A myriad of functions and complex regulation of the CCR7/CCL19/CCL21 chemokine axis in the adaptive immune system

The chemokine receptor CCR7 and its ligands CCL19 and CCL21 control a diverse array of migratory events in adaptive immune function. Most prominently, CCR7 promotes homing of T cells and DCs to T cell areas of lymphoid tissues where T cell priming occurs. However, CCR7 and its ligands also contribute to a multitude of adaptive immune functions including thymocyte development, secondary lymphoid organogenesis, high affinity antibody responses, regulatory and memory T cell function, and lymphocyte egress from tissues. In this survey, we summarise the role of CCR7 in adaptive immunity and describe recent progress in understanding how this axis is regulated. In particular we highlight CCX-CKR, which scavenges both CCR7 ligands, and discuss its emerging significance in the immune system.     

靶向CCL21/CCR7轴治疗肿瘤转移的研究进展

转移是肿瘤患者死亡最常见的原因,而淋巴转移是大多数肿瘤转移的主要途径之一。近年来,CC趋化因子配体21 (CC chemokine ligand 21,CCL21)及其受体CC趋化因子受体7型(CC chemokine receptor type 7,CCR7)在淋巴转移中的作用逐渐受到关注。CCL21主要由淋巴内皮细胞产生,其与树突状细胞(Dendritic cells,DCs)和T细胞等表面CCR7的相互作用是免疫细胞淋巴迁移及淋巴结归巢的主要决定因素。然而,表达CCR7的肿瘤细胞也可以利用类似的机制进入淋巴管进行淋巴转移。如何靶向CCL21/CCR7轴,既能抑制淋巴转移,又不影响抗肿瘤免疫反应已成为肿瘤免疫治疗研究的重要议题。文中将对CCL21/CCR7轴在淋巴转移中的作用及其作为靶点治疗肿瘤转移的临床前和临床试验研究进行综述,为靶向CCL21/CCR7信号轴治疗肿瘤转移的相关研究提供参考。     

CCL21/CCR7轴与乳腺癌转移及治疗的相关研究进展

乳腺癌是发生在乳腺腺上皮组织的恶性肿瘤,是全世界女性死亡的最常见原因之一。趋化因子CCL21是一种具有趋化细胞迁移功能的低分子量蛋白质,CCL21可以与淋巴细胞表面表达的CCR7受体结合,促使其迁移至乳腺肿瘤相关部位;同时还能与乳腺肿瘤细胞上表达的CCR7配体结合,使其不断向正常部位转移和侵袭,从而导致癌细胞恶性发展。CCL21/CCR7轴与乳腺癌的治疗、转移、侵袭与预后有着密切联系。我们简要综述近年来CCL21/CCR7轴与乳腺癌的转移及治疗相关研究进展,以期为乳腺癌的临床治疗提供参考。     

The N-terminal domain of CCL21 reconstitutes high affinity binding, G protein activation, and chemotactic activity, to the C-terminal domain of CCL19

CC chemokine receptor 7 (CCR7), which regulates the trafficking of leucocytes to the secondary lymphoid organs, has two endogenous chemokine ligands: CCL19 and CCL21. Although both ligands possess similar affinities for the receptor and similar abilities to promote G protein activation and chemotaxis, they share only 25% sequence identity. Here, we show that substituting N-terminal six amino acids of CCL21 (SDGGAQ) for the corresponding N-terminal domain of CCL19 (GTNDAE) results in a chimeric chemokine that exhibits high affinity binding and G protein activation of CCR7. These data demonstrate that despite dissimilar sequences, the amino terminal hexapeptide of these two chemokines is capable of performing similar roles resulting in receptor activation.     

Paclitaxel treatment enhances lymphatic metastasis of B16F10 melanoma cells via CCL21/CCR7 axis

Chemotherapeutic drugs have been successfully used to treat several cancers, including melanoma. However, metastasis occasionally occurs after chemotherapy. Here, we reported that paclitaxel (PTX) treatment for B16F10 tumour in mice led to an enhanced lymphatic metastasis of the melanoma cells, although a significant inhibition of tumour growth at the injection site was observed. Further study demonstrated that PTX upregulated the expression of C-C chemokine receptor type 7 (CCR7) in B16F10 cells, enhancing their migration through the activation of JNK and p38 signalling pathways. Loss of CCR7 or blockade of C-C motif chemokine ligand 21 (CCL21)/CCR7 axis abolished the pro-migration effect of PTX on B16F10 melanoma cells. Importantly, combination of PTX and CCR7 mAb could simultaneously delay the tumour growth and reduce the lymphatic metastasis in B16F10 melanoma. The blockade of CCL21/CCR7 axis may collectively serve as a strategy for lymphatic metastasis in some melanoma after chemotherapy.     

C端截短突变体mCCL21-a(1-57)可拮抗CCL21介导的ERK磷酸化及细胞迁移

趋化因子CCL21(CC-chemokine ligand 21,CCL21)与其受体CCR7(CC-chemokine receptor 7,CCR7)的结合可以促进肿瘤的侵袭和转移.本研究旨在构建人趋化因子CCL21的截短突变体,竞争性抑制CCL21与CCR7的结合,从而抑制肿瘤的转移.本研究构建了CCL21的10...     

Matrix metalloproteinase-9 is up-regulated by CCL19/CCR7 interaction via PI3K/Akt pathway and is involved in CCL19-driven BMSCs migration

C–C chemokine receptor 7 (CCR7) and its ligands CCL19 contributes to the directional migration of certain cancer cell lines, but its role in the migration of BMSCs remains vague. The aim of this study was to determine the possible interaction between CCL19-induced conditions and matrix metalloproteinases-9 (MMP9) expression in BMSCs. Cell migration using Transwell assay indicated that activation of CCR7 by its specific ligand, exogenous chemokine ligand 19 (CCL19), was associated with a significant linear increase. Western blot and real-time PCR indicated that CCL19/CCR7 significantly upregulated expression of MMP9, which is related to metastasis-associated genes. The CCL19/CCR7 interaction significantly enhanced phosphorylation of Akt, as measured by Western blot. P-Akt and MMP9 protein expression exhibited a time-dependent pattern, and the peak was at 48 h. LY294002 significantly abolished the effects of exogenous CCL19. These results suggest that CCL19/CCR7 contributes to the migration of BMSCs by upregulating MMP9 potentially via the PI3K/Akt pathway.     

Angiogenic activity of human CC chemokine CCL15 in vitro and in vivo

CCL15 is a novel human CC chemokine and exerts its biological activities on immune cells through CCR1 and CCR3. Because a number of chemokines induce angiogenesis and endothelial cells express CCR1 and CCR3, we investigated the angiogenic activity of CCL15. Both CCL15(1-92) and N-terminal truncated CCL15(25-92) stimulate the chemotactic endothelial cell migration and differentiation, but CCL15(25-92) is at least 100-fold more potent than CCL15(1-92). Treatment with pertussis toxin (PTX), with anti-CCR1, or with anti-CCR3 antibody inhibits the CCL15(25-92)-induced endothelial cell migration. CCL15(25-92) also stimulates sprouting of vessels from aortic rings and mediates angiogenesis in the chick chorioallantoic membrane assay. Our findings demonstrate that CCL15(25-92) has in vitro and in vivo angiogenic activity, and suggest roles of the chemokine in angiogenesis.     

CCR9 and CCL25: A review of their roles in tumor promotion

Chemokines constitute a superfamily of small chemotactic cytokines with functions that are based on interactions with their corresponding receptors. It has been found that, among other functions, chemokines regulate the migratory and invasive abilities of cancer cells. Multiple studies have confirmed that chemokine receptor 9 (CCR9) and its exclusive ligand, chemokine 25 (CCL25), are overexpressed in a variety of malignant tumors and are closely associated with tumor proliferation, apoptosis, invasion, migration and drug resistance. This review evaluates recent advances in understanding the role of CCR9/CCL25 in cancer development. First, we outline the general background of chemokines in cancer and the structure and function of CCR9 and CCL25. Next, we describe the basic function of CCR9/CCL25 in the cancer process. Then, we introduce the role of CCR9/CCL25 and related signaling pathways in various cancers. Finally, future research directions are proposed. In general, this paper is intended to serve as a comprehensive repository of information on this topic and is expected to contribute to the design of other research projects and future efforts to develop treatment strategies for ameliorating the effects of CCR9/CCL25 in cancer.     

小鼠CCR7基因重组腺病毒的构建与鉴定

目的:构建含有小鼠趋化因子受体-7(CCR7)基因的重组腺病毒,为下一步转染不成熟树突状细胞(imDC)诱导免疫耐受研究奠定基础。方法:提取小鼠胸腺总RNA,应用逆转录PCR(RT-PCR)方法,以自行设计的带有酶切位点的引物,扩增获得CCR7基因全部序列,经过T-A克隆,酶切亚克隆到穿梭质粒pAdTrack-CMV上,在BJ5183菌内和pAdeasy-1同源重组,筛选阳性克隆,酶切鉴定,线性化后脂质体法转染HEK293细胞进行包装、PCR鉴定及扩增,得到含有CCR7基因的重组腺病毒,根据报告基因GFP测定病毒滴度。结果:成功构建小鼠CCR7基因重组腺病毒,病毒滴度为1×10~9U/mL。结论:该重组腺病毒载体的成功构建,为进一步研究携带CCR7基因的imDC的趋化迁移性等研究提供了一定的工作基础。     

The pathological effects of CCR2+ inflammatory monocytes are amplified by an IFNAR1-triggered chemokine feedback loop in highly pathogenic influenza infection

Background

Highly pathogenic influenza viruses cause high levels of morbidity, including excessive infiltration of leukocytes into the lungs, high viral loads and a cytokine storm. However, the details of how these pathological features unfold in severe influenza infections remain unclear. Accumulation of Gr1 + CD11b + myeloid cells has been observed in highly pathogenic influenza infections but it is not clear how and why they accumulate in the severely inflamed lung. In this study, we selected this cell population as a target to investigate the extreme inflammatory response during severe influenza infection.

Results

We established H1N1 IAV-infected mouse models using three viruses of varying pathogenicity and noted the accumulation of a defined Gr1 + CD11b + myeloid population correlating with the pathogenicity. Herein, we reported that CCR2+ inflammatory monocytes are the major cell compartments in this population. Of note, impaired clearance of the high pathogenicity virus prolonged IFN expression, leading to CCR2+ inflammatory monocytes amplifying their own recruitment via an interferon-α/β receptor 1 (IFNAR1)-triggered chemokine loop. Blockage of IFNAR1-triggered signaling or inhibition of viral replication by Oseltamivir significantly suppresses the expression of CCR2 ligands and reduced the influx of CCR2+ inflammatory monocytes. Furthermore, trafficking of CCR2+ inflammatory monocytes from the bone marrow to the lung was evidenced by a CCR2-dependent chemotaxis. Importantly, leukocyte infiltration, cytokine storm and expression of iNOS were significantly reduced in CCR2−/− mice lacking infiltrating CCR2+ inflammatory monocytes, enhancing the survival of the infected mice.

Conclusions

Our results indicated that uncontrolled viral replication leads to excessive production of inflammatory innate immune responses by accumulating CCR2+ inflammatory monocytes, which contribute to the fatal outcomes of high pathogenicity virus infections.     

Effect of anti-TNFalpha treatment on circulating endothelial progenitor cells (EPCs) in rheumatoid arthritis

Rheumatoid arthritis (RA) is associated with increased cardiovascular morbidity and mortality, which may be attenuated by anti-inflammatory treatment. Endothelial progenitor cells (EPCs) have the ability to differentiate into mature endothelium and have a potentially reparative role protecting against ischemia and atherosclerosis. OBJECTIVE: To investigate the effect of treatment with infliximab on the number and functional capacity of endothelial progenitor cells (EPCs) in patients with RA, as a possible mechanism for reducing cardiovascular morbidity in this disorder. METHODS: Patients: Active seropositive RA patients (N = 14) considered candidates for starting infliximab treatment, were recruited. Assessment, based on DAS-28, was performed before treatment and 14 days later. Peripheral blood mononuclear cells were isolated and EPC numbers evaluated by the colony-forming unit (CFU) method. Endothelial phenotyping of CFU was performed by immunofluorescence employing antibodies to Tie-2 VEGF-receptor 2, and CD31. EPC Functional properties were evaluated by fibronectin adherance. RESULTS: A significant 33.4% increase (p < 0.001) in EPC levels was observed after infliximab. A 60% increase was noted in the EPC differentiation scale, (p < 0.002) while a 37.6% increase was observed in mean EPC adhesion (p < 0.001). These changes were associated with a 17.5% decrease in the DAS-28 (p < 0.0001). A significant correlation was observed between the clinical response, reflected by changes in DAS-28 and the degree of increase in EPC CFUs. CONCLUSION: A single dose of infliximab improved the number and functional properties of EPCs, in parallel with an early clinical effect, suggesting a possible mechanism by which anti-inflammatory treatment may reduce cardiovascular risk in RA patients.     

Therapeutic effects of isothiocyanate prodrugs on rheumatoid arthritis

Isothiocyanates 7a and 7b have poor stability and aqueous solubility. To address these problems, prodrugs 8a and 8b were synthesized. Prodrugs 8a and 8b were stable in HEPES buffer at pH 4.4, but released the active compounds 7a and 7b in HEPES buffer at pH 7.4 and in mouse plasma, respectively. Compound 8a and especially compound 8b showed anti-inflammatory effects. Compound 8b demonstrated significant efficacy in animal models of traumatic inflammation, acute inflammation and rheumatoid arthritis. Compound 8b also did not cause appreciable toxicity in mice after 5?weeks at a daily dose of 200?mg/kg.     

CCL5/CCR5生物学轴在肿瘤中作用的研究进展

肿瘤因其易转移、易复发的特性成为一大难以治愈的疾病,已严重威胁到人类的生命健康。肿瘤微环境在肿瘤的生长、迁移、免疫逃逸、血管生成等过程中具有明显的促进作用。肿瘤微环境中细胞分泌的CCL5发挥的作用已受到越来越多的关注,且许多研究表明抑制CCL5/CCR5生物学轴可抑制肿瘤迁移、血管生成等,预示着这可能成为一个新的肿瘤治疗策略。本文总结了近年来关于CCL5/CCR5生物学轴的研究,包括CCL5/CCR5生物学轴介导的肿瘤生长迁移、血管生成、免疫逃逸等作用,及CCR5抑制剂在肿瘤治疗中的广阔前景。     

CCR9和CCL25蛋白在上皮性卵巢癌中表达及临床意义

目的:探讨CCR9和CCL25蛋白在不同卵巢组织中的表达及其与上皮性卵巢癌患者临床病理因素之间的关系。方法:通过组织芯片结合免疫组织化学法检测78例上皮性卵巢癌组织和30例正常卵巢组织中CCR9和CCL25表达水平,结合上皮性卵巢癌病人的临床病理资料,进行统计分析。结果:CCR9和CCL25在上皮性卵巢癌中高表达,在正常卵巢组织中低表达,二者的表达与上皮性卵巢癌的组织类型、患者年龄无显著相关(P0.05),而与淋巴结转移、组织学分级和临床分期有显著相关(P0.05);上皮性卵巢癌组织中CCR9与CCL25表达相关(P0.05)。结论:CCR9和CCL25在上皮性卵巢癌的发生发展中可能起重要作用,二者可能是上皮性卵巢癌治疗的一个潜在的分子靶点。     

Molecular targeting of hepatocyte growth factor by an antagonist,NK4, in the treatment of rheumatoid arthritis

Introduction

Hepatocyte growth factor (HGF) is a potent proangiogenic molecule that induces neovascularization. The HGF antagonist, NK4, competitively antagonizes HGF binding to its receptor. In the present study, we determined the inhibitory effect of NK4 in a rheumatoid arthritis (RA) model using SKG mice.

Methods

Arthritis was induced in SKG mice by a single intraperitoneal injection of β-glucan. Recombinant adenovirus containing NK4 cDNA (AdCMV.NK4) was also injected intravenously at the time of or 1 month after β-glucan injection. Ankle bone destruction was examined radiographically. The histopathologic features of joints were examined using hematoxylin and eosin and immunohistochemical staining. Enzyme-linked immunosorbent assays were used to determine the serum levels of HGF, interferon γ (IFN-γ, interleukin 4 (IL-4) and IL-17 production by CD4⁺ T cells stimulated with allogeneic spleen cells.

Results

The intravenous injection of AdCMV.NK4 into SKG mice suppressed the progression of β-glucan-induced arthritis. Bone destruction was also inhibited by NK4 treatment. The histopathologic findings of the ankles revealed that angiogenesis, inflammatory cytokines and RANKL expression in synovial tissues were significantly inhibited by NK4 treatment. Recombinant NK4 (rNK4) proteins inhibited IFN-γ, IL-4 and IL-17 production by CD4⁺ T cells stimulated with allogeneic spleen cells.

Conclusions

These results indicate that NK4 inhibits arthritis by inhibition of angiogenesis and inflammatory cytokine production by CD4⁺ T cells. Therefore, molecular targeting of angiogenic inducers by NK4 can potentially be used as a novel therapeutic approach for the treatment of RA.     

The discovery of BMS-457, a potent and selective CCR1 antagonist

A series of compounds which exhibited good human CCR1 binding and functional potency was modified resulting in the discovery of a novel series of high affinity, functionally potent antagonists of the CCR1 receptor. Issues of PXR activity, ion-channel potency, and poor metabolic stability were addressed by the addition of a hydroxyl group to an otherwise lipophilic area in the molecule resulting in the discovery of preclinical candidate BMS-457 for the treatment of rheumatoid arthritis.     

Chemokine CCL20 enhances the growth of HuH7 cells via phosphorylation of p44/42 MAPK in vitro

CCR6 is the receptor of chemokine CCL20. In the present study, we demonstrated that the surface expression of CCR6 was enhanced on the human HCC cell lines (HuH7, PLC/PRF/5, and HepG2) especially on HuH7 cells, but not on HLE or HLF cells. These HCC cell lines (HuH7, PLC/PRF/5, and HepG2) especially the HuH7 cells secreted a significant amount of CCL20 spontaneously, whereas HLE or HLF did not. Stimulation by CCL20 up-regulated the mRNA expression of CCR6 in HuH7 cells and significantly enhanced the growth of HuH7 cells. CCL20-stimulated growth of HuH7 cells was abrogated by the inhibition of downstream signal transduction pathway mediated by p44/42 MAPK, but not by p38 MAPK or SAPK/JNK. CCR6 expression in human HCC tissues was confirmed by RT-PCR. These results indicate that the growth of a proportion of human HCC cells may be mediated by CCL20-CCR6 axis, like HuH7 cells, in an autocrine or paracrine manner.     

CCR7和B7-2在抗原负载树突状细胞诱导特异性CTL抗肿瘤效应中的表达和意义

目的:研究CCR7(趋化因子受体7)和B7-2(白细胞分化抗原86)与抗原负载树突状细胞(dentritic cell,DC)诱导特异性CTL(细胞毒性T淋巴细胞)抗肿瘤效应的关系.方法:分离和培养DC,制备B16黑色素瘤细胞抗原,进行共培养,即为抗原负载的DC,建立B16黑色素瘤小鼠模型,于肿瘤周围皮下注射抗原负载的DC.应用原位杂交和免疫组织化学方法检测CCR7和B7-2的表达情况.结果:原位杂交和免疫组织化学染色显示,CCR7和B7-2阳性细胞主要分布于肿瘤周围组织,随着注射抗原负载DC时间的进展,CCR7和B7-2呈强阳性表达.结论:CCR7和B7-2的表达与抗原负载树突状细胞诱导特异性CTL抗肿瘤效应有关.