Re-analysis of single cell and spatial transcriptomics data reveals B cell landscape in gastric cancer microenvironment and its potential crosstalk with tumor cells for clinical prognosis.

BACKGROUND: At present, immunotherapy has become a powerful treatment for advanced gastric cancer (AGC), but not all patients can benefit from it. According to the latest research, the impact of B cell subpopulations on the immune microenvironment of gastric cancer (GC) is unknown. Exploring whether the interaction between B cells and tumor cells in GC affects the effectiveness of immunotherapy has attracted our interest. METHODS: This study involved the re-analysis of single-cell RNA (scRNA) and spatial transcriptomics (ST) data from publicly available datasets. The focus was on investigating the subpopulations and differentiation trajectories of B cells in the gastric cancer (GC) tumor immune microenvironment (TIME). Spatial transcriptomics (ST) and multiple immunofluorescence (mIF) revealed a clear co-localization pattern between B cells and tumor cells. Multiple immunotherapy datasets were collected to identify unique immunotherapy biomarkers. The unique immunotherapeutic potential of targeting CCL28 was validated through a mouse gastric cancer model. In addition, flow cytometry revealed changes in the tumor immune microenvironment targeting CCL28. RESULTS: The re-analysis of ST data from multiple cancer types revealed a co-localization pattern between B cells and tumor cells. A significant number of IgA plasma cells were identified in the GC TIME. Five different tumor-infiltrating B cell subpopulations and two unique B cell differentiation trajectories were characterized, along with seven GC-related states. By analyzing the communication between GC cells and B cells, it was further discovered that tumor cells can influence and recruit plasma cells through CCL28-CCR10 signaling. Additionally, there was a crosstalk between GC cells and B cells. Finally, we identified the LAMA/CD44 signaling axis as a potential prognostic marker for immunotherapy through a large amount of immunotherapy data. We also validated through various animal tumor models that targeting CCL28 can significantly promote CD8+T cell infiltration and function in the TME by regulating B cell and plasma cell functions, and has the ability to synergize immunotherapy. CONCLUSION: The co-localization and crosstalk between GC cells and B cells significantly affect the efficacy of immunotherapy, and inhibiting the CCL28-CCR10 signal axis is a potential immunotherapy target for GC. Meanwhile, LAMA/CD44 pair may be a potential adverse indicator for immunotherapy and tumor prognosis.

Environmentally related microcystin-LR-induced ovarian dysfunction via the CCL2-CCR10 axis in mice ameliorated by dietary mulberry.

Microcystin-LR (MC-LR) is a reproductive toxin produced by cyanobacteria in the aquatic environment and can be ingested by humans through drinking water and the food chain, posing a threat to human reproductive health. However, the toxic mechanisms and prospective interventions for MC-LR-induced ovarian dysfunction at environmental doses are unknown. The mulberry fruit is a traditional natural product of plant origin, with various pharmacological effects, such as antioxidant and anti-inflammatory effects. Here, mice were exposed to MC-LR (10, 100 µg/L) in drinking water for 90 days, during which mice were gavage 600 mg/kg/week of mulberry fruit extract (MFE). It was found that MC-LR can accumulate in mouse ovaries, causing sexual hormone disturbance, inflammatory infiltration, and ovarian pathological damage. Results from RNA-seq were shown that CCL2, a chemokine associated with inflammatory response, was significantly increased in mouse ovary after MC-LR exposure. Further investigation revealed that MC-LR exposure aggravates apoptosis of granulosa cells via the CCL2-CCR10 axis-mediated Jak/Stat pathway. Importantly, MFE can significantly ameliorate these ovarian dysfunction phenotypes by inhibiting the activation of the CCL2-CCR10 axis. This study broadened new insights into the ovarian toxicity of MC-LR and clarified the pharmacological effects of mulberry fruit on ovarian function protection.

Inhibition of cc chemokine receptor 10 ameliorates osteoarthritis via inhibition of the phosphoinositide-3-kinase/Akt/mammalian target of rapamycin pathway.

BACKGROUND: Osteoarthritis (OA) is a joint disease characterized by inflammation and progressive cartilage degradation. Chondrocyte apoptosis is the most common pathological feature of OA. Interleukin-1ß (IL-1ß), a major inflammatory cytokine that promotes cartilage degradation in OA, often stimulates primary human chondrocytes in vitro to establish an in vitro OA model. Moreover, IL-1ß is involved in OA pathogenesis by stimulating the phosphoinositide-3-kinase (PI3K)/Akt and mitogen-activated protein kinases pathways. The G-protein-coupled receptor, cc chemokine receptor 10 (CCR10), plays a vital role in the occurrence and development of various malignant tumors. However, the mechanism underlying the role of CCR10 in the pathogenesis of OA remains unclear. We aimed to evaluate the protective effect of CCR10 on IL-1ß-stimulated CHON-001 cells and elucidate the underlying mechanism. METHODS: The CHON-001 cells were transfected with a control small interfering RNA (siRNA) or CCR10-siRNA for 24 h, and stimulated with 10 ng/mL IL-1ß for 12 h to construct an OA model in vitro. The levels of CCR10, cleaved-caspase-3, MMP-3, MMP-13, Collagen II, Aggrecan, p-PI3K, PI3K, p-Akt, Akt, phosphorylated-mammalian target of rapamycin (p-mTOR), and mTOR were detected using quantitative reverse transcription polymerase chain reaction and western blotting. Viability, cytotoxicity, and apoptosis of CHON-001 cells were assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, lactate dehydrogenase assay (LDH), and flow cytometry analysis, respectively. Inflammatory cytokines (TNF-α, IL-6, and IL-8) were assessed using enzyme-linked immunosorbent assay. RESULTS: Level of CCR10 was substantially higher in the IL-1ß-stimulated CHON-001 cells than that in the control group, whereas CCR10 was down-regulated in the CCR10-siRNA transfected CHON-001 cells compared to that in the control-siRNA group. Notably, CCR10 inhibition alleviated IL-1ß-induced inflammatory injury in the CHON-001 cells, as verified by enhanced cell viability, inhibited LDH release, reduced apoptotic cells, and cleaved-caspase-3 expression. Meanwhile, IL-1ß induced the release of tumor necrosis factor alpha, IL-6, and IL-8, increase of MMP-3 and MMP-13, and decrease of Collagen II and Aggrecan in the CHON-001 cells, which were reversed by CCR10-siRNA. However, these effects were reversed upon PI3K agonist 740Y-P treatment. Further, IL-1ß-induced PI3K/Akt/mTOR signaling pathway activation was inhibited by CCR10-siRNA, which was increased by 740Y-P treatment. CONCLUSION: Inhibition of CCR10 alleviates IL-1ß-induced chondrocytes injury via PI3K/Akt/mTOR pathway inhibition, suggesting that CCR10 might be a promising target for novel OA therapeutic strategies.

CCR10-mediated Enhancement of T Cell Trafficking for Improved Tumor Immunotherapy.

BACKGROUND/AIM: The effectiveness of adoptive T cell therapy for solid tumors remains suboptimal, partly attributed to insufficient T cell infiltration into the tumor site. A promising strategy involves directing T cells towards the tumor utilizing tumor-specific chemokine receptors. MATERIALS AND METHODS: We analyzed chemokine receptor expression in activated T cells and chemokine expression in breast and lung cancer using The Cancer Genome Atlas (TCGA) data. Subsequently, we generated 1G4 T cell receptor-engineered T (TCR-T) cells with CCR10 and performed in vitro and in vivo efficacy tests. RESULTS: CCR10 exhibited insufficient expression in various human T cells. Analysis of TCGA RNA sequencing data revealed elevated expression of the chemokine CCL28, the corresponding chemokine for CCR10, in breast and lung cancer. Consequently, we generated CCR10-1G4 TCR-T cells. CCR10-1G4 dual expressing TCR-T cells exhibited comparable cellular cytotoxicity but increased mobility compared to 1G4 TCR-T cells in vitro. Furthermore, injecting CCR10-1G4 dual expressing TCR-T cells into a xenograft tumor model demonstrated enhanced in vivo trafficking and a greater reduction of tumor burden. CONCLUSION: This study highlights the potential of CCR10 for developing efficient adoptive T-cell treatments targeting solid tumors.

Blocking CCR10 Expression Activates m6A Methylation and Alleviates Vascular Endothelial Cell Injury.

BACKGROUND: Cardiovascular disease, one of the most common types of disease in clinical practice today, carries a very high risk of disability and death. This research aims to examine genome-wide changes in injured human dermal microvascular endothelial cells (HDMECs) using the Ribonucleic Acid sequencing (RNA-Seq) technique, and to search for key genes influencing N6-methyladenosine (m6A) methylation, thus gaining new insights into future clinical diagnosis and treatment of cardiovascular diseases (CVDs) and laying a foundation for follow-up research. METHODS: Impaired HDMECs (injury group), established by endotoxin intervention, were analyzed by RNA-Seq for differentially expressed genes (DEGs) relative to normal HDMECs (control group). Then, DEGs that might be associated with m6A methylation were selected for expression blocking to observe m6A methylation alterations. The migration, angiogenesis, and inflammatory response of damaged HDMECs were detected by cell scratch assay, western blotting, and Enzyme-linked Immunosorbent Assay (ELISA) experiments, respectively. RESULTS: In this study, 20 DEGs were screened out from the two groups by RNA-Seq, of which 17 were up-regulated and 3 were down-regulated. The C-C motif chemokine receptor 10 (CCR10) was selected for subsequent analysis. Real-Time Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) identified elevated CCR10 expression and reduced m6A methylation levels in the injury group (p < 0.05). After blocking CCR10 expression in damaged HDMECs by BI6901 (a CCR10 specific blocker) m6A methylation, cell activity, vascular endothelial growth factor A (VEGFA) and CD31 protein expression, as well as relative length and branches of tube formation were found to be increased compared with the injury group, while the levels of inflammatory factors interleukin-1 (IL-1), interleukin-1 (IL-6) and tumor necrosis factor-α (TNF-α) were decreased (p < 0.05). CONCLUSIONS: Blocking CCR10 expression can activate m6A methylation, promote cell activity, inhibit inflammatory reactions and alleviate HDMEC injury, which may become a breakthrough in future diagnosis and treatment of cardiovascular diseases.

Neutralization of excessive CCL28 improves wound healing in diabetic mice.

Introduction: Chronic, non-healing skin wounds such as diabetic foot ulcers (DFUs) are common in patients with type 2 diabetes mellitus (T2DM) and often result in limb amputation and even death. However, mechanisms by which T2DM and inflammation negatively impact skin wound healing remains poorly understood. Here we investigate a mechanism by which an excessive level of chemokine CCL28, through its receptor CCR10, impairs wound healing in patients and mice with T2DM. Methods & Results: Firstly, a higher level of CCL28 was observed in skin and plasma in both patients with T2DM, and in obesity-induced type 2 diabetic db/db mice. Compared with WT mice, adipose tissue from db/db mice released 50% more CCL28, as well as 2- to 3-fold more IL-1ß, IL-6, and TNF-α, and less VEGF, as determined by ELISA measurements. Secondly, overexpression of CCL28 with adenovirus (Adv-CCL28) caused elevation of proinflammatory cytokines as well as CCR10 expression and also reduced eNOS expression in the dorsal skin of WT mice as compared with control Adv. Thirdly, topical application of neutralizing anti-CCL28 Ab dose-dependently accelerated wound closure and eNOS expression, and decreased IL-6 level, with an optimal dose of 1 µg/wound. In addition, mRNA levels of eNOS and anti-inflammatory cytokine IL-4 were increased as shown by real-time RT-PCR. The interaction between eNOS and CCR10 was significantly reduced in diabetic mouse wounds following application of the optimal dose of anti-CCL28 Ab, and eNOS expression increased. Finally, enhanced VEGF production and increased subdermal vessel density as indicated by CD31 immunostaining were also observed with anti-CCL28 Ab. Discussion: Taken together, topical application of neutralizing anti-CCL28 Ab improved dorsal skin wound healing by reducing CCR10 activation and inflammation in part by preventing eNOS downregulation, increasing VEGF production, and restoring angiogenesis. These results indicate anti-CCL28 Ab has significant potential as a therapeutic strategy for treatment of chronic non-healing diabetic skin wounds such as DFUs.

[Role of the CCL28-CCR10 pathway in monocyte migration in rheumatoid arthritis].

OBJECTIVE: To examine the expression of chemokine receptor CCR10 on monocytes/macrophages in the joints of patients with rheumatoid arthritis (RA), and to investigate the role of chemokine CCL28 and its receptor CCR10 in the migration of RA monocytes and its mechanism. METHODS: The expression of CCR10 in synovial tissues from 8 RA patients, 4 osteoarthritis (OA) patients, and 4 normal controls was analyzed by immunohistochemistry, and cell staining was scored on a 0-5 scales. Flow cytometry was used to measure the percentage of CCR10 positive cells in CD14+ monocytes from peripheral blood of 26 RA patients and 20 healthy controls, as well as from synovial fluid of 15 RA patients. The chemotactic migration of monocytes from RA patients and healthy controls in response to CCL28 was evaluated using an in vitro Transwell system. Western blotting was conducted to assess phosphorylation of the extracellular signal-regulated kinase (ERK) and protein kinase B (Akt) pathways in RA monocytes upon CCL28 treatment. RESULTS: CCR10 was predominantly expressed in RA synovial lining cells and sublining macrophages, endothelial cells, and lymphocytes. CCR10 expression was significantly increased on lining cells and sublining macrophages in RA synovial tissue compared with OA and normal synovial tissue (both P < 0.01). The patients with RA had markedly elevated expression of CCR10 on peripheral blood CD14+ monocytes compared with the healthy controls [(15.6±3.0)% vs. (7.7±3.8)%, P < 0.01]. CCR10 expression on synovial fluid monocytes from the RA patients was (32.0±15.0)%, which was significantly higher than that on RA peripheral blood monocytes (P < 0.01). In vitro, CCL28 caused significant migration of CD14+ monocytes from peripheral blood of the RA patients and the healthy controls at concentrations ranging from 10-100 µg/L (all P < 0.01). The presence of neutralizing antibody to CCR10 greatly suppressed CCL28-driven chemotaxis of RA monocytes (P < 0.01). Stimulation of RA monocytes with CCL28 induced a remarkable increase in phosphorylation of ERK and Akt (both P < 0.05). ERK inhibitor (U0126) and phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002) strongly reduced the migration of RA monocytes in response to CCL28 (both P < 0.01). CONCLUSION: RA patients had increased CCR10 expression on peripheral blood, synovial fluid, and synovial tissue monocytes/macrophages. CCL28 ligation to CCR10 promoted RA monocyte migration through activation of the ERK and PI3K/Akt signaling pathways. The CCL28-CCR10 pathway could participate in monocyte recruitment into RA joints, thereby contributing to synovial inflammation and bone destruction.

Roles of CCR10/CCL27-CCL28 axis in tumour development: mechanisms, diagnostic and therapeutic approaches, and perspectives.

Cancer is now one of the major causes of death across the globe. The imbalance of cytokine and chemokine secretion has been reported to be involved in cancer development. Meanwhile, CC chemokines have received considerable interest in cancer research. CCR10, as the latest identified CC chemokine receptor (CCR), has been implicated in the recruitment and infiltration of immune cells, especially lymphocytes, into epithelia such as skin via ligation to two ligands, CCL27 and CCL28. Other than homoeostatic function, several mechanisms have been shown to dysregulate CCR10/CCL27-CCL28 expression in the tumour microenvironment. As such, these receptors and ligands mediate T-cell trafficking in the tumour microenvironment. Depending on the types of lymphocytes recruited, CCR10/CCL27-CCL28 interaction has been shown to play conflicting roles in cancer development. If they were T helper and cytotoxic T cells and natural killer cells, the role of this axis would be tumour-suppressive. In contrast, if CCR10/CCL27-CCL28 recruited regulatory T cells, cancer-associated fibroblasts or myeloid-derived suppressor cells, it would lead to tumour progression. In addition to the trafficking of lymphocytes and immune cells, CCR10 also leads to the migration of tumour cells or endothelial cells (called angiogenesis and lymphangiogenesis) to promote tumour metastasis. Furthermore, CCR10 signalling triggers tumour-promoting signalling such as PI3K/AKT and mitogen-activated protein kinase/extracellular signal-regulated kinase, resulting in tumour cell growth. Since CCR10/CCL27-CCL28 is dysregulated in the tumour tissues, it is suggested that analysis and measurement of them might predict tumour development. Finally, it is hoped using therapeutic approaches based on this axis might increase our knowledge to overcome tumour progression.

CCR10/CCL27 crosstalk regulates cell metastasis via PI3K-Akt signaling axis in non-small-cell lung cancer.

Previous research has shown that CCR10 acts as a vital oncogene in the progression of multiple malignancies. However, its effect on the treatment of non-small-cell lung cancer (NSCLC) has not been investigated. Current research examined the effect of CCR10 on the cellular survival and migration of NSCLC, and the modulation of cell death and found that the expression levels of CCR10 and CCL27 (ligand) were highly upregulated in human NSCLC tissue and cell lines, A549 and H157, compared with adjacent normal lung specimens. MTT and colony formation assays revealed that the blockage of CCR10 inhibited the multiplication and survival of A549 and H157 cells. Further study showed that metastasis-relevant VEGF-C/D, MMP-2/9, TIMP-1/2 were also regulated via CCR10 activation. Notably, increased NF-κB levels were detected in cells with activated CCR10, whereas the levels of NF-κB decreased in cells with blocked CCR10. Finally, the recovery of NF-κB expression counteracted the suppressive influence of CCR10 blockage on NSCLC cell survival, migration, and invasion. These results improved our knowledge understanding the molecular mechanisms of CCR10-CCL27 in progression of NSCLC.

SARS-CoV2 pneumonia recovery is linked to expansion of innate lymphoid cells type 2 expressing CCR10.

Accelerate lung repair in SARS-CoV-2 pneumonia is essential for pandemic handling. Innate lymphoid cells (ILCs) are likely players, given their role in mucosal protection and tissue homeostasis. We studied ILC subpopulations at two time points in a cohort of patients admitted in the hospital due to SARS-CoV-2 infection. COVID-19 patients with moderate/severe respiratory failure featured profound depletion of circulating ILCs at hospital admission, in agreement with overall lymphocyte depletion. However, ILCs recovered in direct correlation with lung function improvement as measured by oxygenation index and in negative association with inflammatory and lung/endothelial damage markers like RAGE. While both ILC1 and ILC2 expanded, ILC2 showed the most striking phenotype changes, with CCR10 upregulation in strong correlation with these parameters. Overall, CCR10+ ILC2 emerge as relevant contributors to SARS-CoV-2 pneumonia recovery.

CCL27 Signaling in the Tumor Microenvironment.

Chemokines are a group of small proteins which play an important role in leukocyte migration and invasion. They are also involved in the cellular proliferation and migration of tumor cells.Chemokine CCL27 (cutaneous T cell-attracting chemokine, CTACK) is mainly expressed by keratinocytes of the normal epidermis. It is well known that this chemokine plays an important role in several inflammatory diseases of the skin, such as atopic dermatitis, contact dermatitis, and psoriasis. Moreover, several studies have shown an association between CCL27 expression and a variety of neoplasms including skin cancer.In this chapter, we address the role of chemokine CCL27 in the tumor microenvironment in the most relevant cancers of the skin and other anatomical locations. We also make a brief comment on future perspectives and the potential relation of CCL27 with different immunotherapeutic modalities.

Systematic Assessment of Chemokine Signaling at Chemokine Receptors CCR4, CCR7 and CCR10.

Chemokines interact with chemokine receptors in a promiscuous network, such that each receptor can be activated by multiple chemokines. Moreover, different chemokines have been reported to preferentially activate different signalling pathways via the same receptor, a phenomenon known as biased agonism. The human CC chemokine receptors (CCRs) CCR4, CCR7 and CCR10 play important roles in T cell trafficking and have been reported to display biased agonism. To systematically characterize these effects, we analysed G protein- and ß-arrestin-mediated signal transduction resulting from stimulation of these receptors by each of their cognate chemokine ligands within the same cellular background. Although the chemokines did not elicit ligand-biased agonism, the three receptors exhibited different arrays of signaling outcomes. Stimulation of CCR4 by either CC chemokine ligand 17 (CCL17) or CCL22 induced ß-arrestin recruitment but not G protein-mediated signaling, suggesting that CCR4 has the potential to act as a scavenger receptor. At CCR7, both CCL19 and CCL21 stimulated G protein signaling and ß-arrestin recruitment, with CCL19 consistently displaying higher potency. At CCR10, CCL27 and CCL28(4-108) stimulated both G protein signaling and ß-arrestin recruitment, whereas CCL28(1-108) was inactive, suggesting that CCL28(4-108) is the biologically relevant form of this chemokine. These comparisons emphasize the intrinsic abilities of different receptors to couple with different downstream signaling pathways. Comparison of these results with previous studies indicates that differential agonism at these receptors may be highly dependent on the cellular context.

Innate lymphoid cells regulate radiation-induced skin damage via CCR10 signaling.

OBJECTIVE: To assess the role of CCR10 in innate lymphoid cells (ILCs) response in radiation-induced skin damage. MATERIAL AND METHODS: CCR10+/- and CCR10-/- mice were treated with either a single dose of 5 Gy or 5 Gy everyday for 6 days with a total dose of 30 Gy with X-ray. ILCs from the skin were isolated and analyzed by flow cytometry 3 and 10 days after irradiation. A mouse model of radio-dermatitis was used to assess the skin damage 10 days after 6 × 5 Gy irradiation. RESULTS: Skin ILCs were decreased in both CCR10+/- and CCR10-/- mice 3 days after single irradiation (p < .05). However, the skin inflammation disappeared and ILCs returned to normal levels 10 days after single irradiation. ILCs of both genotypes were also decreased after 6 × 5 Gy irradiation, but the percentage of skin ILCs in CCR10-/- mice was lower than that in CCR10+/- mice 10 days after irradiation. The immunohistochemistry results showed that CCR10-/- mice had more severe skin inflammation than CCR10+/- mice. CONCLUSION: CCR10-/- mice had lower percentages of ILCs and more skin damage than CCR10+/- mice after irradiation. These findings indicate that skin ILCs are regulated by CCR10, which might be a potential target for reducing the radio-dermatitis.

Comparison of biochemical parameters and chemokine levels in pleural fluid between patients with anergic and non-anergic tuberculous pleural effusion.

Pleural fluid (PF) immune response in anergic tuberculous pleural effusion (TPE) patients is poorly understood. This study aimed to compare PF biochemical parameters and chemokine levels between anergic and non-anergic TPE patients. Chemokine arrays, cytokine measurements, and flow cytometry were performed in 58 patients (TPE [non-anergic (n = 32) and anergic (n = 10)] and malignant pleural effusion (MPE) [n = 16]). PF adenosine deaminase 2 (ADA2) levels were significantly lower in anergic TPE patients than in non-anergic TPE patients (p = 0.048). Among the 40 chemokines tested, PF CCL27 levels were significantly higher in anergic TPE patients than in non-anergic TPE and MPE patients (p < 0.001). The percentage of CD4+CCR10+T cells in PF was higher in anergic TPE patients than in non-anergic TPE and MPE patients (p = 0.001). We reported here that CCL27/CCR10 interactions might contribute to pathophysiology in anergic TPE. PF CCL27 and CD4+CCR10+T cells may help in diagnosing TPE in patients with moderate elevation of PF ADA levels.

CCL28-induced CCR10/eNOS interaction in angiogenesis and skin wound healing.

Chemokines and their receptors play important roles in vascular homeostasis, development, and angiogenesis. Little is known regarding the molecular signaling mechanisms activated by CCL28 chemokine via its primary receptor CCR10 in endothelial cells (ECs). Here, we test the hypothesis that CCL28/CCR10 signaling plays an important role in regulating skin wound angiogenesis through endothelial nitric oxide synthase (eNOS)-dependent Src, PI3K, and MAPK signaling. We observed nitric oxide (NO) production in human primary ECs stimulated with exogenous CCL28, which also induced direct binding of CCR10 and eNOS resulting in inhibition of eNOS activity. Knockdown of CCR10 with siRNA lead to reduced eNOS expression and tube formation suggesting the involvement of CCR10 in EC angiogenesis. Based on this interaction, we engineered a myristoylated 7 amino acid CCR10-binding domain (Myr-CBD7) peptide and showed that this can block eNOS interaction with CCR10, but not with calmodulin, resulting in upregulation of eNOS activity. Importantly, topical administration of Myr-CBD7 peptide on mouse dermal wounds not only blocked CCR10-eNOS interaction, but also enhanced expression of eNOS, CD31, and IL-4 with reduction of CCL28 and IL-6 levels associated with improved wound healing. These results point to a potential therapeutic strategy to upregulate NO bioavailability, enhance angiogenesis, and improve wound healing by disrupting CCL28-activated CCR10-eNOS interaction.

CCR10 expression is required for the adjuvant activity of the mucosal chemokine CCL28 when delivered in the context of an HIV-1 Env DNA vaccine.

An effective prophylactic vaccine targeting HIV must induce a robust humoral response and must direct the bulk of this response to the mucosa-the primary site of HIV transmission. The chemokine, CCL28, is secreted by epithelial cells at mucosal surfaces and recruits' cells expressing its receptor CCR10. CCR10 is predominantly expressed by IgA + ASCs. We hypothesized that co-immunization with plasmid DNA encoding consensus envelope antigens with plasmid-encoded CCL28 would enhance anti-HIV IgA responses at mucosal surfaces. Indeed, animals receiving pCCL28 and pEnvA/C had significantly increased HIV-specific IgA in fecal extract. Surprisingly, CCL28 co-immunization induced a significant increase in anti-HIV IgG in the serum in mice compared to those receiving pEnvA/C alone. These robust antibody responses were not associated with changes in the frequency of germinal center B cells but depended upon the expression of CCR10, as these responses we abolished in CCR10-deficient animals. Finally, immunization with CCL28 led to increased frequencies in HIV-specific CCR10 + and CCR10 + IgA + B cells in the small intestine and Peyer's patches of vaccinated animals as compared to those receiving pEnvA/C alone. These data indicate that CCL28 administration can enhance antigen-specific humoral responses systemically and at mucosal surfaces.

CCL28 promotes locomotor recovery after spinal cord injury via recruiting regulatory T cells.

BACKGROUND: Chemokines play a key role in post-traumatic inflammation and secondary injury after spinal cord injury (SCI). CCL28, the chemokine CC-chemokine ligand 28, is involved in the epithelial and mucosal immunity. However, whether CCL28 participates in the physiopathologic processes after SCI remains unclear. RESULTS: CCL28 is upregulated in the spinal cord after SCI. In addition, neutralizing antibodies against IL-1ß or TNF-α, or treatment of ML120B, a selective inhibitor of IKK-ß, remarkably decrease CCL28 upregulation, suggesting that CCL28 upregulation relies on NF-κB pathway activated by IL-1ß and TNF-α after SCI. Moreover, CD4+CD25+FOXP3+ regulatory T (Treg) cells that express CCR10, a receptor of CCL28, are enriched in the spinal cord after SCI. We further demonstrate that the spinal cord recruits Treg cells through CCL28-CCR10 axis, which in turn function to suppress immune response and promote locomotor recovery after SCI. In contrast, neutralizing CCL28 or CCR10 reduces Treg cell recruitment and delays locomotor recovery. METHODS: The neutralizing antibodies and recombinant CCL28 were injected intraspinally into the mice prior to SCI, which was established via hemitransection. RT-qPCR analysis was performed to determine transcript level, and Western blot analysis and ELISA assay were used to detect protein expression. Immune cells were analyzed by flow cytometry and visualized by immunofluorescence. The chemotaxis was assessed by in vitro transwell migration assay. The mouse locomotor activity was assessed via the Basso Mouse Scale (BMS) system. CONCLUSIONS: These results indicate that NF-κB pathway-regulated CCL28 production plays a protective role after SCI through recruiting CCR10-expressing and immunosuppressive Treg cells, and suggest that interfering CCL28-CCR10 axis might be of potential clinical benefit in improving SCI recovery.

Expression of CCR3 and CCR4 Suggests a Poor Prognosis in Mycosis Fungoides and Sézary Syndrome.

Tumor cells in cutaneous T-cell lymphoma express limited numbers of chemokine receptors. We investigated the expression patterns of CXCR3, CCR3, CCR4 and CCR10 in mycosis fungoides, Sézary syndrome, lym-phomatoid papulosis and anaplastic large cell lymphoma in 121 skin biopsy samples. CXCR3 was expressed in 86% of mycosis fungoides cases but in no anaplastic large cell lymphoma cases. CCR3 was expressed in 73% of cases of CD30+ lymphoproliferative disorders such as lymphomatoid papulosis and anaplastic large cell lymphoma. Mycosis fungoides/Sézary syndrome patients with high CCR3 or CCR4 expression had a poorer survival prognosis than mycosis fungoides/Sézary syndrome patients whose tumor cells did not express these receptors. CCR10 was expressed in 50% of mycosis fungoides/Sézary syndrome cases and in 13% of cases with CD30+ lym-phoproliferative disorders. These results suggest that differential patterns of CXCR3, CCR3, CCR4 and CCR10 expression are useful for the diagnosis of cutaneous T-cell lymphoma. Moreover, expression of CCR3 or CCR4 suggests a poor prognosis in mycosis fungoides/Sézary syndrome.

Evaluation of Lymphocyte Migration to Induced Paederus Dermatitis: An Experimental Study in Rats.

BACKGROUND: Paederus dermatitis (PD) is a blistering disorder that is caused by a small insect of the genus Paederus, especially Paederus fuscipes. This study aimed to investigate the reaction of the adaptive immune system regarding the recruitment of CD3, CCR4, and CCR10 markers, which are specifically expressed on the surface of T lymphocytes. MATERIALS AND METHODS: In this experimental study, 24 female rats were divided into two groups: the test and the negative control. In the test group, PD was induced by making insects in contact with shaved rat skin. Biopsies were obtained 24, 72, and 120 h after induction. In the negative control group, physiological saline was applied. Specimens were evaluated by immunohistochemical staining method. Antibodies against CD3, CCR4, and CCR10 were used. Distribution and staining intensities of CD3, CCR4, and CCR10 markers were estimated by the H-score index and findings were analyzed using the Kruskal-Wallis and Wilcoxon statistical tests. RESULTS: Based on the results of immunohistochemistry, the expression of CD3, CCR4, and CCR10 in the test group at 24, 72, and 120 h compared to the control group showed significant increase (P = 0.0006, P = 0.001, and P < 0.0001), respectively. The peak of expression of all markers was at 72 h after exposure. Hematoxylin and eosin staining also confirmed the fact that the majority of the lymphocyte infiltration occurred at 72 h postexposure. CONCLUSION: The expression of CD3, CCR4, and CCR10 on cells present in PD lesions could indicate that T-lymphocytes are recruited to the site of inflammation by chemokine-chemokine receptor interactions and hence provide evidence for the response by the adaptive immune system following a PD.