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1.
Mol Ther ; 28(3): 901-913, 2020 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-31991109

RESUMO

Esophageal squamous cell carcinoma (ESCC) is a predominant cancer type in developing countries such as China, where ESCC accounts for approximately 90% of esophageal malignancies. Lacking effective and targeted therapy contributes to the poor 5-year survival rate. Recent studies showed that about 30% of ESCC cases have high levels of SOX2. Herein, we aim to target this transcription factor with aptamer. We established a peptide aptamer library and then performed an unbiased screening to identify several peptide aptamers including P42 that can bind and inhibit SOX2 downstream target genes. We further found that P42 overexpression or incubation with a synthetic peptide 42 inhibited the proliferation, migration, and invasion of ESCC cells. Moreover, peptide 42 treatment inhibited the growth and metastasis of ESCC xenografts in mouse and zebrafish. Further analysis revealed that P42 overexpression led to alternations in the levels of proteins that are important for the proliferation and migration of ESCC cells. Taken together, our study identified the peptide 42 as a key inhibitor of SOX2 function, reducing the proliferation and migration of ESCC cells in vitro and in vivo, and thereby offering a potential therapy against ESCC.

3.
J Transl Med ; 17(1): 204, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31215436

RESUMO

BACKGROUND: Neutrophil gelatinase-associated lipocalin (NGAL) is a promising biomarker of early diagnosis and prediction for acute kidney injury (AKI). However, the current program for NGAL detection is not extensively applied in clinics due to the high expense of antibodies. Nucleic acid aptamers are single-strand DNAs or RNAs which could bind to targets with high specificity and affinity, and they have been widely used in the diagnosis and therapy for multiple diseases. It is valuable for us to develop a new method for NGAL detection using aptamers instead of antibodies to achieve increased efficiency and decreased cost. METHODS: Nucleic acid aptamers against NGAL were obtained after SELEX process using magnetic beads, and an enzyme-linked aptamer analysis (ELAA), which can be widely used in clinical diagnosis at low cost, were successfully established. The feasibility of ELAA was further validated with urine samples harvested from 43 AKI patients and 30 healthy people. RESULTS: Three candidate aptamers, including NA36, NA42 and NA53, were obtained after 8 rounds of SELEX process with magnetic beads and verified by quantitative polymerase chain reaction (qPCR), and the Kd value of each aptamer was 43.59, 66.55 and 32.52 nM, respectively. Moreover, the linear relationship was consistent at the range of 125-4000 ng/mL, and the detection limit of ELAA assay was 30.45 ng/mL. We also found that NGAL could be exclusively detected with NA53, and no cross-reaction between NA53 and human albumin or globulin occurred, the coefficient of variation (CV) between inner-plate and inter-plate was less than 15%, and the recovery rate was between 80 and 110%. Moreover, the sensitivity and specificity of ELAA assay in this study are 100% and 90%, respectively. Consistently, these results could also diagnose whether the occurrence of AKI in lots of patients, which has been demonstrated with the ELAA method we established after using NA53. CONCLUSIONS: Taken together, NA53, the best candidate aptamer targeting NGAL protein, can be applied in clinical testing.

4.
Cancer Lett ; 458: 21-28, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31125642

RESUMO

Esophageal cancer (EC) has been a leading cause of cancer death worldwide in part due to late detection and lack of precision treatment. EC includes two major malignancies, esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC). Recent studies reveal that ESCC and EAC have distinct cell of origin and contain cancer stem cells (also known as tumor initiating cells) expressing different cell surface markers. These biomarkers have potentially important values for both early detection and finding effective therapy. In this review we summarize the updated findings for cell of origin and provide an overview of cancer cell biomarkers that have been tested for ESCC and EAC. In addition, we also discuss recent progress in the study of molecular mechanisms leading to these malignancies.

5.
PLoS One ; 14(2): e0212041, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30779754

RESUMO

This study aimed to screen DNA aptamers against the signal molecule C4-HSL of the rhl system for the inhibition of biofilm formation of Pseudomonas aeruginosa using an improved systematic evolution of ligand by exponential enrichment (SELEX) method based on a structure-switching fluorescent activating bead. The aptamers against the C4-HSL with a high affinity and specifity were successfully obtained and evaluated in real-time by this method. Results of biofilm inhibition experiments in vitro showed that the biofilm formation of P. aeruginosa was efficiently reduced to about 1/3 by the aptamers compared with that of the groups without the aptamers. Independent secondary structure simulation and computer-aided tertiary structure prediction (3dRNA) showed that the aptamers contained a highly conserved Y-shaped structural unit. Therefore, this study benefits the search for new methods for the detection and treatment of P. aeruginosa biofilm formation.


Assuntos
4-Butirolactona/análogos & derivados , Aptâmeros de Nucleotídeos/química , Biofilmes/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , 4-Butirolactona/antagonistas & inibidores , 4-Butirolactona/química , Aptâmeros de Nucleotídeos/síntese química , Aptâmeros de Nucleotídeos/farmacologia , Proteínas de Bactérias/metabolismo , Desenho de Drogas , Modelos Moleculares , Conformação de Ácido Nucleico , Ligação Proteica/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , Relação Estrutura-Atividade
6.
Cell Signal ; 51: 222-232, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30102978

RESUMO

Drug repurposing with a better understanding of the underlying mechanism has provided new avenues to find treatment for malignancies. Esophageal adenocarcinoma (EAC) is a rapidly increasing cancer with a dismal 5-year survival rate of <15%. Lack of efficient treatment options contributes to the high mortality rate of EAC. To find new therapy against EAC we performed unbiased drug screening of an FDA-approved drug library and identified that the cardiac glycosides including Ouabain, Digoxin and Digitoxin efficiently inhibit the proliferation of EAC cell lines (OE33 and OE19) both in vitro and in vivo. RNA-Sequencing analysis combined with RNAi screening revealed that Ouabain suppresses the proliferation of EAC cells through downregulation of p38 MAP-Kinase 6 (MAP2K6, also known as MKK6). Consistently, shRNA-mediated knockdown of MKK6 reduced the proliferation of EAC cells and tumor growth. Further analysis demonstrated that MKK6 inhibition leads to the reduced levels of the transcription factor SOX9. In line with this finding, deletion of SOX9 with CRISPR/Cas9 resulted in decreased proliferation of EACs in 3D organoid culture and reduced tumor growth. Together these findings establish a druggable axis that can be harnessed for therapeutic gain against EAC.


Assuntos
Adenocarcinoma/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Neoplasias Esofágicas/tratamento farmacológico , MAP Quinase Quinase 6/antagonistas & inibidores , MAP Quinase Quinase 6/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Fatores de Transcrição SOX9/metabolismo , Adenocarcinoma/enzimologia , Adenocarcinoma/patologia , Animais , Linhagem Celular Tumoral , Digitoxina/farmacologia , Digitoxina/uso terapêutico , Digoxina/farmacologia , Digoxina/uso terapêutico , Neoplasias Esofágicas/enzimologia , Neoplasias Esofágicas/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , MAP Quinase Quinase 6/genética , Camundongos Endogâmicos NOD , Ouabaína/farmacologia , Ouabaína/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Fatores de Transcrição SOX9/genética , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
7.
J Cell Physiol ; 233(5): 3855-3866, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-28777465

RESUMO

The esophagus is a pivotal organ originating from anterior foregut that links the mouth and stomach. Moreover, its development involves precise regulation of multiple signal molecules and signal transduction pathways. After abnormal regulation of these molecules in the basal cells of the esophagus occurs, multiple diseases, including esophageal atresia with or without tracheoesophageal fistula, Barrett esophagus, gastroesophageal reflux, and eosinophilic esophagitis, will take place as a result. Furthermore, expression changes of signal molecules or signal pathways in basal cells and the microenvironment around basal cells both can initiate the switch of malignant transformation. In this review, we highlight the molecular events underlying the transition of normal development to multiple esophageal diseases. Additionally, the animal models of esophageal development and related diseases, challenges, and strategies are extensively discussed.


Assuntos
Esôfago/metabolismo , Refluxo Gastroesofágico/metabolismo , Metástase Neoplásica/patologia , Neoplasias/patologia , Células-Tronco/citologia , Animais , Modelos Animais de Doenças , Esôfago/patologia , Refluxo Gastroesofágico/patologia , Humanos , Neoplasias/metabolismo , Células-Tronco/metabolismo
8.
Nature ; 550(7677): 529-533, 2017 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-29019984

RESUMO

In several organ systems, the transitional zone between different types of epithelium is a hotspot for pre-neoplastic metaplasia and malignancy, but the cells of origin for these metaplastic epithelia and subsequent malignancies remain unknown. In the case of Barrett's oesophagus, intestinal metaplasia occurs at the gastro-oesophageal junction, where stratified squamous epithelium transitions into simple columnar cells. On the basis of a number of experimental models, several alternative cell types have been proposed as the source of this metaplasia but in all cases the evidence is inconclusive: no model completely mimics Barrett's oesophagus in terms of the presence of intestinal goblet cells. Here we describe a transitional columnar epithelium with distinct basal progenitor cells (p63+KRT5+KRT7+) at the squamous-columnar junction of the upper gastrointestinal tract in a mouse model. We use multiple models and lineage tracing strategies to show that this squamous-columnar junction basal cell population serves as a source of progenitors for the transitional epithelium. On ectopic expression of CDX2, these transitional basal progenitors differentiate into intestinal-like epithelium (including goblet cells) and thereby reproduce Barrett's metaplasia. A similar transitional columnar epithelium is present at the transitional zones of other mouse tissues (including the anorectal junction) as well as in the gastro-oesophageal junction in the human gut. Acid reflux-induced oesophagitis and the multilayered epithelium (believed to be a precursor of Barrett's oesophagus) are both characterized by the expansion of the transitional basal progenitor cells. Our findings reveal a previously unidentified transitional zone in the epithelium of the upper gastrointestinal tract and provide evidence that the p63+KRT5+KRT7+ basal cells in this zone are the cells of origin for multi-layered epithelium and Barrett's oesophagus.


Assuntos
Esôfago de Barrett/patologia , Linhagem da Célula , Células Epiteliais/patologia , Epitélio/patologia , Junção Esofagogástrica/patologia , Células-Tronco/patologia , Animais , Esôfago de Barrett/genética , Esôfago de Barrett/metabolismo , Fator de Transcrição CDX2/genética , Fator de Transcrição CDX2/metabolismo , Rastreamento de Células , Esofagite/metabolismo , Esofagite/patologia , Junção Esofagogástrica/metabolismo , Refluxo Gastroesofágico , Células Caliciformes/metabolismo , Células Caliciformes/patologia , Humanos , Queratina-5/metabolismo , Queratina-7/metabolismo , Metaplasia/metabolismo , Metaplasia/patologia , Camundongos , Fosfoproteínas/metabolismo , Células-Tronco/metabolismo , Transativadores/metabolismo
9.
Nutrients ; 9(4)2017 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-28425936

RESUMO

Renal ischemia/reperfusion (I/R) injury continues to be a complicated situation in clinical practice. Genistein, the main isoflavone found in soy products, is known to possess a wide spectrum of biochemical and pharmacological activities. However, the protective effect of genistein on renal I/R injury has not been well investigated. In the current study, we explore whether genistein exhibits its renal-protective effects through SIRT1 (Sirtuin 1) in I/R-induced mice model. We found the treatment of genistein significantly reduced renal I/R-induced cell death, simultaneously stimulating renal cell proliferation. Meanwhile, SIRT1 expression was up-regulated following the administration of genistein in renal region. Furthermore, pharmacological inhibition or shRNA-mediated depletion of SIRT1 significantly reversed the protective effect of genistein on renal dysfunction, cellular damage, apoptosis, and proliferation following I/R injury, suggesting an indispensible role of the increased SIRT1 expression and activity in this process. Meanwhile, the reduced p53 and p21 expression and increased PCNA (Proliferating Cell Nuclear Antigen) expression were blocked after the depletion of SIRT1 compared with the genistein treatment group in the renal I/R process. Hence, our results provided further experimental basis for the potential use of genistein for the treatment of kidney disease with deficiency of SIRT1 activity.


Assuntos
Genisteína/farmacologia , Nefropatias/tratamento farmacológico , Rim/efeitos dos fármacos , Traumatismo por Reperfusão/tratamento farmacológico , Sirtuína 1/metabolismo , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Modelos Animais de Doenças , Rim/fisiopatologia , Camundongos , Camundongos Endogâmicos BALB C , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Traumatismo por Reperfusão/complicações , Sirtuína 1/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
10.
Mol Cancer ; 16(1): 62, 2017 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-28288641

RESUMO

BACKGROUND: High levels of SOX2 protein are correlated with increased dissemination of breast cancer. However, the underlying molecular mechanisms are not fully understood. METHODS: In this study we investigate the role of SOX2 in breast cancer metastasis using multiple in vitro and in vivo assays including cell culture, shRNA-mediated knockdown, wound healing, colony formation, transwell chamber, xenograft and tail vein injection. Moreover, western blot, immunostaining, microarray and real-time PCR were used to determine the change of protein and miRNA levels. Luciferase assays were also used to evaluate activity which TUSC3 is a target of miR-181a-5p and miR-30e-5p, and the clinical survival relevance was analyzed by Kaplan-Meier analysis. RESULTS: We identified a novel pathway involving SOX2 regulation of microRNAs to control the proliferation and migration of breast cancer cells. shRNA-mediated knockdown of SOX2 inhibits breast cancer cell expansion and migration. More importantly, we found that these changes are accompanied by significant reduction in the levels of two microRNAs, miR-181a-5p and miR-30e-5p. Overexpression of these two microRNAs leads to reduced protein levels of Tumor Suppressor Candidate 3 (TUSC3) in breast cancer cells; mutations of the potential binding sites in the 3'-UTR of TUSC3 abrogate the inhibitory effects of the microRNAs. We further found that upregulation of TUSC3 expression leads to reduced proliferation and migration of breast cancer cells. In human breast cancer samples the levels of TUSC3 protein are inversely correlated with those of SOX2 protein. CONCLUSIONS: Taken together, our work reveals a novel SOX2-mediated regulatory axis that plays critical roles in the proliferation, migration and invasiveness of breast cancer cells. Targeting this axis may provide beneficial effect in the treatment of breast cancer.


Assuntos
Neoplasias da Mama/genética , Regulação Neoplásica da Expressão Gênica , Proteínas de Membrana/genética , MicroRNAs/genética , Interferência de RNA , Fatores de Transcrição SOXB1/genética , Proteínas Supressoras de Tumor/genética , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Progressão da Doença , Feminino , Técnicas de Silenciamento de Genes , Xenoenxertos , Humanos , Estimativa de Kaplan-Meier , Camundongos , Modelos Biológicos , Metástase Neoplásica , Prognóstico , Transdução de Sinais
11.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 33(3): 342-346, 2017 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-28274313

RESUMO

Objective To prepare a lentiviral vector expressing LLGL2 and establish KYSE450 and TE-1 cell lines for the stable expression of LLGL2. Methods The full-length LLGL2 sequence was amplified by high-fidelity PCR, and then it was inserted into pCDH-CMV-IRES-GFP-EF1-Puro vectors. The recombinant plasmid was confirmed by double enzyme digestion and sequencing. After co-infection of pCDH-CMV-LLGL2-IRES- GFP-EF1-Puro with vesicular stomatitis virus glycoprotein (VSVG) and PHR into HEK293T cells, the lentivirus was harvested and used for infecting esophageal squamous cell carcinoma cell lines including KYSE450 and TE-1 cells. These two cell lines infected with the lentivirus were screened with puromycin, and the stable cell lines were further confirmed with green fluoresence and Western blotting. Results Dual-enzyme digestion and sequencing confirmed that the pCDH-CMV-LLGL2-IRES-GFP-EF1-Puro vector, a lentiviral expression vector for the overexpression of LLGL2, was successfully constructed through high-fidelity PCR and ligation. Western blotting showed the increased expression level of LLGL2 protein in KYSE450 and TE-1 stable cell lines compared with the controls. Conclusion The experiment successfully established KYSE450 and TE-1 stable cell lines for the overexpression of LLGL2.


Assuntos
Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Proteínas do Citoesqueleto/genética , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas/metabolismo , Proteínas do Citoesqueleto/metabolismo , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas do Esôfago , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Humanos , Lentivirus/genética , Lentivirus/metabolismo , Transfecção
12.
Semin Cell Dev Biol ; 66: 25-35, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28007661

RESUMO

The esophagus is derived from the anterior portion of the developmental intermediate foregut, a structure that also gives rise to other organs including the trachea, lung, and stomach. Genetic studies have shown that multiple signaling pathways (e.g. Bmp) and transcription factors (e.g. SOX2) are required for the separation of the esophagus from the neighboring respiratory system. Notably, some of these signaling pathways and transcription factors continue to play essential roles in the subsequent morphogenesis of the esophageal epithelium which undergoes a simple columnar-to-stratified squamous conversion. Reactivation of the relevant signaling pathways has also been associated with pathogenesis of esophageal diseases that affect the epithelium and its stem cells in adults. In this review we will summarize these findings. We will also discuss new data regarding the cell-of-origin for the striated and smooth muscles surrounding the esophagus and how they are differentiated from the mesenchyme during development.


Assuntos
Esôfago/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento/genética , Células-Tronco/metabolismo , Fatores de Transcrição/metabolismo , Diferenciação Celular , Humanos
13.
Medicine (Baltimore) ; 94(31): e1301, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26252309

RESUMO

As an immunotoxin, diphtheria toxin has been widely used in gene therapy and gene function assays for its roles in protein synthesis inhibition, and the aim of our study is to set up a nonintegrating lentiviral system for specific expression of diphtheria toxin A (DTA) used in cancer gene therapy.Here, we established a lentiviral system that could coordinately express fluorescent protein and DTA driven by the cytomegalovirus (CMV) promoter, which is convenient for us to precisely trace the expression of DTA and monitor the process of lentivirus packaging. To achieve safer cancer therapy, we replaced the CMV promoter with the Survivin promoter, a specific promoter that is dramatically activated in cancer tissues and cells, but not in normal tissues and cells, and that will impose greater therapeutic potential because a significant expression difference occurred between these 2 groups. Meanwhile, we obtained integrase-deficient lentivirus (IDLV) after packaging with the integrase mutant, which expresses defective integrase RRK262263264AAH, to minimize the side effects that derived from the insertional mutagenesis of the host genome.Our results suggest that the IDLV system that we generated possesses therapeutic potential in cancers in vitro and in vivo.


Assuntos
Toxina Diftérica/genética , Vetores Genéticos , Proteínas Inibidoras de Apoptose/genética , Integrases , Lentivirus , Transfecção/métodos , Animais , Técnicas de Cultura de Células , Linhagem Celular , Toxina Diftérica/metabolismo , Feminino , Humanos , Proteínas Inibidoras de Apoptose/metabolismo , Camundongos , Camundongos Nus , RNA Mensageiro/metabolismo , Survivina , Ensaios Antitumorais Modelo de Xenoenxerto
14.
J Clin Invest ; 125(4): 1557-68, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25774506

RESUMO

Tissue homeostasis requires balanced self-renewal and differentiation of stem/progenitor cells, especially in tissues that are constantly replenished like the esophagus. Disruption of this balance is associated with pathological conditions, including eosinophilic esophagitis (EoE), in which basal progenitor cells become hyperplastic upon proinflammatory stimulation. However, how basal cells respond to the inflammatory environment at the molecular level remains undetermined. We previously reported that the bone morphogenetic protein (BMP) signaling pathway is critical for epithelial morphogenesis in the embryonic esophagus. Here, we address how this pathway regulates tissue homeostasis and EoE development in the adult esophagus. BMP signaling was specifically activated in differentiated squamous epithelium, but not in basal progenitor cells, which express the BMP antagonist follistatin. Previous reports indicate that increased BMP activity promotes Barrett's intestinal differentiation; however, in mice, basal progenitor cell-specific expression of constitutively active BMP promoted squamous differentiation. Moreover, BMP activation increased intracellular ROS levels, initiating an NRF2-mediated oxidative response during basal progenitor cell differentiation. In both a mouse EoE model and human biopsies, reduced squamous differentiation was associated with high levels of follistatin and disrupted BMP/NRF2 pathways. We therefore propose a model in which normal squamous differentiation of basal progenitor cells is mediated by BMP-driven NRF2 activation and basal cell hyperplasia is promoted by disruption of BMP signaling in EoE.


Assuntos
Proteínas Morfogenéticas Ósseas/fisiologia , Esofagite Eosinofílica/patologia , Esôfago/citologia , Fator 2 Relacionado a NF-E2/fisiologia , Animais , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/biossíntese , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Proteínas Morfogenéticas Ósseas/biossíntese , Proteínas Morfogenéticas Ósseas/genética , Diferenciação Celular , Células Cultivadas , Esofagite Eosinofílica/metabolismo , Células Epiteliais/metabolismo , Esôfago/crescimento & desenvolvimento , Folistatina/fisiologia , Genes Reporter , Humanos , Hiperplasia , Camundongos , Camundongos Endogâmicos C57BL , Morfogênese , Estresse Oxidativo , Precursores de Proteínas/biossíntese , Precursores de Proteínas/genética , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia
15.
Curr Gene Ther ; 14(5): 352-64, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25174579

RESUMO

Lentiviruses are powerful tools for gene delivery and have been widely used for the dissection of gene functions in both replicating and quiescent cells. Recently, lentiviruses have also been used for delivering target sequences in gene therapy. Although the lentiviral system provides sustained exogenous gene expression, serious concerns have been raised due to its unfavorable insertion-mediated mutagenesis effect, thereby resulting in the silencing or activation of some unexpected genes. Thus, an array of modifications of the original vectors may reduce risks. Here, we briefly review the structure of the integrase protein, which is an essential protein for viral insertion and integration; the mechanisms of integrase-mediated integration; and the effects of the modifications of integrase. Moreover, we discuss the advantages resulting from integrase modifications and their future applications. Taken together, the generation of integrase-deficient lentivirus (IDLV) not only provides us with an opportunity to reduce the risk of virus-mediated insertions, which would improve the safety of gene therapy, but also favors gene correction and vaccine development.


Assuntos
Terapia Genética/métodos , Vetores Genéticos/uso terapêutico , Integrases/deficiência , Lentivirus/genética , Sequência de Aminoácidos , Humanos , Integrases/genética , Dados de Sequência Molecular
16.
Cell Stem Cell ; 12(3): 304-15, 2013 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-23472872

RESUMO

Sox2 regulates the self-renewal of multiple types of stem cells. Recent studies suggest it also plays oncogenic roles in the formation of squamous carcinoma in several organs, including the esophagus where Sox2 is predominantly expressed in the basal progenitor cells of the stratified epithelium. Here, we use mouse genetic models to reveal a mechanism by which Sox2 cooperates with microenvironmental signals to malignantly transform epithelial progenitor cells. Conditional overexpression of Sox2 in basal cells expands the progenitor population in both the esophagus and forestomach. Significantly, carcinoma only develops in the forestomach, where pathological progression correlates with inflammation and nuclear localization of Stat3 in progenitor cells. Importantly, co-overexpression of Sox2 and activated Stat3 (Stat3C) also transforms esophageal basal cells but not the differentiated suprabasal cells. These findings indicate that basal stem/progenitor cells are the cells of origin of squamous carcinoma and that cooperation between Sox2 and microenvironment-activated Stat3 is required for Sox2-driven tumorigenesis.


Assuntos
Inflamação/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Fator de Transcrição STAT3/metabolismo , Células-Tronco/metabolismo , Células-Tronco/patologia , Animais , Células Cultivadas , Humanos , Camundongos , Fatores de Transcrição SOXB1/genética , Fator de Transcrição STAT3/genética
17.
Cell Signal ; 25(5): 1264-71, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23416461

RESUMO

The Sry-containing protein Sox2 initially was known to regulate the self-renewal of the mouse and human embryonic stem cells (ESCs). It is also important for the maintenance of stem cells in multiple adult tissues including the brain and trachea, and it is one of the key transcription factors for establishing induced pluripotent stem cells. Recently, overexpression and gene amplification of Sox2 have been associated with the development of squamous cell carcinoma in multiple tissues such as the lung and esophagus. These different roles for Sox2 involve a complicated regulatory networks consisting of microRNAs, kinases and signaling molecules. While the levels of Sox2 are modulated transcriptionally and translationally, post-translational modification is also important for the various functions of Sox2. In clinics, high levels of Sox2 are correlated with poor prognosis and increased proliferation of cancer stem cells. Therefore targeting Sox2 can be potentially explored for a new therapeutic avenue to treat cancers. This review will focus on the different roles for Sox2 in stem cell maintenance and its oncogenic roles in the context of signal transcription and microRNA regulation. We will also review the main upstream and downstream targets of Sox2, which can be potentially used as therapeutic measures to treat cancer with abnormal levels of Sox2.


Assuntos
Células-Tronco Neoplásicas/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Animais , Transformação Celular Neoplásica , Humanos , MicroRNAs/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Células-Tronco Neoplásicas/citologia , Processamento de Proteína Pós-Traducional , Fatores de Transcrição SOXB1/genética , Transdução de Sinais
18.
J Cell Biochem ; 114(2): 250-5, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22949372

RESUMO

Aptamers are a group of molecules, which can specifically bind, track, and inhibit target molecules, comprising DNA aptamers, RNA aptamers, and peptide aptamers. So far, there are much progress about developing novel aptamers and their expansile applications. This prospect systematically introduces the composition and technological evolution of aptamers, and then focuses on the application of aptamers in cancer diagnosis, imaging, and therapy. Following this, we discuss the potential to harness aptamers in discovering the biomarker of stem cells, which is favorable for us to study the normal developmental or abnormal pathological process of tissue and to deliver drugs into target cells or tissues in the future.


Assuntos
Aptâmeros de Nucleotídeos , Aptâmeros de Peptídeos , Neoplasias , Células-Tronco , Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/uso terapêutico , Aptâmeros de Peptídeos/química , Aptâmeros de Peptídeos/uso terapêutico , Biomarcadores Tumorais/uso terapêutico , Diagnóstico por Imagem , Humanos , Neoplasias/diagnóstico , Neoplasias/patologia , Neoplasias/terapia , RNA Interferente Pequeno/química , RNA Interferente Pequeno/metabolismo , Técnica de Seleção de Aptâmeros/métodos , Células-Tronco/citologia , Células-Tronco/metabolismo
19.
Cell Signal ; 25(1): 349-54, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23085261

RESUMO

CDP, a key transcription regulator encoded by Cutl1 gene, has been demonstrated to be involved in repressing or promoting expression of target genes through its specific DNA-binding, meanwhile, the activity of CDP was influenced by some types of modifications including transcriptional, posttranscriptional, translational and posttranslational modifications. In this review, we systematically analyzed the role of CDP in normal development and tumor progression, and then emphasized its interactors and downstream molecules. Eventually, we concluded that Cut1 could promote cancer progression and its down-regulating expression will be a promising strategy for cancer therapy.


Assuntos
Proteínas de Homeodomínio/metabolismo , Neoplasias/terapia , Proteínas Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Progressão da Doença , Proteínas de Homeodomínio/antagonistas & inibidores , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Nucleares/antagonistas & inibidores , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Transdução de Sinais , Fatores de Transcrição , Fator de Crescimento Transformador beta/metabolismo
20.
J Cell Biochem ; 113(9): 2909-19, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22532014

RESUMO

The site-specific recombination mediated by Cre recombinase has been utilized extensively in genetic engineering and gene function studies. Efficient delivery of a Cre enzyme with enzymatic activity and the ability to monitor the enzyme expression are required in applications, and lentiviral constructs with a fluorescent protein (FP) to report the Cre expression are suitable for most studies. However, the current lentiviral vector systems have some deficiencies in precise reporting the Cre expression through fluorescence. To solve the problem, we generated a lentiviral system with Cre and RFP or EGFP bridged by an FMDV 2A sequence in an open reading frame expressed by a CMV promoter. We then examined the capabilities of the constructs to package with VSVG into infectious virus and to mediate expression of the Cre enzyme and fluorescent reporter. Furthermore, we monitored the bioactivities of the expressed products. We demonstrated the coordinate expression of the enzyme and the reporter. The expressed Cre was efficient at removing LoxP-flanked fragments in cells and did not show obvious cellular toxicity, and the expressed FPs allowed direct observation under fluorescent microscope. Therefore, the conjugation of CMV-Cre-2A-FP represents a significant improvement to the current lentiviral Cre delivery systems for obtaining a required Cre activity while accurately monitoring its presence. Our study also provides information concerning application of the established vector system.


Assuntos
Vírus da Febre Aftosa/genética , Vetores Genéticos/genética , Proteínas de Fluorescência Verde/metabolismo , Integrases/metabolismo , Proteínas de Fluorescência Verde/genética , Células HeLa , Humanos , Integrases/genética , Lentivirus/genética
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