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1.
Proc Natl Acad Sci U S A ; 118(41)2021 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-34625492

RESUMO

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.


Assuntos
Citrobacter rodentium/imunologia , Infecções por Enterobacteriaceae/imunologia , Linfócitos/imunologia , Receptores Tipo II de Peptídeo Intestinal Vasoativo/metabolismo , Peptídeo Intestinal Vasoativo/metabolismo , Animais , Células Dendríticas/imunologia , Microbioma Gastrointestinal/imunologia , Interleucinas/análise , Tecido Linfoide/citologia , Tecido Linfoide/crescimento & desenvolvimento , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR/biossíntese , Receptores Tipo II de Peptídeo Intestinal Vasoativo/genética , Tretinoína/metabolismo , Peptídeo Intestinal Vasoativo/genética , Interleucina 22
2.
J Neuroimmune Pharmacol ; 14(3): 493-502, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31065973

RESUMO

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.


Assuntos
Quimiocinas CC/fisiologia , Encefalomielite Autoimune Experimental/imunologia , Receptores CCR/fisiologia , Células Th17/metabolismo , Receptor 4 Toll-Like/fisiologia , Animais , Células Cultivadas , Quimiotaxia de Leucócito , Encefalomielite Autoimune Experimental/etiologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Regulação da Expressão Gênica/fisiologia , Linfopoese , Camundongos Endogâmicos C57BL , Camundongos Knockout , Glicoproteína Mielina-Oligodendrócito/imunologia , Glicoproteína Mielina-Oligodendrócito/toxicidade , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/toxicidade , Receptores CCR/biossíntese , Transdução de Sinais/fisiologia , Medula Espinal/patologia , Baço/imunologia , Baço/patologia , Receptor 4 Toll-Like/deficiência , Receptor 4 Toll-Like/genética
3.
J Immunol ; 197(4): 1137-47, 2016 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-27402696

RESUMO

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 ∼50% 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.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Quimiotaxia de Leucócito/imunologia , Complexos Multiproteicos/metabolismo , Linfócitos T Reguladores/imunologia , Serina-Treonina Quinases TOR/metabolismo , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Separação Celular , Quimiotaxia de Leucócito/efeitos dos fármacos , Citometria de Fluxo , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores CCR/biossíntese , Transdução de Sinais/fisiologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/metabolismo , Tretinoína/farmacologia
4.
Mol Biol Rep ; 43(7): 583-9, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27168154

RESUMO

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.


Assuntos
Receptores CCR/genética , Expressão Gênica , Células HEK293 , Humanos , RNA Mensageiro/genética , Receptores CCR/biossíntese , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
5.
Arch Oral Biol ; 66: 77-85, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26921718

RESUMO

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.


Assuntos
Quimiocinas CC/farmacologia , Lipopolissacarídeos/farmacologia , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/efeitos dos fármacos , Porphyromonas gingivalis/fisiologia , Animais , Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Quimiocinas CC/biossíntese , Gengiva/citologia , Gengiva/metabolismo , Humanos , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/metabolismo , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoclastos/citologia , Osteoclastos/metabolismo , Osteoclastos/microbiologia , Osteogênese , Porphyromonas gingivalis/efeitos dos fármacos , Células RAW 264.7 , Receptores CCR/biossíntese , Proteínas Recombinantes/farmacologia , Transdução de Sinais/efeitos dos fármacos
6.
Mol Biotechnol ; 58(1): 37-46, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26603441

RESUMO

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.


Assuntos
Sequência de Aminoácidos/genética , Geobacillus/enzimologia , Estrutura Secundária de Proteína , Receptores CCR/química , Clonagem Molecular , Estabilidade Enzimática , Escherichia coli/genética , Geobacillus/química , Modelos Moleculares , Receptores CCR/biossíntese , Receptores CCR/genética , Análise de Sequência de DNA , Especificidade por Substrato , Temperatura
7.
Med Oncol ; 32(11): 254, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26487662

RESUMO

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.


Assuntos
Neoplasias da Mama/metabolismo , Quimiocina CCL2/metabolismo , Quimiotaxia/fisiologia , Receptores CCR/biossíntese , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Quimiocina CCL2/antagonistas & inibidores , Feminino , Humanos , Células MCF-7 , Invasividade Neoplásica/patologia , Invasividade Neoplásica/prevenção & controle , Fosforilação/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores
8.
J Immunol ; 195(7): 3472-81, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26297765

RESUMO

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.


Assuntos
Adenocarcinoma/patologia , Neoplasias do Colo/patologia , Interferon gama/biossíntese , Mucosa Intestinal/imunologia , Linfócitos T/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Granzimas/biossíntese , Humanos , Inflamação/imunologia , Interferon gama/imunologia , Interleucina-17/biossíntese , Interleucina-17/imunologia , Interleucina-2 , Mucosa Intestinal/citologia , Fígado/citologia , Fígado/imunologia , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Receptores CCR/biossíntese , Receptores CCR6/biossíntese , Subpopulações de Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/biossíntese
9.
Nat Biotechnol ; 33(6): 656-60, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26006007

RESUMO

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.


Assuntos
Neoplasias Colorretais/genética , Modelos Animais de Doenças , Neoplasias Hepáticas Experimentais/genética , Receptores CCR/genética , Animais , Blastocisto/patologia , Linhagem Celular Tumoral , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/biossíntese , Neoplasias Hepáticas Experimentais/secundário , Camundongos , Metástase Neoplásica , Receptores CCR/administração & dosagem , Receptores CCR/biossíntese , Receptores Notch/biossíntese , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Bull Exp Biol Med ; 158(2): 192-6, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25430645

RESUMO

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.


Assuntos
Artrite Reumatoide/metabolismo , Quimiocinas CC/biossíntese , Osteoartrite/metabolismo , RNA Mensageiro/metabolismo , Receptores CCR/biossíntese , Membrana Sinovial/metabolismo , Adulto , Quimiocinas CC/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Nefelometria e Turbidimetria , Receptores CCR/metabolismo , Estatísticas não Paramétricas
11.
Oncotarget ; 5(20): 10170-9, 2014 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-25296976

RESUMO

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.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Quimiocinas CC/biossíntese , Neoplasias Pulmonares/metabolismo , Receptores CCR/biossíntese , Adenocarcinoma/sangue , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Adenocarcinoma de Pulmão , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Casos e Controles , Linhagem Celular Tumoral , Quimiocinas CC/sangue , Progressão da Doença , Humanos , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/patologia , Metástase Neoplásica , Prognóstico , Receptores CCR/sangue
12.
Eur J Immunol ; 44(10): 2872-5, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25216053

RESUMO

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.


Assuntos
Diferenciação Celular/imunologia , Células Epiteliais/citologia , Receptores CCR/biossíntese , Timo/citologia , Timo/crescimento & desenvolvimento , Animais
13.
Eur J Immunol ; 44(10): 2918-24, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25070355

RESUMO

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.


Assuntos
Diferenciação Celular/imunologia , Células Epiteliais/citologia , Receptores CCR/biossíntese , Timo/citologia , Timo/crescimento & desenvolvimento , Animais , Biomarcadores/análise , Citometria de Fluxo , Camundongos , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Receptores CCR/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Timo/imunologia , Transcriptoma
14.
BMC Cancer ; 14: 118, 2014 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-24559071

RESUMO

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).


Assuntos
Ligantes , Melanoma/metabolismo , Receptores CCR/metabolismo , Receptores CXCR/metabolismo , Animais , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Quimiotaxia , Modelos Animais de Doenças , Xenoenxertos , Humanos , Imuno-Histoquímica , Espaço Intracelular/metabolismo , Melanoma/genética , Melanoma/patologia , Camundongos , Receptores CCR/biossíntese , Receptores CCR/genética , Receptores CXCR/biossíntese , Receptores CXCR/genética
15.
Oncol Rep ; 31(4): 1629-36, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24481516

RESUMO

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.


Assuntos
Biomarcadores Tumorais/análise , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Receptores CCR/biossíntese , Adulto , Idoso , Western Blotting , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/patologia , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , Receptores CCR/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa
16.
Eur J Immunol ; 44(5): 1320-9, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24500801

RESUMO

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.


Assuntos
Movimento Celular/fisiologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Receptores CCR7/imunologia , Linfócitos T/imunologia , Timo/imunologia , Animais , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Camundongos , Camundongos Knockout , Receptores de Antígenos de Linfócitos T gama-delta/genética , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Receptores CCR/biossíntese , Receptores CCR/genética , Receptores CCR/imunologia , Receptores CCR7/biossíntese , Receptores CCR7/genética , Linfócitos T/citologia , Linfócitos T/metabolismo , Timo/citologia , Timo/metabolismo
17.
Int J Biol Markers ; 29(1): e40-8, 2014 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-24338720

RESUMO

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.


Assuntos
Neoplasias Colorretais/genética , Prognóstico , Receptores CCR/biossíntese , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular Tumoral , Movimento Celular/genética , Neoplasias Colorretais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Metástase Neoplásica , Estadiamento de Neoplasias , Receptores CCR/genética
18.
J Immunol ; 191(3): 1200-9, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-23794633

RESUMO

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.


Assuntos
Células Epiteliais/imunologia , Interleucina-7/biossíntese , Timo/imunologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Antígenos CD/metabolismo , Antígeno B7-1/metabolismo , Complexo CD3/imunologia , Antígenos CD40/metabolismo , Proteínas de Ligação ao Cálcio , Catepsina L/biossíntese , Catepsinas/biossíntese , Diferenciação Celular/imunologia , Movimento Celular , Células Cultivadas , Quimiocinas CC/biossíntese , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Células Epiteliais/metabolismo , Homeostase , Interleucina-7/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Lectinas Tipo C/metabolismo , Ativação Linfocitária , Proteínas de Membrana/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Antígenos de Histocompatibilidade Menor , Técnicas de Cultura de Órgãos , Receptor Ativador de Fator Nuclear kappa-B/biossíntese , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Receptores CCR/biossíntese , Receptores de Superfície Celular/metabolismo , Serina Endopeptidases/biossíntese , Timócitos/metabolismo , Timo/metabolismo , Fatores de Transcrição/biossíntese , Proteína AIRE
19.
J Leukoc Biol ; 94(2): 325-35, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23744644

RESUMO

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.


Assuntos
Imunoglobulina A/imunologia , Switching de Imunoglobulina/efeitos dos fármacos , Fator de Crescimento Transformador beta1/imunologia , Tretinoína/farmacologia , Animais , Células Cultivadas/imunologia , Seleção Clonal Mediada por Antígeno , Endotoxinas/toxicidade , Genes de Imunoglobulinas , Imunidade nas Mucosas/efeitos dos fármacos , Imunoglobulina A/biossíntese , Imunoglobulina G/imunologia , Integrinas/biossíntese , Integrinas/genética , Linfonodos/imunologia , Mesentério/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Nódulos Linfáticos Agregados/imunologia , Receptores CCR/biossíntese , Receptores CCR/genética , Receptores de Retorno de Linfócitos/efeitos dos fármacos , Receptores de Retorno de Linfócitos/imunologia , Receptores do Ácido Retinoico/fisiologia
20.
Clin Exp Immunol ; 174(1): 109-19, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23607934

RESUMO

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.


Assuntos
Diferenciação Celular/imunologia , Movimento Celular/imunologia , Células Dendríticas/imunologia , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/imunologia , Monócitos/imunologia , Tretinoína/farmacologia , Biomarcadores/metabolismo , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/imunologia , Colite Ulcerativa/patologia , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Feminino , Trato Gastrointestinal/patologia , Humanos , Masculino , Monócitos/citologia , Monócitos/efeitos dos fármacos , Especificidade de Órgãos/efeitos dos fármacos , Especificidade de Órgãos/imunologia , Receptores CCR/biossíntese , Receptores CCR7/biossíntese , Tretinoína/uso terapêutico
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