Integrating machine learning algorithms and single-cell analysis to identify gut microbiota-related macrophage biomarkers in atherosclerotic plaques.

Objective: The relationship between macrophages and the gut microbiota in patients with atherosclerosis remains poorly defined, and effective biological markers are lacking. This study aims to elucidate the interplay between gut microbial communities and macrophages, and to identify biomarkers associated with the destabilization of atherosclerotic plaques. The goal is to enhance our understanding of the underlying molecular pathways and to pave new avenues for diagnostic approaches and therapeutic strategies in the disease. Methods: This study employed Weighted Gene Co-expression Network Analysis (WGCNA) and differential expression analysis on atherosclerosis datasets to identify macrophage-associated genes and quantify the correlation between these genes and gut microbiota gene sets. The Random Forest algorithm was utilized to pinpoint PLEK, IRF8, BTK, CCR1, and CD68 as gut microbiota-related macrophage genes, and a nomogram was constructed. Based on the top five genes, a Non-negative Matrix Factorization (NMF) algorithm was applied to construct gut microbiota-related macrophage clusters and analyze their potential biological alterations. Subsequent single-cell analyses were conducted to observe the expression patterns of the top five genes and the interactions between immune cells. Finally, the expression profiles of key molecules were validated using clinical samples from atherosclerosis patients. Results: Utilizing the Random Forest algorithm, we ultimately identified PLEK, IRF8, CD68, CCR1, and BTK as gut microbiota-associated macrophage genes that are upregulated in atherosclerotic plaques. A nomogram based on the expression of these five genes was constructed for use as an auxiliary tool in clinical diagnosis. Single-cell analysis confirmed the specific expression of gut microbiota-associated macrophage genes in macrophages. Clinical samples substantiated the high expression of PLEK in unstable atherosclerotic plaques. Conclusion: Gut microbiota-associated macrophage genes (PLEK, IRF8, CD68, CCR1, and BTK) may be implicated in the pathogenesis of atherosclerotic plaques and could serve as diagnostic markers to aid patients with atherosclerosis.

Novel Findings on CCR1 Receptor in CNS Disorders: A Pathogenic Marker Useful in Controlling Neuroimmune and Neuroinflammatory Mechanisms in Parkinson's Disease.

Parkinson's disease (PD) is recognized as the second most common neurodegenerative disease worldwide. Even if PD etiopathogenesis is not yet fully understood, in recent years, it has been advanced that a chronic state of inflammation could play a decisive role in the development of this pathology, establishing the close link between PD and neuroinflammation. In the broad panorama of inflammation and its several signaling pathways, the C-C chemokine receptor type 1 (CCR1) could play a key pathogenic role in PD progression, and could constitute a valuable target for the development of innovative anti-PD therapies. In this study, we probed the neuroprotective properties of the CCR1 antagonist BX471 compound in a mouse model of MPTP-induced nigrostriatal degeneration. BX471 treatments were performed intraperitoneally at a dose of 3 mg/kg, 10 mg/kg, and 30 mg/kg, starting 24 h after the last injection of MPTP and continuing for 7 days. From our data, BX471 treatment strongly blocked CCR1 and, as a result, decreased PD features, also reducing the neuroinflammatory state by regulating glial activation, NF-κB pathway, proinflammatory enzymes, and cytokines overexpression. Moreover, we showed that BX471's antagonistic action on CCR1 reduced the infiltration of immune cells, including mast cells and lymphocyte T activation. In addition, biochemical analyses carried out on serum revealed a considerable increase in circulating levels of CCR1 following MPTP-induced PD. In light of these findings, CCR1 could represent a useful pathological marker of PD, and its targeting could be a worthy candidate for the future development of new immunotherapies against PD.

Expression of the chemokine receptor CCR1 decreases sensitivity to bortezomib in multiple myeloma cell lines.

BACKGROUND: The proteasome inhibitor bortezomib is one of the primary therapies used for the haematological malignancy multiple myeloma (MM). However, intrinsic or acquired resistance to bortezomib, via mechanisms that are not fully elucidated, is a barrier to successful treatment in many patients. Our previous studies have shown that elevated expression of the chemokine receptor CCR1 in MM plasma cells in newly diagnosed MM patients is associated with poor prognosis. Here, we hypothesised that the poor prognosis conferred by CCR1 expression is, in part, due to a CCR1-mediated decrease in MM plasma cell sensitivity to bortezomib. METHODS: In order to investigate the role of CCR1 in MM cells, CCR1 was knocked out in human myeloma cell lines OPM2 and U266 using CRISPR-Cas9. Additionally, CCR1 was overexpressed in the mouse MM cell line 5TGM1. The effect of bortezomib on CCR1 knockout or CCR1-overexpressing cells was then assessed by WST-1 assay, with or without CCL3 siRNA knockdown or addition of recombinant human CCL3. NSG mice were inoculated intratibially with OPM2-CCR1KO cells and were treated with 0.7 mg/kg bortezomib or vehicle twice per week for 3 weeks and GFP+ tumour cells in the bone marrow were quantitated by flow cytometry. The effect of CCR1 overexpression or knockout on unfolded protein response pathways was assessed using qPCR for ATF4, HSPA5, XBP1, ERN1 and CHOP and Western blot for IRE1α and p-Jnk. RESULTS: Using CCR1 overexpression or CRIPSR-Cas9-mediated CCR1 knockout in MM cell lines, we found that CCR1 expression significantly decreases sensitivity to bortezomib in vitro, independent of the CCR1 ligand CCL3. In addition, CCR1 knockout rendered the human MM cell line OPM2 more sensitive to bortezomib in an intratibial MM model in NSG mice in vivo. Moreover, CCR1 expression negatively regulated the expression of the unfolded protein response receptor IRE1 and downstream target gene XBP1, suggesting this pathway may be responsible for the decreased bortezomib sensitivity of CCR1-expressing cells. CONCLUSIONS: Taken together, these studies suggest that CCR1 expression may be associated with decreased response to bortezomib in MM cell lines.

Role of CCR1/5/7 in hepatocellular carcinoma: a study on prognostic evaluation, molecular subtyping, and association with immune infiltration.

This study aims to assess the prognostic value of the C-C motif chemokine receptor (CCR) gene family in hepatocellular carcinoma (HCC) and its relationship with immune infiltration and molecular subtypes of HCC. The evaluation of the GSE14520 dataset and TCGA database confirmed the prognostic significance of CCR. Building upon the correlation between CCR1, CCR5, and CCR7 and favorable prognosis, we further validated the prognostic importance of CCR1, CCR5, and CCR7 in ICGC database and an independent cohort from Guangxi autonomous region. Then, we constructed a risk prognosis model. Additionally, we observed significant positive correlations between CCR1, CCR5, and CCR7 and the infiltration of B cells, T cells, and macrophages in HCC. Subsequently, we conducted CCK assays, Transwell assays, and colony formation assays to evaluate the molecular biological functions of CCR1, CCR5, and CCR7. These experiments further confirmed that upregulation of CCR1, CCR5, and CCR7 can individually inhibit the proliferation, migration, and stemness of HCC cells. By analyzing the relationship between expression levels and tumor mutation frequency, we discovered that patients with high CCR1 expression were more likely to be classified as non-proliferative HCC. Similar conclusions were observed for CCR5 and CCR7. The association of CCR1, CCR5, and CCR7 with the molecular subtypes of HCC suggests that they may serve as intermediary molecules linking immune status and molecular subtypes in HCC. In summary, CCR1, CCR5, and CCR7 have the potential to serve as prognostic biomarkers for HCC and regulate HCC progression by influencing immune cell infiltration.

CFL1 restores the migratory capacity of bone marrow mesenchymal stem cells in primary Sjögren's syndrome by regulating CCR1 expression.

BACKGROUND: Primary Sjögren's syndrome (pSS) is a chronic systemic autoimmune disease. There is no relevant research on whether the migratory ability of bone marrow mesenchymal stem cells (BM-MSC) is impaired in patients with pSS (pSS-BMMSC). METHODS: Trajectories and velocities of BM-MSC were analyzed. Transwell migration assay and wound healing assay were used to investigate the migratory capacity of BM-MSC. The proliferative capacity of BM-MSC was evaluated by EDU and CCK8 assay. RNA-seq analysis was then performed to identify the underlying mechanism of lentivirus-mediated cofilin-1 overexpression BM-MSC (BMMSCCFL1). The therapeutic efficacy of BMMSCCFL1 was evaluated in NOD mice. RESULTS: The migratory capacity of pSS-BMMSC was significantly reduced compared to normal volunteers (HC-BMMSC). The expression of the motility-related gene CFL1 was decreased in pSS-BMMSC. Lentivirus-mediated CFL1 overexpression of pSS-BMMSC promoted the migration capacity of pSS-BMMSC. Furthermore, RNA-seq revealed that CCR1 was the downstream target gene of CFL1. To further elucidate the mechanism of CFL1 in regulating BM-MSC migration and proliferation via the CCL5/CCR1 axis, we performed a rescue experiment using BX431 (a CCR1-specific inhibitor) to inhibit CCR1. The results showed that CCR1 inhibitors suppressed the migration and proliferation capacity of MSC induced by CFL1. CONCLUSION: The pSS-BMMSC leads to impaired migration and proliferation, and overexpression of CFL1 can rescue the functional deficiency and alleviate disease symptoms in NOD mice. Mechanically, CFL1 can regulate the expression level of the downstream CCL5/CCR1 axis to enhance the migration and proliferation of BM-MSC.

Chemokine receptor CCR1 regulates macrophage activation through mTORC1 signaling in nonalcoholic steatohepatitis.

BACKGROUND AND AIMS: Chemokine (CC motif) receptor 1 (CCR1) promotes liver fibrosis in mice. However, its effects on nonalcoholic steatohepatitis (NASH) remain unclear. Therefore, the present study aimed to investigate the role of CCR1 in the progression of NASH. METHODS: Human serum and liver tissues were obtained from patients with NASH and controls. Systemic (Ccr1-/-) and liver macrophage-knockout Ccr1 (Ccr1LKD) mice were fed a high-cholesterol and high-fat (CL) diet for 12 weeks or a methionine/choline-deficient (MCD) diet for 4 weeks. BX471 was used to pharmacologically inhibit CCR1 in CL-fed mice. RESULTS: CCR1 was significantly upregulated in liver samples from patients with NASH and in animal models of dietary-induced NASH. In the livers of mice fed a CL diet for 12 weeks, the CCR1 protein colocalized with F4/80+ macrophages rather than with hepatic stellate cells. Compared to their wild-type littermates, Ccr1-/- mice fed with the CL or MCD diet showed inhibition of NASH-associated hepatic steatosis, inflammation, and fibrosis. Mechanistically, Ccr1 deficiency suppressed macrophage infiltration and activation by attenuating the mechanistic target of rapamycin complex 1 (mTORC1) signaling. Similar results were observed in Ccr1LKD mice administered the CL diet. Moreover, CCR1 inhibition by BX471 effectively suppressed NASH progression in CL-fed mice. CONCLUSIONS: Ccr1 deficiency mitigated macrophage activity by inhibiting mTORC1 signaling, thereby preventing the development of NASH. Notably, the CCR1 inhibitor BX471 protected against NASH. These findings would help in developing novel strategies for the treatment of NASH.

Inhibition of CCR1 attenuates neuroinflammation via the JAK2/STAT3 signaling pathway after subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: Neuroinflammation is an important mechanism underlying brain injury caused by subarachnoid hemorrhage (SAH). C-C chemokine receptor type 1 (CCR1)-mediated inflammation is involved in the pathology of many central nervous system diseases. Herein, we investigated whether inhibition of CCR1 alleviated neuroinflammation after experimental SAH and aimed to elucidate the mechanisms of its potential protective effects. METHODS: To analyze SAH transcriptome data R studio was used, and a mouse model of SAH was established using endovascular perforations. In this model, the selective CCR1 antagonist Met-RANTES (Met-R) and the CCR1 agonist recombinant CCL5 (rCCL5) were administered 1 h after SAH induction. To investigate the possible downstream mechanisms of CCR1, the JAK2 inhibitor AG490 and the JAK2 activator coumermycin A1 (C-A1) were administered 1 h after SAH induction. Furthermore, post-SAH evaluation, including SAH grading, neurological function tests, Western blot, the terminal deoxynucleotidyl transferase dUTP nick end labeling assay, and Fluoro-Jade B and fluorescent immunohistochemical staining were performed. Cerebrospinal fluid (CSF) samples were detected by ELISA. RESULTS: CCL5 and CCR1 expression levels increased significantly following SAH. Met-R significantly improved neurological deficits in mice, decreased apoptosis and degeneration of ipsilateral cerebral cortex neurons, reduced infiltrating neutrophils, and promoted microglial activation after SAH induction. Furthermore, Met-R inhibited the expression of p-JAK2, p-STAT3, interleukin-1ß, and tumor necrosis factor-α. However, the protective effects of Met-R were abolished by C-A1 treatment. Furthermore, rCCL5 injection aggravated neurological dysfunction and increased the expression of p-JAK2, p-STAT3, interleukin-1ß, and tumor necrosis factor-α in SAH mice, all of which were reversed by the administration of AG490. Finally, the levels of CCL5 and CCR1 were elevate in the CSF of SAH patient and high level of CCL5 and CCR1 levels were associated with poor outcome. CONCLUSION: The present results suggested that inhibition of CCR1 attenuates neuroinflammation after SAH via the JAK2/STAT3 signaling pathway, which may provide a new target for the treatment of SAH.

CCR1 mediates Müller cell activation and photoreceptor cell death in macular and retinal degeneration.

Mononuclear cells are involved in the pathogenesis of retinal diseases, including age-related macular degeneration (AMD). Here, we examined the mechanisms that underlie macrophage-driven retinal cell death. Monocytes were extracted from patients with AMD and differentiated into macrophages (hMdɸs), which were characterized based on proteomics, gene expression, and ex vivo and in vivo properties. Using bioinformatics, we identified the signaling pathway involved in macrophage-driven retinal cell death, and we assessed the therapeutic potential of targeting this pathway. We found that M2a hMdɸs were associated with retinal cell death in retinal explants and following adoptive transfer in a photic injury model. Moreover, M2a hMdɸs express several CCRI (C-C chemokine receptor type 1) ligands. Importantly, CCR1 was upregulated in Müller cells in models of retinal injury and aging, and CCR1 expression was correlated with retinal damage. Lastly, inhibiting CCR1 reduced photic-induced retinal damage, photoreceptor cell apoptosis, and retinal inflammation. These data suggest that hMdɸs, CCR1, and Müller cells work together to drive retinal and macular degeneration, suggesting that CCR1 may serve as a target for treating these sight-threatening conditions.

Structural Insights into Molecular Recognition and Receptor Activation in Chemokine-Chemokine Receptor Complexes.

The chemokine system is a key player in the functioning of the immune system and a sought-after target for drug candidates. The number of experimental structures of chemokines in complex with chemokine receptors has increased rapidly over the past few years, providing essential information for rational development of chemokine receptor ligands. Here, we perform a comparative analysis of all chemokine-chemokine receptor structures, with the aim of characterizing the molecular recognition processes and highlighting the relationships between chemokine structures and functional processes. The structures show conserved interaction patterns between the chemokine core and the receptor N-terminus, while interactions near ECL2 display subfamily-specific features. Detailed analyses of the interactions of the chemokine N-terminal domain in the 7TM cavities reveal activation mechanisms for CCR5, CCR2, and CXCR2 and a mechanism for biased agonism in CCR1.

The Chemokine Receptor CCR1 Mediates Microglia Stimulated Glioma Invasion.

Glioblastoma multiforme (GBM) is the most aggressive form of adult brain tumor which is highly resistant to conventional treatment and therapy. Glioma cells are highly motile resulting in infiltrative tumors with poorly defined borders. Another hallmark of GBM is a high degree of tumor macrophage/microglia infiltration. The level of these tumor-associated macrophages/microglia (TAMs) correlates with higher malignancy and poorer prognosis. We previously demonstrated that inhibition of TAM infiltration into glioma tumors with the CSF-1R antagonist pexidartinib (PLX3397) can inhibit glioma cell invasion in-vitro and in-vivo. In this study, we demonstrate an important role for the chemokine receptor CCR1 in mediating microglia/TAM stimulated glioma invasion. Using two structurally distinct CCR1 antagonists, including a novel inhibitor "MG-1-5", we were able to block microglial activated GL261 glioma cell invasion in a dose dependent manner. Interestingly, treatment of a murine microglia cell line with glioma conditioned media resulted in a strong induction of CCR1 gene and protein expression. This induction was attenuated by inhibition of CSF-1R. In addition, glioma conditioned media treatment of microglia resulted in a rapid upregulation of gene expression of several CCR1 ligands including CCL3, CCL5, CCL6 and CCL9. These data support the existence of tumor stimulated autocrine loop within TAMs which ultimately mediates tumor cell invasion.

Single cell RNA-seq resolution revealed CCR1+/SELL+/XAF+ CD14 monocytes mediated vascular endothelial cell injuries in Kawasaki disease and COVID-19.

INTRODUCTION: The COVID-19 pandemic provide the opportunities to explore the numerous similarities in clinical symptoms with Kawasaki disease (KD), including severe vasculitis. Despite this, the underlying mechanisms of vascular injury in both KD and COVID-19 remain elusive. To identify these mechanisms, this study employs single-cell RNA sequencing to explore the molecular mechanisms of immune responses in vasculitis, and validate the results through in vitro experiments. METHOD: The single-cell RNA sequencing (scRNA-seq) analysis of peripheral blood mononuclear cells (PBMCs) was carried out to investigate the molecular mechanisms of immune responses in vasculitis in KD and COVID-19. The analysis was performed on PBMCs from six children diagnosed with complete KD, three age-matched KD healthy controls (KHC), six COVID-19 patients (COV), three influenza patients (FLU), and four healthy controls (CHC). The results from the scRNA-seq analysis were validated through flow cytometry and immunofluorescence experiments on additional human samples. Subsequently, monocyte adhesion assays, immunofluorescence, and quantitative polymerase chain reaction (qPCR) were used to analyze the damages to endothelial cells post-interaction with monocytes in HUVEC and THP1 cultures. RESULTS: The scRNA-seq analysis revealed the potential cellular types involved and the alterations in genetic transcriptions in the inflammatory responses. The findings indicated that while the immune cell compositions had been altered in KD and COV patients, and the ratio of CD14+ monocytes were both elevated in KD and COV. While the CD14+ monocytes share a large scale of same differentiated expressed geens between KD and COV. The differential activation of CD14 and CD16 monocytes was found to respond to both endothelial and epithelial dysfunctions. Furthermore, SELL+/CCR1+/XAF1+ CD14 monocytes were seen to enhance the adhesion and damage to endothelial cells. The results also showed that different types of B cells were involved in both KD and COV, while only the activation of T cells was recorded in KD. CONCLUSION: In conclusion, our study demonstrated the role of the innate immune response in the regulation of endothelial dysfunction in both KD and COVID-19. Additionally, our findings indicate that the adaptive immunity activation differs between KD and COVID-19. Our results demonstrate that monocytes in COVID-19 exhibit adhesion to both endothelial cells and alveolar epithelial cells, thus providing insight into the mechanisms and shared phenotypes between KD and COVID-19.

Characterizing ligand-receptor interactions and unveiling the pro-tumorigenic role of CCL16-CCR1 axis in the microenvironment of hepatocellular carcinoma.

Background: The heterogeneity of the tumor microenvironment significantly influences the prognosis of hepatocellular carcinoma (HCC) patients, with cell communication through ligand-receptor complexes playing a central role. Methods: We conducted single-cell transcriptomic analysis on ten HCC tissues to identify ligand-receptor genes involved in malignant HCC cell communication using CellChat. Leveraging RNA-Seq data from the TCGA Liver Cancer (TCGA-LIHC) and Liver Cancer - RIKEN, JP (LIRI-JP) cohorts, we employed Cox regression analysis to screen for prognosis-related genes. Prognostic risk models were constructed through unsupervised clustering and differential gene expression analysis. Subsequently, a co-culture system involving tumor cells and macrophages was established. A series of experiments, including Transwell assays, immunofluorescence staining, immunoprecipitation, flow cytometry, and immunohistochemistry, were conducted to elucidate the mechanism through which HCC cells recruit macrophages via the CCL16-CCR1 axis. Results: Single-cell analysis unveiled significant interactions between malignant HCC cells and macrophages, identifying 76 related ligand-receptor genes. Patients were classified into three subtypes based on the expression patterns of eight prognosis-related ligand-receptor genes. The subtype with the worst prognosis exhibited reduced infiltration of T cell-related immune cells, downregulation of immune checkpoint genes, and increased M2-like tumor-associated macrophage scores. In vitro experiments confirmed the pivotal role of the CCL16-CCR1 axis in the recruitment and M2 polarization of tumor-associated macrophages. Clinical samples demonstrated a significant association between CCL16 protein expression levels and advanced stage, lymph node metastasis, and distant metastasis. Immunohistochemistry and immunofluorescence staining further confirmed the correlation between CCL16 and CCR1, CD68, and CD206, as well as CD68+CCR1+ macrophage infiltration. Conclusions: Our study identified molecular subtypes, a prognostic model, and immune microenvironment features based on ligand-receptor interactions in malignant HCC cell communication. Moreover, we revealed the pro-tumorigenic role of HCC cells in recruiting M2-like tumor-associated macrophages through the CCL16-CCR1 axis.

Dysfunction of CCR1+ decidual macrophages is a potential risk factor in the occurrence of unexplained recurrent pregnancy loss.

Recurrent pregnancy loss (RPL) puzzles 1-3% of women of childbearing age worldwide. Immunological factors account for more than 60% of cases of unexplained RPL (URPL); however, the underlying mechanism remains unclear. Here, using single-cell sequencing data and functional experiments with clinical samples, we identified a distinct population of CCR1+ decidual macrophages (dMφ) that were preferentially enriched in the decidua from normal early pregnancies but were substantially decreased in patients with URPL. Specific gene signatures endowed CCR1+ dMφ with immunosuppressive and migration-regulatory properties, which were attenuated in URPL. Additionally, CCR1+ dMφ promoted epithelial-to-mesenchymal transition (EMT) to promote trophoblast migration and invasion by activating the ERK1/2 signaling pathway. Decidual stromal cell (DSC)-derived CCL8 was the key regulator of CCR1+ dMφ as CCL8 recruited peripheral CCR1+ monocytes, induced a CCR1+ dMφ-like phenotype, and reinforced the CCR1+ dMφ-exerted modulation of trophoblasts. In patients with URPL, CCL8 expression in DSCs was decreased and trophoblast EMT was defective. Our findings revealed that CCR1+ dMφ play an important role in immune tolerance and trophoblast functions at the maternal-fetal interface. Additionally, decreased quantity and dysregulated function of CCR1+ dMφ result in URPL. In conclusion, we provide insights into the crosstalk between CCR1+ dMφ, trophoblasts, and DSCs at the maternal-fetal interface and macrophage-targeted interventions of URPL.

Activation of the Chemokine Receptor CCR1 and Preferential Recruitment of Gαi Suppress RSV Replication: Implications for Developing Novel Respiratory Syncytial Virus Treatment Strategies.

Respiratory syncytial virus (RSV) is a major pathogen that can cause acute respiratory infectious diseases of the upper and lower respiratory tract, especially in children, elderly individuals, and immunocompromised people. Generally, following viral infection, respiratory epithelial cells secrete cytokines and chemokines to recruit immune cells and initiate innate and/or adaptive immune responses. However, whether chemokines affect viral replication in nonimmune cells is rarely clear. In this study, we detected that chemokine CCL5 was highly expressed, while expression of its receptor, CCR1, was downregulated in respiratory epithelial cells after RSV infection. When we overexpressed CCR1 on respiratory epithelial cells in vivo or in vitro, viral load was significantly suppressed, which can be restored by the neutralizing antibody for CCR1. Interestingly, the antiviral effect of CCR1 was not related to type I interferon (IFN-I), apoptosis induction, or viral adhesion or entry inhibition. In contrast, it was related to the preferential recruitment and activation of the adaptor Gαi, which promoted inositol 1,4,5-triphosphate receptor type 3 (ITPR3) expression, leading to inhibited STAT3 phosphorylation; explicitly, phosphorylated STAT3 (p-STAT3) was verified to be among the important factors regulating the activity of HSP90, which has been previously reported to be a chaperone of RSV RNA polymerase. In summary, we are the first to reveal that CCR1 on the surface of nonimmune cells regulates RSV replication through a previously unknown mechanism that does not involve IFN-I induction. IMPORTANCE Our results revealed a novel mechanism by which RSV escapes innate immunity. That is, although it induces high CCL5 expression, RSV might attenuate the binding of CCL5 by downregulating the expression of CCR1 in respiratory epithelial cells to weaken the inhibitory effect of CCR1 on HSP90 activity and thereby facilitate RSV replication in nonimmune cells. This study provides a new target for the development of co-antiviral inhibitors against other components of the host and co-molecular chaperone/HSP90 and provides a scientific basis for the search for effective broad-spectrum antiviral drugs.

Evolving polarisation of infiltrating and alveolar macrophages in the lung during metastatic progression of melanoma suggests CCR1 as a therapeutic target.

Metastatic tumour progression is facilitated by tumour associated macrophages (TAMs) that enforce pro-tumour mechanisms and suppress immunity. In pulmonary metastases, it is unclear whether TAMs comprise tissue resident or infiltrating, recruited macrophages; and the different expression patterns of these TAMs are not well established. Using the mouse melanoma B16F10 model of experimental pulmonary metastasis, we show that infiltrating macrophages (IM) change their gene expression from an early pro-inflammatory to a later tumour promoting profile as the lesions grow. In contrast, resident alveolar macrophages (AM) maintain expression of crucial pro-inflammatory/anti-tumour genes with time. During metastatic growth, the pool of macrophages, which initially contains mainly alveolar macrophages, increasingly consists of infiltrating macrophages potentially facilitating metastasis progression. Blocking chemokine receptor mediated macrophage infiltration in the lung revealed a prominent role for CCR2 in Ly6C+ pro-inflammatory monocyte/macrophage recruitment during metastasis progression, while inhibition of CCR2 signalling led to increased metastatic colony burden. CCR1 blockade, in contrast, suppressed late phase pro-tumour MR+Ly6C- monocyte/macrophage infiltration accompanied by expansion of the alveolar macrophage compartment and accumulation of NK cells, leading to reduced metastatic burden. These data indicate that IM has greater plasticity and higher phenotypic responsiveness to tumour challenge than AM. A considerable difference is also confirmed between CCR1 and CCR2 with regard to the recruited IM subsets, with CCR1 presenting a potential therapeutic target in pulmonary metastasis from melanoma.

The rs13075270 and rs13092160 polymorphisms of CCR1 and CCR3 genes on oral aphthous-like lesions in PFAPA syndrome.

Fever is a common symptom of infection in children. Periodic fever syndromes are less common but more complex. One of these Periodic fever syndromes is PFAPA (periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis) syndrome which is known as the most benign syndromes. The cause of this disease is unknown. Various factors, including environmental and genetic factors, are involved in the development of this disease. In this study, the association of rs13075270 and rs13092160 polymorphisms were investigated in CCR1 and CCR3 genes with susceptibility to this syndrome in the Chinese population. In this regard, 38 patients with PFAPA syndrome and 100 healthy individuals were selected. After DNA sampling and extraction, polymorphisms of CCR1 and CCR3 receptor genes were examined by the PCR-RFLP method. Findings were analyzed using SPSS software version 22 with a significant level of P <0.05. The frequency of T/T genotype rs13092160 polymorphism in the patient and control groups was 78.95% and 83%, respectively, C/T genotype was 21.05% and 17% (P = 0.421). The frequency of the C/C genotype was 0 in both groups. Regarding rs13075270 polymorphism, the frequency of T/T genotype in patient and control groups was 15.79% and 81%, C/T genotype was 78.95% and 18% and C/C genotype was 5.26% and 1%, respectively (P<0.05). Thus, in rs13075270 polymorphism, the C/T genotype was associated with the risk of PFAPA syndrome (P<0.05), but rs13092160 polymorphism did not show a significant difference between individuals with PFAPA syndrome and controls.

CCR1 antagonist J-113863 corrects the imbalance of pro- and anti-inflammatory cytokines in a SJL/J mouse model of relapsing-remitting multiple sclerosis.

Multiple sclerosis (MS), an immune-mediated and neurodegenerative disorder of the central nervous system (CNS), is characterized by infiltrating myelin-reactive T lymphocytes and demyelinating lesions. Experimental autoimmune encephalomyelitis (EAE) is a well-established animal model used to study MS. To explore the impact of chemokine receptor CCR1 blockade in EAE and the underlying mechanisms, we used CCR1 antagonist J-113863 in PLP139-151-induced EAE in SJL/J mice. Following EAE induction, mice were treated with J-113863 (10 mg/kg) daily from day 14 until day 25. We investigated the effect of J-113863 on expression levels of GM-CSF, IL-6, IL-10, IL-27 in CD4+ spleen cells, using flow cytometry. We also analyzed the effect of J-113863 on GM-CSF, IL-6, IL-10, IL-27 mRNA and protein expression levels using RT-PCR and Western blot analysis in brain tissues. J-113863 treatment decreased the populations of CD4+GM-CSF+ and CD4+IL-6+ cells and increased CD4+IL-27+ and CD4+IL-10+ cells in the spleen. J-113863 had a suppressive effect on the mRNA and protein expression levels of GM-CSF, and IL-6 in the brain tissue. On the other hand, J-113863 treatment increased the mRNA and protein expression of IL-10 and IL-27 in the brain tissue. Our results highlighted J-113863's potential role in suppressing pro-inflammatory expression and up-regulating anti-inflammatory mediators, which could represent a beneficial alternative approach to MS treatment.

The improvement of nonalcoholic steatohepatitis by Poria cocos polysaccharides associated with gut microbiota and NF-κB/CCL3/CCR1 axis.

BACKGROUND: Nonalcoholic steatohepatitis (NASH) has been linked to inflammation induced by intestinal microbiota. Poria cocos polysaccharides (PCP) possesses anti-inflammation and immunomodulation functions; however, its preventive effects against NASH and potential mechanisms need to be explored. METHODS: The composition of PCP was determined using ion chromatography. C57BL/6 mice were administered the methionine and choline deficient (MCD) diet for 4 weeks to establish the NASH model or methionine-choline-sufficient (MCS) diet to serve as the control. Mice were assigned to the MCS group, MCD group, low-dose PCP (LP) group, and high-dose PCP (HP) group, and were administered the corresponding medications via gavage. Serum biochemical index analysis and liver histopathology examination were performed to verify the successful establishment of NASH model and to evaluate the efficacy of PCP. The composition of intestinal bacteria was profiled through 16S rRNA gene sequencing. Hepatic RNA sequencing (RNA-Seq) was performed to explore the potential mechanisms, which were further confirmed using qPCR, western blot, and immunohistochemistry. RESULTS: PCP consists of glucose, galactose, mannose, D-glucosamine hydrochloride, xylose, arabinose, and fucose. PCP could significantly alleviate symptoms of NASH, including histological liver damage, impaired hepatic function, and increased oxidative stress. Meanwhile, HP could reshape the composition of intestinal bacteria by significantly increasing the relative abundance of Faecalibaculum and decreasing the level of endotoxin load derived from gut bacteria. PCP could also downregulate the expression of pathways associated with immunity and inflammation, including the chemokine signaling pathway, Toll-like receptor signaling pathway, and NF-kappa B signaling pathway. The expression levels of CCL3 and CCR1 (involved in the chemokine signaling pathway), Tlr4, Cd11b, and NF-κb (involved in the NF-kappa B signaling pathway), and Tnf-α (involved in the TNF signaling pathway) were significantly reduced in the HP group compared to the MCD group. CONCLUSIONS: PCP could prevent the development of NASH, which may be associated with the modulation of intestinal microbiota and the downregulation of the NF-κB/CCL3/CCR1 axis.

Expression of the Chemokine Receptor CCR1 in Burkitt Lymphoma Cell Lines Is Linked to the CD10-Negative Cell Phenotype and Co-Expression of the EBV Latent Genes EBNA2, LMP1, and LMP2.

Chemokines and their receptors regulate the migration of immune cells and the dissemination of cancer cells. CCR1, CCR2, CCR3, and CCR5 all belong to a single protein homology cluster and respond to the same inflammatory chemokines. We previously reported that CCR1 and CCR2B are induced upon Epstein-Barr virus (EBV) infection of B cells in vitro. EBV is present in almost all cases of endemic Burkitt lymphoma (BL); however, the contribution of EBV in the pathogenesis of the disease is not fully understood. Here, we analyzed the relation of the expression of CCR1, CCR2, CCR3, and CCR5, the EBV DNA load and expression of EBV latent genes in nine EBV-carrying and four EBV-negative BL cell lines. We revealed that CCR1 is expressed at high mRNA and protein levels in two CD10-negative BL cell lines with co-expression of the EBV latent genes EBNA2, LMP1, and LMP2. Low levels of CCR2 transcripts were found in three BL cell lines. CCR3 and CCR5 transcripts were hardly detectable. Our data suggest that in vivo, CCR1 may be involved in the dissemination of BL cells and in the selection of BL cells with restricted EBV gene expression programs.

Targeting Chemokine Receptor CCR1 as a Potential Therapeutic Approach for Multiple Myeloma.

Multiple myeloma is an incurable plasma B-cell malignancy with 5-year survival rates approximately 10-30% lower than other hematologic cancers. Treatment options include combination chemotherapy followed by autologous stem cell transplantation. However, not all patients are eligible for autologous stem cell transplantation, and current pharmacological agents are limited in their ability to reduce tumor burden and extend multiple myeloma remission times. The "chemokine network" is comprised of chemokines and their cognate receptors, and is a critical component of the normal bone microenvironment as well as the tumor microenvironment of multiple myeloma. Antagonists targeting chemokine-receptor 1 (CCR1) may provide a novel approach for treating multiple myeloma. In vitro CCR1 antagonists display a high degree of specificity, and in some cases signaling bias. In vivo studies have shown they can reduce tumor burden, minimize osteolytic bone damage, deter metastasis, and limit disease progression in multiple myeloma models. While multiple CCR1 antagonists have entered the drug pipeline, none have entered clinical trials for treatment of multiple myeloma. This review will discuss whether current CCR1 antagonists are a viable treatment option for multiple myeloma, and studies aimed at identifying which CCR1 antagonist(s) are most appropriate for this disease.