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1.
Cancer Cell ; 37(2): 200-215.e5, 2020 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-32049046

RESUMO

Deregulation of MYC plays an essential role in T cell acute lymphoblastic leukemia (T-ALL), yet the mechanisms underlying its deregulation remain elusive. Herein, we identify a molecular mechanism responsible for reciprocal activation between Aurora B kinase (AURKB) and MYC. AURKB directly phosphorylates MYC at serine 67, counteracting GSK3ß-directed threonine 58 phosphorylation and subsequent FBXW7-mediated proteasomal degradation. Stabilized MYC, in concert with T cell acute lymphoblastic leukemia 1 (TAL1), directly activates AURKB transcription, constituting a positive feedforward loop that reinforces MYC-regulated oncogenic programs. Therefore, inhibitors of AURKB induce prominent MYC degradation concomitant with robust leukemia cell death. These findings reveal an AURKB-MYC regulatory circuit that underlies T cell leukemogenesis, and provide a rationale for therapeutic targeting of oncogenic MYC via AURKB inhibition.

2.
Blood ; 135(1): 41-55, 2020 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-31697823

RESUMO

To study the mechanisms of relapse in acute lymphoblastic leukemia (ALL), we performed whole-genome sequencing of 103 diagnosis-relapse-germline trios and ultra-deep sequencing of 208 serial samples in 16 patients. Relapse-specific somatic alterations were enriched in 12 genes (NR3C1, NR3C2, TP53, NT5C2, FPGS, CREBBP, MSH2, MSH6, PMS2, WHSC1, PRPS1, and PRPS2) involved in drug response. Their prevalence was 17% in very early relapse (<9 months from diagnosis), 65% in early relapse (9-36 months), and 32% in late relapse (>36 months) groups. Convergent evolution, in which multiple subclones harbor mutations in the same drug resistance gene, was observed in 6 relapses and confirmed by single-cell sequencing in 1 case. Mathematical modeling and mutational signature analysis indicated that early relapse resistance acquisition was frequently a 2-step process in which a persistent clone survived initial therapy and later acquired bona fide resistance mutations during therapy. In contrast, very early relapses arose from preexisting resistant clone(s). Two novel relapse-specific mutational signatures, one of which was caused by thiopurine treatment based on in vitro drug exposure experiments, were identified in early and late relapses but were absent from 2540 pan-cancer diagnosis samples and 129 non-ALL relapses. The novel signatures were detected in 27% of relapsed ALLs and were responsible for 46% of acquired resistance mutations in NT5C2, PRPS1, NR3C1, and TP53. These results suggest that chemotherapy-induced drug resistance mutations facilitate a subset of pediatric ALL relapses.

3.
Cancer Lett ; 469: 151-161, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31669202

RESUMO

Blocking the migration of regulatory T cells (Tregs) to the tumor microenvironment is a promising strategy for tumor immunotherapy. Treg accumulation in the leukemic hematopoietic microenvironment (LHME) has adverse impacts on patient outcomes. The mechanism and effective methods of disrupting Treg accumulation in the LHME have not been well established. Here, we studied the distribution and characteristics of Tregs in the LHME, investigated the effects of Treg ablation on leukemia progression, explored the mechanisms leading to Treg accumulation, and studied whether blocking Treg migration to the LHME delayed leukemia progression in MLL-AF9-induced mouse acute myeloid leukemia (AML) models using wildtype (WT) and Foxp3DTR/GFP mice. Increased accumulation of more activated Tregs was detected in the LHME. Inducible Treg ablation prolonged the survival of AML mice by promoting the antileukemic effects of CD8+ T cells. Furthermore, both local expansion and migration accounted for Treg accumulation in the LHME. Moreover, blocking the CCL3-CCR1/CCR5 and CXCL12-CXCR4 axes inhibited Treg accumulation in the LHME and delayed leukemia progression. Our findings provide laboratory evidence for a potential leukemia immunotherapy by blocking the migration of Tregs.

4.
Stem Cell Res Ther ; 10(1): 407, 2019 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-31864409

RESUMO

BACKGROUND: Imbalance in bone formation and resorption is a crucial component of the pathological process leading to osteoporosis. Electromagnetic fields (EMFs) have been reported to be beneficial to osteogenesis, although the exact mechanism has not been fully clarified. Purinergic receptor P2X7 is expressed in osteoblasts and is reported to participate in the regulation of bone metabolism. OBJECTIVES: To elucidate the link between EMFs and P2X7 expression and investigate its potential as a novel therapeutic target in osteoporosis. METHOD: We investigated the effect of EMFs on P2X7 expression and downstream signaling in human bone marrow mesenchymal stem cells (h-MSCs). We also established an ovariectomized (OVX) osteoporosis rat model to evaluate the therapeutic efficacy of combining EMFs with P2X7 agonists. RESULTS: EMF treatment increased P2X7 expression in h-MSCs under conditions of osteogenic induction but not under regular culture conditions. P2X7 or PI3K/Akt inhibition partially inhibited the pro-osteogenic effect of EMF and lowered the EMF-stimulated activity of the Akt/GSK3ß/ß-catenin axis. No additive effect of this suppression was observed following simultaneous inhibition of P2X7 and PI3K/Akt. EMF treatment in the presence of a P2X7 agonist had a greater effect in increasing osteogenic marker expression than that of EMF treatment alone. In the OVX osteoporosis model, the therapeutic efficacy of combining EMFs with P2X7 agonists was superior to that of EMF treatment alone. CONCLUSIONS: EMF treatment increases P2X7 expression by h-MSCs during osteogenic differentiation, leading to activation of the Akt/GSK3ß/ß-catenin axis, which promotes the osteogenesis. Our findings also indicate that combined EMF and P2X7 agonist treatment may be an effective novel strategy for osteoporosis therapy.

5.
Zhongguo Dang Dai Er Ke Za Zhi ; 21(10): 1016-1021, 2019 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-31642437

RESUMO

OBJECTIVE: To study the clinical features and gene mutation spectrum of children with sideroblastic anemia (SA) and the clinical value of targeted next-generation sequencing in the molecular diagnosis of children with SA. METHODS: Clinical data were collected from 36 children with SA. Targeted next-generation sequencing was used to detect mutations in SA-related pathogenic genes and genes associated with heme synthesis and mitochondrial iron metabolism. The association between genotype and clinical phenotype was analyzed. RESULTS: Of the 36 patients, 32 had congenital sideroblastic anemia (CSA) and 4 had myelodysplastic syndrome with ring sideroblasts (MDS-RS). Mutations in CSA-related genes were detected in 19 children (19/36, 53%), among whom 9 (47%) had ALAS2 mutation, 4 (21%) had SLC25A38 mutation, and 6 (32%) had mitochondrial fragment deletion. No pathogenic gene mutation was detected in 4 children with MDS-RS. Among the 19 mutations, 89% (17/19) were known mutations and 11% (2/19) were novel mutations. The novel mutation of the ALAS2 gene c.1153A>T(p.I385F) was rated as "possibly pathogenic" and the novel mutation of the SLC25A38 gene c.175C>T(p.Q59X) was rated as "pathogenic". CONCLUSIONS: ALAS2 and SLC25A38 gene mutations are commonly seen in children with CSA, but mitochondrial gene fragment deletion also accounts for a relatively high proportion. For children with hypoplastic anemia occurring in infancy, mitochondrial disease should be considered.


Assuntos
Anemia Sideroblástica , Síndromes Mielodisplásicas , 5-Aminolevulinato Sintetase , Anemia Sideroblástica/genética , Criança , Doenças Genéticas Ligadas ao Cromossomo X , Humanos , Proteínas de Transporte da Membrana Mitocondrial , Mutação , Fenótipo
7.
Int J Cancer ; 145(4): 1068-1082, 2019 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-30761524

RESUMO

The P2X7 receptor, an ATP-gated ion channel, is critical for cancer cell growth, invasiveness, and angiogenesis. Previous studies indicate that P2X7 regulates osteoblast proliferation and osteodeposition and that high P2X7 expression has a pro-growth effect in osteosarcoma. However, how it functions in osteosarcoma cell growth and metastasis is not clear. Thus, we elucidated molecular mechanisms of P2X7-dependent positive regulation of osteosarcoma cell proliferation, invasion, migration, epithelial to mesenchymal transition (EMT), and angiogenesis using in vitro and in vivo models. We confirm that P2X7 is highly-expressed in human osteosarcoma tumor tissues and HOS/MNNG, MG63, U2OS, SW1353 and SAOS-2 cell lines. P2X7 receptor stimulation enhanced HOS/MNNG and SAOS-2 cell proliferation, migration and invasion; but knockdown of P2X7 expression or receptor inhibition had opposite effects. P2X7 positively regulated glycogen content, epithelial to mesenchymal transition and stemness of HOS/MNNG cells. P2X7 activation promoted PI3K/Akt/GSK3ß/ß-catenin and mTOR/HIF1α/VEGF signaling, thereby mediating pro-tumor effects of osteosarcoma cells. Consistent with data from in vitro experiments, systemic administration of P2X7 agonist induced tumor growth, metastasis and tumor-associated bone destruction in osteosarcoma-bearing nude mice, whereas a P2X7 antagonist reversed these effects. Thus, the P2X7 receptor participates in regulation of osteosarcoma growth and metastasis and we offer evidence that P2X7 may be a promising therapeutic target for treating osteosarcoma.


Assuntos
Neoplasias Ósseas/genética , Proliferação de Células/genética , Metástase Neoplásica/genética , Osteossarcoma/genética , Receptores Purinérgicos P2X7/genética , Transdução de Sinais/genética , Adolescente , Adulto , Animais , Apoptose/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Criança , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Glicogênio Sintase Quinase 3 beta/genética , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Osteossarcoma/patologia , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Serina-Treonina Quinases TOR/genética , Fator A de Crescimento do Endotélio Vascular/genética , Adulto Jovem , beta Catenina/genética
9.
Stem Cell Res Ther ; 9(1): 143, 2018 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-29784011

RESUMO

BACKGROUND: The ability of mesenchymal stem cells (MSCs) to migrate to the desired tissues or lesions is crucial for stem cell-based regenerative medicine and tissue engineering. Optimal therapeutics for promoting MSC migration are expected to become an effective means for tissue regeneration. Electromagnetic fields (EMF), as a noninvasive therapy, can cause a lot of biological changes in MSCs. However, whether EMF can promote MSC migration has not yet been reported. METHODS: We evaluated the effects of EMF on cell migration in human bone marrow-derived MSCs. With the use of Helmholtz coils and an EMF stimulator, 7.5, 15, 30, 50, and 70 Hz/1 mT EMF was generated. Additionally, we employed the L-type calcium channel blocker verapamil and the focal adhesion kinase (FAK) inhibitor PF-573228 to investigate the role of intracellular calcium content, cell adhesion proteins, and the Rho GTPase protein family (RhoA, Rac1, and Cdc42) in EMF-mediated MSC migration. Cell adhesion proteins (FAK, talin, and vinculin) were detected by Western blot analysis. The Rho GTPase protein family activities were assessed by G-LISA, and F-actin levels, which reflect actin cytoskeletal organization, were detected using immunofluorescence. RESULTS: All the 7.5, 15, 30, 50, and 70 Hz/1 mT EMF promoted MSC migration. EMF increased MSC migration in an intracellular calcium-dependent manner. Notably, EMF-enhanced migration was mediated by FAK activation, which was critical for the formation of focal contacts, as evidenced by increased talin and vinculin expression. Moreover, RhoA, Rac1, and Cdc42 were activated by FAK to increase cytoskeletal organization, thus promoting cell contraction. CONCLUSIONS: EMF promoted MSC migration by increasing intracellular calcium and activating the FAK/Rho GTPase signaling pathways. This study provides insights into the mechanisms of MSC migration and will enable the rational design of targeted therapies to improve MSC engraftment.


Assuntos
Cálcio/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Células-Tronco Mesenquimais/fisiologia , Proteínas rho de Ligação ao GTP/metabolismo , Diferenciação Celular , Movimento Celular , Campos Eletromagnéticos , Humanos , Transdução de Sinais
10.
Blood ; 131(20): 2256-2261, 2018 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-29434033

RESUMO

Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) comprises ∼10% to 15% of childhood ALL cases, many of which respond exquisitely to tyrosine kinase inhibitors (TKIs), for example, imatinib in PDGFRB-rearranged ALL. However, some cases developed drug resistance to TKIs and the mechanisms are poorly understood. In this study, we identified a novel PDGFRB fusion gene, namely AGGF1-PDGFRB, and functionally characterized its oncogenic potential in vitro. Further genomic profiling of longitudinally collected samples during treatment revealed the emergence of a mutation, PDGFRBC843G , which directly conferred resistance to all generations of ABL TKIs, including imatinib, dasatinib, nilotinib, and ponatinib. PDGFRB-mutant leukemia cells are highly sensitive to multitarget kinase inhibitor CHZ868, suggesting potential therapeutic options for some patients resistant to ABL TKIs. In summary, we describe a complex clonal evolution pattern in Ph-like ALL and identified a novel PDGFRB point mutation that drives leukemia relapse after ABL TKI treatment.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Mutação , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Inibidores de Proteínas Quinases/farmacologia , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Proteínas Angiogênicas/genética , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Pré-Escolar , Humanos , Masculino , Proteínas de Fusão Oncogênica , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Inibidores de Proteínas Quinases/uso terapêutico , Recidiva , Resultado do Tratamento , Sequenciamento Completo do Genoma
11.
Cancer Res ; 78(7): 1632-1642, 2018 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-29330145

RESUMO

Aberrant DNA methylation patterns in leukemia might be exploited for therapeutic targeting. In this study, we employed a genetically deficient mouse model to explore the role of the methylated DNA binding protein MBD2 in normal and malignant hematopoiesis. MBD2 ablation led to diminished lymphocytes. Functional defects of the lymphoid compartment were also observed after in vivo reconstitution of MBD2-deficient hematopoietic stem cells (HSC). In an established model of Notch1-driven T-cell acute lymphoblastic leukemia (T-ALL), MBD2 ablation impeded malignant progression and maintenance by attenuating the Wnt signaling pathway. In clinical specimens of human T-ALL, Wnt signaling pathway signatures were significantly enhanced and positively correlated with the expression and function of MBD2. Furthermore, a number of typical Wnt signaling inhibitory genes were abnormally hypermethylated in primary human T-ALL. Abnormal activation of Wnt signaling in T-ALL was switched off by MBD2 deletion, partially by reactivating epigenetically silenced Wnt signaling inhibitors. Taken together, our results define essential roles for MBD2 in lymphopoiesis and T-ALL and suggest MBD2 as a candidate therapeutic target in T-ALL.Significance: This study highlights a methylated DNA binding protein as a candidate therapeutic target to improve the treatment of T-cell acute lymphoblastic leukemias, as a new starting point for developing epigenetic therapy in this and other lymphoid malignancies. Cancer Res; 78(7); 1632-42. ©2018 AACR.


Assuntos
Proteínas de Ligação a DNA/genética , Linfopoese/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Via de Sinalização Wnt/genética , Animais , Linhagem Celular Tumoral , Metilação de DNA/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Células Jurkat , Contagem de Linfócitos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor Notch1/metabolismo
12.
Leuk Lymphoma ; 59(10): 2423-2430, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29043883

RESUMO

The gene, structural maintenance of chromosomes 4 (SMC4) plays important role in chromosomes condensing and mitotic sister chromatid segregation, which has been revealed in regulating multiple cancer development and carcinogenesis. However, the role of SMC4 in acute myeloid leukemia (AML) propagation and its function in regulation of leukemia stem cells (LSCs) is not yet clear. Using an MLL-AF9 induced AML mouse model, we demonstrated that down modulating of SMC4 expression could prolong the survival time of AML mice. Furthermore, we found that knockdown SMC4 expression decreased the proportion of LSCs and affected its leukemia-initiating capacity. Cell cycle assay demonstrated that more LSCs were arrested in G0 phase by SMC4 knockdown. This activity was accompanied by increased expression of the Cdkn1a (P21) and Cdkn1b (P27) as well as decreased expression of CDK4. Therefore, our study revealed the critical role of SMC4 during AML progression and provided new insights into the mechanism of LSC maintenance.


Assuntos
Adenosina Trifosfatases/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Leucemia Experimental/patologia , Leucemia Mieloide Aguda/patologia , Células-Tronco Neoplásicas/patologia , Adenosina Trifosfatases/genética , Adolescente , Animais , Medula Óssea/patologia , Transplante de Medula Óssea , Proteínas Cromossômicas não Histona/genética , Progressão da Doença , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Leucemia Experimental/genética , Leucemia Mieloide Aguda/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Fusão Oncogênica/genética , RNA Interferente Pequeno/metabolismo
13.
Leuk Res ; 65: 20-24, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29253671

RESUMO

Genomic alterations underlying chemotherapy resistance remains poorly characterized in pediatric acute myeloid leukemia (AML). In this study, we used whole exome sequencing to identify gene mutations associated with chemo-resistance in 44 pediatric AML patients. We identified previously unreported mutations involving epigenetic regulators such as KDM5C, SRIT6, CHD4, and PRPF6 in pediatric AML patients. Despite low prevalence in general pediatric AML, mutations involving epigenetic regulators including splicing factors, were collectively enriched as a group in primary chemo-resistance AML patients. In addition, clonal evolution analysis of secondary chemo-resistance AML patients reveals dominant clone at diagnosis could survive several course of intensified chemotherapy. And gain of new mutations in genes such as MVP, TCF3, SS18, and BCL10, may contribute to chemo-resistance at relapse. These results provide novel insights into the genetic basis of treatment failure in pediatric AML.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Resistencia a Medicamentos Antineoplásicos/genética , Epigênese Genética , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Mutação , Sequenciamento Completo do Exoma , Criança , China , Humanos , Leucemia Mieloide Aguda/etnologia
14.
Sheng Wu Gong Cheng Xue Bao ; 33(2): 284-293, 2017 Feb 25.
Artigo em Chinês | MEDLINE | ID: mdl-28956384

RESUMO

To study the biological function of DNAH2 (Homo sapiens dynein, axonemal, heavy chain 2) gene, we constructed human stable U2OS cell line of DNAH2 gene knockout through CRISPR/Cas9n double nick system. The A, B sgRNAs (Single guide RNA) and complementary strands were designed and synthesized. The double-stranded structures were formed during annealing, and connected with BbsⅠ cohesive ends-containing pX462 linear vector to construct the recombinant eukaryotic expression plasmids, including pX462-DNAH2-A and pX462-DNAH2-B. After the co-transfection of the two plasmids into U2OS cells, the addition of puromycin and limiting dilution method were used to obtain positive monoclonal cell line. Western blotting assay was then performed to detect the expression of DNAH2 protein, and PCR-sequencing technology was finally utilized to analyze the mutation feature. The results showed that A, B sgRNAs duplex was successfully inserted into pX462 vector, and DNAH2 protein was not expressed and DNAH2 gene suffered from the frame-shift mutation in U2OS-DNAH2-KO monoclonal cell line. These demonstrated that DNAH2 knockout U2OS stable cell line was successfully constructed through CRISPR/Cas9n double nick system, which providing a useful tool for the study of DNAH2 gene.


Assuntos
Dineínas do Axonema/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Técnicas de Inativação de Genes , Linhagem Celular Tumoral , Vetores Genéticos , Humanos , Plasmídeos , RNA Guia , Transfecção
15.
Sci Rep ; 7(1): 4943, 2017 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-28694518

RESUMO

3-Phosphoinositide-dependent protein kinase 1 (PDK1) is a pivotal regulator in the phosphoinositide 3-kinase (PI3K)-Akt signaling pathway that have been shown to play key roles in the functional development of B and T cells via activation of AGC protein kinases during hematopoiesis. However, the role of PDK1 in HSCs has not been fully defined. Here we specifically deleted the PDK1 gene in the hematopoietic system and found that PDK1-deficient HSCs exhibited impaired function and defective lineage commitment abilities. Lack of PDK1 caused HSCs to be less quiescent and to produce a higher number of phenotypic HSCs and fewer progenitors. PDK1-deficient HSCs were also unable to reconstitute the hematopoietic system. Notably, HSC function was more dependent on PDK1 than on mTORC2, which indicates that PDK1 plays a dominant role in the Akt-mediated regulation of HSC function. PDK1-deficient HSCs also exhibited reduced ROS levels, and treatment of PDK1-deficient HSCs with L-butathioninesulfoximine in vitro elevated the low ROS level and promoted colony formation. Therefore, PDK1 appears to contribute to HSC function partially via regulating ROS levels.


Assuntos
Células-Tronco Hematopoéticas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Apoptose , Ciclo Celular/genética , Doenças Desmielinizantes/genética , Doenças Desmielinizantes/imunologia , Doenças Desmielinizantes/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Técnicas de Silenciamento de Genes , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/ultraestrutura , Imuno-Histoquímica , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Camundongos , Microglia/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Fenótipo , Proteínas Serina-Treonina Quinases/genética , Espécies Reativas de Oxigênio/metabolismo
16.
Genomics Proteomics Bioinformatics ; 15(1): 37-48, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28185911

RESUMO

Primary myelofibrosis (PMF) is a chronic myeloproliferative disorder in human bone marrow. Over 50% of patients with myelofibrosis have mutations in JAK2, MPL, or CALR. However, these mutations are rarely detected in children, suggesting a difference in the pathogenesis of childhood PMF. In this study, we investigated the response to drug treatment of a monozygotic twin pair with typical childhood PMF. The twin exhibited different clinical outcomes despite following the same treatment regimen. The transcriptomic profiles of patient samples after drug treatment (E2 and Y2) were significantly different between the twin pair, which is consistent with the observation that the drug treatment was effective only in the younger brother, despite the twin being genetically identical. Bioinformatics analysis of the drug-responsive genes showed that the JAK-STAT pathway was activated in the cured younger brother, which is opposite to the pathway inhibition observed in adult PMF cases following treatment. Moreover, apoptosis and cell cycle processes were both significantly influenced by drug treatment in the sample of younger brother (Y2), implying their potential association with the pathogenesis of childhood PMF. Gene mutations in JAK2, MPL, or CALR were not observed; however, mutations in genes including SRSF2 and SF3B1 occurred in this twin pair with childhood PMF. Gene fusion events were extensively screened in the twin pair samples and the occurrence of IGLV2-14-IGLL5 gene fusion was confirmed. The current study reported at transcriptomic level the different responses of monozygotic twin brothers with childhood PMF to the same androgen/prednisone treatment regimen providing new insights into the potential pathogenesis of childhood PMF for further research and clinical applications.


Assuntos
Mielofibrose Primária/patologia , Androgênios/uso terapêutico , Medula Óssea/patologia , Pré-Escolar , Cromossomos Humanos Par 19 , Análise Citogenética , Perfilação da Expressão Gênica , Fusão Gênica , Humanos , Cadeias lambda de Imunoglobulina/genética , Masculino , Mutação de Sentido Incorreto , Fosfoproteínas/genética , Prednisona/uso terapêutico , Mielofibrose Primária/tratamento farmacológico , Mielofibrose Primária/genética , RNA/química , RNA/isolamento & purificação , RNA/metabolismo , Fatores de Processamento de RNA/genética , Análise de Sequência de RNA , Fatores de Processamento de Serina-Arginina/genética , Gêmeos Monozigóticos
17.
Nucleic Acids Res ; 45(2): 657-671, 2017 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-28123038

RESUMO

The first intronic mutations in the intron 1 GATA site (int-1-GATA) of 5-aminolevulinate synthase 2 (ALAS2) have been identified in X-linked sideroblastic anemia (XLSA) pedigrees, strongly suggesting it could be causal mutations of XLSA. However, the function of this int-1-GATA site during in vivo development remains largely unknown. Here, we generated mice lacking a 13 bp fragment, including this int-1-GATA site (T AGATAA: AGCCCC) and found that hemizygous deletion led to an embryonic lethal phenotype due to severe anemia resulting from a lack of ALAS2 expression, indicating that this non-coding sequence is indispensable for ALAS2 expression in vivo Further analyses revealed that this int-1-GATA site anchored the GATA site in intron 8 (int-8-GATA) and the proximal promoter, forming a long-range loop to enhance ALAS2 expression by an enhancer complex including GATA1, TAL1, LMO2, LDB1 and Pol II at least, in erythroid cells. However, compared with the int-8-GATA site, the int-1-GATA site is more essential for regulating ALAS2 expression through CRISPR/Cas9-mediated site-specific deletion. Therefore, the int-1-GATA site could serve as a valuable site for diagnosing XLSA in cases with unknown mutations.


Assuntos
5-Aminolevulinato Sintetase/genética , Sítios de Ligação , Diferenciação Celular , Células Eritroides/citologia , Células Eritroides/metabolismo , Fator de Transcrição GATA1/metabolismo , Íntrons , Anemia Sideroblástica/genética , Animais , Sequência de Bases , Sistemas CRISPR-Cas , Modelos Animais de Doenças , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica , Genes Letais , Doenças Genéticas Ligadas ao Cromossomo X/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Hemizigoto , Humanos , Células K562 , Masculino , Mutação , Linhagem , Regiões Promotoras Genéticas , Deleção de Sequência
18.
Bioelectromagnetics ; 38(2): 137-150, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27973686

RESUMO

Optimal therapeutics for hyperthyroidism-induced osteoporosis are still lacking. As a noninvasive treatment, electromagnetic fields (EMF) have been proven to be effective for treating osteoporosis in non-hyperthyroidism conditions. We herein systematically evaluated the reduced effects of EMF on osteoporosis in a hyperthyroidism rat model. With the use of Helmholtz coils and an EMF stimulator, 15 Hz/1 mT EMF was generated. Forty-eight 5-month-old male Sprague-Dawley rats were randomly divided into four different groups: control, levothyroxine treated (L-T4), EMF exposure + levothyroxine (EMF + L-T4), and EMF exposure without levothyroxine administration (EMF). All rats were treated with L-T4 (100 mg/day) except those in control and EMF groups. After 12 weeks, the results obtained from bone mineral density analyses and bone mechanical measurements showed significant differences between L-T4 and EMF + L-T4 groups. Micro CT and bone histomorphometric analyses indicated that trabecular bone mass and architecture in distal femur and proximal tibia were augmented and restored partially in EMF + L-T4 group. In addition, bone thyroid hormone receptors (THR) expression of hyperthyroidism rats was attenuated in EMF + L-T4 group, compared to control group, which was not observed in L-T4 group. According to these results, we concluded that 15 Hz/1 mT EMF significantly inhibited bone loss and micro architecture deterioration in hyperthyroidism rats, which might occur due to reduced THR expression caused by EMF exposure. Bioelectromagnetics. 38:137-150, 2017. © 2016 Wiley Periodicals, Inc.


Assuntos
Hipertireoidismo/complicações , Terapia de Campo Magnético , Osteoporose/etiologia , Osteoporose/terapia , Animais , Densidade Óssea/efeitos da radiação , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos da radiação , Hipertireoidismo/sangue , Hipertireoidismo/urina , Masculino , Osteoclastos/patologia , Osteoclastos/efeitos da radiação , Osteoporose/metabolismo , Osteoporose/fisiopatologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores dos Hormônios Tireóideos/genética , Tíbia/metabolismo , Tíbia/fisiopatologia , Tíbia/efeitos da radiação
19.
Int J Hematol ; 104(4): 430-9, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27329125

RESUMO

Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by a paucity of erythroid progenitors. We summarized the clinical and genetic features of 104 DBA patients in a single-center retrospective study in China. Data of DBA patients who received consultations at our center from 2003 to 2015 were analyzed retrospectively. Genes encoding 10 ribosomal proteins (RPs) and GATA1 were sequenced for mutation detection. Our cohort was composed of 65 males and 39 females. Congenital malformations were observed in 19 patients. Mutations of the RP genes were detected in 58.3 % patients. Twenty different mutations were first reported. Thirty-four patients received prednisone combined with CsA therapy, and improvement was observed in 20 cases. During follow-up for a median 39 months, 33.7 % of the patients achieved remission, 41.3 % of the patients were persistently transfusion independent, 21.7 % of the patients were transfusion dependent, and three patients died. The patient group with detected mutations had a younger age of disease onset, a higher malformation rate, and tended to have a lower remission rate and a higher transfusion-dependence rate. Prednisone in combination with cyclosporine A can be a second-line choice for DBA patients. Differences were detected between DBA patients with and without detectable mutations in the genes studied.


Assuntos
Anemia de Diamond-Blackfan/genética , Mutação , Idade de Início , Anemia de Diamond-Blackfan/terapia , Transfusão de Sangue , China , Quimioterapia Combinada , Feminino , Humanos , Masculino , Estudos Retrospectivos , Proteínas Ribossômicas/genética , Resultado do Tratamento
20.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 24(3): 643-8, 2016 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-27342484

RESUMO

OBJECTIVE: To investigate the effect of ADAR1 on the occurrence and development of mouse T cell acute lymphoblastic leukemia (T-ALL). METHODS: Lck-Cre; ADAR1lox/lox mice were generated through interbreeding. The lineage-cells of Lck-Cre; ADAR1lox/lox mice and the control were enriched respectively by the means of MACS, and the lin- cells were transfected with retrovirus carrying MSCV-ICN1-IRES-GFP fusion gene. Then the transfection efficiency was detected by the means of FACS, and the same number of GFP+ cells were transplanted into lethally irradiated recipient mice to observe the survival of mice in 2 recipient group after transplantation. RESULTS: T cell-specific knockout ADAR1 mice were generated, and Notch1-induced T-ALL mouse model was established successfully. The leukemia with T-ALL characteristics occured in the mice of control group, but did not in the ADAR1 kmockout mice after transplantation. CONCLUSIONS: ADAR1 plays a key role in the incidence and development of Notch1-induced T-ALL.


Assuntos
Adenosina Desaminase/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Receptor Notch1/genética , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Knockout , Linfócitos T
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