RESUMEN
Foxm1 is known as a typical proliferation-associated transcription factor. Here we found that Foxm1 was essential for maintenance of the quiescence and self-renewal capacity of hematopoietic stem cells (HSCs) in vivo in mice. Reducing expression of FOXM1 also decreased the quiescence of human CD34(+) HSCs and progenitor cells, and its downregulation was associated with a subset of myelodysplastic syndrome (MDS). Mechanistically, Foxm1 directly bound to the promoter region of the gene encoding the receptor Nurr1 (Nr4a2; called 'Nurr1' here), inducing transcription, while forced expression of Nurr1 reversed the loss of quiescence observed in Foxm1-deficient cells in vivo. Thus, our studies reveal a previously unrecognized role for Foxm1 as a critical regulator of the quiescence and self-renewal of HSCs mediated at least in part by control of Nurr1 expression.
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Proliferación Celular/genética , Factores de Transcripción Forkhead/genética , Regulación de la Expresión Génica , Células Madre Hematopoyéticas/metabolismo , Animales , Células Cultivadas , Citometría de Flujo , Proteína Forkhead Box M1 , Factores de Transcripción Forkhead/metabolismo , Perfilación de la Expresión Génica , Células HEK293 , Humanos , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/metabolismo , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Regiones Promotoras Genéticas/genética , Unión Proteica , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Leukemia stem cells (LSC) represent a crucial and rare subset of cells present in acute myeloid leukemia (AML); they play a pivotal role in the initiation, maintenance, and relapse of this disease. Targeting LSC holds great promise for preventing AML relapse and improving long-term outcomes. However the precise molecular mechanisms governing LSC self-renewal are still poorly understood. Here, we present compelling evidence that the expression of miR-30e-5p, a potential tumor-suppressive microRNA, is significantly lower in AML samples than in healthy bone marrow samples. Forced expression of miR- 30e effectively inhibits leukemogenesis, impairs LSC self-renewal, and delays leukemia progression. Mechanistically, Cyb561 acts as a direct target of miR-30e-5p in LSC, and its deficiency restricts the self-renewal of LSC by activating reactive oxygen series signaling and markedly prolongs recipients' survival. Moreover, genetic or pharmacological overexpression of miR-30e-5p or knockdown of Cyb561 suppresses the growth of human AML cells. In conclusion, our findings establish the crucial role of the miR-30e-5p/Cyb561/ROS axis in finely regulating LSC self-renewal, highlighting Cyb561 as a potential therapeutic target for LSC-directed therapies.
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Leucemia Mieloide Aguda , MicroARNs , Humanos , Especies Reactivas de Oxígeno , Autorrenovación de las Células/genética , MicroARNs/genética , Transducción de Señal , Recurrencia , Proliferación Celular/genética , Línea Celular TumoralRESUMEN
Bioactive materials have been extensively developed for the adjuvant therapy of cancer. However, few materials can meet the requirements for the postoperative resection of hepatocellular carcinoma (HCC) due to massive bleeding and high recurrence. In particular, combination therapy for HCC has been highly recommended in clinical practice, including surgical resection, interventional therapy, ablation therapy and chemotherapy. Herein, an injectable magnetic colloidal gel (MCG) was developed by controllable electrostatic attraction between clinically available magnetic montmorillonites and amphoteric gelatin nanoparticles. The optimized MCG exhibited an effective magnetic heating effect, remarkable rheological properties, and high gel network stability, realizing the synergistic treatment of postoperative HCC by stimuli-responsive drug delivery, hemostasis and magnetic hyperthermia. Furthermore, a minimal invasive MCG-induced interventional magnetic hyperthermia therapy (MHT) under ultrasound guidance was realized on hepatic tumor rabbits, providing an alternative therapeutics to treat the postoperative recurrence. Overall, MCG is a clinically available injectable formulation for adjuvant therapy after HCC surgical resection.
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Carcinoma Hepatocelular , Hipertermia Inducida , Neoplasias Hepáticas , Animales , Bentonita/uso terapéutico , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/patología , Fenómenos Magnéticos , ConejosRESUMEN
BACKGROUND: PTEN/AKT signaling plays a key role in prostate cancer development and maintenance of prostate cancer stem cells. How other oncogenes or tumor suppressors interact with this pathway remain to be elucidated. SLUG is an zinc finger transcription factor of the Snail superfamily, and it promotes cancer metastasis and determines the mammary stem cell state. METHODS: SLUG was overexpressed in cells by retroviral vector and knockdown of SLUG and PTEN was mediated by shRNAs-expressing lentiviruses. Expression level of SLUG and PTEN was examined by Western blot, RT-PCR, and qPCR analyses. PTEN promoter activity was measured by luciferase reporter assay. ChIP assay was used to measure the binding between SLUG and the PTEN promoter in vivo. RESULT: We showed that overexpression of SLUG decreased expression of PTEN tumor repressor in prostate cancer cell lines 22RV1 and DU145; conversely, knockdown of SLUG expression elevated PTEN expresson at both protein and RNA level in these cells. We demonstrated that SLUG overexpression inhibits PTEN promoter activity through the proximal promoter region in prostate cancer cells. By ChIP assay, we confirmed that SLUG directly binds to the PTEN promoter region covering the E-box sites. We also showed that Slug deficiency leads to an increased expression of PTEN in mouse embryo fibroblasts and prostate tissues. Importantly, we found that overexpression of SLUG increases drug resistance of DU145 prostate cancer cell line and knockdown of SLUG by shRNA sensitizes DU145 cell line to chemotherapeutic drugs. We further demonstrated that PTEN knockdown converts drug sensitivity of DU145 cells expressing SLUG shRNA to anticancer drugs. CONCLUSION: We provide compelling evidence showing that PTEN is a direct functional target of SLUG. Our findings offer new insight in the regulation of the PTEN/AKT pathway and provide a molecular basis for potential targeted therapies of prostate cancer Prostate 75:907-916, 2015. © 2015 Wiley Periodicals, Inc.
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Fosfohidrolasa PTEN/biosíntesis , Neoplasias de la Próstata/metabolismo , Factores de Transcripción/biosíntesis , Animales , Western Blotting , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Fosfohidrolasa PTEN/antagonistas & inhibidores , Fosfohidrolasa PTEN/genética , Regiones Promotoras Genéticas , Neoplasias de la Próstata/genética , ARN Neoplásico/química , ARN Neoplásico/genética , ARN Interferente Pequeño/administración & dosificación , ARN Interferente Pequeño/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Transcripción de la Familia Snail , Factores de Transcripción/genéticaRESUMEN
Interstitial loss of all or part of the long arm of chromosome 5, or del(5q), is a frequent clonal chromosomal abnormality in human myelodysplastic syndrome (MDS, a preleukemic disorder) and acute myeloid leukemia (AML), and is thought to contribute to the pathogenesis of these diseases by deleting one or more tumor-suppressor genes. Although a major commonly deleted region (CDR) has been delineated on chromosome band 5q31.1 (refs. 3-7), attempts to identify tumor suppressors within this band have been unsuccessful. We focused our analysis of gene expression on RNA from primitive leukemia-initiating cells, which harbor 5q deletions, and analyzed 12 genes within the CDR that are expressed by normal hematopoietic stem cells. Here we show that the gene encoding alpha-catenin (CTNNA1) is expressed at a much lower level in leukemia-initiating stem cells from individuals with AML or MDS with a 5q deletion than in individuals with MDS or AML lacking a 5q deletion or in normal hematopoietic stem cells. Analysis of HL-60 cells, a myeloid leukemia line with deletion of the 5q31 region, showed that the CTNNA1 promoter of the retained allele is suppressed by both methylation and histone deacetylation. Restoration of CTNNA1 expression in HL-60 cells resulted in reduced proliferation and apoptotic cell death. Thus, loss of expression of the alpha-catenin tumor suppressor in hematopoietic stem cells may provide a growth advantage that contributes to human MDS or AML with del(5q).
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Transformación Celular Neoplásica , Deleción Cromosómica , Cromosomas Humanos Par 5/genética , Células Progenitoras Mieloides/patología , alfa Catenina/genética , Enfermedad Aguda , Western Blotting , Línea Celular , Línea Celular Tumoral , Metilación de ADN/efectos de los fármacos , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Células HL-60 , Humanos , Ácidos Hidroxámicos/farmacología , Hibridación Fluorescente in Situ/métodos , Células K562 , Leucemia Mieloide/sangre , Leucemia Mieloide/genética , Leucemia Mieloide/patología , Mutación , Síndromes Mielodisplásicos/sangre , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/patología , Células Progenitoras Mieloides/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transfección , Células U937 , alfa Catenina/metabolismoRESUMEN
Liver injury stimulates hepatocyte replication and hepatic stellate cell (HSC) activation, thereby driving liver regeneration. Aberrant HSC activation induces liver fibrosis. However, mechanisms underlying liver regeneration and fibrosis remain poorly understood. Here, we identify hepatic Snai1 and Snai2 as important transcriptional regulators for liver regeneration and fibrosis. Partial hepatectomy or CCl4 treatment increases occupancies of Snai1 and Snai2 on cyclin A2 and D1 promoters in the liver. Snai1 and Snai2 in turn increase promoter H3K27 acetylation and cyclin A2/D1 expressions. Hepatocyte-specific deletion of both Snai1 and Snai2, but not one alone, suppresses liver cyclin A2/D1 expression and regenerative hepatocyte proliferation after hepatectomy or CCl4 treatments but augments CCl4-stimulated HSC activation and liver fibrosis. Conversely, Snai2 overexpression in the liver enhances hepatocyte replication and suppresses liver fibrosis after CCl4 treatment. These results suggest that hepatic Snai1 and Snai2 directly promote, via histone modifications, reparative hepatocyte replication and indirectly inhibit liver fibrosis.
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Ciclina A2 , Regeneración Hepática , Animales , Ratones , Ciclina A2/metabolismo , Hepatectomía , Hígado/metabolismo , Cirrosis Hepática/genética , Cirrosis Hepática/metabolismo , Regeneración Hepática/fisiologíaRESUMEN
Osteosarcoma (OS) is considered the most frequent type of primary malignant bone tumor. Currently, radiotherapy, photodynamic (PDT), and other therapies for osteosarcoma are limited by tumor hypoxia and single efficacy and serve side-effects. Herein, we reported a microalgal drug delivery system (SpiD), doxorubicin (DOX)-loaded Spirulina platensis (Spi) for OS therapy. The specific surface of Spirulina platensis allowed for effective loading of DOX via surface channels and electrostatic interactions. Under 650 nm laser irradiation, SpiD enabled high oxygen production by photosynthesis and enhanced reactive oxygen species (ROS) generation via chlorophyll-assisted photosensitization, synergistically killing tumor cells with the released DOX. Combined chemotherapy and enhanced PDT mediated by SpiD exerted synergic antitumor effects and resulted in potent therapeutic efficacy in orthotopic osteosarcoma mice. Furthermore, SpiD could reduce the side-effects of chemotherapy, showing excellent blood and tissue safety. Taken together, this microalgal drug delivery system provided a natural, efficient, safe, and inexpensive strategy for OS treatment.
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Neoplasias Óseas , Nanopartículas , Osteosarcoma , Fotoquimioterapia , Spirulina , Animales , Ratones , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Osteosarcoma/tratamiento farmacológico , Sistemas de Liberación de Medicamentos , Neoplasias Óseas/tratamiento farmacológico , Línea Celular TumoralRESUMEN
Osteoarthritis (OA) is the most common form of arthritis. However, the exact pathogenesis remains unclear. Emerging evidence shows that N6-methyladenosine (m6A) modification may have an important role in OA pathogenesis. This study aimed to investigate the role of m6A writers and the underlying mechanisms in osteoarthritic cartilage. Among m6A methyltransferases, Wilms tumor 1-associated protein (WTAP) expression most significantly differed in clinical osteoarthritic cartilage. WTAP regulated extracellular matrix (ECM) degradation, inflammation and antioxidation in human chondrocytes. Mechanistically, the m6A modification and relative downstream targets in osteoarthritic cartilage were assessed by methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing, which indicated that the expression of frizzled-related protein (FRZB), a secreted Wnt antagonist, was abnormally decreased and accompanied by high m6A modification in osteoarthritic cartilage. In vitro dysregulated WTAP had positive effects on ß-catenin expression by targeting FRZB, which finally contributed to the cartilage injury phenotype in chondrocytes. Intra-articular injection of adeno-associated virus-WTAP alleviated OA progression in a mouse model, while this protective effect could be reversed by the application of a Wnt/ß-catenin activator. In summary, this study revealed that WTAP-dependent RNA m6A modification contributed to Wnt/ß-catenin pathway activation and OA progression through post-transcriptional regulation of FRZB mRNA, thus providing a potentially effective therapeutic strategy for OA treatment.
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Osteoartritis , beta Catenina , Animales , Humanos , Ratones , beta Catenina/genética , beta Catenina/metabolismo , Cartílago/metabolismo , Proteínas de Ciclo Celular/metabolismo , Condrocitos/metabolismo , Osteoartritis/metabolismo , Factores de Empalme de ARN/metabolismo , ARN Mensajero/genética , Vía de Señalización Wnt/fisiologíaRESUMEN
Percutaneous thermotherapy, a minimally invasive operational procedure, is employed in the ablation of deep tumor lesions by means of target-delivering heat. Conventional thermal ablation methods, such as radiofrequency or microwave ablation, to a certain extent, are subjected to extended ablation time as well as biosafety risks of unwanted overheating. Given its effectiveness and safety, percutaneous thermotherapy gains a fresh perspective, thanks to magnetic hyperthermia. In this respect, an injectable- and magnetic-hydrogel-construct-based thermal ablation agent is likely to be a candidate for the aforementioned clinical translation. Adopting a simple and environment-friendly strategy, a magnetic colloidal hydrogel injection is introduced by a binary system comprising super-paramagnetic Fe3O4 nanoparticles and gelatin nanoparticles. The colloidal hydrogel constructs, unlike conventional bulk hydrogel, can be easily extruded through a percutaneous needle and then self-heal in a reversible manner owing to the unique electrostatic cross-linking. The introduction of magnetic building blocks is exhibited with a rapid magnetothermal response to an alternating magnetic field. Such hydrogel injection is capable of generating heat without limitation of deep penetration. The materials achieve outstanding therapeutic results in mouse and rabbit models. These findings constitute a new class of locoregional interventional thermal therapies with minimal collateral damages.
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Carcinoma Hepatocelular , Coloides , Hidrogeles , Neoplasias Hepáticas , Animales , Conejos , Ratones , Hidrogeles/química , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/patología , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/patología , Coloides/química , Gelatina/química , Humanos , Nanopartículas de Magnetita/química , Nanopartículas de Magnetita/uso terapéutico , Hipertermia Inducida/métodos , Línea Celular Tumoral , Inyecciones , Nanopartículas Magnéticas de Óxido de Hierro/químicaRESUMEN
The severity of osteoarthritis (OA) and cartilage degeneration is highly associated with synovial inflammation. Although recent investigations have revealed a dysregulated crosstalk between fibroblast-like synoviocytes (FLSs) and macrophages in the pathogenesis of synovitis, limited knowledge is available regarding the involvement of exosomes. Here, increased exosome secretion is observed in FLSs from OA patients. Notably, internalization of inflammatory FLS-derived exosomes (inf-exo) can enhance the M1 polarization of macrophages, which further induces an OA-like phenotype in co-cultured chondrocytes. Intra-articular injection of inf-exo induces synovitis and exacerbates OA progression in murine models. In addition, it is demonstrated that inf-exo stimulation triggers the activation of glycolysis. Inhibition of glycolysis using 2-DG successfully attenuates excessive M1 polarization triggered by inf-exo. Mechanistically, HIF1A is identified as the determinant transcription factor, inhibition of which, both pharmacologically or genetically, relieves macrophage inflammation triggered by inf-exo-induced hyperglycolysis. Furthermore, in vivo administration of an HIF1A inhibitor alleviates experimental OA. The results provide novel insights into the involvement of FLS-derived exosomes in OA pathogenesis, suggesting that inf-exo-induced macrophage dysfunction represents an attractive target for OA therapy.
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Exosomas , Osteoartritis , Sinoviocitos , Sinovitis , Humanos , Ratones , Animales , Sinoviocitos/patología , Sinoviocitos/fisiología , Células Cultivadas , Inflamación , Sinovitis/patología , Fibroblastos/patología , Macrófagos/patología , GlucólisisRESUMEN
The canonical Wnt/ß-catenin signaling is activated during development, tumorigenesis, and in adult homeostasis, yet its role in maintenance of hematopoietic stem/progenitor cells is not firmly established. Here, we demonstrate that conditional expression of an active form of ß-catenin in vivo induces a marked increase in the frequency of apoptosis in hematopoietic stem/progenitor cells (HSCs/HPCs). Activation of Wnt/ß-catenin signaling in HPCs in vitro elevates the activity of caspases 3 and 9 and leads to a loss of mitochondrial membrane potential (ΔΨ(m)), indicating that it induces the intrinsic mitochondrial apoptotic pathway. In vivo, expression of activated ß-catenin in HPCs is associated with down-regulation of Bcl2 and expression of Casp3. Bone marrow transplantation assays reveal that enhanced cell survival by a Bcl2 transgene re-establishes the reconstitution capacity of HSCs/HPCs that express activated ß-catenin. In addition, a Bcl2 transgene prevents exhaustion of these HSCs/HPCs in vivo. Our data suggest that activation of the Wnt/ß-catenin pathway contributes to the defective function of HPCs in part by deregulating their survival.
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Apoptosis/fisiología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Mitocondrias/fisiología , Vía de Señalización Wnt/fisiología , beta Catenina/metabolismo , Animales , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Trasplante de Médula Ósea , Supervivencia Celular/fisiología , Células Cultivadas , Potencial de la Membrana Mitocondrial/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , beta Catenina/genéticaRESUMEN
The remarkable biocapacity, injectability, and adaptability of colloidal gels have led to their widespread usage in tissue engineering as irregular defect implants. However, multifunctionalities including electroconductivity and antibacterial property are highly required for colloidal gels. In addition, the inherently weak mechanical property of physically crosslinked colloidal gels limits their application. Herein, we present Ag nanowires (Ag NWs)-reinforced colloidal gels composed of biocompatible gelatin nanoparticles and polydopamine-modified Ag NWs through the controlled electrostatic assembly, which are injectable and conductive. One-dimensional Ag NWs can significantly improve the mechanical and electrical properties of the colloidal gel while maintaining its inherent excellent injectability. Owing to the network of Ag NWs, the storage modulus and conductivity of the optimized Ag NW colloidal gel are 7.5 and 13 times higher, respectively, than those of the colloidal gel made up of polydopamine-modified Ag nanoparticles with equivalent Ag concentration. Furthermore, this Ag NW colloidal gel can adapt to sharp wounds on skin, which accelerates the healing of a MRSA-infected wound via electrical stimulation. This article is protected by copyright. All rights reserved.
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Leptin exerts its biological actions by activating the long-form leptin receptor (LepRb). LepRb signaling impairment and leptin resistance are believed to cause obesity. The transcription factor Slug - also known as Snai2 - recruits epigenetic modifiers and regulates gene expression by an epigenetic mechanism; however, its epigenetic action has not been explored in leptin resistance. Here, we uncover a proobesity function of neuronal Slug. Hypothalamic Slug was upregulated in obese mice. LepRb+ cell-specific Slug-knockout (SlugΔLepRb) mice were resistant to diet-induced obesity, type 2 diabetes, and liver steatosis and experienced decreased food intake and increased fat thermogenesis. Leptin stimulated hypothalamic Stat3 phosphorylation and weight loss to a markedly higher level in SlugΔLepRb than in Slugfl/fl mice, even before their body weight divergence. Conversely, hypothalamic LepRb+ neuron-specific overexpression of Slug, mediated by AAV-hSyn-DIO-Slug transduction, induced leptin resistance, obesity, and metabolic disorders in mice on a chow diet. At the genomic level, Slug bound to and repressed the LepRb promoter, thereby inhibiting LepRb transcription. Consistently, Slug deficiency decreased methylation of LepRb promoter H3K27, a repressive epigenetic mark, and increased LepRb mRNA levels in the hypothalamus. Collectively, these results unravel what we believe to be a previously unrecognized hypothalamic neuronal Slug/epigenetic reprogramming/leptin resistance axis that promotes energy imbalance, obesity, and metabolic disease.
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Enfermedad del Hígado Graso no Alcohólico , Obesidad , Receptores de Leptina , Factores de Transcripción de la Familia Snail , Animales , Ratones , Diabetes Mellitus Tipo 2/metabolismo , Hipotálamo/metabolismo , Leptina/genética , Leptina/metabolismo , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/genética , Obesidad/metabolismo , Receptores de Leptina/genética , Receptores de Leptina/metabolismo , Factores de Transcripción de la Familia Snail/genética , Factores de Transcripción de la Familia Snail/metabolismo , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismoRESUMEN
Aberrant N6-methyladenosine (m6A) modification of mRNAs contributes significantly to the epigenetic tumorigenesis, however, its precise role and the key targets in osteosarcoma (OS) are not defined. Here we reported that selective METTL3 (methyltransferase like 3) elevation and the consequential increase of m6A modification causally affect OS progression. The fast-growing OS cells displayed preferential upregulation of METTL3 and increased m6A modification. Conversely, m6A inhibition by 3-deazaadenosine, siRNA-mediated METTL3 knockdown or a METTL3-selective inhibitor STM2457 effectively inhibits OS cell growth and induced OS cell apoptosis. Further investigation revealed that an oncogenic protein ZBTB7C was likely a critical m6A target that mediated the oncogenic effects. ZBTB7C mRNA contains a typical m6A motif of high confidence and its mRNA and protein were enriched with increased m6A modification in OS samples/cells. In an OS xenograft model, STM2457 or siRNA-mediated METTL3 knockdown effectively lowed ZBTB7C abundance. More importantly, the anti-OS effects of STM2457 were significantly reduced when ZBTB7C was overexpressed by lentivirus. Together, our results demonstrate that the METTL3 aberration and the resultant ZBTB7C m6A modification form an important epigenetic regulatory loop that promotes OS progression, and targeting the METTL3/ZBTB7C axis may provide novel insights into the potential strategies for OS therapy.
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Metiltransferasas , Osteosarcoma , Humanos , Péptidos y Proteínas de Señalización Intracelular , Metiltransferasas/genética , Metiltransferasas/metabolismo , Osteosarcoma/genética , ARN Mensajero/genética , ARN Interferente PequeñoRESUMEN
BACKGROUND: Dysregulation of long non-coding RNAs (lncRNAs) is an important component of tumorigenesis. Aberrant expression of lncRNA taurine upregulated gene 1 (lncTUG1) has been reported in various tumors; however, its precise role and key targets critically involved in osteosarcoma (OS) progression remains unclear. METHODS: The expression profiles of lncRNAs and its regulated miRNAs related to OS progression were assessed by bioinformatics analysis and confirmed by qRT-PCR of OS cells. The miRNA targets were identified by transcriptome sequencing and verified by luciferase reporter and RNA pull-down assays. Several in vivo and in vitro approaches, including CCK8 assay, western blot, qRT-PCR, lentiviral transduction and OS cell xenograft mouse model were established to validate the effects of lncTUG1 regulation of miRNA and the downstream target genes on OS cell growth, apoptosis and progression. RESULTS: We found that lncTUG1 and miR-26a-5p were inversely up or down-regulated in OS cells, and siRNA-mediated lncTUG1 knockdown reversed the miR-26a-5p down-regulation and suppressed proliferation and enhanced apoptosis of OS cells. Further, we identified that an oncoprotein ZBTB7C was also upregulated in OS cells that were subjected to lncTUG1/miR-26a-5p regulation. More importantly, ZBTB7C knockdown reduced the ZBTB7C upregulation and ZBTB7C overexpression diminished the anti-OS effects of lncTUG1 knockdown in the OS xenograft model. CONCLUSIONS: Our data suggest that lncTUG1 acts as a miR-26a-5p sponge and promotes OS progression via up-regulating ZBTB7C, and targeting lncTUG1 might be an effective strategy to treat OS.
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Both extrinsic and intrinsic mechanisms tightly govern hematopoietic stem cell (HSC) decisions of self-renewal and differentiation. However, transcription factors that can selectively regulate HSC self-renewal division after stress remain to be identified. Slug is an evolutionarily conserved zinc-finger transcription factor that is highly expressed in primitive hematopoietic cells and is critical for the radioprotection of these key cells. We studied the effect of Slug in the regulation of HSCs in Slug-deficient mice under normal and stress conditions using serial functional assays. Here, we show that Slug deficiency does not disturb hematopoiesis or alter HSC homeostasis and differentiation in bone marrow but increases the numbers of primitive hematopoietic cells in the extramedullary spleen site. Deletion of Slug enhances HSC repopulating potential but not its homing and differentiation ability. Furthermore, Slug deficiency increases HSC proliferation and repopulating potential in vivo after myelosuppression and accelerates HSC expansion during in vitro culture. Therefore, we propose that Slug is essential for controlling the transition of HSCs from relative quiescence under steady-state condition to rapid proliferation under stress conditions. Our data suggest that inhibition of Slug in HSCs may present a novel strategy for accelerating hematopoietic recovery, thus providing therapeutic benefits for patients after clinical myelosuppressive treatment.
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Hematopoyesis/fisiología , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Regeneración/fisiología , Factores de Transcripción/deficiencia , Animales , Trasplante de Médula Ósea , Diferenciación Celular , Proliferación Celular , Ensayo de Unidades Formadoras de Colonias , Fluorouracilo/toxicidad , Hematopoyesis Extramedular/fisiología , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factores de Transcripción de la Familia Snail , Estrés Fisiológico , Factores de Transcripción/genética , Factores de Transcripción/fisiologíaRESUMEN
Bone marrow injury is a major adverse side effect of radiation and chemotherapy. Attempts to limit such damage are warranted, but their success requires a better understanding of how radiation and anticancer drugs harm the bone marrow. Here, we report one pivotal role of the BH3-only protein Puma in the radiosensitivity of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs). Puma deficiency in mice confers resistance to high-dose radiation in a hematopoietic cell-autonomous manner. Unexpectedly, loss of one Puma allele is sufficient to confer mice radioresistance. Interestingly, null mutation in Puma protects both primitive and differentiated hematopoietic cells from damage caused by low-dose radiation but selectively protects HSCs and HPCs against high-dose radiation, thereby accelerating hematopoietic regeneration. Consistent with these findings, Puma is required for radiation-induced apoptosis in HSCs and HPCs, and Puma is selectively induced by irradiation in primitive hematopoietic cells, and this induction is impaired in Puma-heterozygous cells. Together, our data indicate that selective targeting of p53 downstream apoptotic targets may represent a novel strategy to protecting HSCs and HPCs in patients undergoing intensive cancer radiotherapy and chemotherapy.
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Proteínas Reguladoras de la Apoptosis , Apoptosis/efectos de la radiación , Rayos gamma/efectos adversos , Células Madre Hematopoyéticas/metabolismo , Tolerancia a Radiación/fisiología , Proteínas Supresoras de Tumor , Animales , Apoptosis/fisiología , Relación Dosis-Respuesta en la Radiación , Ratones , Ratones Noqueados , Mutación , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
We show here that a zinc finger transcriptional repressor, Slug, which is aberrantly upregulated by the E2A-HLF oncoprotein in pro-B cell acute leukemia, functions as an antiapoptotic factor in normal hematopoietic progenitor cells. Slug(-/-) mice were much more radiosensitive than wild-type mice, dying earlier and showing accentuated decreases in peripheral blood cell counts, as well as abundant microhemorrhages and widely disseminated bacterial microabscesses throughout the body. Slug expression was detected in diverse subsets of hematopoietic progenitors, but not in more differentiated B and T lymphoid cells, and there was a significant increase in apoptotic (TUNEL-positive) bone marrow progenitor cells in irradiated Slug(-/-) mice compared to wild-type controls. These results implicate Slug in a novel survival pathway that protects hematopoietic progenitors from apoptosis after DNA damage.
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Apoptosis/efectos de la radiación , Células Madre Hematopoyéticas/citología , Factores de Transcripción/fisiología , Dedos de Zinc/fisiología , Animales , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico , Recuento de Células Sanguíneas , Plaquetas/metabolismo , Médula Ósea/metabolismo , Linaje de la Célula , Transformación Celular Neoplásica , Citoprotección , Daño del ADN , Cartilla de ADN/química , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Rayos gamma , Regulación Neoplásica de la Expresión Génica , Hematopoyesis/fisiología , Hematopoyesis/efectos de la radiación , Células Madre Hematopoyéticas/efectos de la radiación , Hemoglobinas/metabolismo , Homocigoto , Etiquetado Corte-Fin in Situ , Leucemia de Células B/genética , Leucemia de Células B/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Reacción en Cadena de la Polimerasa , Recombinación Genética , Factores de Transcripción de la Familia Snail , Bazo/metabolismo , Tasa de Supervivencia , Timo/efectos de la radiación , Proteína p53 Supresora de Tumor/metabolismo , Irradiación Corporal TotalRESUMEN
BACKGROUND: SLUG is a zinc-finger transcription factor of the Snail/Slug zinc-finger family that plays a role in migration and invasion of tumor cells. Mechanisms by which SLUG promotes migration and invasion in prostate cancers remain elusive. METHODS: Expression level of CXCR4 and CXCL12 was examined by Western blot, RT-PCR, and qPCR analyses. Forced expression of SLUG was mediated by retroviruses, and SLUG and CXCL12 was downregulated by shRNAs-expressing lentiviruses. Migration and invasion of prostate cancer were measured by scratch-wound assay and invasion assay, respectively. RESEARCH: We demonstrated that forced expression of SLUG elevated CXCR4 and CXCL12 expression in human prostate cancer cell lines PC3, DU145, 22RV1, and LNCaP; conversely, reduced expression of SLUG by shRNA downregulated CXCR4 and CXCL12 expression at RNA and protein levels in prostate cancer cells. Furthermore, ectopic expression of SLUG increased MMP9 expression and activity in PC3, 22RV1, and DU-145 cells, and SLUG knockdown by shRNA downregulated MMP9 expression. We showed that CXCL12 is required for SLUG-mediated MMP9 expression in prostate cancer cells. Moreover, we found that migration and invasion of prostate cancer cells was increased by ectopic expression of SLUG and decreased by SLUG knockdown. Notably, knockdown of CXCL12 by shRNA impaired SLUG-mediated migration and invasion in prostate cancer cells. Lastly, our data suggest that CXCL12 and SLUG regulate migration and invasion of prostate cancer cells independent of cell growth. CONCLUSION: We provide the first compelling evidence that upregulation of autocrine CXCL12 is a major mechanism underlying SLUG-mediated migration and invasion of prostate cancer cells. Our findings suggest that CXCL12 is a therapeutic target for prostate cancer metastasis.
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Movimiento Celular , Quimiocina CXCL12/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Receptores CXCR4/metabolismo , Factores de Transcripción/genética , Línea Celular Tumoral , Quimiocina CXCL12/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Invasividad Neoplásica , Neoplasias de la Próstata/metabolismo , Receptores CXCR4/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Transcripción de la Familia Snail , Factores de Transcripción/metabolismoRESUMEN
The proteins involved in breast cancer initiation and progression are still largely elusive. To gain insights into these processes, we conducted quantitative proteomic analyses with 21T series of breast cell lines, which include a normal, primary tumor and a metastatic tumor that were isolated from a single patient. Stable isotope labeling of amino acid in cell culture followed by LC-MS/MS analysis was performed and deregulated proteins were identified using statistical analysis. Gene ontology analysis revealed that proteins involved in metabolic processes were the most deregulated in both tumorigenesis and metastasis. Interaction network analysis indicated that ERBB2 signaling played a critical role in tumorigenesis. In addition to known markers such as ERBB2 and E-cadherin, novel markers, including BRP44L, MTHFD2 and TIMM17A, were found to be overexpressed in 21T breast cancer cells and verified in additional breast cell lines. mRNA expression analysis as well as immunohistochemistry analysis in breast cancer tissues indicated that expression level of TIMM17A was directly correlated with tumor progression, and survival analysis suggested that TIMM17A was a powerful prognosis factor in breast cancer. More interestingly, overexpression and siRNA knockdown experiments indicated an oncogenic activity of TIMM17A in breast cancer. Our study provides a list of potential novel markers for breast cancer tumorigenesis and metastasis using a unique cell model. Further studies on TIMM17A as well as other markers on the list may reveal mechanisms that result in more effective therapeutics for cancer treatment.