Expression Of CC Chemokine Receptor Research Articles

Nicotinamide Riboside Augments Human Macrophage Migration via SIRT3-Mediated Prostaglandin E2 Signaling.

NAD+ boosting via nicotinamide riboside (NR) confers anti-inflammatory effects. However, its underlying mechanisms and therapeutic potential remain incompletely defined. Here, we showed that NR increased the expression of CC-chemokine receptor 7 (CCR7) in human M1 macrophages by flow cytometric analysis of cell surface receptors. Consequently, chemokine ligand 19 (CCL19, ligand for CCR7)-induced macrophage migration was enhanced following NR administration. Metabolomics analysis revealed that prostaglandin E2 (PGE2) was increased by NR in human monocytes and in human serum following in vivo NR supplementation. Furthermore, NR-mediated upregulation of macrophage migration through CCL19/CCR7 was dependent on PGE2 synthesis. We also demonstrated that NR upregulated PGE2 synthesis through SIRT3-dependent post-transcriptional regulation of cyclooxygenase 2 (COX-2). The NR/SIRT3/migration axis was further validated using the scratch-test model where NR and SIRT3 promoted more robust migration across a uniformly disrupted macrophage monolayer. Thus, NR-mediated metabolic regulation of macrophage migration and wound healing may have therapeutic potential for the topical management of chronic wound healing.

Association of CCR6 functional polymorphisms with Primary Biliary Cholangitis

The biliary epithelial cells release CC chemokine receptor 6 (CCR6) ligand 20 (CCL20), leading to recruitment of CCR6+ T cells and subsequent infiltration into the biliary epithelium in primary biliary cholangitis patients. Previous genome-wide multi-national meta-analysis, including our Han Chinese cohort, showed significant association of CCR6 and CCL20 single nucleotide polymorphisms (SNP) with PBC. We report here that significantly associated SNPs, identified in the CCR6 locus based on our Han Chinese genome-wide association study, can be separated into “protective” and “risk” groups, but only “risk” SNPs were confirmed using a separate Han Chinese PBC cohort. Only weak association of CCL20 SNPs was observed in Han Chinese PBC cohorts. Fine-mapping and logistical analysis identified a previously defined functional variant that, leads to increased CCR6 expression, which contributed to increased genetic susceptibility to PBC in Han Chinese cohort.

CCR5 regulates Aβ1-42-induced learning and memory deficits in mice

C–C chemokine receptor 5 (CCR5) is a chemokine receptor involved in immune responses and a co-receptor for HIV infection. Recently, CCR5 has also been reported to play a role in synaptic plasticity, learning and memory, and cognitive deficits associated with normal aging, traumatic brain injury (TBI), and HIV-associated neurocognitive disorder (HAND). In contrast, the role of CCR5 in cognitive deficits associated with other disorders, including Alzheimer’s disease (AD), is much less understood. Studies have reported an increase in expression of CCR5 or its ligands in both AD patients and AD rodent models, suggesting a correlation between AD and CCR5 expression. However, whether blocking CCR5 in specific brain regions, such as the hippocampus, could improve memory deficits in AD mouse models is unknown. To study the potential causal role of CCR5 in cognitive deficits in AD, we injected soluble Aβ1-42 or a control (Aβ42-1) oligomers in the dorsal CA1 region of the hippocampus and found that Aβ1-42 injection resulted in severe memory impairment in the object place recognition (OPR) and novel object recognition (NOR) tests. Aβ1-42 injection caused an increase in Ccr5, Ccl3, and Ccl4 in the dorsal hippocampus, and the expression levels of CCR5 and its ligands remained elevated at 2 weeks after Aβ1-42 injection. Knocking down Ccr5 in the CA1 region of dorsal hippocampus reversed the increase in microglia number and size in dorsal CA1 and rescued memory deficits. These results indicate that CCR5 plays an important role in modulating Aβ1-42-induced learning and memory deficits, and suggest that CCR5 antagonists may serve as a potential treatment to improve cognitive deficits associated with AD.

CC chemokine receptors are prognostic indicators of gastric cancer and are associated with immune infiltration

BackgroundCC chemokine receptors are responsible for regulating the tumor microenvironment (TME) and participating in carcinogenesis and tumor advancement. However, no functional study has investigated CC chemokine receptors in gastric cancer (GC) prognosis, risk, immunotherapy, or other treatments.MethodsWe conducted a bioinformatics analysis on GC data using online databases, including the Human Protein Atlas (HPA), Kaplan-Meier (KM) plotter, GeneMANIA, MethSurv, the University of ALabama at Birmingham CANcer (UALCAN) Data Analysis Portal, Gene Set Cancer Analysis (GSCA), cBioportal, and Tumor IMmune Estimation Resource (TIMER).ResultsWe noted that CC chemokine receptor expression correlated with survival in GC. CC chemokine receptor expression was also strongly linked to different tumor-infiltrating immune cells. Additionally, CC chemokine receptors were found to be broadly drug-resistant in GC.ConclusionOur study identifed CC chemokine receptor expression helped in predicting the prognosis of patients diagnosed with GC. The expression level of the CC chemokine receptors was also positively related to multiple tumor-infiltrating lymphocytes (TILs). These findings provide evidence to monitor patients with GC using CC chemokine receptors, which can be used as an effective biomarker for predicting the disease prognosis and be regarded as a therapeutic target for modulating the tumor immune microenvironment.

Chemotherapy-Induced Cellular Senescence Promotes Stemness of B-Cell Non-Hodgkin's Lymphoma Via CCL21/CCR7/NF-Κb Signaling Activation

A bs tract: Resistance to existing therapies is currently the main reason for treatment failure in refractory/relapsed B-NHL patients, and treatment-induced cellular senescence (TIS) is one of the important mechanisms of tumor drug resistance. In our previous studies, we found that the anti-apoptotic and tumorigenic ability of chemotherapy-induced senescent B-NHL cells was significantly enhanced, and the survival period of tumor-bearing mice was significantly shortened, but the mechanism is still unclear. In this study, we identified the C-C chemokine receptor 7 (CCR7) by single-cell RNA-sequencing, which showed significant differential expression in senescent B-NHL cell population and non-senescent cell population. CCL21/CCR7 dependent B-NHL-TIS initiates downstream transcription of NF-κB, thereby enhancing phenotypic and functional stemness features, including the up-regulation of stemness markers, CD34, CD44, LGR5, CD150, and stronger colony forming ability and cell sphere forming ability. The analysis results of clinical patients showed that the expression of CCR7 in the peripheral blood of B-NHL patients was significantly higher than that of healthy normal people and leukemia patients. The expression of CCR7 in peripheral blood of B-NHL patients before treatment was significantly lower than that of relapsed patients, and The expression in abnormal B lymphocyte population is significantly higher than that in normal T and B lymphocyte population, high expression of CCR7 is associated with recurrence and poor prognosis in B-NHL patients. Targeting CCR7/NF-κB signaling has positive significance in inhibiting the acquisition and maintenance of stemness in aging B-NHL cells and preventing tumor drug resistance and recurrence. K ey words: Stress-induced senescence; CCR7; CCL21; senescence related stemness; B-cell non-Hodgkin's lymphoma

Mogamulizumab for Refractory CD3-CD4+ Lymphocytic-Variant Hypereosinophilic Syndrome

OCS Introduction. Lymphocytic hypereosinophilic syndrome (L-HES) is an indolent T-cell lymphoproliferative disorder characterized by chronic blood hypereosinophilia and eosinophil-related organ damage secondary to the production of eosinophilopoietic cytokines (including interleukin-5) by clonal T cells, among which CD3-CD4+ T cells are the most frequent subset. Both high exposure to oral corticosteroids (OCS), failure and/or toxicity of OCS-sparing therapies and the risk of progression to AITL (up to 5 - 10% of patients) underscore the unmet need for novel treatments targeting the clonal T-cells (and not only eosinophils). We previously reported that peripheral CD3-CD4+ T cells express C-C chemokine receptor 4 (CCR4). We hypothesized that mogamulizumab, an afucosylated monoclonal antibody targeting CCR4-positive cells through antibody-dependent cellular cytotoxicity, could be beneficial for refractory L-HES patients. Methods. We assessed both the expression of CCR4 by clonal CD3-CD4+ T cells in the skin and lymph nodes as well as theability of mogamulizumab to induce ADCC in vitro on sorted CD3-CD4+ T cells from two L-HES patients, using homologous sorted NK cells as effector cells. We then set up a compassionate use program of intravenous mogamulizumab (1.0 mg/kg weekly for the first 28-day cycle, then on days 1 and 15 from subsequent cycles) for refractory L-HES patients despite treatment with immunosuppressants and/or biologics targeting the IL-5 pathway. Complete blood response was defined as complete if the subset of CD3-CD4+ T cells was <0.5% of all lymphocytes, or as partial if the number of CD3-CD4+ T cells decreased by >50% without reaching complete response at the time of the last perfusion. Results. Significant CCR4 expression on tissue CD4+ T cells was evidenced, and NK cells indeed led to the lysis of CD3-CD4+ T-cells coated with mogamulizumab (10 μg/ml). Four patients with refractory L-HES (despite treatment with high-dose OCS, n=4; pegylated interferon alpha 2a; n=3; or mepolizumab, a single patient) and active disease entered the compassionate use program (Table 1). Despite substantial tapering in OCS doses, all cases showed clinical response. Blood responses (either partial or complete, two patients each) and sharp decrease in absolute eosinophil counts (AEC) were reported in all patients ( Figure 1). Both patients with lymph node enlargement rapidly improved. Although prolonged blood and clinical responses have been observed after discontinuation of mogamulizumab, relapses still occurred preceded by an increase in T-cell clone size and absolute eosinophil count (AEC). Rechallenging a relapse with a short course of mogamulizumab resulted in a complete remission for a further 12 months in case #1. In terms of safety, mogamulizumab was discontinued in two patients due to adverse events: one for recurrent infusion-related reactions and one for infectious events. Lymphopenia was reported in 3/4 cases but was preexisting in the 3 cases. Case #4 reported a serious adverse event related to a tumor lysis syndrome leading to the massive release of IL-5 and subsequent rebound of AEC. Hence lower doses of mogamulizumab, transient use of OCS and/or mepolizumab should be considered in patients with very high number of CD3-CD4+ T cells who are about to start mogamulizumab. Discussion. Mogamulizumab represents a new therapeutic option for L-HES patients with severe refractory HES, T-cell clone related symptoms and/or large or rapidly increasing clonal T-cell counts. In patients who achieve complete remission under mogamulizumab, whether fixed repeated (e.g., every 6 months) infusions could be beneficial to prevent relapses and transformation into high-grade lymphoma remains to be investigated. Figure legend. Hematological responses to mogamulizumab in four refractory lymphocytic HES patients. oral corticosteroids; pINF-α: pegylated interferon alpha; MEPO: mepolizumab; MOGA: mogamulizumab.

CCR5 promoter polymorphisms associated with nonsmall cell lung cancer.

C-C chemokine receptor 5 (CCR5) plays a crucial role in the regulation of immune cell activation and migration as well as the progression of many cancers. We performed an in silico analysis using public data resources and found that the lung cancer patients with higher CCR5 expression had a notably better overall survival than those with lower CCR5 expression patients and CCR5 expression level is positive correlated with the infiltration of immune cells, such as B, CD8+ T and CD4+ T cells, in both lung adenocarcinoma and lung squamous cell cancer. In the present study, we investigated the association between the promoter polymorphism of CCR5 and nonsmall cell lung cancer (NSCLC). A case-control study of 449 NSCLC patients and 516 controls of Chinese Han population was conducted, along with polymorphism detection using a sequencing method. A dual-luciferase reporter assay system was used to analyse the transcriptional activity of CCR5 promoter variations. Our results showed that the frequency of rs1799987-AA was significantly higher in the NSCLC group than in the controls in recessive model (p=.007, OR=1.66 95% confidence interval [CI]: 1.14-2.40, adjusted by sex and age); the G allele showed a significant associated with NSCLC in dominant model (p=.003, OR=1.64, 95%CI: 1.18-2.28, adjusted by sex and age). Compared with haplotype H1 rs2227010-rs2734648-rs1799987-rs1799988-rs1800023-rs1800024: A-T-G-T-G-C, haplotype H5: A-G-G-T-G-C increased the risk of NSCLC by over 10-fold (p<.0001, OR=16.09, 95%CI: 5.37-48.20, adjusted by sex and age) and notably depressed the transcriptional activity of the CCR5 promoter in 293T, A549, H1299 and HeLa cells. In conclusion, CCR5 promoter polymorphisms are significantly associated with NSCLC by affecting the transcriptional activity of the CCR5 promoter.

Protective effect of Liujing Toutong Tablets on rats with permanent cerebral ischemia via NF-κB signaling pathway

This study investigated the neuroprotective effects and underlying mechanism of Liujing Toutong Tablets(LJTT) on a rat model of permanent middle cerebral artery occlusion(pMCAO). The pMCAO model was established using the suture method. Eighty-four male SPF-grade SD rats were randomly divided into a sham operation group, a model group, a nimodipine group(0.020 g·kg~(-1)), and high-, medium-, and low-dose LJTT groups(2.8, 1.4, and 0.7 g·kg~(-1)). The Longa score, adhesive removal test and laser speckle contrast imaging technique were used to evaluate the degree of neurological functional impairment and changes in local cerebral blood flow. The survival and mortality of rats in each group were recorded daily. After seven days of continuous administration following the model induction, the rats in each group were euthanized, and brain tissue and blood samples were collected for corresponding parameter measurements. Nissl staining was used to examine pathological changes in brain tissue neurons. The levels of tumor necrosis factor-alpha(TNF-α), interleukin-6(IL-6), IL-1β, vascular endothelial growth factor(VEGF), calcitonin gene-related peptide(CGRP), beta-endorphin(β-EP), and endogenous nitric oxide(NO) in rat serum were measured using specific assay kits. The entropy weight method was used to analyze the weights of various indicators. The protein expression levels of nuclear factor kappa-B(NF-κB), inhibitor kappaB alpha(IκBα), phosphorylated IκBα(p-IκBα), and phosphorylated inhibitor of NF-κB kinase alpha(p-IKKα) in brain tissue were determined using Western blot. Immunohistochemistry was used to detect the protein expression of chemokine-like factor 1(CKLF1) and C-C chemokine receptor 5(CCR5) in rat brain tissue. Compared with the sham operation group, the model group showed significantly higher neurological functional impairment scores, prolonged adhesive removal time, decreased cerebral blood flow, increased neuronal damage, reduced survival rate, significantly increased levels of TNF-α, IL-1β, IL-6, CGRP, and NO in serum, significantly decreased levels of VEGF and β-EP, significantly increased expression levels of NF-κB p65, p-IκBα/IκBα, and p-IKKα in rat brain tissue, and significantly upregulated protein expression of CKLF1 and CCR5. Compared with the model group, the high-dose LJTT group significantly improved the neurological functional score of pMCAO rats after oral administration for 7 days. LJTT at all doses significantly reduced adhesive removal time and restored cerebral blood flow. The high-and medium-dose LJTT groups significantly improved neuronal damage. The LJTT groups at all doses showed reduced levels of TNF-α, IL-1β, IL-6, CGRP, and NO in rat serum, increased VEGF and β-EP levels, and significantly decreased expression levels of NF-κB p65, p-IκBα/IκBα, p-IKKα, and CCR5 protein in rat brain tissue. The entropy weight analysis revealed that CGRP and β-EP were significantly affected during the model induction, and LJTT exhibited a strong effect in reducing the release of inflammatory factors such as TNF-α and IL-1β. LJTT may exert a neuroprotective effect on rats with permanent cerebral ischemia by reducing neuroinflammatory damage, and its mechanism may be related to the inhibition of the NF-κB signaling pathway and the regulation of the CKLF1/CCR5 axis. Additionally, LJTT may exert certain analgesic effects by reducing CGRP and NO levels and increasing β-EP levels.

GLUT1 Mediates the Metabolic Reprogramming and Inflammation of CCR2$^+$ Monocytes/Macrophages from Patients with DCM.

Macrophages expressing CC chemokine receptor 2 (CCR2) possess characteristics and performance akin to M1 polarized macrophages, which promote inflammation. Advanced heart failure (HF) patients with higher abundance of CCR2+ macrophages are more likely to experience adverse remodeling. The precise mechanism of CCR2+ macrophages in how they affect the progression of dilated cardiomyopathy remains unknown. Cardiac biopsy samples from dilated cardiomyopathy patients (DCM) were used for immunohistochemistry and immunofluorescence staining. PCR is employed to identify the IL-1β, IL-6, TNF-α, TGF-β, MMP2, MMP9, PKM1, PKM1, GLUT1, GLUT2, GLUT3, GLUT4, PDK1, PFKFB3, PFK1 and HK2 mRNA expression of CCR2+ monocytes/macrophages from the peripheral blood of DCM patients. Seahorse was used to evaluate the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of CCR2+ monocytes/macrophages. 2-DG was used to simulate a lack of glucose. Lentivirus containing GLUT1 inhibitory sequence was used to knockdown GLUT1 gene expression of CCR2+ monocytes/macrophages. Western Blot and immunofluorescence staining was used to evaluate the expression of NLRP3. Immunostaining results of cardiac biopsy tissue from dilated cardiomyopathy (DCM) patients demonstrated that the progression to HF was associated with an increase in the number of CCR2+ macrophages. PCR results demonstrated that CCR2 monocytes and macrophages derived from the blood of DCM patients expressed elevated levels of inflammatory factors and up regulation of glycolysis related genes. In addition, OCR and glucose uptake experiments confirmed that increased glucose uptake of these cells was associated with greater inflammation and correlated with a worsening of cardiac function. limiting the glucose supply to CCR2+ monocytes and macrophages, or suppressing the activity of glucose transporter 1 (GLUT1) could reduce inflammation levels. These results suggest that CCR2+ monocytes and macrophages rely on metabolic reprogramming to trigger inflammatory response and contribute to myocardial injury and the progression of DCM.

Evaluation of Novel Targets, Including CC-Chemokine Receptor 4, in Adult T-Cell Acute Lymphoblastic Leukemia/Lymphoma: A Mayo Clinic Clinical and Pathologic Study.

Unlike B-cell acute lymphoblastic leukemia/lymphoma (ALL/LBL), there have been few therapeutic advances in T-cell ALL (T-ALL)/LBL, an aggressive ALL/LBL subtype. To perform a focused tissue array study to elucidate tumor markers of therapeutic potential in T-ALL/LBL. Using immunohistochemistry, we evaluated expression of leukemic antigens of interest, specifically CC-chemokine receptor 4 (CCR4), among others, on available remnant diagnostic material, including tumor tissue slides obtained from formalin-fixed, paraffin-embedded preserved tissues. Our analysis identified, for the first time, expression of CCR4 in T-ALL/LBL in 11 of 27 cases (40.7%) and confirmed common expression of BCL2, CD38, and CD47, as reported previously. We also identified the expression of CD123 in 4 of 26 cases (15.4%), whereas BCL6 and PDL1 were expressed in a small number of T-ALL/LBL cases. The potential novel target CCR4 was significantly more common in the Pre/Pro-T immunophenotypic subtype, 6 of 9 (66.7%, P = .01). No additional differences in clinical and epidemiologic variables were noted among positive or negative CCR4 cases. These findings support preclinical and clinical testing of therapies targeting CCR4, CD47, BCL2, CD38, and CD123 in T-ALL/LBL, and may help guide the development of targeted clinical trials in T-ALL/LBL, a rare disease in urgent need of novel therapies.

Therapeutic efficacy of the resorcylic acid lactone LL-Z1640-2 for adult T-cell leukaemia/lymphoma.

Adult T-cell leukaemia/lymphoma (ATL) remains incurable. The NF-κB and interferon regulatory factor 4 (IRF4) signalling pathways are among the critical survival pathways for the progression of ATL. TGF-β-activated kinase 1 (TAK1), an IκB kinase-activating kinase, triggers the activation of NF-κB. The resorcylic acid lactone LL-Z1640-2 is a potent irreversible inhibitor of TAK1/extracellular signal-regulated kinase 2 (ERK2). We herein examined the therapeutic efficacy of LL-Z1640-2 against ATL. LL-Z1640-2 effectively suppressed the in vivo growth of ATL cells. It induced in vitro apoptosis and inhibited the nuclear translocation of p65/RelA in ATL cells. The knockdown of IRF4 strongly induced ATL cell death while downregulating MYC. LL-Z1640-2 as well as the NF-κB inhibitor BAY11-7082 decreased the expression of IRF4 and MYC at the protein and mRNA levels, indicating the suppression of the NF-κB-IRF4-MYC axis. The treatment with LL-Z1640-2 also mitigated the phosphorylation of p38 MAPK along with the expression of CC chemokine receptor 4. Furthermore, the inhibition of STAT3/5 potentiated the cytotoxic activity of LL-Z1640-2 against IL-2-responsive ATL cells in the presence of IL-2. Therefore, LL-Z1640-2 appears to be an effective treatment for ATL. Further studies are needed to develop more potent compounds that retain the active motifs of LL-Z1640-2.

Downregulation of CCR5 on brain perivascular macrophages in simian immunodeficiency virus-infected rhesus macaques.

C-C chemokine receptor 5 (CCR5) is a major coreceptor for Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) cell entry; however, its role in brain pathogenesis is largely understudied. Thus, we sought to examine cell type-specific protein expression of CCR5 during SIV infection of the brain. We examined occipital cortical tissue from uninfected rhesus macaques and SIV-infected animals with or without encephalitis using immunohistochemistry and immunofluorescence microscopy to determine the number and distribution of CCR5-positive cells. An increase in the number of CCR5+ cells in the brain of SIV-infected animals with encephalitis was accounted for by increased CD3+CD8+ cells expressing CCR5, but not by increased CCR5+ microglia or perivascular macrophages (PVMs), and a concurrent decrease in the percentage of CCR5+ PVMs was observed. Levels of CCR5 and SIV Gag p28 protein expression were examined on a per-cell basis, and a significant, negative relationship was established indicating decreased CCR5 expression in productively infected cells. While investigating the endocytosis-mediated CCR5 internalization as a mechanism for CCR5 downregulation, we found that phospho-ERK1/2, an indicator of clathrin-mediated endocytosis, was colocalized with infected PVMs and that macrophages from infected animals showed significantly increased expression of clathrin heavy chain 1. These findings show a shift in CCR5-positive cell types in the brain during SIV pathogenesis with an increase in the number of CCR5+ CD8 T cells, and downregulated CCR5 expression on infected PVMs, likely through ERK1/2-driven, clathrin-mediated endocytosis.

CC-Chemokine Receptor-2 Expression in Osteoblasts Contributes to Cartilage and Bone Damage during Post-Traumatic Osteoarthritis.

In osteoarthritis (OA), bone changes are radiological hallmarks and are considered important for disease progression. The C-C chemokine receptor-2 (CCR2) has been shown to play an important role in bone physiology. In this study, we investigated whether Ccr2 osteoblast-specific inactivation at different times during post-traumatic OA (PTOA) progression improves joint structures, bone parameters, and pain. We used a tamoxifen-inducible Ccr2 inactivation in Collagen1α-expressing cells to obtain osteoblasts lacking Ccr2 (CCR2-Col1αKO). We stimulated PTOA changes in CCR2-Col1αKO and CCR2+/+ mice using the destabilization of the meniscus model (DMM), inducing recombination before or after DMM (early- vs. late-inactivation). Joint damage was evaluated at two, four, eight, and twelve weeks post-DMM using multiple scores: articular-cartilage structure (ACS), Safranin-O, histomorphometry, osteophyte size/maturity, subchondral bone thickness and synovial hyperplasia. Spontaneous and evoked pain were assessed for up to 20 weeks. We found that early osteoblast-Ccr2 inactivation delayed articular cartilage damage and matrix degeneration compared to CCR2+/+, as well as DMM-induced bone thickness. Osteophyte formation and maturation were only minimally affected. Late Collagen1α-Ccr2 deletion led to less evident improvements. Osteoblast-Ccr2 deletion also improved static measures of pain, while evoked pain did not change. Our study demonstrates that Ccr2 expression in osteoblasts contributes to PTOA disease progression and pain by affecting both cartilage and bone tissues.

CKLF1 interference alleviates IL‑1β‑induced inflammation, apoptosis and degradation of the extracellular matrix in chondrocytes via CCR5.

Osteoarthritis (OA) is a type of joint disease with a rising prevalence and incidence among the elderly across the global population. Chemokine-like factor 1 (CKLF1) is a human cytokine, which has been demonstrated to be involved in the progression of multiple human diseases. However, little attention has been paid to the impact of CKLF1 on OA. The present study was designed to identify the role of CKLF1 in OA and to clarify the regulatory mechanism. The expression levels of CKLF1 and its receptor CC chemokine receptor 5 (CCR5) were examined by reverse transcription-quantitative PCR (RT-qPCR) and western blotting. A Cell Counting Kit-8 assay was used to estimate cell viability. The levels and expression of inflammatory factors were determined by ELISA and RT-qPCR, respectively. Apoptosis was investigated by TUNEL assays and the protein levels of apoptosis-related factors were analyzed by western blotting. RT-qPCR and western blotting were used to examine the expression of extracellular matrix (ECM) degradation-associated proteins and ECM components. Dimethylmethylene blue analysis was used to analyze the production of soluble glycosamine sulfate additive. A co-immunoprecipitation assay was used to confirm the protein interaction between CKLF1 and CCR5. The results revealed that CKLF1 expression was increased in IL-1β-exposed murine chondrogenic ATDC5 cells. Furthermore, CKLF1 silencing enhanced the viability of IL-1β-induced ATDC5 cells, while inflammation, apoptosis and degradation of the ECM were reduced. Additionally, CKLF1 knockdown led to decreased CCR5 expression in IL-1β-challenged ATDC5 cells, and CKLF1 bound with CCR5. The enhanced viability, as well as the suppressed inflammation, apoptosis and degradation of the ECM, following CKLF1 knockdown in the IL-1β-induced ATDC5 cells were all restored after CCR5 was overexpressed. In conclusion, CKLF1 might serve a detrimental role in the development of OA by targeting its receptor CCR5.

Boosting NAD +in myeloid cells upregulates Prostaglandin E2 (PGE2) and CC-chemokine receptor 7 (CCR7)-mediated migration via Sirt1-dependent transcriptional regulation

Abstract NAD +-boosting via nicotinamide riboside (NR) confer anti-inflammatory effects. However, underlying mechanisms and therapeutic potential remain incompletely defined. Given NAD +is a co-enzyme in oxidoreductive reactions, metabolomics analysis identified two major metabolites that were increased by NR: inosine and Prostaglandin E2 (PGE2). Both inosine and PGE2 supplementation blunted IFNβ release, however NR-mediated monocytic downregulation of IFNβ was dependent on inosine but independent of PGE2. Flow cytometric analysis of cell surface receptors in human M1 macrophage showed NR increased the expression of CC-chemokine receptor 7 (CCR7). Consequently, chemokine ligand 19 (CCL19, ligand for CCR7) - induced macrophage migration was enhanced in response to NR administration. Furthermore, NR-mediated upregulation of macrophage migration through CCL19-CCR7 axis was dependent on PGE2 synthesis. We also demonstrated that NR upregulates PGE2 synthesis through Sirt1-dependent transcriptional regulation of COX2. NR-mediated metabolic regulation appears to modulate disparate myeloid cell functions via distinct signaling intermediates and pathways. NHLBI Division of Intramural Research

RNase P Ribozyme Effectively Inhibits Human CC-Chemokine Receptor 5 Expression and Human Immunodeficiency Virus 1 Infection

Developing novel antiviral agents and approaches is essential for the treatment against human and zoonotic viruses. We had previously produced RNase P-based ribozyme variants capable of efficiently cleaving mRNA in vitro. Here, engineered ribozymes were constructed from an RNase P ribozyme variant to target the mRNA encoding human CC-chemokine receptor 5 (CCR5), an HIV co-receptor. The constructed ribozyme efficiently cleaved the CCR5 mRNA in vitro. In cells expressing the engineered ribozyme, CCR5 expression diminished by more than 90% and the infection of HIV (R5 strain Ba-L) decreased by 200-fold. The ribozyme-expressing cells resistant to R5 strain Ba-L still supported the infection of HIV X4 strain IIIB due to its use of CXCR4 instead of CCR5 as the co-receptor. Thus, the ribozyme is specific against CCR5 but not CXCR4. This indicates that RNase P ribozyme is effective and specific against CCR5 to diminish HIV infection, and also displays the viability of developing engineered RNase P ribozymes against human and zoonotic viruses.

CCR7 Mediates Cell Invasion and Migration in Extrahepatic Cholangiocarcinoma by Inducing Epithelial–Mesenchymal Transition

The epithelial-mesenchymal transition (EMT) contributes to the metastatic cascade in various tumors. C-C chemokine receptor 7 (CCR7) interacts with its ligand, chemokine (C-C motif) ligand 19 (CCL19), to promote EMT. However, the association between EMT and CCR7 in extrahepatic cholangiocarcinoma (EHCC) remains unknown. This study aimed to elucidate the prognostic impact of CCR7 expression and its association with clinicopathological features and EMT in EHCC. The association between CCR7 expression and clinicopathological features and EMT status was examined via the immunohistochemical staining of tumor sections from 181 patients with perihilar cholangiocarcinoma. This association was then investigated in TFK-1 and EGI-1 EHCC cell lines. High-grade CCR7 expression was significantly associated with a large number of tumor buds, low E-cadherin expression, and poor overall survival. TFK-1 showed CCR7 expression, and Western blotting revealed E-cadherin downregulation and vimentin upregulation in response to CCL19 treatment. The wound healing and Transwell invasion assays revealed that the activation of CCR7 by CCL19 enhanced the migration and invasion of TFK-1 cells, which were abrogated by a CCR7 antagonist. These results suggest that a high CCR7 expression is associated with an adverse postoperative prognosis via EMT induction and that CCR7 may be a potential target for adjuvant therapy in EHCC.

Mechanism of artesunate on bone destruction in experimental rheumatoid arthritis based on transcriptomics and network pharmacology

The present study investigated the mechanism of artesunate in the treatment of bone destruction in experimental rheumatoid arthritis(RA) based on transcriptomics and network pharmacology. The transcriptome sequencing data of artesunate in the inhibition of osteoclast differentiation were analyzed to obtain differentially expressed genes(DEGs). GraphPad Prism 8 software was used to plot volcano maps and heat maps were plotted through the website of bioinformatics. GeneCards and OMIM were used to collect information on key targets of bone destruction in RA. The DEGs of artesunate in inhibiting osteoclast differentiation and key target genes of bone destruction in RA were intersected by the Venny 2.1.0 platform, and the intersection target genes were analyzed by Gene Ontology(GO)/Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment. Finally, the receptor activator of nuclear factor-κB(RANKL)-induced osteoclast differentiation model and collagen-induced arthritis(CIA) model were established. Quantitative real time polymerase chain reaction(q-PCR), immunofluorescence, and immunohistochemistry were used to verify the pharmacological effect and molecular mechanism of artesunate in the treatment of bone destruction in RA. In this study, the RANKL-induced osteoclast differentiation model in vitro was established and intervened with artesunate, and transcriptome sequencing data were analyzed to obtain 744 DEGs of artesunate in inhibiting osteoclast differentiation. A total of 1 291 major target genes of bone destruction in RA were obtained from GeneCards and OMIM. The target genes of artesunate in inhibiting osteoclast differentiation and the target genes of bone destruction in RA were intersected to obtain 61 target genes of artesunate against bone destruction in RA. The intersected target genes were analyzed by GO/KEGG enrichment. According to the results previously reported, the cytokine-cytokine receptor interaction signaling pathway was selected for experimental verification. Artesunate intervention in the RANKL-induced osteoclast differentiation model showed that artesunate inhibited CC chemokine receptor 3(CCR3), CC chemokine receptor 1(CCR1) and leukemia inhibitory factor(LIF) mRNA expression in osteoclasts in a dose-dependent manner compared with the RANKL-induced group. Meanwhile, the results of immunofluorescence and immunohistochemistry showed that artesunate could dose-dependently reduce the expression of CCR3 in osteoclasts and joint tissues of the CIA rat model in vitro. This study indicated that artesunate regulated the CCR3 in the cytokine-cytokine receptor interaction signaling pathway in the treatment of bone destruction in RA and provided a new target gene for the treatment of bone destruction in RA.

CC-Chemokine Receptor-2 Expression in Osteoblasts Contributes to Cartilage and Bone Damage during Post-Traumatic Osteoarthritis

CC-Chemokine Receptor-2 Expression in Osteoblasts Contributes to Cartilage and Bone Damage during Post-Traumatic Osteoarthritis

CC chemokine receptor 2 (CCR2) expression promotes diffuse large B-Cell lymphoma survival and invasion

In recent years, CC chemokine receptor 2 (CCR2) has been found to be involved in tumor growth, angiogenesis, epithelial mesenchymal transition, metastasis, and immune escape. CCR2 overexpression was first identified as a poor prognostic predictor in diffuse large B-cell lymphoma (DLBCL) in our published article, but the mechanisms involved remain unknown. In this work, we collected data from another 138 patients with DLBCL data and verified the CCR2 expression level and its relationship to clinicopathological characteristics. Furthermore, we explored the possible mechanisms via in vitro and in vivo experiments. We showed that CCR2 overexpression was an independent prognostic marker and predicted shorter overall survival (OS) and progression-free survival (PFS) in patients with DLBCL. Blockade of CCR2 expression with a CCR2 antagonist inhibited tumor cell proliferation, migration, and anti-apoptosis ability in vitro by affecting the PI3K/Akt signaling pathway and the p38 MAPK signaling pathway. Furthermore, administration of a CCR2 antagonist decreased tumor growth and dissemination of DLBCL cells and increased survival time in the xenograft model. Our study demonstrates that CCR2 expression plays an important role in the development of DLBCL by stimulating cell proliferation, migration, and anti-apoptosis. Therefore, the inhibition of CCR2 may be a potential target for anticancer therapy in DLBCL.The authors show that CC chemokine receptor 2 (CCR2) overexpression is an independent prognostic marker for diffuse large B-cell lymphoma (DLBCL) and predicts overall survival and progression-free survival in these patients. Blockade of CCR2 signaling with a CCR2 antagonist inhibits tumor cell proliferation, migration, and apoptosis inhibition by activating the PI3K/Akt signaling pathway and inhibiting the p38 MAPK signaling pathway. Furthermore, administration of a CCR2 antagonist decreases tumor growth and dissemination of DLBCL cells and increases survival time in a xenograft model.