RESUMO
Activation of the IκB kinase (IKK) complex has recurrently been linked to colorectal cancer (CRC) initiation and progression. However, identification of downstream effectors other than NF-κB has remained elusive. Here, analysis of IKK-dependent substrates in CRC cells after UV treatment revealed that phosphorylation of BRD4 by IKK-α is required for its chromatin-binding at target genes upon DNA damage. Moreover, IKK-α induces the NF-κB-dependent transcription of the cytokine LIF, leading to STAT3 activation, association with BRD4 and recruitment to specific target genes. IKK-α abrogation results in defective BRD4 and STAT3 functions and consequently irreparable DNA damage and apoptotic cell death upon different stimuli. Simultaneous inhibition of BRAF-dependent IKK-α activity, BRD4, and the JAK/STAT pathway enhanced the therapeutic potential of 5-fluorouracil combined with irinotecan in CRC cells and is curative in a chemotherapy-resistant xenograft model. Finally, coordinated expression of LIF and IKK-α is a poor prognosis marker for CRC patients. Our data uncover a functional link between IKK-α, BRD4, and JAK/STAT signaling with clinical relevance.
Assuntos
Quinase I-kappa B , Transdução de Sinais , Humanos , Quinase I-kappa B/metabolismo , NF-kappa B/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Janus Quinases/genética , Fatores de Transcrição STAT , Fosforilação , Fator de Necrose Tumoral alfa/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismoRESUMO
Phosphorylated IKKα(p45) is a nuclear active form of the IKKα kinase that is induced by the MAP kinases BRAF and TAK1 and promotes tumor growth independent of canonical NF-κB signaling. Insights into the sources of IKKα(p45) activation and its downstream substrates in the nucleus remain to be defined. Here, we discover that IKKα(p45) is rapidly activated by DNA damage independent of ATM-ATR, but dependent on BRAF-TAK1-p38-MAPK, and is required for robust ATM activation and efficient DNA repair. Abolishing BRAF or IKKα activity attenuates ATM, Chk1, MDC1, Kap1, and 53BP1 phosphorylation, compromises 53BP1 and RIF1 co-recruitment to sites of DNA lesions, and inhibits 53BP1-dependent fusion of dysfunctional telomeres. Furthermore, IKKα or BRAF inhibition synergistically enhances the therapeutic potential of 5-FU and irinotecan to eradicate chemotherapy-resistant metastatic human tumors in vivo. Our results implicate BRAF and IKKα kinases in the DDR and reveal a combination strategy for cancer treatment.
Assuntos
Dano ao DNA , Resistencia a Medicamentos Antineoplásicos , Fluoruracila/farmacologia , Quinase I-kappa B/metabolismo , Irinotecano/farmacologia , Sistema de Sinalização das MAP Quinases , Proteínas de Neoplasias , Neoplasias , Animais , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Células HCT116 , Humanos , Quinase I-kappa B/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Células MCF-7 , Camundongos , Camundongos Nus , Metástase Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Telômero/genética , Telômero/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Hematopoietic stem cells (HSCs) develop from the hemogenic endothelium in cluster structures that protrude into the embryonic aortic lumen. Although much is known about the molecular characteristics of the developing hematopoietic cells, we lack a complete understanding of their origin and the three-dimensional organization of the niche. Here, we use advanced live imaging techniques of organotypic slice cultures, clonal analysis, and mathematical modeling to show the two-step process of intra-aortic hematopoietic cluster (IACH) formation. First, a hemogenic progenitor buds up from the endothelium and undergoes division forming the monoclonal core of the IAHC. Next, surrounding hemogenic cells are recruited into the IAHC, increasing their size and heterogeneity. We identified the Notch ligand Dll4 as a negative regulator of the recruitment phase of IAHC. Blocking of Dll4 promotes the entrance of new hemogenic Gfi1+ cells into the IAHC and increases the number of cells that acquire HSC activity. Mathematical modeling based on our data provides estimation of the cluster lifetime and the average recruitment time of hemogenic cells to the cluster under physiologic and Dll4-inhibited conditions.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Aorta/embriologia , Proteínas de Ligação ao Cálcio/genética , Divisão Celular , Células Progenitoras Endoteliais/fisiologia , Feminino , Hemangioblastos/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos TeóricosRESUMO
IκBs exert principal functions as cytoplasmic inhibitors of NF-kB transcription factors. Additional roles for IκB homologues have been described, including chromatin association and transcriptional regulation. Phosphorylated and SUMOylated IκBα (pS-IκBα) binds to histones H2A and H4 in the stem cell and progenitor cell compartment of skin and intestine, but the mechanisms controlling its recruitment to chromatin are largely unknown. Here, we show that serine 32-36 phosphorylation of IκBα favors its binding to nucleosomes and demonstrate that p-IκBα association with H4 depends on the acetylation of specific H4 lysine residues. The N-terminal tail of H4 is removed during intestinal cell differentiation by proteolytic cleavage by trypsin or chymotrypsin at residues 17-19, which reduces p-IκBα binding. Inhibition of trypsin and chymotrypsin activity in HT29 cells increases p-IκBα chromatin binding but, paradoxically, impaired goblet cell differentiation, comparable to IκBα deletion. Taken together, our results indicate that dynamic binding of IκBα to chromatin is a requirement for intestinal cell differentiation and provide a molecular basis for the understanding of the restricted nuclear distribution of p-IκBα in specific stem cell compartments.
Assuntos
Cromatina , Histonas , Acetilação , Cromatina/genética , Histonas/metabolismo , Humanos , Inibidor de NF-kappaB alfa/genética , Nucleossomos/genéticaRESUMO
AIMS: Mitochondrial dysfunction and inflammation are at the core of axonal degeneration in several multifactorial neurodegenerative diseases, including multiple sclerosis, Alzheimer's disease, and Parkinson's disease. The transcriptional coregulator RIP140/NRIP1 (receptor-interacting protein 140) modulates these functions in liver and adipose tissue, but its role in the nervous system remains unexplored. Here, we investigated the impact of RIP140 in the Abcd1- mouse model of X-linked adrenoleukodystrophy (X-ALD), a genetic model of chronic axonopathy involving the convergence of redox imbalance, bioenergetic failure, and chronic inflammation. METHODS AND RESULTS: We provide evidence that RIP140 is modulated through a redox-dependent mechanism driven by very long-chain fatty acids (VLCFAs), the levels of which are increased in X-ALD. Genetic inactivation of RIP140 prevented mitochondrial depletion and dysfunction, bioenergetic failure, inflammatory dysregulation, axonal degeneration and associated locomotor disabilities in vivo in X-ALD mouse models. CONCLUSIONS: Together, these findings show that aberrant overactivation of RIP140 promotes neurodegeneration in X-ALD, underscoring its potential as a therapeutic target for X-ALD and other neurodegenerative disorders that present with metabolic and inflammatory dyshomeostasis.
Assuntos
Adrenoleucodistrofia , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Transportadores de Cassetes de Ligação de ATP/uso terapêutico , Adrenoleucodistrofia/genética , Adrenoleucodistrofia/metabolismo , Animais , Modelos Animais de Doenças , Homeostase , Camundongos , Mitocôndrias/metabolismo , Proteína 1 de Interação com Receptor NuclearRESUMO
The intestinal epithelium is a paradigm of adult tissue in constant regeneration that is supported by intestinal stem cells (ISCs). The mechanisms regulating ISC homeostasis after injury are poorly understood. We previously demonstrated that IκBα, the main regulator of NF-κB, exerts alternative nuclear functions as cytokine sensor in a subset of PRC2-regulated genes. Here, we show that nuclear IκBα is present in the ISC compartment. Mice deficient for IκBα show altered intestinal cell differentiation with persistence of a fetal-like ISC phenotype, associated with aberrant PRC2 activity at specific loci. Moreover, IκBα-deficient intestinal cells produce morphologically aberrant organoids carrying a PRC2-dependent fetal-like transcriptional signature. DSS treatment, which induces acute damage in the colonic epithelium of mice, results in a temporary loss of nuclear P-IκBα and its subsequent accumulation in early CD44-positive regenerating areas. Importantly, IκBα-deficient mice show higher resistance to damage, likely due to the persistent fetal-like ISC phenotype. These results highlight intestinal IκBα as a chromatin sensor of inflammation in the ISC compartment.
Assuntos
Intestinos , Células-Tronco , Animais , Mucosa Intestinal , Camundongos , Inibidor de NF-kappaB alfa/genética , FenótipoRESUMO
The t(4;11)(q21;q23) translocation is associated with high-risk infant pro-B-cell acute lymphoblastic leukemia and arises prenatally during embryonic/fetal hematopoiesis. The developmental/pathogenic contribution of the t(4;11)-resulting MLL-AF4 (MA4) and AF4-MLL (A4M) fusions remains unclear; MA4 is always expressed in patients with t(4;11)+ B-cell acute lymphoblastic leukemia, but the reciprocal fusion A4M is expressed in only half of the patients. Because prenatal leukemogenesis manifests as impaired early hematopoietic differentiation, we took advantage of well-established human embryonic stem cell-based hematopoietic differentiation models to study whether the A4M fusion cooperates with MA4 during early human hematopoietic development. Co-expression of A4M and MA4 strongly promoted the emergence of hemato-endothelial precursors, both endothelial- and hemogenic-primed. Double fusion-expressing hemato-endothelial precursors specified into significantly higher numbers of both hematopoietic and endothelial-committed cells, irrespective of the differentiation protocol used and without hijacking survival/proliferation. Functional analysis of differentially expressed genes and differentially enriched H3K79me3 genomic regions by RNA-sequencing and H3K79me3 chromatin immunoprecipitation-sequencing, respectively, confirmed a hematopoietic/endothelial cell differentiation signature in double fusion-expressing hemato-endothelial precursors. Importantly, chromatin immunoprecipitation-sequencing analysis revealed a significant enrichment of H3K79 methylated regions specifically associated with HOX-A cluster genes in double fusion-expressing differentiating hematopoietic cells. Overall, these results establish a functional and molecular cooperation between MA4 and A4M fusions during human hematopoietic development.
Assuntos
Diferenciação Celular/genética , Desenvolvimento Embrionário/genética , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Hematopoese/genética , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Fusão Oncogênica/genética , Animais , Apoptose/genética , Ciclo Celular/genética , Técnicas de Cocultura , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Histonas/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Metilação , Camundongos , Camundongos KnockoutRESUMO
BACKGROUND: Colorectal cancer is a common cause of death in developed countries. Progression from adenoma to invasive carcinoma requires accumulation of mutations starting with the Adenomatous Polyposis Coli (Apc) gene. NF-κB signalling is a key element in cancer, mainly related to the activity of IKKß. IKKα kinase also participates in this process by mechanisms that are primarily unknown. METHODS: We generated a compound mouse model with mutation in Apc and lacking intestinal epithelial IKKα, produced intestinal organoids and tumour spheroids with different genetic backgrounds, and performed immunohistochemistry and RNA-seq analysis. RESULTS: Deficiency of IKKα prevents adenoma formation, with adenomas lacking IKKα showing reduced proliferation. In contrast, IKKα status did not affect normal intestinal function. The same divergent phenotype was found in the organoid-spheroid model. We also found that epithelial IKKα controls stemness, proliferation and apoptosis-related expression. CONCLUSIONS: IKKα is a potential therapeutic target for Apc mutant colorectal cancer patients.
Assuntos
Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/patologia , Quinase I-kappa B/deficiência , Mucosa Intestinal/enzimologia , Mucosa Intestinal/patologia , Células-Tronco Neoplásicas/enzimologia , Células-Tronco Neoplásicas/patologia , Proteína da Polipose Adenomatosa do Colo/genética , Animais , Proliferação de Células/fisiologia , Neoplasias Colorretais/genética , Quinase I-kappa B/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Genetic data indicate that abrogation of Notch-Rbpj or Wnt-ß-catenin pathways results in the loss of the intestinal stem cells (ISCs). However, whether the effect of Notch is direct or due to the aberrant differentiation of the transit-amplifying cells into post-mitotic goblet cells is unknown. To address this issue, we have generated composite tamoxifen-inducible intestine-specific genetic mouse models and analyzed the expression of intestinal differentiation markers. Importantly, we found that activation of ß-catenin partially rescues the differentiation phenotype of Rbpj deletion mutants, but not the loss of the ISC compartment. Moreover, we identified Bmi1, which is expressed in the ISC and progenitor compartments, as a gene that is co-regulated by Notch and ß-catenin. Loss of Bmi1 resulted in reduced proliferation in the ISC compartment accompanied by p16(INK4a) and p19(ARF) (splice variants of Cdkn2a) accumulation, and increased differentiation to the post-mitotic goblet cell lineage that partially mimics Notch loss-of-function defects. Finally, we provide evidence that Bmi1 contributes to ISC self-renewal.
Assuntos
Intestinos/patologia , Complexo Repressor Polycomb 1/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Compartimento Celular , Proliferação de Células , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p19/genética , Inibidor de Quinase Dependente de Ciclina p19/metabolismo , Reparo do DNA , Homeostase , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/deficiência , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Intestinos/anormalidades , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Complexo Repressor Polycomb 1/deficiência , Complexo Repressor Polycomb 1/genética , Proteínas Proto-Oncogênicas/deficiência , Proteínas Proto-Oncogênicas/genética , Receptores Notch/deficiência , Ativação Transcricional/genética , Proteínas Wnt/metabolismo , beta Catenina/metabolismoRESUMO
BACKGROUND AND AIMS: Qigong has been used as a complementary therapy to improve different health-related problems. This study aims to test the effects of Qigong on quality of life, pain and depressive symptoms in older hospitalized patients. METHODS: In this randomized controlled single blind study, we randomized 58 ≥50 years adults admitted to a post-acute intermediate care rehabilitation facility, to receive a 90 min, bi-weekly, 4-week structured Qigong intervention plus usual care and rehabilitation (N = 29) or usual care and rehabilitation alone (N = 29). Outcomes included quality of life (0-100 points visual analogical scale), pain (0-10 points scale), and depressive symptoms (5-item modified Yesavage Geriatric Depression Scale). We also evaluated participants' compliance and safety. RESULTS: Of the enrolled 58 participants (mean age ± SD = 74.3 ± 8.2 years, 88 % women) we dropped-out four in the control group. No statistically significant differences in baseline characteristics were shown between groups, including age, gender, marital status, education, comorbidity and functional status, main diagnostic at admission and number of rehabilitation sessions. In an intention-to-treat analysis (repeated measures ANOVA) the intervention group experienced a significant improvement in quality of life (mean increase of 19 points vs 2.6 points for controls, p = 0.002). Pain and depressive symptoms improved in both groups. Adherence was good (79 % of participants completed the whole program). No adverse events were reported. CONCLUSIONS: According to our results, a structured Qigong intervention, together with usual care, might contribute to improve quality of life of patients admitted to a post-acute intermediate care rehabilitation unit, compared to usual care.
Assuntos
Depressão/prevenção & controle , Dor/prevenção & controle , Qigong , Qualidade de Vida , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Método Simples-CegoRESUMO
BACKGROUND: PTOV1 is an adaptor protein with functions in diverse processes, including gene transcription and protein translation, whose overexpression is associated with a higher proliferation index and tumor grade in prostate cancer (PC) and other neoplasms. Here we report its interaction with the Notch pathway and its involvement in PC progression. METHODS: Stable PTOV1 knockdown or overexpression were performed by lentiviral transduction. Protein interactions were analyzed by co-immunoprecipitation, pull-down and/or immunofluorescence. Endogenous gene expression was analyzed by real time RT-PCR and/or Western blotting. Exogenous promoter activities were studied by luciferase assays. Gene promoter interactions were analyzed by chromatin immunoprecipitation assays (ChIP). In vivo studies were performed in the Drosophila melanogaster wing, the SCID-Beige mouse model, and human prostate cancer tissues and metastasis. The Excel package was used for statistical analysis. RESULTS: Knockdown of PTOV1 in prostate epithelial cells and HaCaT skin keratinocytes caused the upregulation, and overexpression of PTOV1 the downregulation, of the Notch target genes HEY1 and HES1, suggesting that PTOV1 counteracts Notch signaling. Under conditions of inactive Notch signaling, endogenous PTOV1 associated with the HEY1 and HES1 promoters, together with components of the Notch repressor complex. Conversely, expression of active Notch1 provoked the dismissal of PTOV1 from these promoters. The antagonist role of PTOV1 on Notch activity was corroborated in the Drosophila melanogaster wing, where human PTOV1 exacerbated Notch deletion mutant phenotypes and suppressed the effects of constitutively active Notch. PTOV1 was required for optimal in vitro invasiveness and anchorage-independent growth of PC-3 cells, activities counteracted by Notch, and for their efficient growth and metastatic spread in vivo. In prostate tumors, the overexpression of PTOV1 was associated with decreased expression of HEY1 and HES1, and this correlation was significant in metastatic lesions. CONCLUSIONS: High levels of the adaptor protein PTOV1 counteract the transcriptional activity of Notch. Our evidences link the pro-oncogenic and pro-metastatic effects of PTOV1 in prostate cancer to its inhibitory activity on Notch signaling and are supportive of a tumor suppressor role of Notch in prostate cancer progression.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/biossíntese , Biomarcadores Tumorais/genética , Proteínas de Ciclo Celular/biossíntese , Proteínas de Homeodomínio/biossíntese , Proteínas de Neoplasias/genética , Neoplasias da Próstata/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Biomarcadores Tumorais/metabolismo , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Drosophila melanogaster , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Proteínas de Homeodomínio/genética , Humanos , Masculino , Camundongos , Metástase Neoplásica , Proteínas de Neoplasias/metabolismo , Neoplasias da Próstata/patologia , Receptores Notch/biossíntese , Transdução de Sinais/genética , Fatores de Transcrição HES-1 , Ativação Transcricional/genéticaRESUMO
X-linked adrenoleukodystrophy (X-ALD) is an inherited disorder characterized by axonopathy and demyelination in the central nervous system and adrenal insufficiency. Main X-ALD phenotypes are: (i) an adult adrenomyeloneuropathy (AMN) with axonopathy in spinal cords, (ii) cerebral AMN with brain demyelination (cAMN) and (iii) a childhood variant, cALD, characterized by severe cerebral demyelination. Loss of function of the ABCD1 peroxisomal fatty acid transporter and subsequent accumulation of very-long-chain fatty acids (VLCFAs) are the common culprits to all forms of X-ALD, an aberrant microglial activation accounts for the cerebral forms, whereas inflammation allegedly plays no role in AMN. How VLCFA accumulation leads to neurodegeneration and what factors account for the dissimilar clinical outcomes and prognosis of X-ALD variants remain elusive. To gain insights into these questions, we undertook a transcriptomic approach followed by a functional-enrichment analysis in spinal cords of the animal model of AMN, the Abcd1(-) null mice, and in normal-appearing white matter of cAMN and cALD patients. We report that the mouse model shares with cAMN and cALD a common signature comprising dysregulation of oxidative phosphorylation, adipocytokine and insulin signaling pathways, and protein synthesis. Functional validation by quantitative polymerase chain reaction, western blots and assays in spinal cord organotypic cultures confirmed the interplay of these pathways through IkB kinase, being VLCFA in excess a causal, upstream trigger promoting the altered signature. We conclude that X-ALD is, in all its variants, a metabolic/inflammatory syndrome, which may offer new targets in X-ALD therapeutics.
Assuntos
Adipocinas/metabolismo , Adrenoleucodistrofia/genética , Adrenoleucodistrofia/metabolismo , Encéfalo/metabolismo , Ácidos Graxos/metabolismo , Redes e Vias Metabólicas , Fosforilação Oxidativa , Medula Espinal/metabolismo , Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Adiponectina/metabolismo , Adulto , Animais , Vias Biossintéticas , Criança , Modelos Animais de Doenças , Progressão da Doença , Expressão Gênica , Perfilação da Expressão Gênica , Estudos de Associação Genética , Humanos , Quinase I-kappa B/metabolismo , Insulina/metabolismo , Resistência à Insulina , Leptina/metabolismo , Camundongos , NF-kappa B/metabolismo , Estresse Oxidativo , Transdução de Sinais , Receptores Toll-Like/metabolismo , TranscriptomaRESUMO
Notch is a well-conserved signaling pathway and its function in cell fate determination is crucial in embryonic development and in the maintenance of tissue homeostasis during adult life. Notch activation depends on cell-cell interactions that are essential for the generation of cell diversity from initially equivalent cell populations. In the adult hematopoiesis, Notch is undoubtedly a very efficient promoter of T-cell differentiation, and this has masked for a long time the effects of Notch on other blood lineages, which are gradually being identified. However, the adult hematopoietic stem cell (HSC) remains mostly refractory to Notch intervention in experimental systems. In contrast, Notch is essential for the generation of the HSCs, which takes place during embryonic development. This review summarizes the knowledge accumulated in recent years regarding the role of the Notch pathway in the different stages of HSC ontology from embryonic life to fetal and adult bone marrow stem cells. In addition, we briefly examine other systems where Notch regulates specific stem cell capacities, in an attempt to understand how Notch functions in stem cell biology.
Assuntos
Diferenciação Celular , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Desenvolvimento Embrionário , Hematopoese/fisiologia , Humanos , LigantesAssuntos
Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/prevenção & controle , Proteína Jagged-1/antagonistas & inibidores , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Humanos , Terapia de Alvo Molecular/métodos , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Cervical cancer (CC) is a viral-associated tumor caused by the infection with the human papilloma virus. CC is then an immunogenic cancer that expresses viral antigens. Despite being immunogenic, CC does not fully respond to immune checkpoint inhibitors (ICI). LIF is a crucial cytokine in embryo implantation, involved in maternal tolerance that acts as an immunomodulatory factor in cancer. LIF is expressed in CC and high levels of LIF is associated with poor prognosis in CC. METHODS: We evaluated the impact of LIF on the immune response to ICI using primary plasmocytoid dendritic cells (pDCs) and macrophage cultures, syngeneic animals and patient-derived models that recapitulate the human tumor microenvironment. RESULTS: We found that the viral proteins E6 and E7 induce the expression of LIF via the NFκB pathway. The secreted LIF can then repress type I interferon expressed in pDCs, and CXCL9 expressed in tumor associated macrophages. Blockade of LIF promotes the induction of type I interferon and CXCL9 inducing the tumor infiltration of CD8 T cell. This results in the sensitization of the tumor to ICI. Importantly, we observed that patients with CC expressing high levels of LIF tent to be resistant to ICI. CONCLUSION: Our data show that the HPV virus induces the expression of LIF to provide a selective advantage to the tumor cell by generating local immunosuppression via the repression of type I interferon and CXCL9. Combinatory treatment with blocking antibodies against LIF and ICI could be effective against CC expressing high levels of LIF.
RESUMO
BACKGROUND: We have previously shown that non-curative chemotherapy imposes fetal conversion and high metastatic capacity to cancer cells. From the set of genes differentially expressed in Chemotherapy Resistant Cells, we obtained a characteristic fetal intestinal cell signature that is present in a group of untreated tumors and is sufficient to predict patient prognosis. A feature of this fetal signature is the loss of CDX1. METHODS: We have analyzed transcriptomic data in public datasets and performed immunohistochemistry analysis of paraffin embedded tumor samples from two cohorts of colorectal cancer patients. RESULTS: We demonstrated that low levels of CDX1 are sufficient to identify patients with poorest outcome at the early tumor stages II and III. Presence tumor areas that are negative for CDX1 staining in stage I cancers is associated with tumor relapse. CONCLUSIONS: Our results reveal the actual possibility of incorporating CDX1 immunostaining as a valuable biomarker for CRC patients.
Assuntos
Neoplasias Colorretais , Humanos , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/tratamento farmacológico , Perfilação da Expressão Gênica , Transcriptoma , Imuno-Histoquímica , Proteínas de Homeodomínio/genéticaRESUMO
Recent findings suggest that Hematopoietic Stem Cells (HSC) and progenitors arise simultaneously and independently of each other already in the embryonic aorta-gonad mesonephros region, but it is still unknown how their different features are established. Here, we uncover IκBα (Nfkbia, the inhibitor of NF-κB) as a critical regulator of HSC proliferation throughout development. IκBα balances retinoic acid signaling levels together with the epigenetic silencer, PRC2, specifically in HSCs. Loss of IκBα decreases proliferation of HSC and induces a dormancy related gene expression signature instead. Also, IκBα deficient HSCs respond with superior activation to in vitro culture and in serial transplantation. At the molecular level, chromatin regions harboring binding motifs for retinoic acid signaling are hypo-methylated for the PRC2 dependent H3K27me3 mark in IκBα deficient HSCs. Overall, we show that the proliferation index in the developing HSCs is regulated by a IκBα-PRC2 axis, which controls retinoic acid signaling.
Assuntos
Proliferação de Células , Células-Tronco Hematopoéticas , Inibidor de NF-kappaB alfa , Transdução de Sinais , Tretinoína , Animais , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/citologia , Tretinoína/metabolismo , Inibidor de NF-kappaB alfa/metabolismo , Inibidor de NF-kappaB alfa/genética , Camundongos , Desenvolvimento Embrionário/genética , Camundongos Knockout , Complexo Repressor Polycomb 2/metabolismo , Complexo Repressor Polycomb 2/genética , Camundongos Endogâmicos C57BL , Regulação da Expressão Gênica no Desenvolvimento , FemininoRESUMO
Parkinson's disease (PD) is a neurodegenerative disease associated with progressive death of midbrain dopamine (DAn) neurons in the substantia nigra (SN). Since it has been proposed that patients with PD exhibit an overall proinflammatory state, and since astrocytes are key mediators of the inflammation response in the brain, here we sought to address whether astrocyte-mediated inflammatory signaling could contribute to PD neuropathology. For this purpose, we generated astrocytes from induced pluripotent stem cells (iPSCs) representing patients with PD and healthy controls. Transcriptomic analyses identified a unique inflammatory gene expression signature in PD astrocytes compared with controls. In particular, the proinflammatory cytokine IL-6 was found to be highly expressed and released by PD astrocytes and was found to induce toxicity in DAn. Mechanistically, neuronal cell death was mediated by IL-6 receptor (IL-6R) expressed in human PD neurons, leading to downstream activation of STAT3. Blockage of IL-6R by the addition of the FDA-approved anti-IL-6R antibody, Tocilizumab, prevented PD neuronal death. SN neurons overexpressing IL-6R and reactive astrocytes expressing IL-6 were detected in postmortem brain tissue of patients at early stages of PD. Our findings highlight the potential role of astrocyte-mediated inflammatory signaling in neuronal loss in PD and pave the way for the design of future therapeutics.
Assuntos
Células-Tronco Pluripotentes Induzidas , Doenças Neurodegenerativas , Doença de Parkinson , Humanos , Doença de Parkinson/metabolismo , Astrócitos/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Interleucina-6/metabolismo , Doenças Neurodegenerativas/patologia , Neurônios Dopaminérgicos/metabolismoRESUMO
Hematopoietic stem cells (HSCs) develop from the hemogenic endothelium (HE) in the aorta- gonads-and mesonephros (AGM) region and reside within Intra-aortic hematopoietic clusters (IAHC) along with hematopoietic progenitors (HPC). The signalling mechanisms that distinguish HSCs from HPCs are unknown. Notch signaling is essential for arterial specification, IAHC formation and HSC activity, but current studies on how Notch segregates these different fates are inconsistent. We now demonstrate that Notch activity is highest in a subset of, GFI1 + , HSC-primed HE cells, and is gradually lost with HSC maturation. We uncover that the HSC phenotype is maintained due to increasing levels of NOTCH1 and JAG1 interactions on the surface of the same cell (cis) that renders the NOTCH1 receptor from being activated. Forced activation of the NOTCH1 receptor in IAHC activates a hematopoietic differentiation program. Our results indicate that NOTCH1-JAG1 cis-inhibition preserves the HSC phenotype in the hematopoietic clusters of the embryonic aorta.