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Cytokine Aug 2011Chemokines and their receptors control cell migration associated with routine immune surveillance, inflammation and development. They are also implicated in a large...
Chemokines and their receptors control cell migration associated with routine immune surveillance, inflammation and development. They are also implicated in a large number of inflammatory diseases, cancer and HIV. Here we describe a rapid and efficient way to express and purify milligram quantities of multiple chemokine ligands (CCL7/MCP-3, CCL14/HCC-1, CCL3/MIP-1α and CXCL8/IL-8) containing C-terminal modifications to enable coupling to fluorescent dyes or small molecules such as biotin, in vitro. These labeled chemokines display wild-type behavior in both receptor binding and calcium mobilization assays. The ability to rapidly and inexpensively produce labeled chemokines opens the way for their use in many applications, including non-traditional chemokine-receptor interaction studies, both on intact cells and with purified receptor reconstituted in artificial membranes in vitro. Furthermore, the ability to immobilize chemokines to obtain ligand affinity columns aids in efforts to purify chemokine receptors for structural and biophysical studies, by facilitating the separation of functional proteins from their non-functional counterparts.
Topics: Biotin; Chemokine CCL3; Chemokine CCL7; Chemokines; Chemokines, CC; Chromatography, Affinity; Fluorescent Dyes; Humans; Interleukin-8; Ligands; Radioligand Assay; Recombinant Fusion Proteins
PubMed: 21632261
DOI: 10.1016/j.cyto.2011.05.002 -
Frontiers in Immunology 2020Cancer is a significant medical issue, being one of the main causes of mortality around the world. The therapies for this pathology depend on the stage in which the... (Review)
Review
Cancer is a significant medical issue, being one of the main causes of mortality around the world. The therapies for this pathology depend on the stage in which the cancer is found, but it is usually diagnosed at an advanced stage in which the treatment is chemotherapy. Platinum drugs are among the most commonly used in therapy, unfortunately, one of the main obstacles to this treatment is the development of chemoresistance, which is the ability of cancer cells to evade the effects of drugs. Although some molecular mechanisms involved in resistance to platinum drugs are described, elucidation is still required of others. Secretion of inflammatory mediators such as cytokines and chemokines, by tumor microenvironment components or tumor cells, show direct influence on proliferation, metastasis and progression of cancer and are related to chemoresistance and poor prognosis. In this review, the general mechanisms associated with resistance to platinum drugs, inflammation on cancer development and chemoresistance in various types of cancer will be approached with special emphasis on the current history of CC chemokines subfamily-mediated chemoresistance.
Topics: Cell Proliferation; Chemokines, CC; Drug Resistance, Neoplasm; Humans; Inflammation; Neoplasms; Platinum; Signal Transduction; Tumor Microenvironment
PubMed: 32499779
DOI: 10.3389/fimmu.2020.00901 -
Arthritis & Rheumatology (Hoboken, N.J.) Mar 2014Systemic sclerosis (SSc)-related interstitial lung disease (ILD) is one of the leading causes of mortality. We undertook this study to analyze the gene expression of...
OBJECTIVE
Systemic sclerosis (SSc)-related interstitial lung disease (ILD) is one of the leading causes of mortality. We undertook this study to analyze the gene expression of lung tissue in a prospective cohort of patients with SSc-related ILD and to compare it with that in control lungs and with 2 prospective clinical parameters in order to understand the molecular pathways implicated in progressive lung disease.
METHODS
Lung tissue was obtained by open lung biopsy in 28 consecutive patients with SSc-related ILD and in 4 controls. High-resolution computed tomography (HRCT) and pulmonary function testing (PFT) were performed at baseline and 2-3 years after treatment based on lung histologic classification. Microarray analysis was performed, and the results were correlated with changes in the HRCT score (FibMax) and PFT values. Quantitative polymerase chain reaction (qPCR) and immunohistochemistry were used to confirm differential levels of messenger RNA and protein.
RESULTS
Lung microarray data distinguished patients with SSc-related ILD from healthy controls. In the lungs of patients with SSc-related ILD who had nonspecific interstitial pneumonia (NSIP), expressed genes included macrophage markers, chemokines, collagen, and transforming growth factor β (TGFβ)- and interferon (IFN)-regulated genes. Expression of these genes correlated with progressive lung fibrosis defined by the change in FibMax. Immunohistochemistry confirmed increased markers of collagen (COL1A1), IFN (OAS1 and IFI44), and macrophages (CCL18 and CD163), and the positive correlation with the change in FibMax was confirmed by qPCR in a larger group of SSc patients with NSIP. Several genes correlated with both the change in FibMax (r > 0.4) and the change in % predicted forced vital capacity (r < -0.1), including IFN and macrophage markers, chemokines, and heat-shock proteins.
CONCLUSION
These results highlight major pathogenic pathways relevant to progressive pulmonary fibrosis in SSc-related ILD: macrophage emigration and activation, and up-regulated expression of TGFβ- and IFN-regulated genes.
Topics: 2',5'-Oligoadenylate Synthetase; Adult; Antigens; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Chemokines, CC; Collagen Type I; Collagen Type I, alpha 1 Chain; Cytoskeletal Proteins; Disease Progression; Female; Humans; Lung; Macrophage Activation; Male; Middle Aged; Pulmonary Fibrosis; Receptors, Cell Surface; Respiratory Function Tests; Scleroderma, Systemic; Transforming Growth Factor beta
PubMed: 24574232
DOI: 10.1002/art.38288 -
PloS One Jul 2007CCL25/TECK and CCL28/MEC are CC chemokines primarily expressed in thymic dendritic cells and mucosal epithelial cells. Their receptors, CCR9 and CCR10, are mainly...
BACKGROUND
CCL25/TECK and CCL28/MEC are CC chemokines primarily expressed in thymic dendritic cells and mucosal epithelial cells. Their receptors, CCR9 and CCR10, are mainly expressed on T and B lymphocytes. In human, mouse, pig and sheep CCL25 and CCL28 play an important role in the segregation and the compartmentalization of the mucosal immune system. As evidenced by early comparisons of germ-free and conventional animals, the intestinal bacterial microflora has a marked effect on host intestinal immune functions. However, little is known about the impact of bacterial colonization on constitutive and induced chemokine expressions as well as on the generation of anti-inflammatory mechanisms.
METHODOLOGY/PRINCIPAL FINDINGS
Therefore, we decided to focus by qPCR on the mRNA expression of two main gut chemokines, CCL25 and CCL28, their receptors CCR9 and CCR10, the Tregs marker Foxp3 and anti-inflammatory cytokines TGF-beta and IL-10 following colonization with different bacterial species within the small intestine. To accomplish this we used an original germ-free neonatal pig model and monoassociated pigs with a representative Gram-negative (Escherichia coli) or Gram-positive (Lactobacillus fermentum) commensal bacteria commonly isolated from the neonatal pig intestine. Our results show a consistent and marked effect of microbial colonization on the mRNA expression of intestinal chemokines, chemokine receptors, Foxp3 and TGF-beta. Moreover, as evidenced by in vitro experiments using two different cell lines, the pattern of regulation of CCL25 and CCL28 expression in the gut appears complex and suggests an additional role for in vivo factors.
CONCLUSIONS/SIGNIFICANCE
Taken together, the results highlight the key role of bacterial microflora in the development of a functional intestinal immune system in an elegant and relevant model for human immune system development.
Topics: Animals; Chemokines; Chemokines, CC; Escherichia coli; Forkhead Transcription Factors; Gene Expression Regulation; Humans; Intestine, Small; Mice; RNA, Messenger; Receptors, CCR; Receptors, CCR10; Sheep; Swine; T-Lymphocytes, Regulatory; Transcription, Genetic
PubMed: 17653288
DOI: 10.1371/journal.pone.0000677 -
Scientific Reports Mar 2019Despite recent advances, breast cancer (BrCa) still affects many women and the impact is disproportional in African Americans (AA) compared to European Americans (EA)....
Despite recent advances, breast cancer (BrCa) still affects many women and the impact is disproportional in African Americans (AA) compared to European Americans (EA). Addressing socioeconomic and behavioral status has not been enough to reduce disparity, suggesting contribution of biological differences in BrCa disparity. Our laboratory was first to show involvement of CC chemokines in BrCa. In this study, using ONCOMINE, TCGA, bc-GenExMiner and KMplotter, we examined the association of CC chemokines in BrCa outcomes and disparity. We show over-expression of CCL5, -7, -11, -17, -20, -22 and -25 in BrCa tissues. High mRNA levels of CCL7, -8, -17, -20 and -25 predicted a decrease in overall survival (OS). CCL7 and CCL8 were associated with decreased relapse-free survival. Expression of CCL17 and CCL25 was associated with decreased OS in AA. In EA, CCL8 was associated with decreased OS. Expression of CCL5, -7, -8, -17, -20 and -25 was highest in TNBC. Expression of CCL11 and CCL22 was associated with HER2. CCL7, -8, -17, -20 and -25 were elevated in AAs. In conclusion, our analysis suggests significant association of CC-chemokines in BrCa progression, OS and disparate disease outcome in AA compared to EA patients.
Topics: Breast Neoplasms; Chemokines, CC; Disease Progression; Female; Humans; RNA, Messenger; White People
PubMed: 30850664
DOI: 10.1038/s41598-019-40514-9 -
Clinical and Experimental Immunology Nov 2002
Topics: Anti-HIV Agents; Antiretroviral Therapy, Highly Active; Chemokines, CC; HIV Infections; HIV-1; Humans
PubMed: 12390302
DOI: 10.1046/j.1365-2249.2002.02010.x -
Biomolecules Oct 2022Human C-C motif ligand 16 (CCL16) is a chemokine that is distinguished by a large cleavable C-terminal extension of unknown significance. Conflicting data have been...
Human C-C motif ligand 16 (CCL16) is a chemokine that is distinguished by a large cleavable C-terminal extension of unknown significance. Conflicting data have been reported concerning its tissue distribution and modulation of expression, rendering the biological function of CCL16 enigmatic. Here, we report an integrated approach to the characterisation of this chemokine, including a re-assessment of its expression characteristics as well as a biophysical investigation with respect to its structure and dynamics. Our data indicate that CCL16 is chiefly synthesised by hepatocytes, without an appreciable response to mediators of inflammation, and circulates in the blood as a full-length protein. While the crystal structure of CCL16 confirms the presence of a canonical chemokine domain, molecular dynamics simulations support the view that the C-terminal extension impairs the accessibility of the glycosaminoglycan binding sites and may thus serve as an intrinsic modulator of biological activity.
Topics: Humans; Chemokines, CC; Chemokines; Ligands; Glycosaminoglycans
PubMed: 36358937
DOI: 10.3390/biom12111588 -
Theranostics 2021Considerable evidence suggests that breast cancer metastasis and recurrence occur due to emergence of cancer stem cells (CSCs). In our previous study, we designed a...
Considerable evidence suggests that breast cancer metastasis and recurrence occur due to emergence of cancer stem cells (CSCs). In our previous study, we designed a high-throughput siRNA screening platform that identifies inflammation genes involved in the regulation of cancer cell stemness. We reported that CCL16 protein decreases OCT4 expression and reduces the ALDH+ subpopulation. However, the mechanism by which CCL16 maintains stem cell-like properties remains unclear. Tissue microarrays were used to evaluate CCL16 expression. Cancer stemness assays were performed in CCL16 knockdown and overexpressing cells and in a xenograft model . Human phosphokinase array, immunofluorescence and chromatin immunoprecipitation assays were performed to explore the underlying mechanism. We report that CCL16 was overexpressed in breast tumors and significantly correlated with clinical progression. We found that silencing CCL16 in MDA-MB-231 and BT549 cells diminished CSC properties including ALDH+ subpopulation, side population, chemo-resistance, and sphere formation. Furthermore, mice bearing CCL16-silenced MDA-MB-231 xenografts had lower tumorigenic frequency and developed smaller tumors. Exploration of the underlying mechanism found that CCL16 selects CCR2 to activate p-AKT/GSK3β signaling and facilitate β-catenin nuclear translocation. Further, CCL16 binds to the OCT4 promoter and promotes OCT4 expression. In addition, shRNAs targeting CCR2 and XAV939 targeting β-catenin abolished CCL16-mediated cancer stemness. Upstream, IL10 mediates STAT3 activation, which binds to the CCL16 promoter and enhances its expression. The STAT3-targeted inhibitor Stattic suppressed CCL16 expression and restrained tumor progression . We identified a potential CSC regulator and suggest a novel mechanism for how CCL16 governs cancer cell stemness. We propose that CCL16 could be an effective target for breast cancer therapy.
Topics: Animals; Apoptosis; Biomarkers, Tumor; Breast Neoplasms; Cell Proliferation; Chemokines, CC; Female; Gene Expression Regulation, Neoplastic; Glycogen Synthase Kinase 3 beta; Humans; Mice; Mice, Inbred NOD; Mice, SCID; Neoplastic Stem Cells; Octamer Transcription Factor-3; Receptors, CCR2; Tumor Cells, Cultured; Xenograft Model Antitumor Assays; beta Catenin
PubMed: 33500726
DOI: 10.7150/thno.51000 -
Infectious Diseases in Obstetrics and... 2001The mechanism whereby the placental cells of a human immunodeficiency virus (HIV)-1-infected mother protect the fetus from HIV-1 infection is unclear. Interferons (IFNs)...
OBJECTIVE
The mechanism whereby the placental cells of a human immunodeficiency virus (HIV)-1-infected mother protect the fetus from HIV-1 infection is unclear. Interferons (IFNs) inhibit the replication of viruses by acting at various stages of the life cycle and may play a role in protecting against vertical transmission of HIV-1. In addition the beta-chemokines RANTES (regulated on activation T cell expressed and secreted), macrophage inflammatory protein-1-alpha (MIP-1alpha), and MIP-1beta can block HIV-1 entry into cells by preventing the binding of the macrophage-trophic HIV-1 strains to the coreceptor CCR5. In this study the production of IFNs and beta-chemokines by placental trophoblasts of HIV-1-infected women who were HIV-1 non-transmitters was examined.
METHODS
Placental trophoblastic cells were isolated from 29 HIV-1-infected and 10 control subjects. Supernatants of trophoblast cultures were tested for the production of IFNs and beta-chemokines by enzyme linked immunosorbent assay (ELISA). Additionally, HIV-1-gag and IFN-beta transcripts were determined by a semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) assay.
RESULTS
All placental trophoblasts of HIV-1-infected women contained HIV-1-gag transcripts. There were no statistical differences in the median constitutive levels of IFN-alpha and IFN-gamma produced by trophoblasts of HIV-1 infected and control subjects. In contrast, trophoblasts of HIV-1-infected women constitutively produced significantly higher levels of IFN-beta protein than trophoblasts of control subjects. Furthermore, the median levels of beta-chemokines produced by trophoblasts of HIV-infected and control women were similar.
CONCLUSIONS
Since there was no correlation between the placental HIV load and the production of interferons or beta-chemokines, the role of trophoblast-derived IFNs and beta-chemokines in protecting the fetus from infection with HIV-1 is not clear.
Topics: CD4 Lymphocyte Count; Chemokines, CC; Enzyme-Linked Immunosorbent Assay; Female; HIV Infections; HIV-1; Humans; Infant, Newborn; Infectious Disease Transmission, Vertical; Interferon-beta; Interferon-gamma; Interferons; Pregnancy; Pregnancy Complications, Infectious; Pregnancy Outcome; Reverse Transcriptase Polymerase Chain Reaction; Trophoblasts; Viral Load; Virus Replication
PubMed: 11495560
DOI: 10.1155/S1064744901000175 -
Nature Communications Dec 2023The C-C motif chemokine receptor 8 (CCR8) is a class A G-protein coupled receptor that has emerged as a promising therapeutic target in cancer. Targeting CCR8 with an...
The C-C motif chemokine receptor 8 (CCR8) is a class A G-protein coupled receptor that has emerged as a promising therapeutic target in cancer. Targeting CCR8 with an antibody has appeared to be an attractive therapeutic approach, but the molecular basis for chemokine-mediated activation and antibody-mediated inhibition of CCR8 are not fully elucidated. Here, we obtain an antagonist antibody against human CCR8 and determine structures of CCR8 in complex with either the antibody or the endogenous agonist ligand CCL1. Our studies reveal characteristic antibody features allowing recognition of the CCR8 extracellular loops and CCL1-CCR8 interaction modes that are distinct from other chemokine receptor - ligand pairs. Informed by these structural insights, we demonstrate that CCL1 follows a two-step, two-site binding sequence to CCR8 and that antibody-mediated inhibition of CCL1 signaling can occur by preventing the second binding event. Together, our results provide a detailed structural and mechanistic framework of CCR8 activation and inhibition that expands our molecular understanding of chemokine - receptor interactions and offers insight into the development of therapeutic antibodies targeting chemokine GPCRs.
Topics: Humans; Chemokines, CC; Receptors, CCR8; Ligands; Chemokine CCL1; Receptors, Chemokine; Antibodies
PubMed: 38040762
DOI: 10.1038/s41467-023-43601-8