The C-C Chemokines CCL17 and CCL22 and Their Receptor CCR4 in CNS Autoimmunity.

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS). It affects more than two million people worldwide, mainly young adults, and may lead to progressive neurological disability. Chemokines and their receptors have been shown to play critical roles in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), a murine disease model induced by active immunization with myelin proteins or transfer of encephalitogenic CD4⁺ T cells that recapitulates clinical and neuropathological features of MS. Chemokine ligand-receptor interactions orchestrate leukocyte trafficking and influence multiple pathophysiological cellular processes, including antigen presentation and cytokine production by dendritic cells (DCs). The C-C class chemokines 17 (CCL17) and 22 (CCL22) and their C-C chemokine receptor 4 (CCR4) have been shown to play an important role in homeostasis and inflammatory responses. Here, we provide an overview of the involvement of CCR4 and its ligands in CNS autoimmunity. We review key clinical studies of MS together with experimental studies in animals that have demonstrated functional roles of CCR4, CCL17, and CCL22 in EAE pathogenesis. Finally, we discuss the therapeutic potential of newly developed CCR4 antagonists and a humanized anti-CCR4 antibody for treatment of MS.

Macrophage Subset Expressing CD169 in Peritoneal Cavity-Regulated Mucosal Inflammation Together with Lower Levels of CCL22.

Crohn's disease (CD) and ulcerative colitis (UC) are the most widely known types of inflammatory bowel diseases (IBD) and have been paid more attention due to their increasing incidence and a substantial increase in the risk of colorectal cancer (CRC). However, the phenotype and, more importantly, the function in the regulation of mucosal inflammation by different macrophages are poorly understood, even though macrophages constitute a major subset of intestinal myeloid cells. The results firstly showed that the subset of peritoneal CD11b+CD169+ macrophages increased and CCL22 expression level decreased significantly during the DSS-induced colitis. DSS-induced colitis was alleviated in CD169-DTR mice at least partially due to the deletion CD169+ macrophages. Moreover, the CCL22 expression level in peritoneal macrophages from CD169-DTR mice was much higher than that from WT mice with DSS-induced colitis. And, the cell-sorting result revealed that CD11b+CD169+ macrophage cells did not express CCL22 dominantly. Further experiment in vivo demonstrated that treatment with recombinant murine CCL22 (rmCCL22) ameliorated the clinical symptoms of DSS-induced colitis. All these data indicated that macrophage subset of CD11b+CD169+ from peritoneal cavity played critical role probably together with low levels of CCL22 in DSS-induced colitis.

Functional reprogramming of human prostate cancer to promote local attraction of effector CD8(+) T cells.

BACKGROUND: Local infiltration of CD8(+) T cells (CTLs) in tumor lesions predicts overall clinical outcomes and the clinical benefit of cancer patients from immune checkpoint blockade. In the current study, we evaluated local production of different classes of chemokines in prostate cancer lesions, and the feasibility of their modulation to promote selective entry of CTLs into prostate tumors. METHODS: Chemokine expression in prostate cancer lesion was analyzed by TaqMan-based quantitative PCR, confocal fluorescence microscopy and ELISA. For ex vivo chemokine modulation analysis, prostate tumor explants from patients undergoing primary prostate cancer resections were cultured for 24 hr, in the absence or presence of the combination of poly-I:C, IFNα, and celecoxib (PAC). The numbers of cells producing defined chemokines in the tissues were analyzed by confocal microscopy. Chemotaxis of effector CD8(+) T cells towards the untreated and PAC-treated tumor explant supernatants were evaluated in a standard in vitro migration assays, using 24 well trans-well plates. The number of effector cells that migrated was enumerated by flow cytometry. Pearson (r) correlation was used for analyzing correlations between chemokines and immune filtrate, while paired two tailed students t-test was used for comparison between treatment groups. RESULTS: Prostate tumors showed uniformly low levels of CTL/NK/Th1-recruiting chemokines (CCL5, CXCL9, CXCL10) but expressed high levels of chemokines implicated in the attraction of myeloid derived suppressor cells (MDSC) and regulatory T cells (Treg ): CCL2, CCL22, and CXCL12. Strong positive correlations were observed between CXCL9 and CXCL10 and local CD8 expression. Tumor expression levels of CCL2, CCL22, and CXCL12 were correlated with intratumoral expression of MDSC/Treg markers: FOXP3, CD33, and NCF2. Treatment with PAC suppressed intratumoral production of the Treg -attractant CCL22 and Treg /MDSC-attractant, CXCL12, while increasing the production of the CTL attractant, CXCL10. These changes in local chemokine production were accompanied by the reduced ability of the ex vivo-treated tumors to attract CD4(+) FOXP3(+) Treg cells, and strongly enhanced attraction of the CD8(+) Granzyme B(+) CTLs. CONCLUSIONS: Our data demonstrate that the chemokine environment in prostate cancer can be reprogrammed to selectively enhance the attraction of type-1 effector immune cells and reduce local attraction of MDSCs and Tregs . Prostate 76:1095-1105, 2016. © 2016 Wiley Periodicals, Inc.

Increased production of IL-5 and dominant Th2-type response in airways of Churg-Strauss syndrome patients.

OBJECTIVE: Churg-Strauss syndrome (CSS) is a rare systemic vasculitis associated with eosinophilia and asthma. We assessed the local immune response in airways of CSS patients with different activity of the disease. METHODS: Concentration of IL-5, CCL17, CCL22 and CCL26 (ELISA) together with cell expression of T-helper-related genes (real-time PCR array) were measured in bronchoalveolar lavage fluid (BALF) sampled from 11 patients with active CSS, 11 patients with CSS in remission and 9 control subjects with bronchial asthma. RESULTS: In active CSS, both BALF and blood eosinophil counts were increased (P<0.01). BALF cells in active disease were characterized by an increased expression of Th2 and regulatory-type transcripts: STAT6, STAT3, GATA3, IL4, IL5 and IL10 as compared with asthmatics, and STAT5A, CCR4, FOXP3, IL4, IL5 and IL10 when compared with inactive CSS. There was significant increase in BALF concentration of IL-5 and CCL26 in exacerbation of CSS. CCR4-active chemokines were detected more frequently in active disease. We found a strong positive correlation between clinical parameters of disease activity (BVAS, eosinophilia) and expression of IL4, IL5, IL10 and STAT5A. CONCLUSION: These results indicate that as compared with asthma, active-CSS patients have much stronger local Th2 response in the airways. Airway cells may contribute to lung eosinophilia in CSS by producing IL-5 and eosinophil active chemokines.

Increased frequencies of CD4+ CD25+ FOXP3+ regulatory T cells in human nasal inverted papilloma.

BACKGROUND: The purpose of this study was to investigate the presence of CD4(+) CD25(+) FOXP3(+) regulatory T (Treg) cells both in peripheral blood and local tumors in patients with nasal inverted papilloma (NIP). METHODS: By using flow cytometry, the frequencies of CD4(+) CD25(+) FOXP3(+) Treg cells in both peripheral blood and tissues from 18 patients with NIP and 8 control subjects were determined. CCL22 and CCL17 proteins in NIP tumors were analyzed by using enzyme-linked immunosorbent assay. The suppressive capacity of Treg cells was estimated by WST-8 and enzyme-linked immunosorbent assay (ELISA; interferon-gamma [IFN-γ] and interleukin-4 [IL-4]) analysis. RESULTS: Patients with NIP showed increased CD4(+) CD25(+) FOXP3(+) Treg cell frequencies in tumor tissues and CD4(+) T cell fraction rather than in peripheral blood. CCL22 increased in NIP tumors. Phytohemagglutinin (PHA)-induced proliferation and cytokine production of CD4(+) CD25(-) T cells were suppressed equally well by CD4(+) CD25(high) cells from both patients with NIP and controls. CONCLUSION: Our study demonstrated the increased frequencies of CD4(+) CD25(+) FOXP3(+) Treg cells in NIP tumors, which might be influenced by CCL22.

Cytokine analysis as a tool to understand tumour-host interaction in ovarian cancer.

Epithelial ovarian cancer (EOC) is an immunogenic tumour and exploits many suppressive ways to escape immune eradication. EOC is known to spread primarily by tumour cell implantations in peritoneal cavity. Therefore, ascites may be an ideal fluid compartment to unravel the immune status of the peritoneal cavity. We analysed the expression of IL-1ß, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IFN-γ, TNF-α, TNF-ß, TGF-ß and CCL22 in ovarian cancer ascites, representing immune activating and suppressing cytokines. We observed high expression of pro-inflammatory cytokines IL-6, IL-8 and immune suppressive cytokines IL-10, CCL22 and TGF-ß in most samples whereas Th1 (IL-12p70, IFN-γ) and Th2 (IL-4, IL-5) cytokines were only detectable in 13% of the samples. TGF-ß was only detected in latent form, questioning its immune suppressive role. CCL22 was in similar levels present in early stage compared to advanced stage tumours. At advanced stage, we observed a negative correlation with CCL22 levels and Th1/2 cytokine expression. We found a positive correlation between IL-6 concentration in ascites and residual disease after debulking. Additionally, IL-6 levels were remarkably higher at recurrence compared to primary advanced disease, which opens an opportunity for inhibition of IL-6 expression in the prevention of recurrence. Despite the heterogeneity of EOC and the complexity of cytokine functions, our results show that cytokine analysis in ascites may aid in understanding tumour-host interaction in EOC.

Inhibitory effects of heartwood extracts of Broussonetia kazinoki Sieb on the development of atopic dermatitis in NC/Nga mice.

We investigated the effects of a topically applied extract of the heartwood of Broussonetia kazinoki Sieb (B. kazinoki) on atopic dermatitis (AD)-like skin lesions induced by an extract of the house-dust mite Dermatophagoides farina in NC/Nga mice. We found that topically applied B. kazinoki extract suppressed the histological manifestations of AD-like skin lesions, and decreased the levels of plasma immunoglobulin E (IgE) and interleukin-4 (IL-4) in the mice. Moreover, B. kazinoki inhibited the induction of thymus-and-activation-regulated chemokine (TARC/CCL17), macrophage-derived chemokine (MDC/CCL22), and regulated-on-activation-normal T cell-expressed-and-secreted chemokine (RANTES/CCL5) in HaCaT cells activated by tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). In conclusion, our results suggest that B. kazinoki extract has therapeutic advantages in the treatment of AD.

Permanent up-regulation of regulatory T-lymphocytes in patients with head and neck cancer.

Various immune functions of different types of immune cells are strongly impaired in patients with head and neck squamous cell carcinoma (HNSCC). Regulatory T-lymphocyte cells (Tregs) have been suggested to be involved in the immunomodulation of immune responses and contribute to HNSCC progression and immune escape. 'Naturally' occurring CD4+ CD25+ Tregs represent a small fraction within the different subsets of regulatory T cells, which are known to inhibit numerous immune functions of different types of immune cells. In this study, the cellular ratio of CD4+ CD25(high) Tregs to the entire population of CD4+ T-lymphocytes was analyzed with respect to different stages of tumor progression and disease. Our data indicate a significantly high increased abundance of CD4+ CD25(high) CD127(low) Tregs in the peripheral blood of patients with HNSCC, which in addition show modulated expression levels of various functional proteins. Surprisingly, increased Treg levels were found even in patients with no active disease several years after tumor resection, with no significant correlation to the individual tumor stage. Additionally, increased levels of chemokine CCL22, which mediates migration of Tregs to the tumor, and upregulation of the corresponding receptor protein CCR4 were observed in HNSCC. Our data strongly suggest that HNSCC leads to a permanent shift of Treg levels with hardly recognizable recovery rates.

Morphometric analysis of the balance between CXCR3+ T cells and FOXP3+ regulatory T cells in lymphocyte-rich and conventional gastric cancers.

Lymphocyte-rich gastric carcinomas (Ly-rich GCs) are characterized by the formation of lymphoid stroma. Our previous study has shown abundant infiltration of CXCR3(+) T cells and their frequent clustering with CXCL9 (monokine induced by interferon-gamma)-expressing stromal cells in the lymphoid stroma of Ly-rich GCs. Foxp3(+) regulatory T cells (Tregs) suppress immune responses in the peripheral tissues and play a role in the immunosuppression of cancer tissues. The present study was therefore undertaken to evaluate the significance of the balance between CXCR3(+) T cells and Tregs in 44 Ly-rich and 37 conventional GCs by morphometrical analyses of immunohistochemistry. Compared with the pronounced infiltration of CXCR3(+) T cells in the lymphoid stroma, the numbers of Foxp3(+) cells were relatively low in Ly-rich GCs. Therefore, the ratios of CXCR3(+)/Foxp3(+) cells were much higher in Ly-rich GCs than in conventional GCs. This suggests the occurrence of T-helper type 1 (Th1)-shifted immune responses in Ly-rich GCs. On the other hand, conventional GCs were characterized by a paucity of CXCR3(+) T cells and a relative abundance of Tregs. Furthermore, the stroma inside the cancer was characterized by even less CXCR3(+) cells, suggesting a strongly immunosuppressive microenvironment. Since Tregs are known to express CCR4, we also examined the tissue distribution of cells expressing its ligand CCL22. CCL22 was not detected in conventional GCs and only sparsely detected in dendritic cells but not in cancer cells in Ly-rich GCs. To conclude, Tregs may play a more important role in conventional GCs than in Ly-rich GCs from the viewpoint of immunosuppression.