Introduction

Chemokines are small molecules that induce chemotaxis and activation of certain subsets of leukocytes. Most chemokines are 8–10 kDa in size, although CX3CL1 is around 90 kDa in its soluble form (Bazan et al. 1997). They are classified into four types: CC chemokines, CXC chemokines, C chemokines, and CX3C chemokine. The classification is based on the pattern of cysteine residues in each protein (Charo and Ransohoff 2006). Chemokines are promiscuous in their receptor binding. For example, CCL26 can bind CX3CR1 as well as CCR3 and thus may play a dual role in allergic disease, attracting eosinophils and T cells and monocytes (Nakayama et al. 2010). Accumulating evidence supports that chemokines have broader functions than just chemotaxis. The expression pattern of chemokines and chemokine receptors is specific to certain organs and cells. Therefore, chemokines are important to elucidate the mechanism of organ-specific human diseases. Many chemokines expressed in the skin, such as CXCL8, CXCL9, CXCL10, CCL11, CCL17, CCL26, and CCL27, mediate recruitment of leukocytes to the skin from the blood. There are also chemokines that mediate egress of leukocytes via the lymphatics such as CCL21, CXCL12, and CX3CL1. Both types of chemokines are important for maintaining skin homeostasis and their disruption can result in skin pathologies. The present review highlights roles of four chemokines that we have been studying these years: CCL17, CCL11, CCL26, and CX3CL1.

CCL17 and Skin Diseases

CCL17, also called thymus and activation-regulated chemokine, is a member of the CC chemokine family. It is a ligand of CCR4 and CCR8 and serves to recruit cells expressing these chemokine receptors (Inngjerdingen et al. 2000). CCL17 is mainly produced by dendritic cells (Imai et al. 1996; Sallusto et al. 1998b), fibroblasts (Yu et al. 2002), and blood endothelial cells (Campbell et al. 1999). Langerhans cells, skin-resident dendritic cells, produce high levels of CCL17 constitutively during culture even without exogenous stimuli. Stimulation with transforming growth factor (TGF)-β, tumor necrosis factor (TNF)-α, interleukin (IL)-4, IL-13 and anti-CD40 antibody enhanced the expression. In contrast, its production level decreased in the presence of granulocyte–macrophage colony-stimulating factor, interferon (IFN)-γ, or IL-10 (Fujita et al. 2005; Xiao et al. 2003). CCR4 is predominantly expressed on Th2 lymphocytes, basophils, and natural killer (NK) cells (Godiska et al. 1997; Nickel et al. 1999; Sallusto et al. 1998a).

CCL17-transgenic (Tg) mice which overexpress CCL17 in the epidermis showed enhanced Th2-type and decreased Th1-type contact hypersensitivity (CHS). An increased number of CCR4+ cells infiltrated in Tg mice. IL-4 mRNA expression level was higher and that of IFN-γ was lower in Tg mice in both acute and chronic CHS. Moreover, higher levels of serum IgE were observed in Tg mice after CHS (Tsunemi et al. 2006). Thus, interactions between CCL17 and CCR4 are important for skin inflammation by Th2-polarized cells. The importance of CCR4 in macrophages has also been shown using knockout mice. CCR4−/− mice exhibited significantly decreased mortality on administration of high or low dose bacterial lipopolysaccharide compared with wild-type (WT) mice (Chvatchko et al. 2000). After high-dose lipopolysaccharide treatment, peritoneal lavage contained significantly smaller numbers of macrophages in CCR4−/− mice. Intrapulmonary administration of bleomycin sulfate provoked lethal inflammatory and fibrotic responses in WT mice, while such responses were absent in CCR4−/− mice (Trujillo et al. 2008). Transcript and protein analyses of alveolar and bone marrow-derived macrophages showed that cells isolated from CCR4−/− mice did not exhibit CCL17-dependent M1 activation in response to bleomycin. CCR4−/− mice were similarly resistant to challenge with Toll-like receptor agonists as well as bacterial peritonitis (Ness et al. 2006). Macrophages from CCR4−/− mice exhibited many features consistent with alternative activation. When the effects of CCR4 deficiency in oxazolone-induced CHS were examined, the inflammatory response in the skin at 24 h post-elicitation was stronger in CCR4−/− mice compared to WT mice. There were enhanced ear swelling and increased inflammatory cell infiltration, probably due to deficient migration of suppressive T cells (Lehtimäki et al. 2010). Thus, interactions between CCL17 and CCR4 play critical roles in skin homing of Th2 cells, classical activation pathways of macrophages, and attraction of suppressive T cells into the skin.

CCL17 and CCR4 play important roles in the pathogenesis of inflammatory skin diseases such as atopic dermatitis (AD), bullous pemphigoid, dermatomyositis, papuloerythroderma syndrome, and angioedema (Kawashima et al. 2010; Saeki and Tamaki 2006; Tamaki et al. 2006; Yokobayashi et al. 2008). Among them, AD showed variable and higher serum CCL17 levels than other skin diseases. In AD, serum CCL17 levels sharply reflected disease activity and decreased after the treatment in accordance with the improvement in clinical symptoms (Kakinuma et al. 2001). CCR4 expression levels on CD4+ peripheral blood mononuclear cells in AD patients were significantly higher than those seen in healthy controls (Wakugawa et al. 2001). Moreover, serum CCL17 levels are superior to those of other chemokines as a disease marker of AD (Kimura et al. 2014). Thus, CCL17 plays critical roles in various Th2-type skin diseases, especially in AD.

CCL17 is also important in hematological malignancy (Sugaya 2010). In patients with mycosis fungoides (MF) and Sézary syndrome (SS), both of which are cutaneous T-cell lymphoma, serum CCL17 levels were elevated, reflecting the disease activity (Kakinuma et al. 2003). We and others have shown that transformed large cells in tumor stage MF and SS express CCR4 (Ferenczi et al. 2002; Kakinuma et al. 2003; Sokolowska-Wojdylo et al. 2005; Yagi et al. 2006). Tumor cells of adult T-cell leukemia/lymphoma (ATLL) also express CCR4 (Yoshie et al. 2002). Mogamulizumab, a humanized defucosylated anti-CCR4 monoclonal antibody, showed potent antitumor activity mediated by highly enhanced antibody-dependent cellular cytotoxicity against primary ATLL cells both in vitro and ex vivo (Ishii et al. 2010). This drug demonstrated clinically meaningful antitumor activity in patients with relapsed ATLL with an acceptable toxicity profile (Ishida et al. 2012). It is now approved for use by the Japanese government in the treatment of recurrent ATLL and MF/SS. Taken together, CCL17 expressed by Langerhans cells, blood endothelial cells, and fibroblasts plays a key role in attracting Th2 cells and tumor cells of ATLL and MF/SS into the skin, developing various Th2-type inflammatory skin diseases as well as cutaneous lymphoma (Fig. 1).

Fig. 1
figure 1

CCL17 expression by epidermal Langerhans cells, fibroblasts, and blood endothelial cells and infiltration of CCR4-expressing Th2 cells. CCR4 expression on ATLL/MF/SS tumor cells is important for their skin homing

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CCL11, CCL26, and Skin Diseases

CCL11 and CCL26 belong to the CC chemokine family. The former is also called eotaxin-1 and the latter is called eotaxin-3. Both of them have potent chemotactic activity for CCR3+ cells such as eosinophils (Forssmann et al. 1997) as well as subpopulations of Th2 cells (Sallusto et al. 1997). In cutaneous tissue components, CCL11 is produced by dermal fibroblasts (Bartels et al. 1996), blood endothelial cells (Rothenberg et al. 1995), and dendritic cells (Beaulieu et al. 2002). CCL26 is reported to be expressed by epidermal keratinocytes (Nishi et al. 2008), dermal fibroblasts (Hoeck and Woisetschläger 2001), and blood endothelial cells (Shinkai et al. 1999). CCR3, a receptor for CCL11 and CCL26, is expressed on eosinophils, basophils, and subpopulations of T cells (Forssmann et al. 1997; Sallusto et al. 1997; Uguccioni et al. 1997).

In vivo function of CCL11 and CCR3 has been studied using knockout mice. Eosinophils and the eosinophil product major basic protein were absent from the skin of ovalbumin-sensitized CCR3−/− mice (Ma et al. 2002). Recruitment of eosinophils to lung parenchyma and bronchoalveolar lavage fluid was also severely impaired in CCR3−/− mice, suggesting that CCR3 plays an essential role in eosinophil recruitment to the skin and the lung and in the development of airway hyperresponsiveness. Analysis of airway eosinophilia revealed a dominant role for CCL24 and a synergistic reduction in CCL11/24 double-knockout mice as well as CCR3-knockout mice (Pope et al. 2005). Moreover, bleomycin-induced lung injury was significantly reduced in CCL11−/− mice (Huaux et al. 2005). Expression of profibrotic cytokines such as TGF-β was diminished in the absence of CCL11, suggesting that CCL11 and CCR3 are important in the pulmonary recruitment of granulocytes and play significant pathogenic roles in lung fibrosis. Thus, interactions between CCL11 and CCR3 play important roles in skin and lung inflammation and fibrosis mainly mediated by eosinophils.

CCL11 protein is upregulated in a variety of inflammatory diseases that are characterized by massive infiltration of eosinophils, such as allergic asthma (Campbell et al. 1998; Ganzalo et al. 1996), allergic rhinitis (Baraniuk 1997), and AD (Kaburagi et al. 2001; Yawalkar et al. 1999), and is thought to be a key player in the pathogenesis of these conditions. CCL26 also plays important roles in allergic diseases such as AD and allergic asthma (Berkman et al. 2001; Kagami et al. 2003). Moreover, we and others reported association of CCL26 with angioedema (Yokobayashi et al. 2008), MF/SS (Miyagaki et al. 2010), bullous pemphigoid (Kagami et al. 2012), eosinophilic folliculitis (Nakahigashi et al. 2012; Yokobayashi et al. 2013), and eosinophilic cellulitis (Fujimura et al. 2014), all of which are regarded as Th2-type diseases.

Thus, both CCL11 and CCL26 play important roles in establishing Th2-dominant microenvironment in the skin.

There are emerging data suggesting that signaling through chemokine/chemokine receptor interactions is implicated with tumor growth and invasion of several cancer types, including lymphomas, in an autocrine/paracrine manner. In most cases of anaplastic large cell lymphoma (ALCL), tumor cells express CCR3 (Kleinhans et al. 2003; Yamaguchi et al. 2006). ALCL cells also express CCL11 (Kleinhans et al. 2003). Therefore, CCL11 may act in autocrine manner in ALCL. Indeed, we demonstrated cell survival effects of CCL11–CCR3 interactions in ALCL cells (Miyagaki et al. 2011). CCL11 increased cell survival rates and promoted proliferation of CCR3+ lymphoma cells via extracellular signal-regulated kinase pathway. We also showed Bcl-xL and survivin expression in CCL11-stimulated lymphoma cells in vitro and ALCL tumor cells in situ. Taken together, CCL11 and CCL26 expressed by skin-resident cells such as fibroblasts, blood endothelial cells, and keratinocytes induce infiltration of CCR3-expressing cells such as Th2 cells and eosinophils (Fig. 2). CCL11 may serve as an autocrine as well as a paracrine in ALCL.

Fig. 2
figure 2

CCL11/CCL26 expression by fibroblasts, blood endothelial cells, and keratinocytes and infiltration of CCR3-expressing cells such as Th2 cells and eosinophils. CCR3 expression on ALCL tumor cells is important for their proliferation

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CX3CL1 and Skin Diseases

CX3CL1, also called fractalkine, and its receptor CX3CR1 have been identified as key mediators of transmigration of immune cells such as T cells, NK cells, and monocytes (Bazan et al. 1997; White and Greaves 2012). CX3CL1 is expressed by inflamed endothelial cells, epithelial cells, including keratinocytes, macrophages, and vascular smooth muscle cells, while CX3CR1 is mainly expressed on T cells, NK cells, neutrophils, monocytes, and mast cells (Imai et al. 1997; Sugaya et al. 2003; Yang et al. 2007). CX3CL1 exists in two forms, a membrane-bound and a soluble form, which is an unusual feature of this chemokine. In its membrane-bound form, CX3CL1 acts as an adhesion molecule, whereas in its soluble form, it acts as a chemoattractant for T cells, NK cells, and monocytes (White and Greaves 2012). Monocytes can be classified into two functional subsets based on their chemokine receptor expression pattern: a short-lived CCR2+ subset that is actively recruited into inflamed tissues, and a CX3CR1+ subset characterized as residents within non-inflamed tissues (Geissmann et al. 2003). Ly6C+CCR2+CX3CR1− inflammatory and Ly6C−CCR2−CX3CR1+ resident monocytes are generally thought to differentiate into M1 and M2 macrophages, respectively. Recent studies have revealed that CX3CR1 is expressed on cells of the monocyte-macrophage dendritic cell lineage (Auffray et al. 2009; Jung et al. 2000) and that CX3CL1 promotes trafficking of dendritic cells through inflamed lymphatics (Johnson and Jackson 2013).

CX3CR1−/− mice have been utilized to investigate roles of CX3CL1 and CX3CR1 in various disease models (Ishida et al. 2008; Morimura et al. 2013; Yu et al. 2007). In the setting of wound healing, CX3CL1 is expressed by macrophages and endothelial cells, while CX3CR1 is expressed by macrophages and fibroblasts (Ishida et al. 2008). Decreased expression of macrophage-related cytokines such as TGF-β and vascular endothelial growth factor and reduced deposition of α-smooth muscle actin and collagen were shown in the wounded skin of CX3CR1−/− mice (Ishida et al. 2008). The roles of CX3CL1 in tumor immunity was also studied using CX3CR1−/− mice. CX3CR1−/− mice injected with B16 melanoma cells had increased lung tumor burden together with selective reduction of monocytes and NK cells in the lungs (Yu et al. 2007). Moreover, using mouse vasculitis model, we showed that the development of leukocytoclastic vasculitis was significantly reduced in CX3CR1−/− mice compared with WT mice (Morimura et al. 2013). Infiltration of neutrophils and mast cells as well as expression of IL-6 and TNF-α were decreased in CX3CR1−/− mice. Thus, knockout mouse models showed critical roles of CX3CL1 and CX3CR1 in wound healing, tumor immunity, and vasculitis.

The importance of interaction between CX3CL1 and CX3CR1 has been postulated in many inflammatory diseases, including rheumatoid arthritis, vasculitis, systemic sclerosis, and AD (Echigo et al. 2004; Hasegawa et al. 2005; Morimura et al. 2013; Murphy et al. 2008). Serum CX3CL1 levels and the frequencies of CX3CR1+ T cells in peripheral blood were significantly higher in patients with rheumatoid arthritis than in healthy controls (Murphy et al. 2008). Moreover, CX3CL1 levels in synovial fluid were elevated in rheumatoid arthritis, compared to osteoarthritis or healthy subjects. Serum CX3CL1 levels were also significantly higher in patients with vasculitis including polyarteritis nodosa than in healthy controls (Morimura et al. 2013). Upregulated expression of CX3CL1 and CX3CR1 in the affected tissue of systemic sclerosis may cooperatively augment the recruitment of mononuclear cells, leading to inflammation and vascular injury (Hasegawa et al. 2005). CX3CR1+ cells in the affected skin of patients with AD increased compared with those in normal skin and serum CX3CL1 levels were increased in AD patients, which were associated with the disease severity (Echigo et al. 2004). Thus, interaction between CX3CL1 and CX3CR1 plays important roles in the development of rheumatoid arthritis, vasculitis, and AD. CX3CL1 expressed on blood endothelial cells can be a therapeutic target to prevent infiltration of CX3CR1+ immune cells such as mast cells, neutrophils, and macrophages (Fig. 3).

Fig. 3
figure 3

CX3CL1 expression on blood endothelial cells and infiltration of CX3CR1-expressing mast cells and neutrophils in the settings of vasculitis. CX3CL1-CX3CR1 interaction is also important for migration of resident macrophages

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Conclusion

Many chemokines are involved in the development of skin diseases. They play important roles in establishing microenvironment in which migratory immune cells, together with skin-resident cells, cause prolonged inflammation. Some chemokines can also serve as an autocrine as well as a paracrine in skin malignancy. Biologics targeting chemokines and their receptors are promising strategies for various skin diseases that are resistant to the current therapy. Indeed, in Japan, anti-CCR4 monoclonal antibody is already approved in the treatment of recurrent ATLL and MF/SS. Although we cannot overlook potential side effects and relatively high prices, we should keep working hard to develop new drugs targeting chemokines or chemokine receptors to improve patients’ life expectancy and quality of life.