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Background: Cancer patients with hepatitis B virus (HBV) and hepatitis C virus (HCV) infections are at high risk of viral reactivation after cancer treatment. However, there is a paucity of data regarding HBV or HCV reactivation in cancer patients who receive newer anticancer drugs such as immune checkpoint inhibitors; Bruton tyrosine kinase (BTK) inhibitors; agents targeting CD22, CD38, and CC chemokine receptor 4 (CCR4); and chimeric antigen receptor (CAR) T-cell therapies., Objectives: In this narrative review article, we describe the rate, characteristics, and outcomes of HBV and HCV reactivation in patients receiving novel systemic anticancer therapies., Sources: We searched MEDLINE for all original research articles, case reports, and systematic reviews published in English between July 2013 and December 2021 on cancer patients with HBV or HCV infection receiving novel systemic anticancer therapy., Content: The risk of HBV or HCV reactivation is not well defined in cancer patients receiving immune checkpoint inhibitors (durvalumab, atezolizumab, nivolumab, pembrolizumab, ipilimumab, and tremelimumab); BTK inhibitors (ibrutinib and acalabrutinib); agents targeting CD22 (inotuzumab ozogamicin), CD38 (daratumumab, isatuximab), and CCR4 (mogamulizumab); and CAR T-cell therapy (axicabtagene-ciloleucel). However, screening for chronic HBV and HCV infections and routine monitoring of patients with such infections during novel anticancer therapy are recommended for early identification of viral reactivation, which can impact outcomes of oncologic treatment or be fatal., Implications: Specific strategies for risk assessment, monitoring, and management should be designed to reduce the risk of reactivation after novel anticancer therapy in patients with chronic HBV or HCV infections., (Copyright © 2022 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

This study described the first impedimetric immunosensor reported for the determination of CCR4, a new prostate cancer biomarker. This impedimetric immunosensor was constructed through the modification of disposable indium tin oxide (ITO) sheet with a conjugated pyrrole polymer P(Pyr-Pac) and subsequent immobilization of anti-CC chemokine receptor 4 (CCR4) antibodies. Acid-substituted poly(pyrrole) P(Pyr-Pac) polymer contained a lot of carboxyl groups on its end site, which were suitable for attachment of anti-CCR4 antibodies. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were chosen to investigate electrode preparation stages and, EIS was chosen to detect the CCR4 concentration. Anti-CCR4 antibody attached biosensing surface was highly selective to CCR4 antigen, the specific interaction resulted changes in electrochemical signal. Optimization studies containing polymer amount, anti-CCR4 antibody concentration, anti-CCR4 antibody immobilization time and anti-CCR4 antibody-CCR4 antigen interaction time were studied. The developed immunosensor displayed a linear increase with concentrations of CCR4 antigen (0.02-8 pg/mL) and a low detection limit of 6.4 fg/mL. In addition, this biosensor had great reproducibility and repeatability. Moreover, the designed biosensor was successfully used for the quantification of CCR4 antigen in serum samples. The recovery for the spiked serum samples was between 98.25% and 103.99%. The suggested immunosensor illustrated a good selectivity towards some interferents including different biomarkers. This study could establish a new approach for future cancer biomarker detection., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Bronchial asthma is characterized by allergic airway inflammation involving C-C chemokine receptor type 4 (CCR4)-positive Th2 cells. As such, we hypothesize that the disease can be alleviated by targeted-elimination of CCR4⁺ cells. Thymus and activation-regulated chemokine (TARC)-PE38, a TARC fused the exotoxin fragment PE38 from Pseudomonas aeruginosa, has been shown to efficiently kill CCR4⁺ cells by delivering the exotoxin fragment PE38 into CCR4⁺ cells. To test our hypothesis, we examined whether TARC-PE38 could suppress allergic airway inflammation in a mouse model of house dust mite (HDM)-induced allergic airway inflammation., Methods: We evaluated the effect of TARC-PE38 on the major characteristics of HDM-induced allergic airway inflammation. Airway hyperresponsiveness, lung histopathology, lung Th1/Th2 cell populations, and concentrations of Th1/Th2 cytokines in the lungs were assessed in HDM-sensitized and challenged mice in the presence and absence of TARC-PE38., Results: TARC-PE38 efficiently suppressed allergic airway inflammation by significantly reducing airway hyperresponsiveness, the overall area of inflammation, and goblet cell hyperplasia. In HDM-sensitized and challenged mice, TARC-PE38 specifically reduced the numbers of CCR4⁺ cells. This reduction was associated with a significant decrease in the production of Th2 cytokines in the airway,and a decrease in the number of leukocytes, including macrophages, eosinophils and lymphocytes, within the subepithelial area of the lungs and airway lumen. TARC-PE38 had noeffect on Th1 cells., Conclusion: Our data suggest that the elimination of CCR4⁺ cells via TARC-PE38 treatment is sufficient to control allergic airway inflammation and airway hyperresponsiveness., (Copyright © 2013. Published by Elsevier B.V.)

Little is known about the role of chemokines and their receptors interaction, which are essential for recruitment of selective lymphocyte subsets during inflammation, in the pathogenesis of idiopathic nonspecific interstitial pneumonia (NSIP). Recent studies have revealed Th1 and Th2 cells preferentially employ the chemokine receptors, CXCR3 and CCR4, respectively, in the process of accumulation into inflammatory sites. We evaluated the CXCR3 and CCR4 expression on infiltrated lymphocytes in lung tissues of 12 NSIP cases and 10 idiopathic pulmonary fibrosis (IPF) cases in our previous study. The number of CXCR3 positive lymphocytes of NSIP patients was significantly greater than that of IPF patients (261.1+/-145.1 vs. 64.9+/-27.0, P<0.01). The number of CCR4 positive lymphocytes of NSIP patients was significantly lower than that of IPF (9.5+/-8.3 vs.62.6+/-26.9, P<0.01). The CXCR3 to CCR4 ratio of NSIP patients was significantly greater than that of IPF patients (47.9+/-45.9 vs. 1.11+/-0.40, P<0.01). The differences of CXCR3 positive, CCR4 positive lymphocyte counts, and of CXCR3/CCR4 ratio between cellular and fibrosing NSIP were not significant. These results suggest that a Th1 pattern of chemokine receptor expression predominates in the lung interstitium of patients with NSIP but, in IPF patients, CCR4 might be relatively predominant, in contrast to the finding in NSIP patients, and that Th1/Th2 balance might be an important factor in the pathogenesis of NSIP.

Background: Fructooligosaccharides (FOS) in prebiotic foods can alter intestinal immune responses. The combination of probiotics with oligosaccharides has been reported to alter intestinal flora and suggested to be beneficial against food allergy in humans., Methods: All male Nc/jic mice used in this 8-week study were 6 weeks of age and were allotted to the following three groups: (1) the nonsensitization group; (2) the ovalbumin (OVA) sensitization +5% fructose-containing control food administration group; and (3) the OVA sensitization +5% FOS-containing food administration group. Duodenal tissues were collected and then immunohistochemically stained with monoclonal antibodies to CCR4 and CCR5. The number of mast cells and the villus edema formation rate in the duodenum were determined by image analysis., Results: The number of CCR4-positive cells increased significantly in Group 2 as compared with Group 1 and tended to decrease in Group 3 as compared with Group 2. Relatively few CCR5-positive cells were observed in the duodenum. FOS tended to reduce the number of CCR4-positive cells but significantly reduced the number of mast cells and the edema formation rate in the duodenum., Conclusions: This study demonstrated a correlation between the number of CCR4-positive cells and villus edema formation rate. Therefore, FOS, which we inferred to show antiallergic activity for food allergy in this study and which has already been established to be safe for use as food in humans, can be considered to be potentially useful for the prevention of food allergy in pediatric patients with allergy.

Regulatory T cells (Tregs) are crucial for the induction and maintenance of self-tolerance and are present in peripheral tissues such as skin and gut under normal, noninflamed conditions. We report isolation and expansion of the Treg population resident in normal human skin. Cutaneous Tregs expressed high levels of CD25, L-selectin, GITR, FOXP3, and intracellular CTLA-4, low levels of CD69, and high levels of the skin-homing addressins CLA, CCR4, and CCR6. Skin Tregs suppressed the proliferation of CD25(lo) T cells from the same skin sample in response to CD3 and CD28 antibodies. Suppression was dependent on cell contact and not affected by neutralizing antibodies to interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta). Surprisingly, cutaneous Tregs proliferated in an antigen-independent manner when cultured in contact with dermal fibroblasts and IL-15, conditions similar to those found in chronically inflamed skin. We hypothesize that local proliferation of Tregs may occur within inflamed skin and could serve as a brake for cutaneous inflammation as well as a mechanism for the homeostatic proliferation of natural Tregs that has been observed within intact organisms.

T cells resident in normal skin likely conduct immunosurveillance and are implicated in the development of inflammatory disorders such as psoriasis. This population of cells is difficult to study because existing techniques allow isolation of only few cells. We report here a novel method of isolating T cells from both normal and diseased human skin. Explants of skin cultured on three-dimensional matrices led to the outgrowth of dermal fibroblasts that elaborated T cell chemoattractant factors. These factors led to the migration of skin resident T cells out of skin explants where they could be collected and studied. Skin resident T cells isolated from explant cultures were CD45RO(+) memory T cells and expressed high levels of cutaneous lymphocyte antigen (CLA) and chemokine receptor (CCR)4. Inclusion of IL-2 and IL-15 in explant cultures produced up to a 10-fold expansion of skin-resident T cells, while maintaining the CLA(+)CCR4(+) skin-homing phenotype as well as a diverse T cell repertoire. This method also allowed efficient isolation of malignant T cells from the skin lesions of cutaneous T cell lymphoma and the isolation of tumor-infiltrating lymphocytes from primary squamous cell carcinomas and melanoma metastases.

Skin inflammation in atopic dermatitis (AD) is characterized by the predominant infiltration of T-helper (Th)2-cells in lesional skin. However, the mechanism of recruitment of these cells in lesional skin of AD is not yet fully elucidated. In this study, we investigated the role of IL-13-stimulated human primary keratinocytes (HPKs) in the recruitment of lymphocytes and further delineated the mechanism of enrichment of these cells. In the migration assays, we observed preferential enrichment of CD4(+)CCR4(+) T cells towards IL-13-stimulated HPKs. Interestingly, CD4(+)CCR4(+) T cells from AD showed a higher chemotactic response than those from healthy individuals. We observed a significant increase in the expression of CCL22 in IL-13-stimulated HPKs as compared to unstimulated cells. Blocking of CCL22 in IL-13-stimulated HPKs by a neutralizing antibody resulted in 70-90% inhibition in migration of CD4(+)CCR4(+) T cells. Moreover, IL-13 upregulated IFN-gamma-induced chemokines, CCL2 and CCL5, in HPKs. Taken together, our data suggest that IL-13-stimulated HPKs participate in a positive feedback loop by preferentially enriching Th2-cells in lesional skin of acute AD patients. However, in chronic phase, IL-13 may act in synergy with IFN-gamma resulting in lymphocytes recruitment of a mixed phenotype at the site of inflammation, thus contributing to the chronification of eczema.

Background: Ketoprofen is well known to evoke the allergic type of photocontact dermatitis when it is applied to the skin and irradiated with ultraviolet A (UVA) light., Objective: We aimed to establish a murine model of this photosensitivity and to characterize pathogenic T cells concerned with the sensitivity., Methods: Various strains of mice were sensitized on two consecutive days by application of ketoprofen to the shaved abdomen and irradiation of the skin with UVA. Five days later, they were elicited with ketoprofen plus UVA on the earlobes. Immune lymph node cells and epidermal cells from the challenged sites were analyzed by RT-PCR., Results: Mice were successfully sensitized and challenged with 4% and 2% ketoprofen, respective, plus UVA at 20J/cm2. The responses in H-2k mice were higher than those in the other strains examined. Immune lymph node CD4+ or CD8+ cells from ketoprofen-photosensitized H-2k mice were transferred i.v. to naïve syngeneic recipients. Mice receiving CD4+ but not CD8+ cells exhibited ketoprofen photosensitivity, but transference of both CD4+ and CD8+ cell populations was more effective. Lymph node cells from photosensitized mice expressed high levels of mRNA for Th2 cytokine (IL-4) and Th2 chemokine receptor (CCR4) as well as Th1 cytokine (IFN-gamma) and Th1 chemokine receptor (CXCR3), as assessed by RT-PCR. In addition, epidermal cells from challenged earlobes expressed increased levels of both Th1 (TARC) and Th2 (Mig) chemokines., Conclusion: It is considered that not only Th1 but also Th2 cells participate in the pathogenesis of murine photocontact dermatitis to ketoprofen.

Chemokine-like factor 1 (CKLF1) exhibits chemotactic effects on leukocytes. Its amino acid sequence shares similarity with those of TARC/CCL17 and MDC/CCL22, the cognate ligands for CCR4. The chemotactic effects of CKLF1 for CCR4-transfected cells could be desensitized by TARC/CCL17 and markedly inhibited by PTX. CKLF1 induced a calcium flux in CCR4-transfected cells and fully desensitized a subsequent response to TARC/CCL17, and TARC/CCL17 could partly desensitize the response to CKLF1. CKLF1 caused significant receptor internalization in pCCR4-EGFP transfected cells. Taken together, CKLF1 is a novel functional ligand for CCR4.

Skin-infiltrating T lymphocytes are thought to play a major role in the pathogenesis of cutaneous lupus erythematosus (CLE). In this study, we investigated the role of the chemokine receptor 4 (CCR4) and its ligand thymus- and activation-regulated chemokine (TARC/CCL17) for the recruitment of T cells in inflamed skin of patients with CLE. We found significant numbers of CCR4+ T lymphocytes in the skin of all patients with CLE. Interestingly, a subset of patients with disseminated scarring skin involvement were characterized by both lesional and circulating CD8+ T cells expressing CCR4. Destruction of epidermal and adnexal structures was histomorphologically associated with CCR4+ cytotoxic T cells invading basal layers of the epidermis where keratinocytes showed apoptotic death. The CCR4 ligand TARC/CCL17 was strongly expressed in skin lesions and elevated in the serum of CLE patients. The functional relevance of lymphocytic CCR4 expression could be confirmed by TARC/CCL17-specific in vitro migration assays. Our investigations suggest that CCR4 and TARC/CCL17 play a role in the pathophysiology of CLE. In particular, cytotoxic CD8+ T cells expressing CCR4 appear to be involved in scarring subtypes of CLE.

Cancer metastasis is the end result of a complex series of biologic events that leads to the formation of clinically significant secondary tumors at distant sites. The sites of distant metastasis are not random since certain tumors show a tendency to develop metastases in specific organs. Human melanoma, for example, demonstrates frequent metastasis to brain, lungs, lymph nodes, and skin. Herein, we review the evidence that suggests that a limited number of chemokine receptors may play critical roles in determining organ-selective metastasis in melanoma by regulating diverse processes such as chemoattraction, adhesion, and survival. In particular, we describe roles for CC chemokine receptor 7 (CCR7) in lymph node metastasis, CXC chemokine receptor 4 (CXCR4) in pulmonary metastasis, and CCR10 in skin metastasis, using a mouse model of melanoma. Preliminary evidence in this preclinical model suggests that inhibiting the function of these receptors may decrease the ability of cancer cells to disseminate to other sites and/or block their ability to survive and form tumors. Therefore, manipulation of the chemokine network could have therapeutic potential in human malignancies.

Controversy persists pertaining to the role of CCR4 ligands, namely CCL17 (or thymus and activation regulated chemokine; TARC) and CCL22 (or macrophage-derived chemokine; MDC), in Th2-type cytokine-dominated responses in the lung. Accordingly, the present study addressed the relative role of each of these CC chemokines during an evolving pulmonary granulomatous response elicited by the intrapulmonary embolization of live Schistosoma mansoni eggs into S. mansoni-sensitized mice. CCL22 protein expression peaked at day 4, but CCL17 levels were not increased significantly at any time after egg challenge. CCR4 transcript and protein expression were highest at day 8 after egg embolization and CCR4 protein was prominently expressed in macrophages surrounding S. mansoni eggs. Systemic immunoneutralization of CCL22 from the time of egg injection into S. mansoni-sensitized mice for 8 days significantly decreased CCR4 protein expression, the eosinophil content, the overall size of the egg granuloma, and its hydroxyproline content. Whole lung levels of interferon-gamma were also significantly increased at day 8 in anti-CCL22-treated mice. The systemic immunoneutralization of CCL17 had a lesser effect on all of the granuloma parameters listed above, but this antibody treatment significantly decreased granuloma hydroxyproline content to a greater extent than the anti-CCL22 antibody treatment. In addition, the immunoneutralization of CCL17 significantly increased whole lung levels of interleukin (IL)-4, IL-5, IL-13, transforming growth factor-beta, IL-12, and tumor necrosis factor-alpha at day 8 after egg infusion. Thus, these studies demonstrate a major role for CCL22 and a lesser role for CCL17 during an evolving S. mansoni egg granuloma in the lung.

Background: The expression of CXCR4 has been implicated in metastatic dissemination in different models of breast cancer and melanoma. In the present study, we evaluated CXCR4 expression in non-small-cell lung cancer (NSCLC) and the relationship between CXCR4 expression and the prognosis of stage I disease., Patients and Methods: Using immunohistochemical analysis, we retrospectively analyzed CXCR4 expression in specimens from 61 patients with completely resected pathologically confirmed stage I NSCLC for whom clinical follow-up data were available., Results: In the present study, we have shown that: CXCR4 is expressed by tumor cells in stage I NSCLC; CXCR4 is located in the nucleus and/or in the cytoplasm of tumor cells; strong nuclear staining was observed in 17 cases (29.8%); patients whose tumors had CXCR4-positive nuclear staining had a significantly longer duration of survival than patients whose tumors had no nuclear expression (P = 0.039, log-rank test). Interestingly, the 5-year metastasis rates were 23.5% and 34.1% in patients with CXCR4-positive and CXCR4-negative nuclear expression, respectively (P = 0.2)., Conclusion: Strong CXCR4-positive nuclear staining was associated with a significantly better outcome in early-stage NSCLC. The mechanisms underlying this clinically and biologically important finding need to be further explored.

Detailed longitudinal studies of patients with X-linked lymphoproliferative disease (XLP) may increase our understanding of the immunologic defects that contribute to the development of lymphoma and hypogammaglobulinemia in XLP. We describe progressive changes observed in immunoglobulin concentrations, lymphocyte subsets, and Epstein-Barr virus (EBV) loads occurring in a 2-year period in a newly infected, but otherwise healthy, carrier (patient 9). We compare these findings with those observed in the patient's brother, who had hypogammaglobulinemia and XLP (patient 4). Immunoglobulin G (IgG), IgM, and IgA concentrations increased in patient 9 during acute EBV infection, but thereafter they decreased steadily to concentrations consistent with hypogammaglobulinemia, reaching a plateau 5 months after infection. In both patients, CD19+ B-lymphocyte rates remained lower than 3%, with a contraction of the B-cell memory compartment (CD27+ CD19+/CD19+) to 20% (normal range, 32%-56%). T-lymphocyte subpopulations showed a reduction in CD4+ T-cell counts and a permanent CD8+ T-cell expansion. Interestingly, CXCR3 memory TH1 cells were expanded and CCR4+ TH2 lymphocytes were reduced, suggesting that abnormal skewing of memory T-cell subsets might contribute to reduced antibody synthesis. Despite an expanded number of CD3+CD8+ lymphocytes, increased EBV loads occurred in both patients without overt clinical symptoms of mononucleosis, lymphoproliferative disease, or lymphoma.

Roxithromycin (RXM), an anti-bacterial macrolide, has various immunomodulatory activities. To investigate the ability of RXM to downregulate skin-infiltration of T-lymphocytes, we examined the effects of RXM on keratinocyte production of chemokines and T cell expression of chemokine receptors. Normal human and HaCaT keratinocytes were cultured with RXM and stimulants. RXM at 1 or 10 microM significantly suppressed the production/expression of Th2 chemokines MDC and TARC in these keratinocytes, but the production of IP-10 was not affected. The effect of RXM on T-cell expression of the corresponding chemokine receptors was also tested in Th2-rich peripheral blood lymphocytes. The IL-2-enhanced expression level of Th2 chemokine receptor CCR4 was decreased by RXM at 10 microM, whereas the expression of CXCR3 was unchanged. Thus, RXM downmodulates both the production and receptor expression of Th2 but not Th1 chemokines involved in cutaneous immunity, suggesting its beneficial therapeutic effects on Th2-mediated or allergic skin disorders.

Lichen planus is a chronic inflammatory disease of the skin and oral mucosa in which the cell-mediated cytotoxicity is regarded as a major mechanism of pathogenesis. To understand its pathophysiology further, the present study examined the in situ expression of chemokines and chemokine receptors in oral lichen planus. Immunohistochemical analysis of 15 cases has consistently revealed that infiltrating CD4(+) and CD8(+) T cells in the submucosa predominantly expressed CCR5 and CXCR3. Furthermore, infiltrating T cells, particularly CD8(+) T cells, were positive for RANTES/CCL5 and IP-10/CXCL10, the ligands of CCR5 and CXCR3, respectively. By immunoelectron microscopy, these chemokines were localized in the cytolytic granules of CD8(+) T cells. Lesional keratinocytes also overexpressed the ligands of CXCR3, namely, MIG/CXCL9, CXCL10, and I-TAC/CXCL11. Our data suggest that the chemokines signaling via CCR5 and CXCR3, which are known to be selectively expressed by type 1 T cells, are predominantly involved in T-cell infiltration of oral lichen planus. Furthermore, the presence of CCL5 and CXCL10 in the cytolytic granules of tissue-infiltrating CD8(+) T cells expressing CCR5 and CXCR3 reveals a potential self-recruiting mechanism involving activated effector cytotoxic T cells.

Glomerular monocyte/macrophage (Mo/M phi) infiltrates play a role in many forms of glomerulonephritis (GN), and the intensity of Mo/M phi trafficking correlates with the loss of renal function and histological damage. We analyzed the functional role of macrophage-derived chemokine (MDC), a potent mononuclear cell chemoattractant, during the progression of anti-glomerular basement membrane (GBM) antibody (Ab) GN, a model of crescentic GN in the WKY rat, and whether the effects of MDC were dependent on its receptor CCR4. MDC mRNA and protein expression were markedly induced in nephritic glomeruli throughout the disease. Blocking the function of MDC did not affect the developing of the disease from days 2 to 7, but it dramatically blocked M omicron/M phi infiltration in the glomeruli, prevented crescent formation, and reversed renal function impairment during days 7 to 14 of the anti-GBM GN. In this study, we also found that MDC activity on M omicron/M phi in this GN was at least partly dependent on a new variant of CCR4. These results suggest that MDC is critically involved in the development of anti-GBM GN from acute glomerular injury to irreversible tissue damage. In addition, an antagonist to MDC may represent a prime drug target for therapeutic application to intervene in the progression of anti-GBM GN and in other M omicron/M phi-dominant GN.

The chemokine receptors (CCRs) CCR4 and CCR10, and the cutaneous lymphocyte antigen (CLA), have each been proposed as critical mediators of skin-specific TH lymphocyte homing in mice and humans. CLA initiates skin homing by mediating E-selectin-dependent tethering and rolling within cutaneous venules, but the specific roles of CCR4 and CCR10 are unclear. We have generated an antihuman CCR10 monoclonal antibody (mAb; 1B5) to illuminate the individual contributions of these molecules. This mAb allows us to compare CCR10, CCR4, and CLA expression within human TH populations. The mAb 1B5 recognizes functional CCR10 expression, as chemotactic responsiveness to cutaneous T-cell-attracting chemokine (CTACK)/CCL27 (a CCR10 ligand) parallels the staining of TH subsets. We find CCR10 expressed by only a minority (approximately 30%) of blood-borne, skin-homing (CLA+/CCR4+) TH cells. However, essentially all members of the relatively small "effector" (CLA+/CCR4+/CD27-/CCR7-) skin-homing TH population express CCR10. Most skin-infiltrating lymphocytes in allergic delayed-type hypersensitivity (DTH) and bacterial chancroid skin lesions express both CCR4 and CLA, but only about 10% express CCR10. This suggests for the 2 models of TH skin homing studied here that CCR10+ TH cells have no advantage over other CLA+/CCR4+ TH cells in homing to cutaneous sites. We conclude that the skin-homing TH compartment is itself divided into distinct subpopulations, the smaller of which expresses both CCR4 and CCR10, and the larger of which expresses only CCR4. Thus, CCR10 is unlikely to be necessary for cutaneous homing of TH cells in the models studied here. CCR10 may instead play a role in the movement of specialized "effector" cutaneous TH cells to and/or within epidermal microenvironments.

Chemokines are critical molecules in leukocyte trafficking, promoting site-specific migration to various tissues. The chemokine receptor CCR4 has recently been associated with skin-homing T cells. In view of the potential importance of CCR4 in skin homing of T cells, we investigated the expression pattern of CCR4 and its ligands TARC/CCL17 and MDC/CCL22 in the peripheral blood and skin of patients with cutaneous T cell lymphoma, a putative malignancy of the skin-homing T cells. In this study we analyzed the pattern of coexpression of the skin-homing molecules cutaneous lymphocyte antigen (CLA) and CCR4 in the blood and skin of patients with cutaneous T cell lymphoma. In the blood of cutaneous T cell lymphoma patients with peripheral blood involvement we found significantly increased percentages of T cells displaying the skin-homing phenotype (CLA+CCR4+) compared with healthy individuals. T cells expressing CLA and CCR4 were also found at high levels in cutaneous T cell lymphoma lesions along with abundant expression of the two CCR4 ligands TARC/CCL17 and MDC/CCL22. These data may explain, in part, why these T cells accumulate in the skin, a diagnostic feature of cutaneous T cell lymphomas.

Mast cells (MCs) accumulate at sites of allergic mucosal inflammation where they act as central effector and regulatory cells. Because chemokines are of vital importance in directing inflammatory leukocytes to the sites of inflammations, we have investigated the expression and function of CC-chemokine receptor (CCR) on human MCs. Two previously unrecognized MC-chemokine receptors, CCR1 and CCR4, could be identified on cord blood-derived MCs (CBMCs). CCR1 and CCR4 expressed on CBMCs exhibited a unique response profile. Of seven CCR1 and CCR4 agonists tested, only CCL5/RANTES act as an agonist inducing chemotaxis. The migration could be partially blocked by specific antibodies against CCR1 or CCR4, while a complete inhibition was achieved when both CCR1 and CCR4 were blocked. These results demonstrate that both CCR1 and CCR4 are functional receptors on human mast cells with capacity to mediate migration towards CCL5.

Development of allergic contact dermatitis to haptens depends upon a balance between CD8(+) T lymphocytes with pathogenic activity and CD4(+) T cells, which comprise both effector and regulatory cells. Thus, differential recruitment of CD8(+) and CD4(+) lymphocytes to sites of hapten challenge may have considerable impact on disease expression. Here the migration of cutaneous lymphocyte-associated antigen+, nickel-specific CD8(+) and CD4(+) T cell lines were compared with a panel of chemokines produced in the skin during allergic contact dermatitis. CCL17/TARC and CCL22/MDC induced a 3-fold higher migration of CD4(+) compared with CD8(+) lymphocytes. In contrast, CXCL10/IP-10 was 2-fold more potent in attracting CD8(+) cells. These findings were consistent with the higher expression of CCR4 and CXCR3 on CD4(+) and CD8(+) T cell lines, respectively. Moreover, CCR4 expression was high on nickel-specific T helper 2, intermediate on T helper 1 and T cytotoxic 2, and almost undetectable on T cytotoxic 1 clones. On the contrary, CXCR3 was expressed by T cytotoxic 1 and 2 and T helper 1, but not T helper 2 clones. Reverse transcription-polymerase chain reaction analysis of the skin before and after hapten challenge revealed the constitutive presence of TARC, and the early appearance of CCL2/MCP-1, followed by IP-10, CCL4/MIP-1beta, and MDC mRNA. Supernatants from activated keratinocytes induced a strong migration of CD8(+) lymphocytes, which was blocked by neutralization of IP-10. Conversely, supernatants from immature and mature dendritic cells attracted mostly CD4(+) lymphocytes in a TARC- and MDC-dependent manner. Our data indicate that distinct chemokines and cell types control the accumulation of CD8(+) and CD4(+) T cells within inflamed skin.

Chemokines and chemokine receptors play important roles in migration and tissue localization of various lymphocyte subsets. Here, we report the highly frequent expression of CCR4 in adult T-cell leukemia (ATL) and human T-cell leukemia virus type 1 (HTLV-1)-immortalized T cells. Flow cytometric analysis revealed that ATL and HTLV-1-immortalized T-cell lines consistently expressed CCR4. Inducible expression of HTLV-1 transcriptional activator tax in a human T-cell line Jurkat did not, however, up-regulate CCR4 mRNA. In vitro immortalization of peripheral blood T cells led to preferential outgrowth of CD4(+) T cells expressing CCR4. We further demonstrated highly frequent expression of CCR4 in fresh ATL cells by (1) reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of CCR4 expression in peripheral blood mononuclear cells (PBMCs) from patients with ATL and healthy controls; (2) flow cytometric analysis of CCR4-expressing cells in PBMCs from patients with ATL and healthy controls; (3) CCR4 staining of routine blood smears from patients with ATL; and (4) an efficient migration of fresh ATL cells to the CCR4 ligands, TARC/CCL17 and MDC/CCL22, in chemotaxis assays. Furthermore, we detected strong signals for CCR4, TARC, and MDC in ATL skin lesions by RT-PCR. Collectively, most ATL cases have apparently derived from CD4(+) T cells expressing CCR4. It is now known that circulating CCR4(+) T cells are mostly polarized to Th2 and also contain essentially all skin-seeking memory T cells. Thus, HTLV-1-infected CCR4(+) T cells may have growth advantages by deviating host immune responses to Th2. CCR4 expression may also account for frequent infiltration of ATL into tissues such as skin and lymph nodes.

Chemokines and chemokine receptors are key mediators for regulating cell traffic and positioning in both homeostatic and inflammatory conditions. It is also presumed that chemokines and their receptors are likely to play a critical role in the localization of malignant hematopoietic cells in their target organs. This study analyzed chemokine and chemokine receptor expression in several Hodgkin disease (HD)-derived cell lines and in HD tumors. All HD-derived cell lines expressed functional CCR7 and CXCR4 receptors. CCR7 up-regulation was mediated by constitutive NF-kappaB activity. Lymphoid tissues in HD revealed differential expression levels of CCR7, CXCR4, and CXCR5, depending on the distinct subtypes of HD. HD of the classical subtypes, predominantly located in the interfollicular zone, showed strong CCR7 and CXCR4 expression and moderate CXCR5 expression. In contrast, the nodular lymphocyte-predominant HD (NLP) subtype, regularly associated with follicular structures, exhibited no CCR7 reactivity but abundant CXCR4 staining. Their respective chemokine ligands showed marked expression by reactive cells within the tumors of classical HD and outside of the tumor nodules in NLPHD. Functionally, such differential chemokine receptor expression might contribute to specific localization and confinement of neoplastic cells within the target organs in the distinct HD entities.

Differential expression of adhesion molecules and chemokine receptors has been useful for identification of peripheral blood memory lymphocyte subsets with distinct tissue and microenvironmental tropisms. Expression of CCR4 by circulating memory CD4(+) lymphocytes is associated with cutaneous and other systemic populations while expression of CCR9 is associated with a small intestine-homing subset. CCR5 and CXCR3 are also expressed by discrete memory CD4(+) populations in blood, as well as by tissue-infiltrating lymphocytes from a number of sites. To characterize the similarities and differences among tissue-infiltrating lymphocytes, and to shed light on the specialization of lymphocyte subsets that mediate inflammation and immune surveillance in particular tissues, we have examined the expression of CCR4, CXCR3, and CCR5 on CD4(+) lymphocytes directly isolated from a wide variety of normal and inflamed tissues. Extra-lymphoid tissues contained only memory lymphocytes, many of which were activated (CD69(+)). As predicted by classical studies, skin lymphocytes were enriched in CLA expression whereas intestinal lymphocytes were enriched in alpha(4)beta(7) expression. CCR4 was expressed at high levels by skin-infiltrating lymphocytes, at lower levels by lung and synovial fluid lymphocytes, but never by intestinal lymphocytes. Only the high CCR4 levels characteristic of skin lymphocytes were associated with robust chemotactic and adhesive responses to TARC, consistent with a selective role for CCR4 in skin lymphocyte homing. In contrast, CXCR3 and CCR5 were present on the majority of lymphocytes from each non-lymphoid tissue examined, suggesting that these receptors are unlikely to determine tissue specificity, but rather, may play a wider role in tissue inflammation.

Recent studies indicate that Th1 and Th2 cells differ in their chemokine receptor expression and their responsiveness to various chemokines. Therefore, selective Th2 cell recruitment in Th2-predominant inflammatory diseases such as atopic dermatitis may be under the influence of some chemokines. It is reported that CC chemokine receptor (CCR) 4 is selectively expressed on Th2 cells whereas CXC chemokine receptor (CXCR) 3 is selectively expressed on Th1 cells. In this study we examined CCR4 and CXCR3 expression on peripheral blood CD4+ and CD8+ T cells obtained from adult atopic dermatitis subjects, and compared the results with those from patients with psoriasis vulgaris and healthy controls. CCR4 was preferentially expressed on CD4+ T cells from atopic dermatitis subjects and CXCR3 was preferentially expressed on CD4+ T cells from psoriasis vulgaris subjects. This CCR4 expression was prominent especially in severe atopic dermatitis subjects. CCR4 expression on CD4+ T cells in severe atopic dermatitis subjects decreased on improvement of disease activity. CD25 was preferentially expressed on CCR4+CD4+ T cells but not on CXCR3+CD4+ T cells in atopic dermatitis subjects. Cutaneous lymphocyte-associated antigen was also preferentially expressed on CCR4+CD4+ T cells but not on CXCR3+CD4+ T cells in atopic dermatitis subjects. CD4+ T cells in atopic dermatitis skin lesions were predominantly CCR4+ cells. Taken together, this study strongly indicates that CCR4+CD4+ T cells reflect disease activity and suggests that CCR4 expression is important for T cell infiltration into atopic dermatitis lesions. Thus, CCR4 may be a possible target for therapy of atopic dermatitis in the future.

Our previous study demonstrated formation of T cell-dendritic cell (DC) clusters in inflamed dermis of intraorally autotransplanted skin flaps. Such T cell-DC clusters are supposed to be important for close interactions between T cells and DCs including the specific antigen presentation. Here we show the involvement of the macrophage-derived chemokine (MDC/CCL22) and its specific receptor CC chemokine receptor 4 (CCR4) in the formation of T cell-DC clusters. Reverse transcriptase-polymerase chain reaction analysis revealed high levels of mRNA expression for MDC and CCR4 in inflamed skin and neck lymph nodes (LNs), but not in normal skin. Immunohistochemically, MDC(+) cells and CCR4(+) cells were mainly located within the T cell-DC clusters both in the dermis of inflamed skin and the T cell area of LNs. MDC(+) cells were identified to be DCs both in inflamed skin and LNs. The majority of CCR4(+) cells were CD4(+) T cells, accounting for approximately one-third of total CD4(+) T cells in the inflamed skin. Our data suggest that the MDC-CCR4 system plays an important role in the formation of T cell-DC clusters both in inflamed skin and LNs.

Platelets are known to contain platelet factor 4 and beta-thromboglobulin, alpha-chemokines containing the CXC motif, but recent studies extended the range to the beta-family characterized by the CC motif, including RANTES and Gro-alpha. There is also evidence for expression of chemokine receptors CCR4 and CXCR4 in platelets. This study shows that platelets have functional CCR1, CCR3, CCR4, and CXCR4 chemokine receptors. Polymerase chain reaction detected chemokine receptor messenger RNA in platelet RNA. CCR1, CCR3, and especially CCR4 gave strong signals; CXCR1 and CXCR4 were weakly positive. Flow cytometry with specific antibodies showed the presence of a clear signal for CXCR4 and weak signals for CCR1 and CCR3, whereas CXCR1, CXCR2, CXCR3, and CCR5 were all negative. Immunoprecipitation and Western blotting with polyclonal antibodies to cytoplasmic peptides clearly showed the presence of CCR1 and CCR4 in platelets in amounts comparable to monocytes and CCR4 transfected cells, respectively. Chemokines specific for these receptors, including monocyte chemotactic protein 1, macrophage inflammatory peptide 1alpha, eotaxin, RANTES, TARC, macrophage-derived chemokine, and stromal cell-derived factor 1, activate platelets to give Ca(++) signals, aggregation, and release of granule contents. Platelet aggregation was dependent on release of adenosine diphosphate (ADP) and its interaction with platelet ADP receptors. Part, but not all, of the Ca(++) signal was due to ADP release feeding back to its receptors. Platelet activation also involved heparan or chondroitin sulfate associated with the platelet surface and was inhibited by cleavage of these glycosaminoglycans or by heparin or low molecular weight heparin. These platelet receptors may be involved in inflammatory or allergic responses or in platelet activation in human immunodeficiency virus infection.

Atopic dermatitis is an inflammatory skin disease in which the inflammation is characterized by the influx of lymphocytes into the dermis. It is generally believed that atopic dermatitis is a Th2-type disease, i.e., the T lymphocytes produce interleukin-4, interleukin-5, interleukin-10, and interleukin-13, although it has become evident in recent years that the cytokine profile in the skin changes during the course of the disease towards a Th1-Th2 mixed cytokine profile (interferon-gamma, tumor necrosis factor alpha, and interleukin-2). The lymphocytes that home into the skin express cutaneous lymphocyte-associated antigen, and it has recently been shown that most of the lymphocytes in this population express the chemokine receptor CCR4. CCR4 is the receptor for the CC chemokine TARC (thymus and activation regulated chemokine), and this chemokine is expressed predominantly by keratinocytes in the basal layer of the epidermis of lesional atopic dermatitis skin in mice. In humans, however, it was shown to be expressed in the endothelial cells of the dermis. We have examined the peripheral blood mononuclear cells of atopic dermatitis patients for the expression of cutaneous lymphocyte-associated antigen and CCR4 and compared them with peripheral blood mononuclear cells from normal controls. We found that the proportion of CLA+CCR4+ lymphocytes is upregulated in atopic dermatitis patients. In addition we have examined skin biopsies of lesional and non-lesional skin from atopic dermatitis patients and found that the keratinocytes, but not the endothelial cells, produce TARC in the lesional but not in the nonlesional skin. To gain insight in the stimulatory mechanisms for TARC production in keratinocytes, as previously observed in mice, we cultured HaCaT cells and found that interferon-gamma and tumor necrosis factor alpha work synergistically to induce TARC production. These observations suggest that the induction of TARC production in keratinocytes plays an important role in the late phase skin invasion by CCR4+CLA+ Th2-type lymphocytes in atopic dermatitis.

Chemokine receptors mediate the migration of lymphocytes through the binding of soluble ligands, and their expression is differentially regulated in lymphocyte subsets. The pattern of chemokine receptor expression in T-cell non-Hodgkin lymphoma has not been previously studied. Using a panel of mouse monoclonal antibodies, we studied the immunohistochemical expression of the Th1-associated chemokine receptor CXCR3 in 141 patients with T-cell lymphoma, and we studied the receptors CCR4 and CCR5 and some of their ligands in a subset of these tumors. Expression of CXCR3 was typical of the smaller T cells in angioimmunoblastic lymphoma (15 of 18 patients), angiocentric lymphoma (3 of 3 patients), histiocyte-rich tumors (4 of 5 patients), and unspecified T-cell lymphomas (17 of 39 patients). CXCR3 expression was seen in only 1 of 15 patients with anaplastic lymphoma kinase (ALK)-positive anaplastic large-cell lymphoma. In contrast, all ALK-positive tumors showed diffuse reactivity for the Th2-associated receptor CCR4 (5 of 5 patients). CCR4 expression was also a consistent feature of the large-cell transformation of mycosis fungoides. CCR5 expression showed no consistent association with any T-cell tumor type. The chemokines Mig (CXCR3 ligand), TARC (CCR4 ligand), and MCP-2 (CCR5 ligand) were detected in intratumoral blood vessels and histiocytes. Mig was also coexpressed by a subset of CXCR3-positive tumor cells in 6 of 20 lymphomas. MCP-2 was highly expressed in stromal cells in 3 patients with nodal involvement by cutaneous T-cell lymphoma. As with normal T-cell subsets, we demonstrated that there is frequent differential expression of chemokine receptors in T-cell tumors, which may explain, in part, the distinctive patterns of spread in different tumor subtypes. (Blood. 2000;96:685-690)

Platelets play roles in both thrombosis and inflammation, and chemokines that are released at sites of inflammation could potentially activate platelets. Among the chemokine receptors expressed on platelets, the CXCR4 is the receptor for chemokine stromal cell-derived factor-1 (SDF-1), and the CCR4 is the receptor for macrophage-derived chemokine (MDC). Of the chemokines tested, SDF-1 and MDC were the only 2 that activated platelets. Both are weak agonists, but they enhanced response to low-dose adenosine 5'-diphosphate (ADP), epinephrine, or serotonin. When SDF-1 and MDC were added together, full and brisk platelet aggregation occurred. Platelet activation by these 2 chemokines appears to involve distinct pathways: SDF-1 inhibited an increase in cyclic adenosine monophosphate (cAMP) following prostaglandin (PG) I(2), while MDC had no effect. In contrast, MDC, but not SDF-1, lead to Ca(++) mobilization by platelets. Further, second-wave aggregation induced by MDC in platelet-rich plasma was inhibited by aspirin, ADP scavenger creatine phosphate/creative phosphokinase (CP/CPK), and ARL-66096, an antagonist of the ADP P2T(AC) receptor involved in adenylyl cyclase inhibition. But the aggregation was not affected by A3P5PS, an inhibitor of the ADP P2Y receptor. SDF-1-induced aggregation was inhibited by aspirin, but it was only slightly affected by CP/CPK, ARL-66096, or A3P5PS. Finally, the presence of chemokines in platelets was determined. Reverse transcriptase-polymerase chain reaction studies with platelet RNA did not detect the presence of SDF-1 or MDC. In summary, SDF-1 and MDC are platelet agonists that activate distinct intracellular pathways. Their importance in the development of thrombosis at sites of inflammation needs to be further evaluated.

Kaposi's sarcoma-associated herpesvirus (KSHV) encodes 3 genes that are homologous to cellular chemokines. vMIP-III, the product of open reading frame K4.1, is the most distantly related to human chemokines and has yet to be characterized. We have examined the interaction of vMIP-III with chemokine receptors, its expression in KS lesions, and its in ovo angiogenic properties. We show expression of vMIP-III in KS lesions and demonstrate the stimulation of angiogenesis by this chemokine, like vMIP-I and vMIP-II, in the chick chorioallantoic membrane assay. vMIP-III does not block human immunodeficiency virus entry through the coreceptors CCR3, CCR5, or CXCR4. However, vMIP-III is an agonist for the cellular chemokine receptor CCR4. CCR4 is expressed by TH2-type T cells. Consistent with this, vMIP-III preferentially chemoattracts this cell type. Because of these biologic properties and because it is expressed in KS lesions, vMIP-III may play an important role in the pathobiology of KS. (Blood. 2000;95:1151-1157)

Hodgkin's lymphoma is characterized by the combination of Reed-Sternberg (R-S) cells and a prominent inflammatory cell infiltrate. One of the intriguing questions regarding this disease is what is causing the influx of T lymphocytes into the involved tissues. We applied the serial analysis of gene expression (SAGE) technique on the Hodgkin's lymphoma-derived cell line L428 and on an Epstein-Barr virus (EBV)-transformed lymphoblastoid B-cell line. A frequently expressed tag in L428 corresponded to the T-cell-directed CC chemokine TARC. Reverse transcription polymerase chain reaction analyses demonstrated expression of TARC in nodular sclerosis (NS) and mixed cellularity (MC) classical Hodgkin's lymphomas but not in NLP Hodgkin's lymphoma, anaplastic large-cell lymphomas, and large-B-cell lymphomas with CD30 positivity. Two of five cases of T-cell-rich B-cell lymphoma (TCRBCL) were TARC positive. RNA in situ hybridization (ISH) showed a strong signal for TARC in the cytoplasm of R-S cells, and immunohistochemical staining confirmed the presence of the TARC protein in the R-S cells of NS and MC Hodgkin's lymphomas. The lymphocytic and histiocytic (L&H)-type cells of nodular lymphocyte predominance Hodgkin's lymphoma and the neoplastic cells of non-Hodgkin's lymphomas with the exception of two cases of TCRBCL did not stain for TARC. TARC is known to bind to the CCR4 receptor, which is expressed on activated Th2 lymphocytes. The immunophenotype of lymphocytes surrounding R-S cells is indeed Th2-like, and by RNA ISH these lymphocytes showed a positive signal for the chemokine receptor CCR4. The findings suggest that production of TARC by the R-S cells may explain the characteristic T-cell infiltrate in classical Hodgkin's lymphoma.