Chemokine receptor CCR5 deficiency exacerbates cerulein-induced acute pancreatitis in mice

Acute pancreatitis (AP) is an inflammatory disease involving the production of different cytokines and chemokines and is characterized by leukocyte infiltration. Because the chemokine receptor CCR5 and its ligands [the CC chemokines CCL3/MIP-1alpha, CCL4/MIP-1beta, and CCL5/regulated upon activation, normal T cell expressed and secreted (RANTES)] regulate leukocyte chemotaxis and activation, we investigated the expression of CCR5 ligands and the role of CCR5 and its ligands in experimental AP in mice. AP was induced by hourly intraperitoneal injections of cerulein in CCR5-deficient (CCR5(-/-)) or wild-type (WT) mice. Induction of AP by cerulein resulted in an early increase of pancreatic CCL2, CCL3, and CCL4 mRNA expression, whereas CCL5 mRNA expression occurred later. CCR5(-/-) mice developed a more severe pancreatic injury than WT mice during cerulein-induced AP, as assessed by a more pronounced increase in serum amylase and lipase levels and by more severe pancreatic edema, inflammatory infiltrates (mainly neutrophils), and necrosis. CCR5(-/-) mice also exhibited increased production of CCL2/MCP-1, CCL3/MIP-1alpha, and CCL4/MIP-1beta during the course of cerulein-induced AP. In vivo simultaneous neutralization of CC chemokines with monoclonal antibodies in CCR5(-/-) mice reduced the severity of cerulein-induced AP, indicating a role of CC chemokines in exacerbating the course of AP in the absence of CCR5. Moreover, simultaneous neutralization of CCR5 ligands in WT mice also reduced the severity of cerulein-induced AP. In conclusion, lack of the chemokine receptor CCR5 exacerbates experimental cerulein-induced AP and leads to increased levels of CC chemokines and a more pronounced pancreatic inflammatory infiltrate, suggesting that CCR5 expression can modulate severity of AP.     

Chemokine Receptor CCR8 Is Required for Lipopolysaccharide-Triggered Cytokine Production in Mouse Peritoneal Macrophages

Chemokine (C-C motif) receptor 8 (CCR8), the chemokine receptor for chemokine (C-C motif) ligand 1 (CCL1), is expressed in T-helper type-2 lymphocytes and peritoneal macrophages (PMφ) and is involved in various pathological conditions, including peritoneal adhesions. However, the role of CCR8 in inflammatory responses is not fully elucidated. To investigate the function of CCR8 in macrophages, we compared cytokine secretion from mouse PMφ or bone marrow-derived macrophages (BMMφ) stimulated with various Toll-like receptor (TLR) ligands in CCR8 deficient (CCR8^{- /-}) and wild-type (WT) mice. We found that CCR8^-/- PMφ demonstrated attenuated secretion of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-10 when stimulated with lipopolysaccharide (LPS). In particular, LPS-induced IL-10 production absolutely required CCR8. CCR8-dependent cytokine secretion was characteristic of PMφ but not BMMφ. To further investigate this result, we selected the small molecule compound R243 from a library of compounds with CCR8-antagonistic effects on CCL1-induced Ca²⁺ flux and CCL1-driven PMφ aggregation. Similar to CCR8^-/- PMφ, R243 attenuated secretion of TNF-α, IL-6, and most strikingly IL-10 from WT PMφ, but not BMMφ. CCR8^-/- PMφ and R243-treated WT PMφ both showed suppressed c-jun N-terminal kinase activity and nuclear factor-κB signaling after LPS treatment when compared with WT PMφ. A c-Jun signaling pathway inhibitor also produced an inhibitory effect on LPS-induced cytokine secretion that was similar to that of CCR8 deficiency or R243 treatment. As seen in CCR8^-/- mice, administration of R243 attenuated peritoneal adhesions in vivo. R243 also prevented hapten-induced colitis. These results are indicative of cross talk between signaling pathways downstream of CCR8 and TLR-4 that induces cytokine production by PMφ. Through use of CCR8^-/- mice and the new CCR8 inhibitor, R243, we identified a novel macrophage innate immune response pathway that involves a chemokine receptor.     

CCR1 and CC chemokine ligand 5 interactions exacerbate innate immune responses during sepsis

CCR1 has previously been shown to play important roles in leukocyte trafficking, pathogen clearance, and the type 1/type 2 cytokine balance, although very little is known about its role in the host response during sepsis. In a cecal ligation and puncture model of septic peritonitis, CCR1-deficient (CCR1(-/-)) mice were significantly protected from the lethal effects of sepsis when compared with wild-type (WT) controls. The peritoneal and systemic cytokine profile in CCR1(-/-) mice was characterized by a robust, but short-lived and regulated antibacterial response. CCR1 expression was not required for leukocyte recruitment, suggesting critical differences extant in the activation of WT and CCR1(-/-) resident or recruited peritoneal cells during sepsis. Peritoneal macrophages isolated from naive CCR1(-/-) mice clearly demonstrated enhanced cytokine/chemokine generation and antibacterial responses compared with similarly treated WT macrophages. CCR1 and CCL5 interactions markedly altered the inflammatory response in vivo and in vitro. Administration of CCL5 increased sepsis-induced lethality in WT mice, whereas neutralization of CCL5 improved survival. CCL5 acted in a CCR1-dependent manner to augment production of IFN-gamma and MIP-2 to damaging levels. These data illustrate that the interaction between CCR1 and CCL5 modulates the innate immune response during sepsis, and both represent potential targets for therapeutic intervention.     

CCR5 deficiency aggravates crescentic glomerulonephritis in mice

The chemokine receptor CCR5 is predominantly expressed on monocytes and Th1-polarized T cells, and plays an important role in T cell and monocyte recruitment in inflammatory diseases. To investigate the functional role of CCR5 in renal inflammation, we induced a T cell-dependent model of glomerulonephritis (nephrotoxic serum nephritis) in CCR5(-/-) mice. Induction of nephritis in wild-type mice resulted in up-regulation of renal mRNA expression of the three CCR5 chemokine ligands, CCL5 (15-fold), CCL3 (4.9-fold), and CCL4 (3.4-fold), in the autologous phase of the disease at day 10. The up-regulated chemokine expression was paralleled by infiltration of monocytes and T cells, followed by renal tissue injury, albuminuria, and loss of renal function. Nephritic CCR5(-/-) mice showed a 3- to 4-fold increased renal expression of CCL5 (61.6-fold vs controls) and CCL3 (14.1-fold vs controls), but not of CCL4, in comparison with nephritic wild-type mice, which was accompanied by augmented renal T cell and monocyte recruitment and increased lethality due to uremia. Furthermore, CCR5(-/-) mice showed an increased renal Th1 response, whereas their systemic humoral and cellular immune responses were unaltered. Because the CCR5 ligands CCL5 and CCL3 also act via CCR1, we investigated the effects of the pharmacological CCR1 antagonist BX471. CCR1 blockade in CCR5(-/-) mice significantly reduced renal chemokine expression, T cell infiltration, and glomerular crescent formation, indicating that increased renal leukocyte recruitment and consecutive tissue damage in nephritic CCR5(-/-) mice depended on functional CCR1. In conclusion, this study shows that CCR5 deficiency aggravates glomerulonephritis via enhanced CCL3/CCL5-CCR1-driven renal T cell recruitment.     

Differential role of CCR2 in islet and heart allograft rejection: tissue specificity of chemokine/chemokine receptor function in vivo

Chemokines have a pivotal role in the mobilization and activation of specific leukocyte subsets in acute allograft rejection. However, the role of specific chemokines and chemokine receptors in islet allograft rejection has not been fully elucidated. We now show that islet allograft rejection is associated with a steady increase in intragraft expression of the chemokines CCL8 (monocyte chemoattractant protein-2), CCL9 (monocyte chemoattractant protein-5), CCL5 (RANTES), CXCL-10 (IFN-gamma-inducible protein-10), and CXCL9 (monokine induced by IFN-gamma) and their corresponding chemokine receptors CCR2, CCR5, CCR1, and CXCR3. Because CCR2 was found to be highly induced, we tested the specific role of CCR2 in islet allograft rejection by transplanting fully MHC mismatched islets from BALB/c mice into C57BL/6 wild-type (WT) and CCR2-deficient mice (CCR2-/-). A significant prolongation of islet allograft survival was noted in CCR2-/- recipients, with median survival time of 24 and 12 days for CCR2-/- and WT recipients, respectively (p < 0.0001). This was associated with reduction in the generation of CD8+, but not CD4+ effector alloreactive T cells (CD62L(low)CD44(high)) in CCR2-/- compared with WT recipients. In addition, CCR2-/- recipients had a reduced Th1 and increased Th2 alloresponse in the periphery (by ELISPOT analysis) as well as in the grafts (by RT-PCR). However, these changes were only transient in CCR2-/- recipients that ultimately rejected their grafts. Furthermore, in contrast to the islet transplants, CCR2 deficiency offered only marginal prolongation of heart allograft survival. This study demonstrates the important role for CCR2 in early islet allograft rejection and highlights the tissue specificity of the chemokine/chemokine receptor system in vivo in regulating allograft rejection.     

Regulation of inflammatory chemokine receptors on blood T cells associated to the circulating versus liver chemokines in dengue fever

Little is known about the role of chemokines/chemokines receptors on T cells in natural DENV infection. Patients from DENV-2 and -3- outbreaks were studied prospectively during the acute or convalescent phases. Expression of chemokine receptor and activation markers on lymphocyte subpopulations were determined by flow cytometry analysis, plasma chemokine ligands concentrations were measured by ELISA and quantification of CCL5/RANTES(+) cells in liver tissues from fatal dengue cases was performed by immunochemistry. In the acute DENV-infection, T-helper/T-cytotoxic type-1 cell (Th1/Tc1)-related CCR5 is significantly higher expressed on both CD4 and CD8 T cells. The Th1-related CXCR3 is up-regulated among CD4 T cells and Tc2-related CCR4 is up-regulated among CD8 T cells. In the convalescent phase, all chemokine receptor or chemokine ligand expression tends to reestablish control healthy levels. Increased CCL2/MCP-1 and CCL4/MIP-1β but decreased CCL5/RANTES levels were observed in DENV-patients during acute infection. Moreover, we showed an increased CD107a expression on CCR5 or CXCR3-expressing T cells and higher expression of CD29, CD44(HIGH) and CD127(LOW) markers on CCR4-expressing CD8 T cells in DENV-patients when compared to controls. Finally, liver from dengue fatal patients showed increased number of cells expressing CCL5/RANTES in three out of four cases compared to three death from a non-dengue patient. In conclusion, both Th1-related CCR5 and CXCR3 among CD4 T cells have a potential ability to exert cytotoxicity function. Moreover, Tc1-related CCR5 and Tc2-related CCR4 among CD8 T cells have a potential ability to exert effector function and migration based on cell markers evaluated. The CCR5 expression would be promoting an enhanced T cell recruitment into liver, a hypothesis that is corroborated by the CCL5/RANTES increase detected in hepatic tissue from dengue fatal cases. The balance between protective and pathogenic immune response mediated by chemokines during dengue fever will be discussed.     

The Absence of CCR7 Results in Dysregulated Monocyte Migration and Immunosuppression Facilitating Chronic Cutaneous Leishmaniasis

The protozoan parasite Leishmania major causes cutaneous lesions to develop at the site of infection, which are resolved with a strong Th1 immune response in resistant hosts, such as C57BL/6 mice. In contrast, the lesions ulcerate in susceptible hosts which display a Th2 response, such as BALB/c mice. The migration of cells in the immune response to L. major is regulated by chemokines and their receptors. The chemokine receptor CCR7 is expressed on activated DCs and naïve T cells, allowing them to migrate to the correct micro-anatomical positions within secondary lymphoid organs. While there have been many studies on the function of CCR7 during homeostasis or using model antigens, there are very few studies on the role of CCR7 during infection. In this study, we show that B6.CCR7^-/- mice were unable to resolve the lesion and developed a chronic disease. The composition of the local infiltrate at the lesion was significantly skewed toward neutrophils while the proportion of CCR2⁺ monocytes was reduced. Furthermore, a greater percentage of CCR2⁺ monocytes expressed CCR7 in the footpad than in the lymph node or spleen of B6.WT mice. We also found an increased percentage of regulatory T cells in the draining lymph node of B6.CCR7^-/- mice throughout infection. Additionally, the cytokine milieu of the lymph node showed a Th2 bias, rather than the resistant Th1 phenotype. This data shows that CCR7 is required for a protective immune response to intracellular L. major infection.     

CCR5 Controls Immune and Metabolic Functions during Toxoplasma gondii Infection

CCR5, an important receptor related to cell recruitment and inflammation, is expressed during experimental Toxoplasma gondii infection. However, its role in the immunopathology of toxoplasmosis is not clearly defined yet. Thus, we inoculated WT and CCR5^-/- mice with a sub lethal dose of the parasite by oral route. CCR5^-/- mice were extremely susceptible to infection, presenting higher parasite load and lower tissue expression of IL-12p40, IFN-γ, TNF, IL-6, iNOS, Foxp3, T-bet, GATA-3 and PPARα. Although both groups presented inflammation in the liver with prominent neutrophil infiltration, CCR5^-/- mice had extensive tissue damage with hepatocyte vacuolization, steatosis, elevated serum triglycerides and transaminases. PPARα agonist Gemfibrozil improved the vacuolization but did not rescue CCR5^-/- infected mice from high serum triglycerides levels and enhanced mortality. We also found intense inflammation in the ileum of CCR5^-/- infected mice, with epithelial ulceration, augmented CD4 and decreased frequency of NK cells in the gut lamina propria. Most interestingly, these findings were accompanied by an outstanding accumulation of neutrophils in the ileum, which seemed to be involved in the gut immunopathology, once the depletion of these cells was accompanied by reduced local damage. Altogether, these data demonstrated that CCR5 is essential to the control of T. gondii infection and to maintain the metabolic, hepatic and intestinal integrity. These findings add novel information on the disease pathogenesis and may be relevant for directing future approaches to the treatment of multi-deregulated diseases.     

CCL25/CCR9 interactions regulate large intestinal inflammation in a murine model of acute colitis

Background & Aims

CCL25/CCR9 is a non-promiscuous chemokine/receptor pair and a key regulator of leukocyte migration to the small intestine. We investigated here whether CCL25/CCR9 interactions also play a role in the regulation of inflammatory responses in the large intestine.

Methods

Acute inflammation and recovery in wild-type (WT) and CCR9^−/− mice was studied in a model of dextran sulfate sodium (DSS)-induced colitis. Distribution studies and phenotypic characterization of dendritic cell subsets and macrophage were performed by flow cytometry. Inflammatory bowel disease (IBD) scores were assessed and expression of inflammatory cytokines was studied at the mRNA and the protein level.

Results

CCL25 and CCR9 are both expressed in the large intestine and are upregulated during DSS colitis. CCR9^−/− mice are more susceptible to DSS colitis than WT littermate controls as shown by higher mortality, increased IBD score and delayed recovery. During recovery, the CCR9^−/− colonic mucosa is characterized by the accumulation of activated macrophages and elevated levels of Th1/Th17 inflammatory cytokines. Activated plasmacytoid dendritic cells (DCs) accumulate in mesenteric lymph nodes (MLNs) of CCR9^−/− animals, altering the local ratio of DC subsets. Upon re-stimulation, T cells isolated from these MLNs secrete significantly higher levels of TNFα, IFNγ, IL2, IL-6 and IL-17A while down modulating IL-10 production.

Conclusions

Our results demonstrate that CCL25/CCR9 interactions regulate inflammatory immune responses in the large intestinal mucosa by balancing different subsets of dendritic cells. These findings have important implications for the use of CCR9-inhibitors in therapy of human IBD as they indicate a potential risk for patients with large intestinal inflammation.     

Role of CC chemokine receptor 1 and two of its ligands in human dengue infection. Three approaches under the Cuban situation

Any of the four dengue serotypes can cause a severe disease, partly due to systemic inflammation orchestrated by mediators like cytokines and chemokines. We addressed the role of CCR1 and its ligands CCL3/MIP-1α and CCL5/RANTES in dengue infection using three different approaches: an ex vivo model exploring memory immune response in subjects with a well characterized dengue immune background, an in vivo study in patients with primary or secondary dengue infection, and an approach in fatal dengue. CCR1 and CCL3/MIP-1α gene expression showed differences after homotypic and heterotypic challenge according to dengue immune background of subjects, in correspondence with previous observations in Cuban dengue outbreaks. CCL5/RANTES gene expression was higher after homotypic challenge. CCR1 and CCL3/MIP-1α gene expression was higher in patients with secondary infection during critical days of the dengue disease, while the increase in RANTES expression started earlier than the observed for CCR1 and CCL3/MIP-1α. CCR1 and CCL3/MIP-1α gene expression was as high in brain as in spleen tissue from necropsy. Our results confirm the strong influence of previous immunity in subsequent dengue infections, and confer a possible pathogenic role to CCR1 and CCL3/MIP-1α in dengue disease and a possible protective role for CCL5/RANTES, probably through CCR5 interaction.     

Stimulation of oral fibroblast chemokine receptors identifies CCR3 and CCR4 as potential wound healing targets

The focus of this study was to determine which chemokine receptors are present on oral fibroblasts and whether these receptors influence proliferation, migration, and/or the release of wound healing mediators. This information may provide insight into the superior wound healing characteristics of the oral mucosa. The gingiva fibroblasts expressed 12 different chemokine receptors (CCR3, CCR4, CCR6, CCR9, CCR10, CXCR1, CXCR2, CXCR4, CXCR5, CXCR7, CX3CR1, and XCR1), as analyzed by flow cytometry. Fourteen corresponding chemokines (CCL5, CCL15, CCL20, CCL22, CCL25, CCL27, CCL28, CXCL1, CXCL8, CXCL11, CXCL12, CXCL13, CX3CL1, and XCL1) were used to study the activation of these receptors on gingiva fibroblasts. Twelve of these fourteen chemokines stimulated gingiva fibroblast migration (all except for CXCL8 and CXCL12). Five of the chemokines stimulated proliferation (CCL5/CCR3, CCL15/CCR3, CCL22/CCR4, CCL28/CCR3/CCR10, and XCL1/XCR1). Furthermore, CCL28/CCR3/CCR10 and CCL22/CCR4 stimulation increased IL‐6 secretion and CCL28/CCR3/CCR10 together with CCL27/CCR10 upregulated HGF secretion. Moreover, TIMP‐1 secretion was reduced by CCL15/CCR3. In conclusion, this in‐vitro study identifies chemokine receptor‐ligand pairs which may be used in future targeted wound healing strategies. In particular, we identified the chemokine receptors CCR3 and CCR4, and the mucosa specific chemokine CCL28, as having an predominant role in oral wound healing by increasing human gingiva fibroblast proliferation, migration, and the secretion of IL‐6 and HGF and reducing the secretion of TIMP‐1.     

Targeting Spare CC Chemokine Receptor 5 (CCR5) as a Principle to Inhibit HIV-1 Entry

CCR5 binds the chemokines CCL3, CCL4, and CCL5 and is the major coreceptor for HIV-1 entry into target cells. Chemokines are supposed to form a natural barrier against human immunodeficiency virus, type 1 (HIV-1) infection. However, we showed that their antiviral activity is limited by CCR5 adopting low-chemokine affinity conformations at the cell surface. Here, we investigated whether a pool of CCR5 that is not stabilized by chemokines could represent a target for inhibiting HIV infection. We exploited the characteristics of the chemokine analog PSC-RANTES (N-α-(n-nonanoyl)-des-Ser(1)-[l-thioprolyl(2), l-cyclohexylglycyl(3)]-RANTES(4-68)), which displays potent anti-HIV-1 activity. We show that native chemokines fail to prevent high-affinity binding of PSC-RANTES, analog-mediated calcium release (in desensitization assays), and analog-mediated CCR5 internalization. These results indicate that a pool of spare CCR5 may bind PSC-RANTES but not native chemokines. Improved recognition of CCR5 by PSC-RANTES may explain why the analog promotes higher amounts of β-arrestin 2·CCR5 complexes, thereby increasing CCR5 down-regulation and HIV-1 inhibition. Together, these results highlight that spare CCR5, which might permit HIV-1 to escape from chemokines, should be targeted for efficient viral blockade.     

Lack of CCR2 results in increased mortality and impaired leukocyte activation and trafficking following infection of the central nervous system with a neurotropic coronavirus

In the present study, we evaluated the role of CCR2 in a model of viral-induced neurologic disease. An orchestrated expression of chemokines, including the CCR2 ligands monocyte chemoattractant protein-1/CCL2 and monocyte chemoattractant protein-3/CCL7, occurs within the CNS following infection with mouse hepatitis virus (MHV). Infection of mice lacking CCR2 (CCR2(-/-)) with MHV resulted in increased mortality and enhanced viral recovery from the brain that correlated with reduced (p < or = 0.04) T cell and macrophage/microglial (determined by F4/80 Ag expression, p < or = 0.004) infiltration into the CNS. Moreover, MHV-infected CCR2(-/-) mice displayed a significant decrease in Th1-associated factors IFN-gamma (p < or = 0.001) and RANTES/CCL5 (p < or = 0.002) within the CNS as compared with CCR2(+/+) mice. Further, peripheral CD4(+) and CD8(+) T cells from immunized CCR2(-/-) mice displayed a marked reduction in IFN-gamma production in response to viral Ag and did not migrate into the CNS of MHV-infected recombination-activating gene (RAG)1(-/-) mice following adoptive transfer. In addition, macrophage/microglial infiltration into the CNS of RAG1(-/-) mice receiving CCR2(-/-) splenocytes was reduced (p < or = 0.05), which correlated with a reduction in the severity of demyelination (p < or = 0.001) as compared with RAG1(-/-) mice receiving splenocytes from CCR2(+/+) mice. Collectively, these results indicate an important role for CCR2 in host defense and disease by regulating leukocyte activation and trafficking.     

Nitric oxide synthase-2 modulates chemokine production by Trypanosoma cruzi-infected cardiac myocytes

An intense inflammatory process is associated with Trypanosoma cruzi infection. We investigated the mediators that trigger leukocyte activation and migration to the heart of infected mice. It is known that nitric oxide (NO) modulates the inflammatory response. During T. cruzi infection, increased concentrations of NO are produced by cardiac myocytes (CMs) in response to IFN-gamma and TNF. Here, we investigated whether NO, IFN-gamma and TNF regulate chemokine production by T. cruzi-infected CMs. In addition, we examined the effects of the NOS2 deficiency on chemokine expression both in cultured CMs and in hearts obtained from infected mice. After infection of cultured WT CMs with T. cruzi, the addition of IFN-gamma and TNF increased both mRNA and protein levels of the chemokines CXCL1, CXCL2, CCL2, CCL3, CCL4 and CCL5. Interestingly, T. cruzi-infected NOS2-deficient CMs produced significantly higher levels of CCL2, CCL4, CCL5 and CXL2 in the presence of IFN-gamma and TNF. Infection of NOS2-null mice resulted in a significant increase in the expression of both chemokine mRNA and protein levels in the heart of, compared with hearts obtained from, infected WT mice. Our data indicate that NOS2 is a potent modulator of chemokine expression which is critical to triggering the generation of the inflammatory infiltrate in the heart during T. cruzi infection.     

The chemokine receptor, CCR5

The chemokine receptor, CCR5, is a G protein coupled receptor responsible for some of the effects of the chemokines CCL3, CCL4 and CCL5. It is also one of the co-receptors for the entry of human immunodeficiency virus-1 (HIV-1) into cells. Regulation of CCR5 number on cells is, therefore, important for determining the infection rate by HIV-1.     

Exacerbation of facial motoneuron loss after facial nerve axotomy in CCR3-deficient mice

We have previously demonstrated a neuroprotective mechanism of FMN (facial motoneuron) survival after facial nerve axotomy that is dependent on CD4⁺ Th2 cell interaction with peripheral antigen-presenting cells, as well as CNS (central nervous system)-resident microglia. PACAP (pituitary adenylate cyclase-activating polypeptide) is expressed by injured FMN and increases Th2-associated chemokine expression in cultured murine microglia. Collectively, these results suggest a model involving CD4⁺ Th2 cell migration to the facial motor nucleus after injury via microglial expression of Th2-associated chemokines. However, to respond to Th2-associated chemokines, Th2 cells must express the appropriate Th2-associated chemokine receptors. In the present study, we tested the hypothesis that Th2-associated chemokine receptors increase in the facial motor nucleus after facial nerve axotomy at timepoints consistent with significant T-cell infiltration. Microarray analysis of Th2-associated chemokine receptors was followed up with real-time PCR for CCR3, which indicated that facial nerve injury increases CCR3 mRNA levels in mouse facial motor nucleus. Unexpectedly, quantitative- and co-immunofluorescence revealed increased CCR3 expression localizing to FMN in the facial motor nucleus after facial nerve axotomy. Compared with WT (wild-type), a significant decrease in FMN survival 4 weeks after axotomy was observed in CCR3^−/− mice. Additionally, compared with WT, a significant decrease in FMN survival 4 weeks after axotomy was observed in Rag2^−/− (recombination activating gene-2-deficient) mice adoptively transferred CD4⁺ T-cells isolated from CCR3^−/− mice, but not in CCR3^−/− mice adoptively transferred CD4⁺ T-cells derived from WT mice. These results provide a basis for further investigation into the co-operation between CD4⁺ T-cell- and CCR3-mediated neuroprotection after FMN injury.     

CCR2 Regulates the Uptake of Bone Marrow-Derived Fibroblasts in Renal Fibrosis

Recent studies have shown that bone marrow-derived fibroblasts contribute significantly to the pathogenesis of renal fibrosis. However, the molecular mechanisms underlying the recruitment of bone marrow-derived fibroblasts into the kidney are incompletely understood. Bone marrow-derived fibroblasts express the chemokine receptor - CCR2. In this study, we tested the hypothesis that CCR2 participates in the recruitment of fibroblasts into the kidney during the development of renal fibrosis. Bone marrow-derived collagen-expressing GFP⁺ fibroblasts were detected in the obstructed kidneys of chimeric mice transplanted with donor bone marrow from collagen α1(I)-GFP reporter mice. These bone marrow-derived fibroblasts expressed PDGFR-β and CCR2. CCR2 knockout mice accumulated significantly fewer bone marrow-derived fibroblast precursors expressing the hematopoietic marker-CD45 and the mesenchymal markers-PDGFR-β or procollagen I in the obstructed kidneys compared with wild-type mice. Furthermore, CCR2 knockout mice displayed fewer bone marrow-derived myofibroblasts and expressed less α-SMA or FSP-1 in the obstructed kidneys compared with wild-type mice. Consistent with these findings, genetic deletion of CCR2 inhibited total collagen deposition and suppressed expression of collagen I and fibronectin. Moreover, genetic deletion of CCR2 inhibits MCP-1 and CXCL16 gene expression associated with a reduction of inflammatory cytokine expression and macrophage infiltration, suggesting a linear interaction between two chemokines/ligand receptors in tubular epithelial cells. Taken together, our results demonstrate that CCR2 signaling plays an important role in the pathogenesis of renal fibrosis through regulation of bone marrow-derived fibroblasts. These data suggest that inhibition of CCR2 signaling could constitute a novel therapeutic approach for fibrotic kidney disease.     

CCL2/CCR2 Regulates the Tumor Microenvironment in HER-2/neu-Driven Mammary Carcinomas in Mice

Chronic inflammation is a hallmark of cancer. Inflammatory chemokines, such as C-C chemokine ligand 2 (CCL2), are often present in tumors but their roles in cancer initiation and maintenance are not clear. Here we report that CCL2 promotes mammary carcinoma development in a clinically relevant murine model of breast cancer. Targeted disruption of Ccl2 slowed the growth of activated Her2/neu-driven mammary tumors and prolonged host survival. Disruption of Ccl2 was associated with a decrease in the development and mobilization of endothelial precursor cells (EPCs) which can contribute to tumor neovascularization. In contrast, disruption of Ccr2, which encodes CCL2’s sole signaling receptor, accelerated tumor development, shortened host survival, and mobilized EPCs. However, pharmacological inhibition of CCR2 phenocopied Ccl2 disruption rather than Ccr2 disruption, suggesting that the Ccr2^-/- phenotype is a consequence of unanticipated alterations not linked to intact CCL2/CCR2 signaling. Consistent with this explanation, Ccr2^-/- monocytes are more divergent from wild type monocytes than Ccl2^-/- monocytes in their expression of genes involved in key developmental and functional pathways. Taken together, our data suggest a tumor-promoting role for CCL2 acting through CCR2 on the tumor microenvironment and support the targeting of this chemokine/receptor pair in breast cancer.