RESUMO
BACKGROUND: To evaluate the apoptosis inhibitor of macrophage (AIM) deposition patterns on the kidneys of children with IgA nephropathy (IgAN) and Henoch-Schönlein purpura nephritis (HSPN) and to investigate the clinical usefulness of serum and/or urinary AIM levels as biomarkers for the disease activity. METHODS: Immunohistochemical study was performed in the kidneys of 37 patients with IgAN and 10 patients with HSPN. Serum and urinary AIM levels in the patients and 20 healthy controls (HCs) were quantified by enzyme-linked immunosorbent assay. The results were compared with clinical features. RESULTS: In patients with IgAN and HSPN, AIM expression was observed in various areas, including the glomerular mesangial and capillary areas, the proximal and distal tubular epithelial cells, and on infiltrating macrophages in the glomeruli and interstitial areas. Serum and urinary AIM levels were significantly elevated in these patients compared with the HCs. Urinary AIM levels were positively correlated with the histological severity and degree of proteinuria and hematuria as well as urinary ß2 microglobulin and urinary N-acetyl-ß-D-glucosaminidase levels. CONCLUSIONS: AIM plays an important role in the pathogenesis of IgAN and HSPN. Urinary AIM levels can potentially reflect active renal inflammation in these diseases and may represent a useful biomarker for disease activity. IMPACT: Urinary AIM levels may represent a useful biomarker for disease activity of IgAN and HSPN. AIM expression was observed in the glomeruli, tubular epithelial cells, and infiltrating macrophages in glomeruli and interstitial area. U-AIM/Cr were significantly correlated not only with proteinuria, hematuria, and u-ß2MG and u-NAG levels but also with the activity index of histological findings in kidney biopsy specimens. Our results can emphasize the important role of AIM in the pathogenesis of IgAN and HSPN.
Assuntos
Proteínas Reguladoras de Apoptose/biossíntese , Biomarcadores/metabolismo , Glomerulonefrite por IGA/genética , Vasculite por IgA/genética , Receptores Depuradores/biossíntese , Adolescente , Apoptose , Biópsia , Estudos de Casos e Controles , Criança , Pré-Escolar , Feminino , Glomerulonefrite por IGA/metabolismo , Humanos , Vasculite por IgA/metabolismo , Imuno-Histoquímica , Inflamação , Japão , Rim/patologia , Glomérulos Renais/metabolismo , Contagem de Leucócitos , Macrófagos/metabolismo , MasculinoRESUMO
Apoptosis inhibitor of macrophages (AIM) is a protein which plays important roles in controlling the immune response and inflammation in human and mice. In dogs, AIM is reported to be expressed in cancerated macrophages and regulate the survival of these tumor cells. In this study, to elucidate the physiological expression pattern of AIM in dogs, systemic expression and distribution of AIM of dogs were investigated. Mature healthy Beagles were used. Various tissues, peripheral blood cells, and bone marrow cells of normal dogs were collected for in situ hybridization, real-time RT-PCR, and immunohistochemistry. AIM mRNA and protein were expressed in tissue macrophages of the spleen, liver, lungs, and lymph nodes, but not in the microglia of the cerebrum. Proximal tubules in the kidney also expressed AIM protein. Monocytes and B lymphocytes in circulating blood and a part of microvasculature endothelial cells showed AIM expression at both the mRNA and protein levels. In the bone marrow, early-stage monocyte progenitor-like cells expressed AIM mRNA and protein. These results clarified that AIM is expressed in more cell types than previously reported in human and mice. These data spread the possibility of AIM physiological functions and implies the relationship of AIM to the maturation of macrophage-strain cells in dogs and other species.
Assuntos
Linfócitos B/metabolismo , Células Endoteliais/metabolismo , Regulação da Expressão Gênica , Macrófagos/metabolismo , Receptores Depuradores/biossíntese , Animais , Linfócitos B/citologia , Cães , Células Endoteliais/citologia , Humanos , Macrófagos/citologia , Camundongos , Especificidade de Órgãos , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Especificidade da EspécieRESUMO
BACKGROUND AIMS: Few studies have examined the migration pattern of natural killer (NK) cells, especially after radiation treatment for cancer. We investigated whether irradiation can modulate the expression of chemokines in cancer cells and the migration of NK cells to irradiated tumor cells. METHODS: The expression of chemokine receptors (CXCR3, CXCR4 and CXCR6) on interleukin-2 (IL-2)/IL-15-activated NK cells was assessed using flow cytometry. Related chemokine ligands (CXCL11, CXCL12 and CXCL16) in human breast cancer cell lines (MCF7, SKBR3 and MDA-MB231) irradiated at various doses were assessed using reverse transcription-polymerase chain reaction (RT-PCR), fluorescence-activated cell sorting (FACS) and enzyme-linked immunosorbent assay (ELISA). The cell-free culture supernatant was collected 96 h after irradiation of breast cancer cell lines for migration and blocking assays. RESULTS: The activated NK cells expressed CXCR6. Expression of the CXCR6 ligand CXCL16 increased in a time- and dose-dependent manner in all analyzed cancer cell lines. CXCL16 expression was statistically significantly enhanced in all breast cancer cell lines on day 3 after 20 Gy irradiation. Activated NK cells migration correlated with CXCL16 concentration (R2 = 0.91; P <0.0001). Significantly enhanced migration of NK cells to irradiated cancer cells was observed for a dose of 20 Gy in MCF7 (P = 0.043) and SKBR3 (P = 0.043) cells, but not in MDA-MB231 (P = 0.225) cells. A blocking assay using a CXCR6 antibody showed a significant decrease in the migration of activated NK cells in all cancer cell lines. CONCLUSIONS: Our data indicate that irradiation induces CXCL16 chemokine expression in cancer cells and enhances the migration of activated NK cells expressing CXCR6 to irradiated breast cancer cells. These results suggest that radiation would improve the anti-tumor effect of NK cells through enhanced migration of NK cells to tumor site for the treatment of patients with breast cancer.
Assuntos
Neoplasias da Mama/radioterapia , Movimento Celular/efeitos da radiação , Quimiocinas CXC/biossíntese , Células Matadoras Naturais/imunologia , Receptores de Quimiocinas/biossíntese , Receptores Depuradores/biossíntese , Receptores Virais/biossíntese , Anticorpos Bloqueadores/farmacologia , Linhagem Celular Tumoral , Quimiocina CXCL12/biossíntese , Quimiocina CXCL16 , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Humanos , Interleucina-15/metabolismo , Interleucina-2/metabolismo , Células Matadoras Naturais/metabolismo , Células MCF-7 , Receptores CXCR3/biossíntese , Receptores CXCR4/biossíntese , Receptores CXCR6 , Receptores de Quimiocinas/imunologia , Receptores Virais/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/imunologiaRESUMO
Chemokine (C-X-C motif) ligand 16 (CXCL16) is a new angiogenic factor inducing angiogenesis via extracellular signal-regulated kinases pathway. To further understand the molecular mechanism underlying CXCL16induced angiogenesis, we explored involvement of other relevant pathways in CXCL16-induced angiogenesis. In the present study, we investigated the mechanisms underlying CXCL16-induced angiogenesis in human umbilical vein endothelial cells (HUVECs). CXCL16 promoted HUVEC proliferation, tube formation and migration. Enzyme-linked immunosorbent assay revealed that CXCL16 induced vascular endothelial growth factor secretion from HUVECs. Western blot analysis showed that CXCL16 increased the level of hypoxiainducible factor 1α, p-extracellular signal-regulated kinases (ERK), p-p38 and p-Akt dose- and time-dependently. ERK-, p38- and Akt-selective inhibitors significantly suppressed HUVEC proliferation, migration, tube formation and hypoxia-inducible factor 1α (HIF-1α) expression induced by CXCL16. Furthermore, CXCL16 peptides induced CXCL16 secretion via ERK, p38 and Akt pathways, which was suppressed by HIF-1α-selective inhibitor PX12. Our data suggest that CXCL16 induces angiogenesis in autocrine manner via ERK, Akt, p38 pathways and HIF-1α modulation.
Assuntos
Comunicação Autócrina/genética , Quimiocinas CXC/biossíntese , Subunidade alfa do Fator 1 Induzível por Hipóxia/biossíntese , Neovascularização Patológica/genética , Receptores Depuradores/biossíntese , Movimento Celular/genética , Proliferação de Células/genética , Quimiocina CXCL16 , Quimiocinas CXC/genética , Regulação Neoplásica da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Sistema de Sinalização das MAP Quinases/genética , Neovascularização Patológica/patologia , Proteínas Proto-Oncogênicas c-akt/genética , Receptores Depuradores/genética , Fator A de Crescimento do Endotélio Vascular/biossíntese , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
Nasopharyngeal carcinoma (NPC) is a common malignancy in southern China and Southeast Asia. NPC frequently metastasizes to the bone in advanced patients resulting in high mortality. The molecular mechanisms for NPC development and cancer-induced bone lesions are unclear. In this study, we firstly determined chemokine receptor CCR2 and CXCR6 expressions in clinical specimens and CNE2, SUNE1, CNE1, and HK1 cell lines. Then, we measured chemokine CCL2 and CXCL16 production in these NPC cell lines by ELISA. Expression levels of these chemokines and their receptors were observed to positively correlate with tumor aggressiveness. Furthermore, U0126 (MEK inhibitor) was used to treat these NPC cell lines. CCL2 and CXCL16 expression levels and cell proliferation were significantly inhibited by U0126 in a dose- and time-dependent manner. Finally, we collected conditioned medium (CM) from NPC cell cultures in the presence of U0126 treatment. When mouse bone marrow non-adherent cells were treated with the CM, the numbers of multinucleated osteoclast formation were dramatically diminished. These results indicate that MEK inhibitor diminishes NPC cell proliferation and NPC-induced osteoclastogenesis via modulating CCL2 and CXCL16 expressions. This study provides novel therapeutic targets such as CCL2/CCR2 and CXCL16/CXCR6 for advanced NPC patients.
Assuntos
Quimiocina CCL2/biossíntese , Quimiocinas CXC/biossíntese , MAP Quinase Quinase Quinases/genética , Neoplasias Nasofaríngeas/genética , Receptores Depuradores/biossíntese , Animais , Células da Medula Óssea/efeitos dos fármacos , Butadienos/administração & dosagem , Carcinoma , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quimiocina CCL2/genética , Quimiocina CXCL16 , Quimiocinas CXC/genética , Meios de Cultivo Condicionados/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , MAP Quinase Quinase Quinases/antagonistas & inibidores , Camundongos , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/tratamento farmacológico , Neoplasias Nasofaríngeas/patologia , Nitrilas/administração & dosagem , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Inibidores de Proteínas Quinases/administração & dosagem , Receptores Depuradores/genéticaRESUMO
BACKGROUND: The chemokine CXCL16 and its receptor CXCR6 are expressed by a variety of immune cells and have been shown to influence angiogenesis. The expression of CXCR6 and CXCL16 has been examined in numerous human cancers; however no studies have yet investigated their influence on prognosis in non-small cell lung cancer (NSCLC). We aimed to explore their prognostic significance in NSCLC, in addition to examining associations with previously investigated markers. METHODS: Resected tumor tissue from 335 consecutive unselected stage I-IIIA NSCLC patients (1990-2005) were collected. Immunohistochemistry was used to evaluate the expression of CXCR6 and CXCL16 on tissue microarrays. In vitro, NSCLC cells (NCI-H460, A549 cells) were transfected with CXCL16 siRNA to examine effects on proliferation. RESULTS: In univariate analysis, ↑ stromal cell CXCL16 expression was a significant positive prognostic factor (P = 0.016). CXCR6 was expressed in cancer cells, but did not show any prognostic impact. In the multivariate analysis, combined ↑cancer, and ↑stromal cell CXCL16 expression was an independent positive prognostic factor when compared to ↓stromal and ↓cancer cell expression (HR: 0.42; 95 % CI: 0.20-0.88; P = 0.022). Knockdown of CXCL16 by siRNA resulted in accelerated proliferation of NSCLC cell lines. CONCLUSION: We have shown that combined ↑cancer and ↑stromal cell CXCL16 expression is an independent positive prognostic factor in NSCLC. Further studies are warranted to elucidate the biological mechanism underlying this finding.
Assuntos
Biomarcadores Tumorais/biossíntese , Carcinoma Pulmonar de Células não Pequenas/genética , Quimiocinas CXC/biossíntese , Receptores de Quimiocinas/biossíntese , Receptores Depuradores/biossíntese , Receptores Virais/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Quimiocina CXCL16 , Quimiocinas CXC/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neovascularização Patológica/genética , Neovascularização Patológica/patologia , Prognóstico , Receptores CXCR6 , Receptores de Quimiocinas/genética , Receptores Depuradores/genética , Receptores Virais/genéticaRESUMO
Our previous studies demonstrate that CXCL6/CXCR6 chemokine axis induces prostate cancer progression by the AKT/mTOR signaling pathway; however, its role and mechanisms underlying invasiveness and metastasis of breast cancer are yet to be elucidated. In this investigation, CXCR6 protein expression was examined using high-density tissue microarrays and immunohistochemistry. Expression of CXCR6 shows a higher epithelial staining in breast cancer nest site and metastatic lymph node than the normal breast tissue, suggesting that CXCR6 may be involved in breast cancer (BC) development. In vitro and in vivo experiments indicate that overexpression of CXCR6 in BC cells has a marked effect on increasing cell migration, invasion and metastasis. In contrast, reduction of CXCR6 expression by shRNAs in these cells greatly reduce its invasion and metastasis ability. Mechanistic analyses show that CXCL16/CXCR6 chemokine axis is capable of modulating activation of RhoA through activating ERK1/2 signaling pathway, which then inhibits the activity of cofilin, thereby enhancing the stability of F-actin, responsible for invasiveness and metastasis of BC. Taken together, our data shows for the first time that the CXCR6 / ERK1/2/ RhoA / cofilin /F-actin pathway plays a central role in the development of BC. Targeting the signaling pathway may prove beneficial to prevent metastasis and provide a more effective therapeutic strategy for BC.
Assuntos
Neoplasias da Mama/metabolismo , Quimiocinas CXC/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Receptores de Quimiocinas/metabolismo , Receptores Depuradores/metabolismo , Receptores Virais/metabolismo , Fatores de Despolimerização de Actina/metabolismo , Actinas/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Quimiocina CXCL16 , Quimiocinas CXC/biossíntese , Quimiocinas CXC/genética , Progressão da Doença , Feminino , Humanos , Células MCF-7 , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/genética , Invasividade Neoplásica , Fosforilação , Receptores CXCR6 , Receptores de Quimiocinas/biossíntese , Receptores de Quimiocinas/genética , Receptores Depuradores/biossíntese , Receptores Depuradores/genética , Receptores Virais/biossíntese , Receptores Virais/genética , Transdução de Sinais , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
Non-alcoholic fatty liver disease (NAFLD) is a major cause of morbidity and mortality in developed countries, resulting in steatohepatitis (NASH), fibrosis and eventually cirrhosis. Modulating inflammatory mediators such as chemokines may represent a novel therapeutic strategy for NAFLD. We recently demonstrated that the chemokine receptor CXCR6 promotes hepatic NKT cell accumulation, thereby controlling inflammation in experimental NAFLD. In this study, we first investigated human biopsies (n = 20), confirming that accumulation of inflammatory cells such as macrophages is a hallmark of progressive NAFLD. Moreover, CXCR6 gene expression correlated with the inflammatory activity (ALT levels) in human NAFLD. We then tested the hypothesis that pharmacological inhibition of CXCL16 might hold therapeutic potential in NAFLD, using mouse models of acute carbon tetrachloride (CCl4)- and chronic methionine-choline-deficient (MCD) diet-induced hepatic injury. Neutralizing CXCL16 by i.p. injection of anti-CXCL16 antibody inhibited the early intrahepatic NKT cell accumulation upon acute toxic injury in vivo. Weekly therapeutic anti-CXCL16 administrations during the last 3 weeks of 6 weeks MCD diet significantly decreased the infiltration of inflammatory macrophages into the liver and intrahepatic levels of inflammatory cytokines like TNF or MCP-1. Importantly, anti-CXCL16 treatment significantly reduced fatty liver degeneration upon MCD diet, as assessed by hepatic triglyceride levels, histological steatosis scoring and quantification of lipid droplets. Moreover, injured hepatocytes up-regulated CXCL16 expression, indicating that scavenging functions of CXCL16 might be additionally involved in the pathogenesis of NAFLD. Targeting CXCL16 might therefore represent a promising novel therapeutic approach for liver inflammation and steatohepatitis.
Assuntos
Quimiocina CXCL6/biossíntese , Quimiocinas CXC/biossíntese , Fígado/metabolismo , Macrófagos/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Receptores Depuradores/biossíntese , Regulação para Cima , Animais , Intoxicação por Tetracloreto de Carbono/metabolismo , Intoxicação por Tetracloreto de Carbono/patologia , Quimiocina CXCL16 , Doença Crônica , Feminino , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Fígado/patologia , Macrófagos/patologia , Masculino , Camundongos , Hepatopatia Gordurosa não Alcoólica/patologiaRESUMO
BACKGROUND: Cardiac inflammation has been suggested to play a critical role in the pathogenesis of inflammatory cardiomyopathy as well as in progressive heart failure (HF). CXC motif ligand 16 (CXCL16) is a recently discovered chemokine produced by several inflammatory cells and representing an important pathogenic mediator in the development of HF. The present study evaluates the diagnostic and prognostic relevance of CXCL16 expression in endomyocardial biopsies of consecutive patients with congestive HF. METHODS AND RESULTS: 174 patients (age 54.4±14.6 years) with congestive HF undergoing endomyocardial biopsy for diagnostic reasons were prospectively enrolled. Biopsies were analyzed using established histopathological and immunohistological criteria together with CXCL16 staining. CXCL16 was significantly enhanced in patients with inflammatory cardiomyopathy (78/127, 61.4%) as compared to patients with non-inflammatory cardiomyopathy (17/47, 36.2%, p=0.003). During a mean follow-up of 27.5 months, 20 patients (11.5%) reached the primary endpoint (death of all causes). Of all clinical (age, gender, NYHA functional class, systolic pulmonary artery pressure, left ventricular ejection fraction), laboratory (brain natriuretic peptide) and immunohistological (CXCL16) parameters tested, CXCL16 was the only independent predictor of death (hazard ratio 5.4; 95% confidence interval 1.2 to 24.0; p=0.027). Subgroup analysis revealed CXCL16 as a predictor of death in both patients with inflammatory and with non-inflammatory cardiomyopathy. CONCLUSION: According to the present observations CXCL16 is enhanced in inflammatory cardiomyopathy and turned out as an independent predictor of death in patients with HF undergoing endomyocardial biopsy.
Assuntos
Cardiomiopatias/metabolismo , Cardiomiopatias/mortalidade , Quimiocinas CXC/biossíntese , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/mortalidade , Receptores Depuradores/biossíntese , Adulto , Idoso , Biomarcadores/metabolismo , Cardiomiopatias/diagnóstico , Quimiocina CXCL16 , Estudos de Coortes , Feminino , Seguimentos , Insuficiência Cardíaca/diagnóstico , Humanos , Inflamação/diagnóstico , Inflamação/metabolismo , Inflamação/mortalidade , Masculino , Pessoa de Meia-Idade , Mortalidade/tendências , Valor Preditivo dos Testes , Estudos ProspectivosRESUMO
BACKGROUND: In certain cancers, expression of CXCL16 and its receptor CXCR6 associate with lymphocyte infiltration, possibly aiding anti-tumour immune response. In other cancers, CXCL16 and CXCR6 associate with pro-metastatic activity. In the current study, we aimed to characterise the role of CXCL16, sCXCL16, and CXCR6 in ovarian cancer (OC). METHODS: CXCL16/CXCR6 expression was analysed on tissue microarray containing 306 OC patient samples. Pre-treatment serum sCXCL16 was determined in 118 patients using ELISA. In vitro, (primary) OC cells were treated with an ADAM-10/ADAM-17 inhibitor (TAPI-2) and an ADAM-10-specific inhibitor (GI254023x), whereupon CXCL16 levels were evaluated on the cell membrane (immunofluorescent analysis, western blots) and in culture supernatants (ELISA). In addition, cell migration was assessed using scratch assays. RESULTS: sCXCL16 independently predicted for poor survival (hazard ratio=2.28, 95% confidence interval=1.29-4.02, P=0.005), whereas neither CXCL16 nor CXCR6 expression correlated with survival. Further, CXCL16/CXCR6 expression and serum sCXCL16 levels did not associate with lymphocyte infiltration. In vitro inhibition of both ADAM-17 and ADAM-10, but especially the latter, decreased CXCL16 membrane shedding and strongly reduced cell migration of A2780 and cultured primary OC-derived malignant cells. CONCLUSIONS: High serum sCXCL16 is a prognostic marker for poor survival of OC patients, possibly reflecting ADAM-10 and ADAM-17 pro-metastatic activity. Therefore, serum sCXCL16 levels may be a pseudomarker that identifies patients with highly metastatic tumours.
Assuntos
Proteínas ADAM/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Quimiocinas CXC/sangue , Proteínas de Membrana/metabolismo , Neoplasias Ovarianas/sangue , Receptores Depuradores/sangue , Proteína ADAM10 , Proteína ADAM17 , Quimiocina CXCL16 , Quimiocinas CXC/biossíntese , Feminino , Humanos , Imuno-Histoquímica , Metástase Neoplásica , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/patologia , Prognóstico , Estudos Prospectivos , Receptores CXCR6 , Receptores de Quimiocinas/biossíntese , Receptores de Quimiocinas/sangue , Receptores Depuradores/biossíntese , Receptores Virais/biossíntese , Receptores Virais/sangue , Análise de Sobrevida , Análise Serial de TecidosRESUMO
Chemokines are a family of peptide mediators that play an essential role in cellular migration and intracellular communication in tumor cells as well as immune cells. We hypothesized that the CXCL16-CXCR6 ligand-receptor system plays an important role in Ewing sarcoma (ES) family tumor (ESFT) progression. Using real-time quantitative reverse transcription-polymerase chain reaction, we investigated the mRNA expression of CXCL16, CXCR6, and ADAM 10 in various cell lines. We also investigated the expression of CXCL16, CXCR6, ADAM 10, and ADAM 17 in tissue samples from 61 ESFT patients using immunohistochemistry. The mRNA expression levels of CXCL16 and CXCR6 in the ES cell line were higher than those in the other cell lines. Immunohistochemical staining revealed that CXCL16 and CXCR6 were highly expressed in tumor cells of ESFT and showed a positive correlation between them. The expression of CXCL16 and CXCR6 was associated with the occurrence of lung metastasis. Univariate analysis revealed that CXCL16 or CXCR6 expression was associated with worse prognosis of ESFT patients. In addition, CXCL16 and CXCR6 expression was associated with shorter overall survival irrespective of other prognostic factors. Our results suggest that the CXCL16/CXCR6 axis appears to be important in the progression of ESFT, resulting in more aggressive clinical behavior. Furthermore, there may be a decrease in the overall survival in ESFT patients who have tumors that stain strongly for CXCL16 and CXCR6.
Assuntos
Biomarcadores Tumorais/análise , Neoplasias Ósseas/metabolismo , Quimiocinas CXC/biossíntese , Receptores de Quimiocinas/biossíntese , Receptores Depuradores/biossíntese , Receptores Virais/biossíntese , Sarcoma de Ewing/metabolismo , Adolescente , Adulto , Neoplasias Ósseas/mortalidade , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Quimiocina CXCL16 , Quimiocinas CXC/análise , Criança , Pré-Escolar , Suscetibilidade a Doenças , Feminino , Humanos , Imuno-Histoquímica , Lactente , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real , Receptores CXCR6 , Receptores de Quimiocinas/análise , Receptores Depuradores/análise , Receptores Virais/análise , Sarcoma de Ewing/mortalidade , Sarcoma de Ewing/patologia , Transcriptoma , Adulto JovemRESUMO
OBJECTIVE: Cannabinoid receptors are activated in murine macrophages upon exposure to oxidized low-density lipoproteins (oxLDL), and type-1 cannabinoid receptor (CB1R) is considered as a risk factor in atherosclerosis, because it promotes cholesterol accumulation and release of inflammatory mediators. Conversely, accumulated evidence suggests a protective role for type-2 cannabinoid receptor (CB2R). Here, we sought to ascertain whether different elements of the endocannabinoid system (ECS) were activated in human lipid-laden macrophages, and whether CB2R played any role in atherogenesis and inflammation of these cells. METHODS AND RESULTS: Human macrophages were exposed to oxLDL in order to obtain lipid-laden foam cells. Liquid chromatography/mass spectrometry (LC/MS) was used to measure the production of the endocannabinoids in both macrophages and foam cells, and radiometric assays were performed to measure cannabinoid receptor binding and activity of endocannabinoid metabolizing enzymes. OxLDL accumulation was investigated by confocal imaging, and cytokine production and release were measured by means of flow cytometry and ELISA. The results showed that human macrophages possess a fully functional ECS, which was modulated by oxLDL. Selective CB2R activation reduced cellular oxLDL accumulation, which was associated with decreased expression of CD36 scavenger receptor, and decreased production of TNFα, IL-12 and IL-10. These anti-atherogenic and anti-inflammatory effects were reverted by the selective CB2R antagonist SR144528. CONCLUSIONS: A fully active ECS is present in human macrophages and macrophage-derived foam cells. Selective activation of CB2R reduces CD36-dependent oxLDL accumulation and modulates production of inflammatory cytokines, thus representing a potential therapeutic strategy to combat atherosclerosis.
Assuntos
Endocanabinoides/fisiologia , Células Espumosas/metabolismo , Macrófagos/metabolismo , Receptor CB2 de Canabinoide/fisiologia , Canfanos/farmacologia , Linhagem Celular , Humanos , Lipoproteínas LDL/metabolismo , Proteínas de Membrana Lisossomal/biossíntese , Pirazóis/farmacologia , Receptor CB2 de Canabinoide/antagonistas & inibidores , Receptores Depuradores/biossínteseRESUMO
Autophagy and phagocytosis are conserved cellular functions involved in innate immunity. However, the nature of their interactions remains unclear. We evaluated the role of autophagy in regulating phagocytosis in macrophages from myeloid-specific autophagy-related gene 7-deficient (Atg7â»/â») mice. Atg7â»/â» macrophages exhibited higher bacterial uptake when infected with Mycobacterium tuberculosis (Mtb) or with M. tuberculosis var. bovis BCG (BCG). In addition, BCG-infected Atg7â»/â» mice showed increased bacterial loads and exacerbated lung inflammatory responses. Atg7â»/â» macrophages had increased expression of two class A scavenger receptors: macrophage receptor with collagenous structure (MARCO) and macrophage scavenger receptor 1 (MSR1). The increase in scavenger receptors was caused by increased activity of the nuclear factor (erythroid-derived 2)-like 2 (NFE2L2) transcription factor resulting from accumulated sequestosome 1 (SQSTM1 or p62) in Atg7â»/â» macrophages. These insights increase our understanding of the host-pathogen relationship and suggest that therapeutic strategies should be designed to include modulation of both phagocytosis and autophagy.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Autofagia/imunologia , Proteínas de Choque Térmico/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Fagocitose/imunologia , Receptores Imunológicos/biossíntese , Receptores Depuradores Classe A/biossíntese , Animais , Proteína 7 Relacionada à Autofagia , Carga Bacteriana/imunologia , Células Cultivadas , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/genética , Infecções por Mycobacterium/imunologia , Mycobacterium bovis/crescimento & desenvolvimento , Mycobacterium bovis/imunologia , Mycobacterium tuberculosis/crescimento & desenvolvimento , Mycobacterium tuberculosis/imunologia , Pneumonia/imunologia , Pneumonia/microbiologia , Receptores Depuradores/biossíntese , Receptores Depuradores/imunologia , Proteína Sequestossoma-1 , Regulação para CimaRESUMO
BACKGROUND: Human enterovirus type 71 (EV71) and Coxsackievirus A group type 16 (CA16) belong to human Enterovirus species A of the family Picornaviridae. These viruses are recognized as the major pathogens responsible for epidemics of hand-foot-mouth disease (HFMD), which presents with fever and vesicular eruptions of palms, soles of the feet or mouth. Human scavenger receptor class B, member 2 (SCARB2) has been identified as the receptor for both EV71 and CA16, as overexpression of SCARB2 in cells can enhance virus replication significantly. METHODS: In this study, we used a lentivirus packaging vector to transduce the SCARB2 gene into human embryonic kidney cells (293), human rhabdomyosarcoma cells (RD) and African green monkey kidney cells (Vero) to create stable expression lines. Expression of SCARB2 in the resulting three transgenic cell lines was confirmed by real-time RT-PCR, immunofluorescence and flow cytometry. RESULTS: Levels of SCARB2 mRNA determined by real-time RT-PCR in 293-SCARB2 (293S) or RD-SCARB2 (RDS) transgenic cell lines were approximately 2 × 10(2) times higher than those in 293 and RD cells, respectively, and three times higher in Vero-SCARB2 (VeroS) than in Vero cells. Furthermore, EV71 and CA16 virus titers in 293S and RDS cells were 10(2)-10(3)-fold higher (detected in RD cell) than those in the parental cells, and a 10-fold higher titer of EV71 was achieved in VeroS cells compared with that in Vero cells. CONCLUSIONS: We established for the first time three cell lines stably overexpressing SCARB2, which showed drastic increases in susceptibility to EV71/CA16 infection. These optimal cell lines may be utilized to develop inactivated vaccines for EV71/CA16 and facilitate rapid detection and isolation of HFMD pathogens or other Enterovirus serotypes. Furthermore, these stable cell lines also can serve as tools to facilitate drug screenings as well as molecular studies on virus-host interactions and pathogenesis of causative agents for HFMD.
Assuntos
Enterovirus Humano A/crescimento & desenvolvimento , Enterovirus/crescimento & desenvolvimento , Expressão Gênica , Proteínas de Membrana Lisossomal/biossíntese , Receptores Depuradores/biossíntese , Receptores Virais/biossíntese , Animais , Linhagem Celular , Chlorocebus aethiops , Humanos , Proteínas de Membrana Lisossomal/genética , Receptores Depuradores/genética , Receptores Virais/genética , Transdução Genética , Cultura de Vírus/métodosRESUMO
The present study examined a correlation between CXC chemokine ligand 16 (CXCL16) and cell differentiation antigen 99 (CD99) expression and investigated the role of CXCL16 in human lymphoma cell lines and clinical samples. Cytokine antibody arrays were used to measure the differentially expressed cytokines in tumor tissues. The expression of CXCL16 and CD99 was analyzed by quantitative PCR (qPCR) and western blotting, while the pathways involved were assessed by western blotting and enzyme-linked immunosorbent assay (ELISA). The expression of CXCL16 was investigated in 9 lymphoma cell lines (L428, RPMI-8226, KM3, Jurkat, OCI-Ly1, OCI-Ly8, OCI-Ly10, Karpass299 and Raji) as well as in clinical lymphoma samples using qPCR, western blotting and immunochemistry. Soluble CXCL16 (sCXCL16) levels were measured by ELISA and proliferation was analyzed by CCK8 proliferation assays. CXCL16 was one of the upregulated chemokines when lymphoma cells where transferred from in vitro to in vivo conditions. The increased expression and secretion of CXCL16 paralleled with a decrease of mCD99L2 and was accompanied by NF-κB pathway activation and vice versa. CXCL16 was expressed in all 9 lymphoma cell lines with the highest levels in the Hodgkin lymphoma (HL) cell line L428, the plasma cell-derived cell lines RPMI8226 and KM3 and the T leukemia-derived cell line Jurkat. Higher levels of sCXCL16 were secreted by L428 cells, the diffuse large B-cell lymphoma (DLBCL)-derived cell lines (OCI-Ly1, OCI-Ly8 and OCI-Ly10) and Jurkat cells. CXCL16 was widely expressed in clinical samples of lymphoma patients with higher levels in HL compared to non-Hodgkin lymphoma. Human recombinant CXCL16 had no significant effect on L428 cell proliferation, but was able to stimulate CD4+ T lymphocytes to proliferate. CXCL16, inversely correlated with CD99 expression in Hodgkin Reed-Sternberg (H/RS) cells, is widely expressed in diverse types of lymphomas.
Assuntos
Antígenos CD/biossíntese , Moléculas de Adesão Celular/biossíntese , Quimiocinas CXC/biossíntese , Linfoma/metabolismo , Receptores Depuradores/biossíntese , Células de Reed-Sternberg/metabolismo , Antígeno 12E7 , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Quimiocina CXCL16 , Quimiocinas CXC/genética , Quimiocinas CXC/metabolismo , Citocinas/genética , Citocinas/metabolismo , Regulação para Baixo , Feminino , Humanos , Células Jurkat , Linfoma/genética , Linfoma/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/genética , NF-kappa B/metabolismo , Receptores Depuradores/genética , Receptores Depuradores/metabolismo , Regulação para CimaRESUMO
In chronic inflammatory diseases, cardiovascular disease risk is increased and is the main cause of increased mortality. Oxidized LDL (oxLDL) and scavenger receptors participate in atherogenesis. Using human arterial endothelial cells (HAECs), we evaluated the effect of IL-6 and TNF-α on the expression of scavenger receptors. IL-6 induced expression of SR-A mRNA and TNF-α induced both SR-A and LOX-1 mRNA. Both did induce either CD36 or CD68. To assess the function of scavenger receptors, MCP-1 production by oxLDL from cytokine-pretreated HAEC was examined. In accordance with scavenger receptor expression, oxLDL-induced MCP-1 production was increased in IL-6- or TNF-α-pretreatment. Serum from rheumatoid arthritis patients but not from healthy subjects increased mRNA expressions of SR-A, LOX-1 and CD36 in HAEC. SR-A expression was inhibited by both anti-IL-6 receptor antibody (α-IL-6R Ab) and TNF-α receptor (p75)-Fc (TNFR-Fc) and LOX-1 expression was inhibited by TNFR-Fc. CD36 expression was affected by neither. Serum from rheumatoid arthritis patients augmented oxLDL-induced MCP-1 production. Both α-IL-6R Ab and TNFR-Fc partially inhibited this MCP-1 production. In conclusion, our results strongly support that blocking therapy of IL-6 and TNF-α might be beneficial to reduce atherosclerosis risk in chronic inflammatory diseases.
Assuntos
Quimiocina CCL2/antagonistas & inibidores , Interleucina-6/antagonistas & inibidores , Lipoproteínas LDL/antagonistas & inibidores , Receptores Depuradores/biossíntese , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Artrite Reumatoide/sangue , Artrite Reumatoide/metabolismo , Células Cultivadas , Quimiocina CCL2/biossíntese , Células Endoteliais/metabolismo , Humanos , Interleucina-6/fisiologia , Lipoproteínas LDL/biossíntese , Receptores Depuradores/fisiologia , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Staphylococcal nuclease domain-containing 1 (SND1) is a multifunctional protein that is overexpressed in multiple cancers, including hepatocellular carcinoma (HCC). Stable overexpression of SND1 in Hep3B cells expressing a low level of SND1 augments, whereas stable knockdown of SND1 in QGY-7703 cells expressing a high level of SND1 inhibits establishment of xenografts in nude mice, indicating that SND1 promotes an aggressive tumorigenic phenotype. In this study we analyzed the role of SND1 in regulating tumor angiogenesis, a hallmark of cancer. Conditioned medium from Hep3B-SND1 cells stably overexpressing SND1 augmented, whereas that from QGY-SND1si cells stably overexpressing SND1 siRNA significantly inhibited angiogenesis, as analyzed by a chicken chorioallantoic membrane assay and a human umbilical vein endothelial cell differentiation assay. We unraveled a linear pathway in which SND1-induced activation of NF-κB resulted in induction of miR-221 and subsequent induction of angiogenic factors Angiogenin and CXCL16. Inhibition of either of these components resulted in significant inhibition of SND1-induced angiogenesis, thus highlighting the importance of this molecular cascade in regulating SND1 function. Because SND1 regulates NF-κB and miR-221, two important determinants of HCC controlling the aggressive phenotype, SND1 inhibition might be an effective strategy to counteract this fatal malady.
Assuntos
Carcinoma Hepatocelular/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Proteínas Nucleares/metabolismo , Núcleo Celular/metabolismo , Quimiocina CXCL16 , Quimiocinas CXC/biossíntese , Endonucleases , Ensaio de Imunoadsorção Enzimática/métodos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Metástase Neoplásica , Neovascularização Patológica , RNA Interferente Pequeno/metabolismo , Receptores Depuradores/biossíntese , Ribonuclease Pancreático/biossínteseRESUMO
BACKGROUND: Coronary artery disease is the single leading cause of death in the United States. Commonly it is treated with coronary bypass grafting using the saphenous vein (SV) or internal mammary artery (IMA) as a conduit. Unfortunately, the SV has much lower patency rates compared with the IMA. Several hypotheses exist as to why occlusion occurs more commonly in SV grafts than in IMA grafts. However detailed studies in this area have been limited. This study investigates the effects of pressure distention on inflammation in SV conduit used in coronary artery bypass grafting (CABG). METHODS: Saphenous vein distention pressure was measured intraoperatively during 48 CABG procedures. A segment of SV was excised from the conduit before distention. Because the vein was used for coronary artery grafting, sequential pieces were archived for evaluation. Real-time polymerase chain reaction (RT-PCR) and immunohistochemical analyses were performed to investigate a change in the expression of biomarkers. RESULTS: Upregulation of various biomarkers occurred. These biomarkers included scavenger receptors A and B (SR-A, SR-B), toll-like receptors 2 and 4 (TLR2, TLR4), platelet endothelial cell adhesion molecule (PECAM), vascular cell adhesion molecule (VCAM), and intercellular cell adhesion molecule (ICAM) in segments of SV that were subjected to distention. Immunohistochemical results mirrored RT-PCR findings. A significant correlation was observed between biomarkers and pressure values. CONCLUSIONS: These studies demonstrate that markers of inflammation are upregulated in response to SV distention. The data suggest that the pressure used in graft preparation procedures should be regulated to avoid inflammation and its potential to induce graft failure.
Assuntos
Moléculas de Adesão Celular/análise , Ponte de Artéria Coronária/métodos , Oclusão de Enxerto Vascular/etiologia , Receptores Depuradores/análise , Veia Safena/transplante , Coleta de Tecidos e Órgãos/efeitos adversos , Receptores Toll-Like/análise , Idoso , Biomarcadores , Moléculas de Adesão Celular/biossíntese , Moléculas de Adesão Celular/genética , Doença das Coronárias/cirurgia , Feminino , Oclusão de Enxerto Vascular/metabolismo , Humanos , Hiperplasia , Masculino , Pessoa de Meia-Idade , Flebite/etiologia , Flebite/metabolismo , Pressão/efeitos adversos , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real , Receptores Depuradores/biossíntese , Receptores Depuradores/genética , Veia Safena/metabolismo , Veia Safena/patologia , Coleta de Tecidos e Órgãos/métodos , Receptores Toll-Like/biossíntese , Receptores Toll-Like/genética , Regulação para Cima , Grau de Desobstrução VascularRESUMO
BACKGROUND: Oxidative stress-induced endothelial dysfunction and oxidized low-density lipoprotein (LDL) might play a key role in the pathogenesis of atherosclerosis. We recently identified a vascular endothelial scavenger receptor, collectin placenta 1 (CL-P1), which acts as a receptor for oxidized LDL as well as for microbes. METHODS: We demonstrate how hypoxic and oxidative stress induced CL-P1 expression and compared their effects with the expression of lectin-like oxidized low-density lipoprotein receptor 1 (LOX-1), an endothelial scavenger receptor expressed by oxidative stress. RESULTS: Hypoxia/reoxygenation induced CL-P1 mRNA and protein expression in human umbilical vein endothelial cells (HUVECs). The expression of LOX-1 mRNA in these cells peaked slightly at 24 h, while the expression of CL-P1 had an onset at 72 h and was sustained for 120 h after reoxygenation. Furthermore, the exposure of rat carotid artery endothelium to ischemia/reperfusion increased the maximal CL-P1 mRNA expression at 72 h and expression of its protein peaked at 7 days after this treatment. We demonstrate that CL-P1 up-regulation is induced in vitro and in vivo by oxidative stress. GENERAL SIGNIFICANCE: The inducible expression of CL-P1 by oxidative stress might play a crucial role in endothelial dysfunction or chronic activation leading to the pathogenesis of atherosclerosis.