (TECK), a response lost in later stages of B cell development. B cell-attracting chemokine 1 (BCA-1) responses correlate with CXC chemokine receptor (CXCR)5 expression, are first displayed by a pro-B cell subset, are lost in pre-B cells, and then are regained just before and after egress from the marrow. All peripheral B cell subsets, including follicular and germinal center as well as marginal zone and peritoneal B1 B cells, respond to BCA-1, implying that responsiveness to this follicular chemokine is not sufficient to predict follicle localization. Responses to the CC chemokine receptor (CCR)7 ligands secondary lymphoid tissue chemoattractant (SLC) and macrophage inflammatory protein (MIP)-3beta, implicated in homing to lymphoid tissues, are upregulated before B cell exit from the marrow, but increase further in the periphery and are shared by all peripheral B cells. In contrast, responsiveness to MIP-3alpha and expression of CCR6 are acquired only after emigration to the periphery and during maturation into the recirculating B cell pool. Chemotaxis to stromal cell-derived factor 1alpha is observed at all stages of B cell differentiation. Thus, unique patterns of chemokine responses may help define developing B cell populations and direct their maturation in the marrow and migration to the periphery.     

Recruitment of Foxp3+ T regulatory cells mediating allograft tolerance depends on the CCR4 chemokine receptor

Although certain chemokines and their receptors guide homeostatic recirculation of T cells and others promote recruitment of activated T cells to inflammatory sites, little is known of the mechanisms underlying a third function, migration of Foxp3(+) regulatory T (T reg) cells to sites where they maintain unresponsiveness. We studied how T reg cells are recruited to cardiac allografts in recipients tolerized with CD154 monoclonal antibody (mAb) plus donor-specific transfusion (DST). Real-time polymerase chain reaction showed that intragraft Foxp3 levels in tolerized recipients were approximately 100-fold higher than rejecting allografts or allografts associated with other therapies inducing prolonged survival but not tolerance. Foxp3(+) cells were essential for tolerance because pretransplant thymectomy or peritransplant depletion of CD25(+) cells prevented long-term survival, as did CD25 mAb therapy in well-functioning allografts after CD154/DST therapy. Analysis of multiple chemokine pathways showed that tolerance was accompanied by intragraft up-regulation of CCR4 and one of its ligands, macrophage-derived chemokine (CCL22), and that tolerance induction could not be achieved in CCR4(-/-) recipients. We conclude that Foxp3 expression is specifically up-regulated within allografts of mice displaying donor-specific tolerance, that recruitment of Foxp3-expressing T reg cells to an allograft tissue is dependent on the chemokine receptor, CCR4, and that, in the absence of such recruitment, tolerizing strategies such as CD154 mAb therapy are ineffectual.     

Cooperating mechanisms of CXCR5 and CCR7 in development and organization of secondary lymphoid organs

Homeostatic chemokines participate in the development of secondary lymphoid organs and later on in the functional organization of these tissues. The development of lymph nodes (LNs) and Peyer's patches depends on the recruitment of CD3- CD4+ interleukin (IL)-7R alpha hi cells to sites of future organ development. CD3- CD4+ IL-7R alpha hi cells express the chemokine receptor CXCR5 and might be attracted by its ligand CXCL13, which is secreted by mesenchymal cells. Mesenchymal cells also secrete CCL19, a ligand for CCR7, yet it is not clear whether CCR7 and CCL19 are important for secondary lymphoid organ development. Analyzing CXCR5-/- CCR7-/- double deficient mice we now show that these mice lack all examined peripheral LNs suggesting a profound role for both receptors in secondary lymphoid organ development. We demonstrate that CD3- CD4+ IL-7R alpha hi cells express CXCR5 as well as CCR7 indicating that both receptors cooperate during an early step of secondary lymphoid organ development. Furthermore, CXCR5-/- CCR7-/- mice display a severely disturbed architecture of mesenteric LN and spleen. Due to an impaired migration of B cells into the white pulp, CXCR5-/- CCR7-/- mice fail to develop B cell follicles but show small clusters of unorganized lymphocytes in the spleen. These data demonstrate a cooperative function of CXCR5 and CCR7 in lymphoid organ organogenesis and organization.