RESUMO
Approximately 25% of cancers are preceded by chronic inflammation that occurs at the site of tumor development. However, whether this multifactorial oncogenic process, which commonly occurs in the intestines, can be initiated by a specific immune cell population is unclear. Here, we show that an intestinal T cell subset, derived from interleukin-17 (IL-17)-producing helper T (TH17) cells, induces the spontaneous transformation of the intestinal epithelium. This subset produces inflammatory cytokines, and its tumorigenic potential is not dependent on IL-17 production but on the transcription factors KLF6 and T-BET and interferon-γ. The development of this cell type is inhibited by transforming growth factor-ß1 (TGFß1) produced by intestinal epithelial cells. TGFß signaling acts on the pretumorigenic TH17 cell subset, preventing its progression to the tumorigenic stage by inhibiting KLF6-dependent T-BET expression. This study therefore identifies an intestinal T cell subset initiating cancer.
RESUMO
Primary cilia (PC) are important signaling hubs, and we here explored their role in colonic pathology. In the colon, PC are mostly present on fibroblasts, and exposure of mice to either chemically induced colitis-associated colon carcinogenesis (CAC) or dextran sodium sulfate (DSS)-induced acute colitis decreases PC numbers. We generated conditional knockout mice with reduced numbers of PC on colonic fibroblasts. These mice show increased susceptibility to CAC, as well as DSS-induced colitis. Secretome and immunohistochemical analyses of DSS-treated mice display an elevated production of the proinflammatory cytokine IL-6 in PC-deficient colons. An inflammatory environment diminishes PC presence in primary fibroblast cultures, which is triggered by IL-6 as identified by RNA-seq analysis together with blocking experiments. These findings suggest an activation loop between IL-6 production and PC loss. An analysis of PC presence on biopsies of patients with ulcerative colitis or colorectal cancer (CRC) reveals decreased numbers of PC on colonic fibroblasts in pathological compared with surrounding normal tissue. Taken together, we provide evidence that a decrease in colonic PC numbers promotes colitis and CRC.
Assuntos
Cílios , Interleucina-6 , Camundongos , Animais , Interleucina-6/genéticaRESUMO
Lyl1 encodes a hematopoietic- and endothelial-specific bHLH transcription factor. Lyl1-deficient mice are viable, but they display mild hematopoietic and vascular defects. Specifically, LYL1 is required for the maturation and stabilization of blood vessel endothelial adherens junctions. Here, we report that young adult Lyl1-/- mice exhibit transient overweight associated with general expansion of adipose tissue, without signs of metabolic disorder and unrelated to food intake. The increased fat tissue development in Lyl1-/- mice resulted from earlier differentiation of adipose stem cells (ASCs) into adipocytes through noncell autonomous mechanisms. Specifically, we found that in Lyl1-/- mice, the adipose tissue vascular structures are immature, as indicated by their high permeability, reduced coverage by pericytes, lower recruitment of VE-cadherin and ZO1 at cell junctions, and more prone to angiogenesis. Together, our data show that in Lyl1-/- mice, the impaired vascular compartment of the adipose niche promotes ASC differentiation, leading to early adipocyte expansion and premature ASC depletion. Our study highlights the major structural role of the adipose tissue vascular niche in coordinating stem cell self-renewal and differentiation into adipocytes.
Assuntos
Tecido Adiposo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/deficiência , Proteínas de Neoplasias/deficiência , Neovascularização Patológica , Nicho de Células-Tronco , Células-Tronco/metabolismo , Tecido Adiposo/irrigação sanguínea , Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/metabolismo , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Células-Tronco/patologiaRESUMO
Maturation of newly formed vessels is a multistep phenomenon during which functional endothelial barriers are established. Disruption of vessel integrity is an important feature in many physiological and pathological processes. We previously reported that lymphoblastic leukemia-derived sequence 1 (LYL1) is required for the late stages of postnatal angiogenesis to limit the formation of new blood vessels, notably by regulating the activity of the small GTPase Rap1. In this study, we show that LYL1 is also required during the formation of the mature endothelial barrier in the lungs of adult mice. Specifically, LYL1 knockdown in human endothelial cells downregulated the expression of ARHGAP21 and ARHGAP24, which encode two Rho GTPase-activating proteins, and this was correlated with increased RhoA activity and reorganization of the actin cytoskeleton into stress fibers. Importantly, in lungs of Lyl1-deficient mice, both vascular endothelial (VE)-cadherin and p120-catenin were poorly recruited to endothelial adherens junctions, indicative of defective cell-cell junctions. Consistent with this, higher Evans blue dye extravasation, edema, and leukocyte infiltration in the lung parenchyma of Lyl1-/- mice than in wild-type littermates confirmed that lung vascular permeability is constitutively elevated in Lyl1-/- adult mice. Our data show that LYL1 acts as a stabilizing signal for adherens junction formation by operating upstream of VE-cadherin and of the two GTPases Rap1 and RhoA. As increased vascular permeability is a key feature and a major mechanism of acute respiratory distress syndrome, molecules that regulate LYL1 activity could represent additional tools to modify the endothelial barrier permeability.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Permeabilidade da Membrana Celular , Endotélio Vascular/patologia , Células Endoteliais da Veia Umbilical Humana/patologia , Pulmão/patologia , Proteínas de Neoplasias/fisiologia , Actinas/genética , Actinas/metabolismo , Animais , Western Blotting , Células Cultivadas , Endotélio Vascular/metabolismo , Imunofluorescência , Proteínas Ativadoras de GTPase/genética , Proteínas Ativadoras de GTPase/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Técnicas Imunoenzimáticas , Pulmão/irrigação sanguínea , Pulmão/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fibras de Estresse , Proteínas rap1 de Ligação ao GTP/genética , Proteínas rap1 de Ligação ao GTP/metabolismo , Proteína rhoA de Ligação ao GTP/genética , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
The 2 related basic helix loop helix genes, LYL1 and TAL-1 are active in hematopoietic and endothelial lineages. While Tal-1 is essential for both hematopoietic and vascular development, the role of Lyl1 appears to be distinct as deficient mice are viable and display modest hematopoietic defects. Here, we reveal a role for Lyl1 as a major regulator of adult neovascularization. Tumors implanted into Lyl1-deficient mice showed higher proliferation and angiogenesis, as evidenced by enlarged lumens, reduced pericyte coverage and increased permeability, compared with wild type littermates. Of note, Lyl1-deficient tumor vessels exhibited an up-regulation of Tal-1, the VE-Cadherin target gene, as well as Angiopoietin-2, 3 major actors in angiogenesis. Hematopoietic reconstitution experiments demonstrated that this sustained tumor angiogenesis was of endothelial origin. Moreover, the angiogenic phenotype observed in the absence of Lyl1 function was not tumor-restricted as microvessels forming in Matrigel or originating from aortic explants were also more numerous and larger than their wild-type counterparts. Finally, LYL1 depletion in human endothelial cells revealed that LYL1 controls the expression of molecules involved in the stabilization of vascular structures. Together, our data show a role for LYL1 in the postnatal maturation of newly formed blood vessels.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/irrigação sanguínea , Neoplasias Experimentais/metabolismo , Neovascularização Patológica/metabolismo , Angiopoietina-2/biossíntese , Angiopoietina-2/genética , Animais , Antígenos CD/biossíntese , Antígenos CD/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Caderinas/biossíntese , Caderinas/genética , Regulação Neoplásica da Expressão Gênica/genética , Hematopoese/genética , Humanos , Camundongos , Camundongos Mutantes , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Neoplasias Experimentais/genética , Neovascularização Patológica/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteína 1 de Leucemia Linfocítica Aguda de Células T , Regulação para Cima/genéticaRESUMO
The R2TP chaperone cooperates with HSP90 to integrate newly synthesized proteins into multi-subunit complexes, yet its role in tissue homeostasis is unknown. Here, we generated conditional, inducible knock-out mice for Rpap3 to inactivate this core component of R2TP in the intestinal epithelium. In adult mice, Rpap3 invalidation caused destruction of the small intestinal epithelium and death within 10 days. Levels of R2TP substrates decreased, with strong effects on mTOR, ATM and ATR. Proliferative stem cells and progenitors deficient for Rpap3 failed to import RNA polymerase II into the nucleus and they induced p53, cell cycle arrest and apoptosis. Post-mitotic, differentiated cells did not display these alterations, suggesting that R2TP clients are preferentially built in actively proliferating cells. In addition, high RPAP3 levels in colorectal tumors from patients correlate with bad prognosis. Here, we show that, in the intestine, the R2TP chaperone plays essential roles in normal and tumoral proliferation.
Assuntos
Proliferação de Células , Células Epiteliais/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Mucosa Intestinal/metabolismo , Chaperonas Moleculares/metabolismo , Animais , Células Cultivadas , Células Epiteliais/citologia , Humanos , Mucosa Intestinal/citologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microscopia Confocal , Ligação Proteica , Células-Tronco/citologia , Células-Tronco/metabolismoRESUMO
To clarify the function of cyclin A2 in colon homeostasis and colorectal cancer (CRC), we generated mice deficient for cyclin A2 in colonic epithelial cells (CECs). Colons of these mice displayed architectural changes in the mucosa and signs of inflammation, as well as increased proliferation of CECs associated with the appearance of low- and high-grade dysplasias. The main initial events triggering those alterations in cyclin A2-deficient CECs appeared to be abnormal mitoses and DNA damage. Cyclin A2 deletion in CECs promoted the development of dysplasia and adenocarcinomas in a murine colitis-associated cancer model. We next explored the status of cyclin A2 expression in clinical CRC samples at the mRNA and protein levels and found higher expression in tumors of patients with stage 1 or 2 CRC compared with those of patients with stage 3 or 4 CRC. A meta-analysis of 11 transcriptome data sets comprising 2239 primary CRC tumors revealed different expression levels of CCNA2 (the mRNA coding for cyclin A2) among the CRC tumor subtypes, with the highest expression detected in consensus molecular subtype 1 (CMS1) and the lowest in CMS4 tumors. Moreover, we found high expression of CCNA2 to be a new, independent prognosis factor for CRC tumors.
Assuntos
Colo/metabolismo , Neoplasias Colorretais/metabolismo , Ciclina A2/metabolismo , Homeostase , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/metabolismo , Animais , Colo/patologia , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Ciclina A2/genética , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Estadiamento de Neoplasias , Neoplasias Experimentais/diagnóstico , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , PrognósticoRESUMO
OBJECTIVE: The percentage and the activity of natural killer (NK) cells are known to be decreased in HIV-infected patients. However, the mechanisms responsible for this NK deficiency are poorly understood. Because of the role of NK cells in the host defence against microbial infections, this defect contributes to the virus-induced immune deficiency. The aim of the present study was to better understand this defect in order to be able to restore NK function in HIV infection. DESIGN AND METHODS: The expression of the cytolytic mediators perforin and granzyme A was analysed by flow cytometry, the lytic activity of peripheral blood NK cells of HIV-infected patients was analysed by cytotoxic assay, and the expression of perforin was followed during administration of interferon (IFN)alpha attached to polyethylene glycol (PEG)-IFNalpha. RESULTS: The lytic activity and the expression of perforin and granzyme A was low in NK cells of infected individuals in comparison with normal control volunteers. In both groups NK cytotoxic capacity was linked to perforin expression. The low perforin expression in HIV-infected subjects negatively correlated with HIV RNA plasma level. administration of PEG-IFNalpha restored perforin expression even in patients whose viral load was not reduced by this treatment. CONCLUSIONS: These results suggest that HIV-induced NK deficiency could be partly mediated by a defect in perforin and granzyme A expression, and that PEG-IFNalpha could be used in infected subjects to directly improve their natural immunity in addition to eventually reducing their viraemia.
Assuntos
Infecções por HIV/tratamento farmacológico , HIV-1 , Interferon-alfa/uso terapêutico , Células Matadoras Naturais/metabolismo , Glicoproteínas de Membrana/metabolismo , Adulto , Contagem de Linfócito CD4 , Estudos de Casos e Controles , Testes Imunológicos de Citotoxicidade , Feminino , Granzimas , Infecções por HIV/imunologia , Humanos , Líquido Intracelular/metabolismo , Células Matadoras Naturais/imunologia , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Perforina , Proteínas Citotóxicas Formadoras de Poros , Serina Endopeptidases/metabolismo , ViremiaRESUMO
The two related basic helix-loop-helix, TAL1 and LYL1, and their cofactor LIM-only-2 protein (LMO2) are present in blood and endothelial cells. While their crucial role in early hematopoiesis is well established, their function in endothelial cells and especially in angiogenesis is less understood. Here, we identified ANGIOPOIETIN-2 (ANG-2), which encodes a major regulator of angiogenesis, as a direct transcriptional target of TAL1, LYL1 and LMO2. Knockdown of any of the three transcription factors in human blood and lymphatic endothelial cells caused ANG-2 mRNA and protein down-regulation. Transient transfections showed that the full activity of the ANG-2 promoter required the integrity of a highly conserved Ebox-GATA composite element. Accordingly, chromatin immunoprecipitation assays demonstrated that TAL1, LYL1, LMO2 and GATA2 occupied this region of ANG-2 promoter in human endothelial cells. Furthermore, we showed that LMO2 played a central role in assembling TAL1-E47, LYL1-LYL1 or/and LYL1-TAL1 dimers with GATA2. The resulting complexes were able to activate endogenous ANG-2 expression in endothelial cells as well as in non-endothelial cells. Finally, we showed that ANG-2 gene activation during angiogenesis concurred with the up-regulation of TAL1 and LMO2. Altogether, we identified ANG-2 as a bona fide target gene of LMO2-complexes with TAL1 and/or LYL1, highlighting a new function of the three hematopoietic factors in the endothelial lineage.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Angiopoietina-2/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/metabolismo , Proteínas com Domínio LIM/fisiologia , Proteínas de Neoplasias/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Angiopoietina-2/metabolismo , Sequência de Bases , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Sequência Conservada , Fator de Transcrição GATA2/fisiologia , Técnicas de Silenciamento de Genes , Genes Reporter , Células HEK293 , Humanos , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Luciferases de Vaga-Lume/biossíntese , Luciferases de Vaga-Lume/genética , Microvasos/citologia , Dados de Sequência Molecular , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Multimerização Proteica , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , RNA Interferente Pequeno/genética , Proteína 1 de Leucemia Linfocítica Aguda de Células T , Transcrição GênicaRESUMO
Tumor cells are known to interact closely with nontumoral infiltrating cells in order to grow and proliferate. Monocyte-derived cells constitute a major component of the tumoral infiltrate and a high level of these cells has been associated with increased tumor growth and poor prognosis in patients with breast cancer. For their growth and metastatic propagation, solid tumors are dependant on angiogenesis and accumulated evidences suggest that monocyte-derived cells could also play an important role in this phenomenon. However, the precise nature of proangiogenic factors secreted by these cells in breast carcinomas, and their direct influence on vessel formation, has not been determined. In the present study, we show that soluble factors secreted by breast tumor cells induce monocytes to produce a variety of proangiogenic CXC chemokines without secretion of angiostatic CXC chemokines. Using in vitro tubule formation in Matrigel, we demonstrated that the CXC chemokines secreted by MTSs (monocytes cultured with tumor cell supernatants) were able to induce microvessel formation. The profile of secreted CXC chemokines was characteristic for each tumor cell line or fresh tumor cells. This last result points out that a precise profiling of secreted proangiogenic factors inside the tumor, by tumor cells themselves or tumor-infiltrating monocyte-derived cells, is important for a precise targeting of therapeutic agents against neovascularization.
Assuntos
Indutores da Angiogênese/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Quimiocinas CXC/metabolismo , Monócitos/metabolismo , Neovascularização Patológica/fisiopatologia , Mama/metabolismo , Mama/patologia , Neoplasias da Mama/irrigação sanguínea , Diferenciação Celular , Células Cultivadas , Feminino , HumanosRESUMO
The quantity of envelope glycoprotein molecules (Env) on HIV-1 particles is still an issue of debate and, depending on the strain of virus and the nature of the producer cells, it can vary greatly. Here, we have attempted to address how Env density influences HIV-1 fitness. To this aim, we have produced HIV-1-derived viral particles with various amounts of R5 Env (low Env: Envlo; high Env: Envhi), using a regulatable expression system. The infectivity was assayed on human cells, engineered to express the HIV receptor CD4 and the co-receptor CCR5, as well as on peripheral blood lymphocytes and macrophages. In these experiments, low levels of Env were sufficient for cell infection, albeit at low efficiency. Increasing the amount of Env resulted in cooperatively improved infectivity, but a threshold was rapidly attained, indicating that only a fraction of Env was required for efficient infection. Unexpectedly, Env incorporation beyond what gives maximal infection transiently stimulated the expression of proviral genes, as well as retrovirus production, in newly infected cells. This was likely a consequence of induced NF-kappaB activity, as this transcription factor is triggered by Envhi, but not by Envlo, virions. Thus, our data suggest that one major effect of high Env density on the surface of HIV may not be better infection yields but rather improved viral production by newly infected cells.