RESUMEN
Group 3 innate lymphoid cells (ILC3s) control the formation of intestinal lymphoid tissues and play key roles in intestinal defense. They express neuropeptide vasoactive intestinal peptide (VIP) receptor 2 (VPAC2), through which VIP modulates their function, but whether VIP exerts other effects on ILC3 remains unclear. We show that VIP promotes ILC3 recruitment to the intestine through VPAC1 independent of the microbiota or adaptive immunity. VIP is also required for postnatal formation of lymphoid tissues as well as the maintenance of local populations of retinoic acid (RA)-producing dendritic cells, with RA up-regulating gut-homing receptor CCR9 expression by ILC3s. Correspondingly, mice deficient in VIP or VPAC1 suffer a paucity of intestinal ILC3s along with impaired production of the cytokine IL-22, rendering them highly susceptible to the enteric pathogen Citrobacter rodentium This heightened susceptibility to C. rodentium infection was ameliorated by RA supplementation, adoptive transfer of ILC3s, or by recombinant IL-22. Thus, VIP regulates the recruitment of intestinal ILC3s and formation of postnatal intestinal lymphoid tissues, offering protection against enteric pathogens.
Asunto(s)
Citrobacter rodentium/inmunología , Infecciones por Enterobacteriaceae/inmunología , Linfocitos/inmunología , Receptores de Tipo II del Péptido Intestinal Vasoactivo/metabolismo , Péptido Intestinal Vasoactivo/metabolismo , Animales , Células Dendríticas/inmunología , Microbioma Gastrointestinal/inmunología , Interleucinas/análisis , Tejido Linfoide/citología , Tejido Linfoide/crecimiento & desarrollo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores CCR/biosíntesis , Receptores de Tipo II del Péptido Intestinal Vasoactivo/genética , Tretinoina/metabolismo , Péptido Intestinal Vasoactivo/genética , Interleucina-22RESUMEN
Toll-like receptor 4 (TLR4) is a key component in innate immunity and has been linked to central nervous system (CNS) inflammation diseases, such as multiple sclerosis (MS), an inflammatory disorder induced by autoreactive Th17 cells. In our study, we found that TLR4 deficient (TLR4-/-) mice were inadequate to induce experimental autoimmune encephalomyelitis (EAE), characterized by low clinic score and weight loss, alleviative demyelinating, as well as decreased inflammatory cell infiltration in the spinal cord. In the lesion area of EAE mice, loss of TLR4 down-regulated the secretion of inflammatory cytokines and chemokine CCL25. Furthermore, the expression of CCR9 was decreased and chemotactic migration was attenuated in TLR4-/- Th17 cells. Our results demonstrate that TLR4 may mediate Th17 infiltration through CCL25/CCR9 signal during pathogenesis of EAE. Graphical Abstract Immunofluorescent staining of RORγt (green) and CCR9 (red) in spinal cords. TLR4 deficiency down-regulates CCR9 expression in infiltrating lymphocytes.
Asunto(s)
Quimiocinas CC/fisiología , Encefalomielitis Autoinmune Experimental/inmunología , Receptores CCR/fisiología , Células Th17/metabolismo , Receptor Toll-Like 4/fisiología , Animales , Células Cultivadas , Quimiotaxis de Leucocito , Encefalomielitis Autoinmune Experimental/etiología , Encefalomielitis Autoinmune Experimental/patología , Femenino , Regulación de la Expresión Génica/fisiología , Linfopoyesis , Ratones Endogámicos C57BL , Ratones Noqueados , Glicoproteína Mielina-Oligodendrócito/inmunología , Glicoproteína Mielina-Oligodendrócito/toxicidad , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/toxicidad , Receptores CCR/biosíntesis , Transducción de Señal/fisiología , Médula Espinal/patología , Bazo/inmunología , Bazo/patología , Receptor Toll-Like 4/deficiencia , Receptor Toll-Like 4/genéticaRESUMEN
The vitamin A metabolite all-trans retinoic acid (ATRA) induces a gut-homing phenotype in activated CD4(+) conventional T cells (Tconv) by upregulating the integrin α4ß7 and the chemokine receptor CCR9. We report that, in contrast to mouse Tconv, only â¼50% of regulatory T cells (Treg) upregulate CCR9 when stimulated by physiological levels of ATRA, even though Tconv and Treg express similar levels of the retinoic acid receptor (RAR). The resulting bimodal CCR9 expression is not associated with differences in the extent of their proliferation, level of Foxp3 expression, or affiliation with naturally occurring Treg or induced Treg in the circulating Treg pool. Furthermore, we find that exposure of Treg to the mechanistic target of rapamycin (mTOR) inhibitor rapamycin suppresses upregulation of both CCR9 and α4ß7, an effect that is not evident with Tconv. This suggests that in Treg, ATRA-induced upregulation of CCR9 and α4ß7 is dependent on activation of a mTOR signaling pathway. The involvement of mTOR is independent of Akt activity, because specific inhibition of Akt, pyruvate dehydrogenase kinase-1, or its downstream target glycogen synthase kinase-3 did not prevent CCR9 expression. Additionally, Rictor (mTOR complex [mTORC]2)-deficient Treg showed unaltered ability to express CCR9, whereas Raptor (mTORC1)-deficient Treg were unable to upregulate CCR9, suggesting the selective participation of mTORC1. These findings reveal a novel difference between ATRA signaling and chemokine receptor induction in Treg versus Tconv and provide a framework via which the migratory behavior of Treg versus Tconv might be regulated differentially for therapeutic purposes.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Quimiotaxis de Leucocito/inmunología , Complejos Multiproteicos/metabolismo , Linfocitos T Reguladores/inmunología , Serina-Treonina Quinasas TOR/metabolismo , Animales , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Separación Celular , Quimiotaxis de Leucocito/efectos de los fármacos , Citometría de Flujo , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores CCR/biosíntesis , Transducción de Señal/fisiología , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/metabolismo , Tretinoina/farmacologíaRESUMEN
ACKR4 also called CCX-CKR, CCRL1 as a member of atypical chemokine receptors, regulates the biological responses by clearance or transporting homeostatic chemokines such as CCL19, CCL21, CCL25, and CXCL13. Since these chemokines are involved in cancer development and metastasis, ACKR4 could have inhibition roles in cancer cell proliferation and invasion. Forming complexes with chemokine receptors by ACKR4 as in the case of hCXCR3 which lead to chemotaxis prevention is the other function of this protein is. However, as an atypical chemokine receptor, ACKR4 is less well-characterized compared to other members. Here, as the first step in understanding the molecular mechanisms of ACKR4 action, transfectants in HEK293T cell, was generated. In this study, ACKR4 coding sequence was cloned and human embryonic kidney 293T cells were used for recombinant production of ACKR4 protein. The liposome-mediated transfection with ACKR4 CDs, were detected in ACKR4 positive cells as early as 48 h post-transfection. The production of ACKR4 protein was confirmed using RT-PCR, dot blot, western blot, and flow cytometry. ACKR4 may represent a novel molecular target in cancer therapy, which might provide a chance for new therapeutic strategy. Therefore, the first step in the understanding of the molecular mechanisms of ACKR4 action is generation ACKR4-HEK293T recombinant cells.
Asunto(s)
Receptores CCR/genética , Expresión Génica , Células HEK293 , Humanos , ARN Mensajero/genética , Receptores CCR/biosíntesis , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
OBJECTIVE: P. gingivalis is a gram-negative anaerobic bacterium and a major periodontal pathogen. LPS produced by P. gingivalis promotes osteoclast formation. TECK is a CC chemokine whose expression is increased in gingival epithelial cells exposed to P. gingivalis LPS. In this study, we investigated the effect of TECK in osteoclastogenesis induced by P. gingivalis LPS. DESIGNS: Real time reverse transcriptase polymerase chain reaction (RTPCR) analysis and western blotting were performed to confirm TECK in MG63, human osteoblast cell line and primary murine osteoblasts and CCR9 in RAW 264.7 cells and murine bone marrow macrophages (BMMs) as osteoclast precursors. P. gingivalis LPS-treated BMMs and Raw 264.7 cells were cultured with or without TECK or TECK antibody to examine the effect of TECK on osteoclast formation. Cocultures with murine osteoblasts and bone marrow cells were also treated with or without TECK or TECK antibody. Luciferase assay and western blotting were used to determine whether TECK-CCR9 induced osteoclastogenesis was mediated through NFATc1 or NF-kB signaling. RESULTS: TECK was shown to be expressed by osteoblasts, and its receptor, CCR9, by osteoclast precursors. TECK increased P. gingivalis LPS-induced osteoclast numbers in an in vitro osteoclast formation assay using osteoclast precursors. The enhanced osteoclast formation by TECK was mediated by NFATc1, but not by NF-kB signaling. CONCLUSION: TECK may be a novel regulator of osteoclast formation induced by P. gingivalis LPS in periodontitis.
Asunto(s)
Quimiocinas CC/farmacología , Lipopolisacáridos/farmacología , Factores de Transcripción NFATC/metabolismo , Osteoclastos/efectos de los fármacos , Porphyromonas gingivalis/fisiología , Animales , Células de la Médula Ósea/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular , Quimiocinas CC/biosíntesis , Encía/citología , Encía/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , FN-kappa B/metabolismo , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteoclastos/citología , Osteoclastos/metabolismo , Osteoclastos/microbiología , Osteogénesis , Porphyromonas gingivalis/efectos de los fármacos , Células RAW 264.7 , Receptores CCR/biosíntesis , Proteínas Recombinantes/farmacología , Transducción de Señal/efectos de los fármacosRESUMEN
A gene encoding a carboxylesterase produced by Geobacillus thermoleovoras CCR11 was cloned in the pET-3b cloning vector, sequenced and expressed in Escherichia coli BL21(DE3). Gene sequence analysis revealed an open reading frame of 750 bp that encodes a polypeptide of 250 amino acid residues (27.3 kDa) named CaesCCR11. The enzyme showed its maximum activity at 50 °C and pH 5-8, with preference for C4 substrates, confirming its esterase nature. It displayed good resistance to temperature, pH, and the presence of organic solvents and detergents, that makes this enzyme biotechnologically applicable in the industries such as fine and oleo-chemicals, cosmetics, pharmaceuticals, organic synthesis, biodiesel production, detergents, and food industries. A 3D model of CaesCCR11 was predicted using the Bacillus sp. monoacyl glycerol lipase bMGL H-257 structure as template (PBD code 3RM3, 99 % residue identity with CaesCCR11). Based on its canonical α/ß hydrolase fold composed of 7 ß-strands and 6 α-helices, the α/ß architecture of the cap domain, the GLSTG pentapeptide, and the formation of distinctive salt bridges, we are proposing CaesCCR11 as a new member of family XV of lipolytic enzymes.
Asunto(s)
Secuencia de Aminoácidos/genética , Geobacillus/enzimología , Estructura Secundaria de Proteína , Receptores CCR/química , Clonación Molecular , Estabilidad de Enzimas , Escherichia coli/genética , Geobacillus/química , Modelos Moleculares , Receptores CCR/biosíntesis , Receptores CCR/genética , Análisis de Secuencia de ADN , Especificidad por Sustrato , TemperaturaRESUMEN
The human chemokine receptor CCRL2 is a member of the atypical chemokine receptor family. CCRL2 is unable to couple with G-proteins and fails to induce classical chemokine signaling for the highly conserved DRYLAIV motif essential for signaling has been changed to QRYLVFL. We investigated whether CCRL2 is involved in the chemotaxis, invasion, and proliferation of human breast cancer cells. Firstly, expression of CCRL2 was determined in six breast cancer cell lines by real-time RT-PCR and Western blot. Then, we established stable cell lines overexpressing CCRL2 to explore the function of CCRL2 in chemotaxis and invasion by transwell assays, and the signaling downstream was further investigated. The effect of CCRL2 on proliferation was detected by colony formation assays and tumor xenograft study. We found that stable overexpression of CCRL2 in MDA-MB-231 and BT-549 cells attenuated the chemotaxis and invasion stimulated by its ligand CCL2. CCRL2 inhibits p38 MAPK (p38) phosphorylation and up-regulates the expression of E-cadherin. This effect was eliminated by the inhibitor of p38 MAPK. CCRL2 inhibited the growth of breast cancer cells in vitro and in vivo. Our results suggest that CCRL2 functions as a tumor suppressor in human breast cancer cells.
Asunto(s)
Neoplasias de la Mama/metabolismo , Quimiocina CCL2/metabolismo , Quimiotaxis/fisiología , Receptores CCR/biosíntesis , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Quimiocina CCL2/antagonistas & inhibidores , Femenino , Humanos , Células MCF-7 , Invasividad Neoplásica/patología , Invasividad Neoplásica/prevención & control , Fosforilación/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidoresRESUMEN
Mucosa-associated invariant T (MAIT) cells are innate-like T cells with a conserved TCR α-chain recognizing bacterial metabolites presented on the invariant MHC-related 1 molecule. MAIT cells are present in intestinal tissues and liver, and they rapidly secrete IFN-γ and IL-17 in response to bacterial insult. In colon cancer, IL-17-driven inflammation promotes tumor progression, whereas IFN-γ production is essential for antitumor immunity. Thus, tumor-associated MAIT cells may affect antitumor immune responses by their secreted cytokines. However, the knowledge of MAIT cell presence and function in tumors is virtually absent. In this study, we determined the frequency, phenotype, and functional capacity of MAIT cells in colon adenocarcinomas and unaffected colon lamina propria. Flow cytometric analyses showed significant accumulation of MAIT cells in tumor tissue, irrespective of tumor stage or localization. Colonic MAIT cells displayed an activated memory phenotype and expression of chemokine receptors CCR6 and CCR9. Most MAIT cells in unaffected colon tissues produced IFN-γ, whereas only few produced IL-17. Colonic MAIT cells also produced TNF-α, IL-2, and granzyme B. In the tumors, significantly lower frequencies of IFN-γ-producing MAIT cells were seen, whereas there were no differences in the other cytokines analyzed, and in vitro studies showed that secreted factors from tumor tissue reduced IFN-γ production from MAIT cells. In conclusion, MAIT cells infiltrate colon tumors but their ability to produce IFN-γ is substantially reduced. We suggest that MAIT cells have the capacity to promote local immune responses to tumors, but factors in the tumor microenvironment act to reduce MAIT cell IFN-γ production.
Asunto(s)
Adenocarcinoma/patología , Neoplasias del Colon/patología , Interferón gamma/biosíntesis , Mucosa Intestinal/inmunología , Linfocitos T/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Granzimas/biosíntesis , Humanos , Inflamación/inmunología , Interferón gamma/inmunología , Interleucina-17/biosíntesis , Interleucina-17/inmunología , Interleucina-2 , Mucosa Intestinal/citología , Hígado/citología , Hígado/inmunología , Activación de Linfocitos/inmunología , Masculino , Persona de Mediana Edad , Receptores CCR/biosíntesis , Receptores CCR6/biosíntesis , Subgrupos de Linfocitos T/inmunología , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Current orthotopic xenograft models of human colorectal cancer (CRC) require surgery and do not robustly form metastases in the liver, the most common site clinically. CCR9 traffics lymphocytes to intestine and colorectum. We engineered use of the chemokine receptor CCR9 in CRC cell lines and patient-derived cells to create primary gastrointestinal (GI) tumors in immunodeficient mice by tail-vein injection rather than surgery. The tumors metastasize inducibly and robustly to the liver. Metastases have higher DKK4 and NOTCH signaling levels and are more chemoresistant than paired subcutaneous xenografts. Using this approach, we generated 17 chemokine-targeted mouse models (CTMMs) that recapitulate the majority of common human somatic CRC mutations. We also show that primary tumors can be modeled in immunocompetent mice by microinjecting CCR9-expressing cancer cell lines into early-stage mouse blastocysts, which induces central immune tolerance. We expect that CTMMs will facilitate investigation of the biology of CRC metastasis and drug screening.
Asunto(s)
Neoplasias Colorrectales/genética , Modelos Animales de Enfermedad , Neoplasias Hepáticas Experimentales/genética , Receptores CCR/genética , Animales , Blastocisto/patología , Línea Celular Tumoral , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intercelular/biosíntesis , Neoplasias Hepáticas Experimentales/secundario , Ratones , Metástasis de la Neoplasia , Receptores CCR/administración & dosificación , Receptores CCR/biosíntesis , Receptores Notch/biosíntesis , Transducción de Señal , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We studied the expression of some CC chemokines and their receptors in the synovium of patients with rheumatoid arthritis, osteoarthrosis, and a history of joint injury. In patients with rheumatoid arthritis, the levels mRNA for some angiogenic and proinflammatory chemokines (CCL5/RANTES, CCL11/eotaxin, CCL24/eotaxin-2, and CCL26/eotaxin-3) and their receptors (CCR1, CCR2, CCR3, CCR4, and CCR5) was elevated. mRNA expression correlated with activity, stage, and serological status of rheumatoid arthritis. Obtained data confirm the importance of CC chemokines as mediators of angiogenesis and inflammation in the synovium in rheumatoid arthritis.
Asunto(s)
Artritis Reumatoide/metabolismo , Quimiocinas CC/biosíntesis , Osteoartritis/metabolismo , ARN Mensajero/metabolismo , Receptores CCR/biosíntesis , Membrana Sinovial/metabolismo , Adulto , Quimiocinas CC/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Nefelometría y Turbidimetría , Receptores CCR/metabolismo , Estadísticas no ParamétricasRESUMEN
Poor clinical outcome of lung cancer (LuCa) is primarily due to lack of knowledge about specific molecules involved in its progression and metastasis. In this study, we for the first time show the clinical and biological significance of CC chemokine receptor-9 (CCR9) in non-small cell lung cancer (NSCLC). Expression of CCR9 and CCL25, the only natural ligand of CCR9, was significantly higher (p<0.0001) in NSCLC tissues and serum respectively, compared to their respective controls. Interestingly, expression of both CCR9 and CCL25 was significantly higher in adenocarcinomas (ACs) compared to squamous cell carcinomas (SCCs) (p = 0.04, and p< 0.0001). Similar to tissues, AC and SCC cell lines were positive for CCR9 expression. Despite of marginal difference in CCR9 expression, AC cells showed higher migratory and invasive potential in response to CCL25, compared to SCC cells. This differential biological response of AC cells was primarily due to differential expression of matrix metalloproteinases and tissue inhibitor of metalloproteinases under the influence of CCL25. Our results suggest CCR9 as a potential target for developing new treatment modality for NSCLC. Additionally, differential serum CCL25 level in ACs and SCCs, two NSCLC subtypes, suggest its potential as a non-invasive diagnostic/prognostic biomarker.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Quimiocinas CC/biosíntesis , Neoplasias Pulmonares/metabolismo , Receptores CCR/biosíntesis , Adenocarcinoma/sangre , Adenocarcinoma/metabolismo , Adenocarcinoma/patología , Adenocarcinoma del Pulmón , Carcinoma de Pulmón de Células no Pequeñas/sangre , Carcinoma de Pulmón de Células no Pequeñas/patología , Estudios de Casos y Controles , Línea Celular Tumoral , Quimiocinas CC/sangre , Progresión de la Enfermedad , Humanos , Neoplasias Pulmonares/sangre , Neoplasias Pulmonares/patología , Metástasis de la Neoplasia , Pronóstico , Receptores CCR/sangreRESUMEN
Cortical thymic epithelial cells (cTECs) and medullary thymic epithelial cells (mTECs), which play essential roles in the establishment of a functionally competent and self-tolerant repertoire of T cells, are derived from common thymic epithelial progenitor cells (pTECs). Recent findings indicate that mTECs are derived from cells that express molecules that are abundant in cTECs rather than mTECs, and provide fresh insight into the characteristics of pTECs and their diversification pathways into TEC subpopulations. In this issue of the European Journal of Immunology, Ribeiro et al. [Eur. J. Immunol. 2014. 44: 2918-2924] focus on CCRL1, an atypical chemokine receptor that is highly expressed by cTECs rather than mTECs, and show that CCRL1-expressing embryonic TECs can give rise to mTECs. Interestingly, Ribeiro et al. further report that a fraction of postnatal mTECs express CCRL1 at a low level, suggesting novel complexity in mTECs.
Asunto(s)
Diferenciación Celular/inmunología , Células Epiteliales/citología , Receptores CCR/biosíntesis , Timo/citología , Timo/crecimiento & desarrollo , AnimalesRESUMEN
Cortical and medullary thymic epithelial cells (cTECs and mTECs, respectively) provide inductive microenvironments for T-cell development and selection. The differentiation pathway of cTEC/mTEC lineages downstream of common bipotent progenitors at discrete stages of development remains unresolved. Using IL-7/CCRL1 dual reporter mice that identify specialized TEC subsets, we show that the stepwise acquisition of chemokine (C-C motif) receptor-like 1 (CCRL1) is a late determinant of cTEC differentiation. Although cTECs expressing high CCRL1 levels (CCRL1(hi) ) develop normally in immunocompetent and Rag2(-/-) thymi, their differentiation is partially blocked in Rag2(-/-) Il2rg(-/-) counterparts. These results unravel a novel checkpoint in cTEC maturation that is regulated by the cross-talk between TECs and immature thymocytes. Additionally, we identify new Ulex europaeus agglutinin 1 (UEA)(+) mTEC subtypes expressing intermediate CCRL1 levels (CCRL1(int) ) that conspicuously emerge in the postnatal thymus and differentially express Tnfrsf11a, Ccl21, and Aire. While rare in fetal and in Rag2(-/-) thymi, CCRL1(int) mTECs are restored in Rag2(-/-) Marilyn TCR-Tg mice, indicating that the appearance of postnatal-restricted mTECs is closely linked with T-cell selection. Our findings suggest that alternative temporally restricted routes of new mTEC differentiation contribute to the establishment of the medullary niche in the postnatal thymus.
Asunto(s)
Diferenciación Celular/inmunología , Células Epiteliales/citología , Receptores CCR/biosíntesis , Timo/citología , Timo/crecimiento & desarrollo , Animales , Biomarcadores/análisis , Citometría de Flujo , Ratones , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos , Receptores CCR/inmunología , Linfocitos T/citología , Linfocitos T/inmunología , Timo/inmunología , TranscriptomaRESUMEN
BACKGROUND: Chemokines have been implicated in tumor progression and metastasis. In melanoma, chemokine receptors have been implicated in organ selective metastasis by regulating processes such as chemoattraction, adhesion and survival. METHODS: In this study we have analyzed, using flow cytometry, the systems formed by the chemokine receptors CXCR3, CXCR4, CXCR7, CCR7 and CCR10 and their ligands in thirteen human melanoma cell lines (five established from primary tumors and eight established from metastasis from different tissues). WM-115 and WM-266.4 melanoma cell lines (obtained from a primary and a metastatic melanoma respectively) were xenografted in nude mice and the tumors and cell lines derived from them were also analyzed. RESULTS: Our results show that the melanoma cell lines do not express or express in a low degree the chemokine receptors on their cell surface. However, melanoma cell lines show intracellular expression of all the aforementioned receptors and most of their respective ligands. When analyzing the xenografts and the cell lines obtained from them we found variations in the intracellular expression of chemokines and chemokine receptors that differed between the primary and metastatic cell lines. However, as well as in the original cell lines, minute or no expression of the chemokine receptors was observed at the cell surface. CONCLUSIONS: Coexpression of chemokine receptors and their ligands was found in human melanoma cell lines. However, this expression is intracellular and receptors are not found at the cell membrane nor chemokines are secreted to the cell medium. The levels of expressed chemokine receptors and their ligands show dynamic variations after xenotransplantation that differ depending on the origin of the cell line (from primary tumor or from metastasis).
Asunto(s)
Ligandos , Melanoma/metabolismo , Receptores CCR/metabolismo , Receptores CXCR/metabolismo , Animales , Línea Celular Tumoral , Membrana Celular/metabolismo , Quimiotaxis , Modelos Animales de Enfermedad , Xenoinjertos , Humanos , Inmunohistoquímica , Espacio Intracelular/metabolismo , Melanoma/genética , Melanoma/patología , Ratones , Receptores CCR/biosíntesis , Receptores CCR/genética , Receptores CXCR/biosíntesis , Receptores CXCR/genéticaRESUMEN
αß T-cell development and selection proceed while thymocytes successively migrate through distinct regions of the thymus. For γδ T cells, the interplay of intrathymic migration and cell differentiation is less well understood. Here, we crossed C-C chemokine receptor (CCR)7-deficient (Ccr7(-/-) ) and CCR9-deficient mice (Ccr9(-/-) ) to mice with a TcrdH2BeGFP reporter background to investigate the impact of thymic localization on γδ T-cell development. γδ T-cell frequencies and numbers were decreased in CCR7-deficient and increased in CCR9-deficient mice. Transfer of CCR7- or CCR9-deficient BM into irradiated C57BL/6 WT recipients reproduced these phenotypes, pointing toward cell-intrinsic migration defects. Monitoring recent thymic emigrants by intrathymic labeling allowed us to identify decreased thymic γδ T-cell output in CCR7-deficient mice. In vitro, CCR7-deficient precursors showed normal γδ T-cell development. Immunohistology revealed that CCR7 and CCR9 expression was important for γδ T-cell localization within thymic medulla or cortex, respectively. However, γδ T-cell motility was unaltered in CCR7- or CCR9-deficient thymi. Together, our results suggest that proper intrathymic localization is important for normal γδ T-cell development.
Asunto(s)
Movimiento Celular/fisiología , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Receptores CCR7/inmunología , Linfocitos T/inmunología , Timo/inmunología , Animales , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/inmunología , Ratones , Ratones Noqueados , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Receptores CCR/biosíntesis , Receptores CCR/genética , Receptores CCR/inmunología , Receptores CCR7/biosíntesis , Receptores CCR7/genética , Linfocitos T/citología , Linfocitos T/metabolismo , Timo/citología , Timo/metabolismoRESUMEN
The chemokine receptor CCR9 was recently implicated in tumor biology. In the present study, our objective was to evaluate the clinical significance and potential role of CCR9 in hepatocellular carcinoma (HCC). CCR9 expression was detected by immunohistochemistry, quantitative PCR (qPCR) and western blotting in HCC patients. The prognostic significance of CCR9 expression was assessed. The functional roles of CCR9 in HCC were investigated using MTT, BrdU, colony formation assay and flow cytometry. CCR9 was significantly elevated in HCC tissue samples. High CCR9 expression was correlated with multiple tumor nodes, high Edmondson-Steiner grade and vascular invasion. Multivariate analysis showed that CCR9 expression was an independent prognostic factor for the overall survival (OS) of HCC patients. Further investigations revealed that ectopic expression of CCR9 enhanced cell proliferation and tumorigenicity in HCC cells, whereas CCR9 silencing impaired cell proliferation and tumorigenicity, which was mediated through downregulation of the cell cycle regulators p21, p27 as well as upregulation of cyclin D1. These results suggest that CCR9 can act as a novel prognostic marker and therapeutic target for HCC.
Asunto(s)
Biomarcadores de Tumor/análisis , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Receptores CCR/biosíntesis , Adulto , Anciano , Western Blotting , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/patología , Femenino , Citometría de Flujo , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Pronóstico , Modelos de Riesgos Proporcionales , Receptores CCR/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Colorectal cancer is one of the most common malignant cancers, with bad prognosis when distal metastasis occurs. The current study aimed to investigate the potential value of using CCX-CKR expression for the prognosis of colorectal cancer patients. The results showed that CCX-CKR expression was a negative predictor of cancer metastasis, and that it was positively correlated to the patients’ survival rate. Finally, we found that CCX-CKR expression in vitro could modulate cellular migration and invasion abilities, potentially via the regulation of other chemotactic factors/receptors.
Asunto(s)
Neoplasias Colorrectales/genética , Pronóstico , Receptores CCR/biosíntesis , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Movimiento Celular/genética , Neoplasias Colorrectales/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Invasividad Neoplásica/genética , Metástasis de la Neoplasia , Estadificación de Neoplasias , Receptores CCR/genéticaRESUMEN
The present study demonstrates that RA has activity of an IgA switch factor and is more specific than TGF-ß1. RA independently caused only IgA switching, whereas TGF-ß1 caused IgA and IgG2b switching. We found that RA increased IgA production and that this was a result of its ability to increase the frequency of IgA-secreting B cell clones. Increased IgA production was accompanied by an increase of GLTα. RA activity was abrogated by an antagonist of the RAR. Additionally, RA affected intestinal IgA production in mice. Surprisingly, RA, in combination with TGF-ß1, notably enhanced not only IgA production and GLTα expression but also CCR9 and α4ß7 expression on B cells. These results suggest that RA selectively induces IgA isotype switching through RAR and that RA and TGF-ß have important effects on the overall gut IgA antibody response.
Asunto(s)
Inmunoglobulina A/inmunología , Cambio de Clase de Inmunoglobulina/efectos de los fármacos , Factor de Crecimiento Transformador beta1/inmunología , Tretinoina/farmacología , Animales , Células Cultivadas/inmunología , Selección Clonal Mediada por Antígenos , Endotoxinas/toxicidad , Genes de Inmunoglobulinas , Inmunidad Mucosa/efectos de los fármacos , Inmunoglobulina A/biosíntesis , Inmunoglobulina G/inmunología , Integrinas/biosíntesis , Integrinas/genética , Ganglios Linfáticos/inmunología , Mesenterio/inmunología , Ratones , Ratones Endogámicos BALB C , Ganglios Linfáticos Agregados/inmunología , Receptores CCR/biosíntesis , Receptores CCR/genética , Receptores Mensajeros de Linfocitos/efectos de los fármacos , Receptores Mensajeros de Linfocitos/inmunología , Receptores de Ácido Retinoico/fisiologíaRESUMEN
Thymic epithelial cells (TECs) help orchestrate thymopoiesis, and TEC differentiation relies on bidirectional interactions with thymocytes. Although the molecular mediators that stimulate medullary thymic epithelial cell (mTEC) maturation are partially elucidated, the signals that regulate cortical thymic epithelial cell (cTEC) homeostasis remain elusive. Using IL-7 reporter mice, we show that TECs coexpressing high levels of IL-7 (Il7(YFP+) TECs) reside within a subset of CD205(+)Ly51(+)CD40(low) cTECs that coexpresses Dll4, Ccl25, Ccrl1, Ctsl, Psmb11, and Prss16 and segregates from CD80(+)CD40(high) mTECs expressing Tnfrsf11a, Ctss, and Aire. As the frequency of Il7(YFP+) TECs gradually declines as mTEC development unfolds, we explored the relationship between Il7(YFP+) TECs and mTECs. In thymic organotypic cultures, the thymocyte-induced reduction in Il7(YFP+) TECs dissociates from the receptor activator of NF-κB-mediated differentiation of CD80(+) mTECs. Still, Il7(YFP+) TECs can generate some CD80(+) mTECs in a stepwise differentiation process via YFP(-)Ly51(low)CD80(low) intermediates. Il7(YFP+) TECs are sustained in Rag2(-/-) mice, even following in vivo anti-CD3ε treatment that mimics the process of pre-TCR ß-selection of thymocytes to the double positive (DP) stage. Using Marilyn-Rag2(-/-) TCR transgenic, we find that positive selection into the CD4 lineage moderately reduces the frequency of Il7(YFP+) TECs, whereas negative selection provokes a striking loss of Il7(YFP+) TECs. These results imply that the strength of MHC/peptide-TCR interactions between TECs and thymocytes during selection constitutes a novel rheostat that controls the maintenance of IL-7-expressing cTECs.
Asunto(s)
Células Epiteliales/inmunología , Interleucina-7/biosíntesis , Timo/inmunología , Proteínas Adaptadoras Transductoras de Señales , Animales , Antígenos CD/metabolismo , Antígeno B7-1/metabolismo , Complejo CD3/inmunología , Antígenos CD40/metabolismo , Proteínas de Unión al Calcio , Catepsina L/biosíntesis , Catepsinas/biosíntesis , Diferenciación Celular/inmunología , Movimiento Celular , Células Cultivadas , Quimiocinas CC/biosíntesis , Proteínas de Unión al ADN/deficiencia , Proteínas de Unión al ADN/genética , Células Epiteliales/metabolismo , Homeostasis , Interleucina-7/metabolismo , Péptidos y Proteínas de Señalización Intracelular/biosíntesis , Lectinas Tipo C/metabolismo , Activación de Linfocitos , Proteínas de la Membrana/biosíntesis , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Antígenos de Histocompatibilidad Menor , Técnicas de Cultivo de Órganos , Receptor Activador del Factor Nuclear kappa-B/biosíntesis , Receptor Activador del Factor Nuclear kappa-B/metabolismo , Receptores CCR/biosíntesis , Receptores de Superficie Celular/metabolismo , Serina Endopeptidasas/biosíntesis , Timocitos/metabolismo , Timo/metabolismo , Factores de Transcripción/biosíntesis , Proteína AIRERESUMEN
Human monocyte-derived dendritic cells (DC) (MoDC) are utilized for immunotherapy. However, in-vitro immunological effects are often not mirrored in vivo. We studied the tissue-homing potential of MoDC. Circulating monocytes and DC expressed different tissue-homing markers and, during in-vitro development of MoDC, homing marker expression was lost resulting in a 'homeless' phenotype. Retinoic acid (RA) induced gut-homing markers (ß7 and CCR9) and a regulatory phenotype and function [decreased human leucocyte antigen D-related (HLA-DR) and increased ILT3 and fluorescein isothiocyanate (FITC-dextran uptake) in MoDC]. RA-MoDC were less stimulatory and primed conditioned T cells with a gut-homing profile (ß7(+)CLA(-)). Unlike the normal intestinal microenvironment, that from inflamed colon of ulcerative colitis (UC) patients did not induce regulatory properties in MoDC. However, RA-MoDC maintained their regulatory gut-specific properties even in the presence of UC microenvironment. Therefore, MoDC may be ineffectual for immunotherapy because they lack tissue-homing and tissue-imprinting specificity. However, MoDC rehabilitation with gut-homing potential by RA could be useful in promoting immunotherapy in pathologies such as UC.