Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 4.593
Filtrar
1.
Life Sci ; 234: 116783, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31442552

RESUMO

Breast cancer (BCa) is the most commonly diagnosed lethal cancer in women worldwide. Notch signaling pathway is directly linked to BCa recurrence and aggressiveness. Natural remedies are becoming a prime choice to overcome against cancer due to lesser side effect and cost-effectiveness. Bulbine frutescens (Asphodelaceae), a traditional medicinal plant in South Africa possess bioactive flavonoids and terpenoids. Polar (methanol) and non-polar (hexane) B. frutescens plant extracts were prepared. GC-MS analysis revealed the differential presence of secondary metabolites in both methanolic and hexane extracts. We hereby first time evaluated the anticancer potential of B. frutescens methanolic and hexane extract in triple-negative and luminal BCa cells. B. frutescens extracts significantly decreased cell viability (IC50 4.8-28.4 µg/ml) and induced cell cycle arrest at G1 phase in MDA-MB-231 and T47D cells as confirmed by spectrophotometry and flow cytometry technique. RT-PCR analysis of cell cycle (cyclin D1, CDK4, and p21) and apoptosis modulating genes (caspase 3, Bcl2 and survivin) revealed upexpression of p21, and caspase 3, and down expression of cyclin D1, CDK4, Bcl2 and survivin genes in extract-treated BCa cells. Fluorescence spectrophotometry and confocal microscopy showed B. frutescens induced nuclear morphology and mitochondrial integrity disruption, and increased reactive oxygen species production in MDA-MB-231 and T47D cells. Flow cytometric apoptosis analysis of B. frutescens extracts treated MDA-MB-231 cells showed ≈13% increase in early apoptotic population in comparison to non-treated cells. Dual-Luciferase Reporter assay confirmed notch promoter inhibitory activity of B. frutescens extracts. Moreover, RTPCR analysis showed down regulation of notch responsive genes (Hes1 and Hey1) at transcription levels in extract-treated BCa cells. Western Blot analysis showed increased procaspase 3 protein expression in extract-treated BCa cells. In all the assays methanolic extract showed better anti-cancer properties. Literature-based identification of methanol soluble phytochemicals in B. frutescens and in silico docking study revealed Bulbineloneside D as a potent ϒ-secretase enzyme inhibitor. In comparison to standard notch inhibitor, lead phytochemical showed two additional hydrophobic interactions with Ala80 and Leu81 amino acids. In conclusion, B. frutescens phytochemicals have cell cycle arrest, ROS production, apoptosis induction, and mitochondria membrane potential disruption efficacy in breast cancer cells. B. frutescens phytochemicals have the ability to downregulate the notch signaling pathway in triple-negative and luminal breast cancer cells.


Assuntos
Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos , Xanthorrhoeaceae/química , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Secretases da Proteína Precursora do Amiloide/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
2.
Zhonghua Gan Zang Bing Za Zhi ; 27(7): 527-532, 2019 Jul 20.
Artigo em Chinês | MEDLINE | ID: mdl-31357779

RESUMO

Objective: To observe the expressional changes in Notch signaling pathway and toll-like receptor 4 (TLR4) and their interactions on the functions of CD14(+) monocytes in chronic hepatitis C patients. Methods: A total of 24 treatment-naïve chronic hepatitis C cases and 10 healthy individuals, who visited Shaanxi Provincial People's Hospital from August to October 2017, were enrolled. Selected CD14(+) monocytes were stimulated by the Notch signaling pathway inhibitor DAPT or transfected with TLR4 siRNA, and the levels of Notch1, Notch2, Hes1 and Hes5 mRNA were detected by real-time quantitative PCR. TLR4 protein levels and phosphorylation of NF-κB was detected by Western blot. ELISA was used to detect the level of cytokines secreted from CD14(+) monocytes. A t-test or paired t-test was used for comparison between groups. Results: The relative expression of Notch1 mRNA (3.97 ± 2.03 vs. 0.91 ± 0.76, P < 0.01) and downstream of Notch signaling pathway (5.96 ± 2.31 vs. 0.99 ± 0.45, P < 0.01), Hes1 mRNA and Hes5 mRNA (4.31 ± 1.05 vs. 0.84 ± 0.20, P < 0.01) in CD14+ monocytes of chronic hepatitis C patients was significantly higher than that of healthy individuals. The relative expression of TLR4 mRNA (5.14 ± 1.09 vs. 1.27 ± 0.39) and protein level in CD14(+) monocytes of chronic hepatitis C patients were significantly higher than those of healthy individuals (P < 0.01). An inhibition of Notch signaling pathway with DAPT had reduced the relative expression level of TLR4 mRNA (2.58 ± 1.36 vs. 4.34 ± 1.88, P < 0.05), protein expression and phosphorylation of NF-B in CD14(+) monocytes of chronic hepatitis C patients. Furthermore, the secretion level of MCP-1 [(94.32 ± 23.59) pg/ml vs. (64.07 ± 9.39) pg/ml, P < 0.01] and IL-8 [(12.54 ± 4.89) pg/ml vs. (7.92 ± 3.01) pg/ml, P < 0.05] was significantly reduced. TLR4 siRNA transfection reduced the expressions of Notch1 mRNA (2.09 ± 1.72 vs. 3.73 ± 1.75, P < 0.05), Hes1 (2.87 ± 0.84 vs. 5.54 ± 0.97, P < 0.01), and Hes5 (2.89 ± 0.93 vs. 4.51 ± 1.54, P < 0.01) in CD14(+) monocytes of chronic hepatitis C patients. Conclusion: Interaction of Notch signaling pathway with TLR4 can promote the function of CD14(+) monocytes in chronic hepatitis C patients.


Assuntos
Hepatite C Crônica , Monócitos/citologia , Receptores Notch/metabolismo , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Estudos de Casos e Controles , Células Cultivadas , Humanos , Receptores de Lipopolissacarídeos , NF-kappa B/metabolismo
3.
Life Sci ; 232: 116630, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31279783

RESUMO

AIMS: Lung adenocarcinoma consists of multiple therapeutic targets, however, patients will inevitably progress to later stage diagnosis with Tyrosine Kinase Inhibitor treatment resistance. We aim to investigate the roles of non-coding TUSC7 in ordering the cell division tendency, helping to sensitize the resistance in a miRNA incorporating way. MATERIALS AND METHODS: Online study of bioinformatics analysis, molecular experiments of luciferase test, immunofluorescence staining and qRT-PCR were applied to dig out the mechanistic regulations. KEY FINDINGS: TUSC-7 inhibited the renewal ability of adenocarcinoma stem cells, yielding to asymmetric cell splitting. Informatics analysis and the luciferase testing confirmed the 3'UTR binding site, and revealed the post-transcriptional regulation of NUMB referring to miR-146. TUSC-7 sponged miR-146 and abolished its degradation toward to NUMB, and this integrated cascade made several genes become tangled to full functionality. SIGNIFICANCE: TUSC-7 was proved to be one strong suppressive lnc-RNA in lung adenocarcinoma stem cells, functioning through inactivating NOTCH signaling, and the turbulence on division modes precisely pointed to the key mechanisms of stem cells' renewal. The decreasing of tumor suppressive miR-146 was necessary in TUSC-7 conducted renewal repression, despite it alone could also reduce the renewal efficiency, indicating that more complicated non-coding genes may be involved in its regulation.


Assuntos
Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/patologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Células-Tronco Neoplásicas/metabolismo , RNA Longo não Codificante/fisiologia , Regiões 3' não Traduzidas , Adenocarcinoma de Pulmão/genética , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Humanos , Neoplasias Pulmonares/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Receptores Notch/genética , Receptores Notch/metabolismo
4.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 35(5): 405-411, 2019 May.
Artigo em Chinês | MEDLINE | ID: mdl-31223109

RESUMO

Objective To investigate the effect of miR-25-3p targeting a disintegrin and metalloproteinase 10 (ADAM10) on the differentiation of P19 cells into cardiomyocytes by regulating Notch signaling pathway. Methods P19 cells were induced to differentiate into cardiomyocytes with dimethyl sulfoxide (DMSO) and collected at 0, 5, and 10 days. The mRNA levels of myocardial differentiation markers GATA4, cTnT, atrial natriuretic polypeptide (ANP) and the level of miR-25-3p were detected by real-time quantitative PCR (qRT-PCR). The protein level of ADAM10 was assessed by Western blot analysis. The miR-25-3p was over-expressed in P19 cells by infected with retrovirus, and the expression levels of miR-25-3p and ADAM10 in the infected P19 cells were detected by qRT-PCR and Western blotting. Bioinformatics were used to predict the targeted matching relationship between miR-25-3p and ADAM10 gene, which was then verified by the luciferase reporter gene system. After infection, P19 cells were induced to differentiate by DMSO for 10 days. Then the protein expression of cTnI was detected by immunofluorescence assay to calculate the differentiation rate of cardiomyocytes, and the proteins expression of myocardial differentiation markers GATA4, cTnT, ANP and Notch signaling pathway-related molecules Notch1, hes family bHLH transcription factor 1 (Hes1), Hey1, and Hey2 were detected by Western blotting. Results During the 0, 5 and 10 days of the differentiation of P19 cells into myocardial cells, the mRNA expression levels of GATA4, cTnT, ANP and the protein expression level of ADAM10 gradually increased, while the expression level of miR-25-3p gradually decreased. After retrovirus infection, the expression level of miR-25-3p in the infected P19 cells went up significantly, while the protein expression level of ADAM10 went down significantly. Subsequently, ADAM10 was confirmed as a target gene of miR-25-3p. After the 10 days of differentiation, over-expression of miR-25-3p significantly decreased the differentiation rate of cardiomyocytes, and down-regulated the levels of the markers of myocardial differentiation-related proteins GATA4, cTnT, ANP, and the Notch signaling pathway related-proteins, including Notch1, Hes1, Hey1 and Hey2. Conclusion The miR-25-3p can significantly inhibit the differentiation of P19 cells into cardiomyocytes, and the mechanism may be related to inhibite the activation of Notch signaling pathway by depressing ADAM10 expression.


Assuntos
Proteína ADAM10/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Diferenciação Celular , Proteínas de Membrana/metabolismo , MicroRNAs/metabolismo , Miócitos Cardíacos/citologia , Transdução de Sinais , Animais , Linhagem Celular Tumoral , Camundongos , Receptores Notch/metabolismo
5.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 35(5): 441-446, 2019 May.
Artigo em Chinês | MEDLINE | ID: mdl-31223113

RESUMO

Objective To investigate the effect of baicalin on the viability and cell cycle of psoriatic keratinocytes and its possible mechanism. Methods MTT assay was used to detect the viability of keratinocytes treated by 0, 10, 50, 100, 200, 300 µg/mL baicalin for 48 hours. The cell cycle and apoptotic rate were detected by flow cytometry. The mRNA expression levels of ki67, Fas, and caspase-3 were analyzed by real-time quantitative PCR and the protein expression of Notch 1, Jagged 1 and Hes l in the keratinocytes were observed by Western blot analysis. Results The viability of keratinocytes was inhibited by baicalin in a dose-dependent manner. Baicalin (200 µg/mL) significantly promoted the apoptosis of keratinocytes, arrested the S phase, inhibited the mRNA expression of ki67, increased the Fas and caspase-3 levels, down-regulated the protein expression of Jagged 1, and up-regulated the Notch 1 and Hes protein levels. Conclusion Baicalin can significantly inhibit the viability of keratinocytes and promote cell apoptosis, probably by activating Notch signaling pathway.


Assuntos
Flavonoides/farmacologia , Queratinócitos/efeitos dos fármacos , Psoríase/patologia , Receptores Notch/metabolismo , Transdução de Sinais , Apoptose , Células Cultivadas , Humanos , Queratinócitos/citologia
6.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 35(2): 165-168, 2019 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-31250610

RESUMO

OBJECTIVE: To investigate the effects of Notch signal on hypoxic induction factor (HIF-1α) and autophagy-associated genes Beclin1, LC3I, LC3II in oxygen-glucose deprivation (OGD) induced myocardial cell injury. METHODS: The OGD model was established using hypoxic culture box and hypoglycemic DMEM medium. The cells were divided into normal control group, OGD group, OGD + NC siRNA group, OGD + Notch1 siRNA group and OGD + HIF-1α siRNA group. Western blot was used to detect the interference effects of HIF-1α siRNA and Notch1 siRNA. The effects of Notch1 siRNA and HIF-1α siRNA on the activity of myocardial cells in OGD model were detected by the CCK-8 assay. The effects of Notch1 siRNA and HIF-1α siRNA on autophage-associated genes Beclin1, LC3I and LC3II expression were detected by Western blot. RESULTS: The results of Western blot showed that HIF-1α siRNA could effectively knock down the expression of HIF-1α in myocardial cells in OGD model, and Notch1 siRNA could effectively knock down the expression of Notch1 and HIF-1α in myocardial cells in OGD model. The result of CCK-8 assay showed that Notch1 siRNA and HIF-1α siRNA reduced the activity of myocardial cells in OGD model, and there was no statistical difference between the two groups. Western blot results showed that Notch1 siRNA and HIF-1α siRNA could reduce the expressions of the autophagy-associated genes Beclin1, LC3I and LC3II, and reduce the ratio of LC3II to LC3I at mRNA level. CONCLUSION: Notch1 plays a role in myocardial protection by regulating the expression of HIF-1α to regulate the autophagy in OGD model cells.


Assuntos
Autofagia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Miócitos Cardíacos/citologia , Receptores Notch/metabolismo , Transdução de Sinais , Proteína Beclina-1/metabolismo , Hipóxia Celular , Células Cultivadas , Glucose , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Miócitos Cardíacos/patologia , Oxigênio
7.
Cell Prolif ; 52(4): e12637, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31168899

RESUMO

OBJECTIVES: Chondrocyte proliferation and differentiation are crucial for endochondral ossification, but their regulatory mechanism remains unclear. The present study aimed to determine the physiological function of TGFß1 signalling in the proliferation and differentiation of antler chondrocytes and explore its relationship with Notch, Shh signalling and Foxa. MATERIALS AND METHODS: Immunofluorescence, Western blot, MTS assay, flow cytometry, RNA interference and real-time PCR were used to analyse the function and regulatory mechanisms of TGFß1 signalling in antler chondrocyte proliferation and differentiation. RESULTS: TGFß1, TGFBR1 and TGFBR2 were highly expressed in antler cartilage. TGFß1 promoted chondrocyte proliferation, increased the proportion of S-phase cells and induced the expression of hypertrophic chondrocyte markers Col X, Runx2 and Alpl. However, this induction was weakened by TGFß receptor inhibitor SB431542 and Smad3 inhibitor SIS3. Simultaneously, TGFß1 activated Notch and Shh signalling whose blockage attenuated the above effects of rTGFß1, whereas addition of rShh rescued the defects in chondrocyte proliferation and differentiation elicited by SB431542 and SIS3. Further analysis revealed that inhibition of Notch signalling impeded TGFß1 activation of the Shh pathway. Knockdown of Foxa1, Foxa2 and Foxa3 abrogated the effects of TGFß1 on chondrocyte differentiation. Notch and Shh signalling mediated the regulation of Foxa transcription factors by TGFß1. CONCLUSIONS: TGFß1 signalling could induce the proliferation and differentiation of antler chondrocytes through Notch-Shh-Foxa pathway.


Assuntos
Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta1/metabolismo , Animais , Chifres de Veado , Benzamidas/farmacologia , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Dioxóis/farmacologia , Proteínas Hedgehog/metabolismo , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Isoquinolinas/farmacologia , Piridinas/farmacologia , Pirróis/farmacologia , Receptores Notch/metabolismo , Fase S/efeitos dos fármacos , Fase S/fisiologia , Transdução de Sinais/efeitos dos fármacos
8.
Mol Med Rep ; 19(6): 5227-5236, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059052

RESUMO

Psoriasis is a chronic inflammatory disease characterized by the abnormal differentiation and hyperproliferation of epidermal keratinocytes. The aim of the present study was to investigate the mechanism by which microRNA­125b (miR­125b) inhibits the activation of the bromodomain­containing protein 4 (BRD4)/Notch signaling pathway in psoriasis. The contents of associated miRNAs in serum samples from 32 patients with psoriasis were detected by reverse transcription­quantitative polymerase chain reaction (RT­qPCR). The most significantly downregulated miRNA, miR­125b, was screened out. In experiments using HaCaT cells, the association between miR­125b and cell proliferation was observed using a Cell Counting Kit­8 assay, that between miR­125b and the Notch signaling pathway was observed by western blotting and RT­qPCR, and that between miR­125b and the upstream molecule BRD4 of the Notch signaling pathway was observed by luciferase reporter assay and western blotting. The proliferation of HaCaT cells became apparent following miR­125b inhibition. The Jagged­1 ligand in the Notch signaling pathway was upregulated, the active intracellular domain of the Notch1 receptor was increasingly truncated, and the Notch signaling pathway was activated. Furthermore, the inhibited miR­125b contributed directly toward the upstream protein BRD4 3'­UTR of Jagged­1, ultimately activating the Notch signaling pathway with the upregulation of Jagged­1. In conclusion, the proliferation of HaCaT cells mediated by the Jagged­1/Notch signaling pathway was decreased with the miR­125b­mediated inhibition of BRD4 expression. Therefore, miR­125b may be a biomarker and potential therapeutic target for psoriasis treatment.


Assuntos
Proliferação de Células , Proteína Jagged-1/metabolismo , MicroRNAs/metabolismo , Proteínas Nucleares/metabolismo , Psoríase/patologia , Receptores Notch/metabolismo , Fatores de Transcrição/metabolismo , Regiões 3' não Traduzidas , Adolescente , Adulto , Antagomirs/metabolismo , Linhagem Celular , Sobrevivência Celular , Feminino , Humanos , Masculino , MicroRNAs/antagonistas & inibidores , MicroRNAs/sangue , MicroRNAs/genética , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Psoríase/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Adulto Jovem
9.
Biomed Res Int ; 2019: 4647252, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31093499

RESUMO

Small-cell lung cancer (SCLC) is a highly malignant type of lung cancer with no effective second-line chemotherapy drugs. Arsenic trioxide (As2O3) was reported to exert antiangiogenesis activities against lung cancer and induce poor development of vessel structures, similar to the effect observed following the blockade of Notch signaling. However, there are no direct evidences on the inhibitory effects of As2O3 on tumor growth and angiogenesis via blockade of Notch signaling in SCLC. Here, we found that As2O3 significantly inhibited the tumor growth and angiogenesis in SCLC and reduced the microvessel density. As2O3 disturbed the morphological development of tumor vessels and downregulated the protein levels of delta-like canonical Notch ligand 4 (Dll4), Notch1, and Hes1 in vivo. DAPT, a Notch signaling inhibitor, exerted similar effects in SCLC. We found that both As2O3 treatment and Notch1 expression knockdown resulted in the interruption of tube formation by human umbilical vein endothelial cells (HUVECs) on Matrigel. As2O3 had no effects on Dll4 level in HUVECs but significantly inhibited the expression of Notch1 and its downstream gene Hes1 regardless of Dll4 overexpression or Notch1 knockdown. These findings suggest that the antitumor activity of As2O3 in SCLC was mediated via its antiangiogenic effect through the blockade of Notch signaling, probably owing to Notch1 targeting.


Assuntos
Inibidores da Angiogênese/farmacologia , Trióxido de Arsênio/farmacologia , Neoplasias Pulmonares/patologia , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos , Carcinoma de Pequenas Células do Pulmão/patologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Colágeno/farmacologia , Regulação para Baixo/efeitos dos fármacos , Combinação de Medicamentos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Laminina/farmacologia , Lentivirus/metabolismo , Neoplasias Pulmonares/irrigação sanguínea , Masculino , Camundongos Nus , Proteoglicanas/farmacologia , Carcinoma de Pequenas Células do Pulmão/irrigação sanguínea , Fatores de Transcrição HES-1/metabolismo , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Nat Commun ; 10(1): 2016, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31043605

RESUMO

Appropriate therapeutic modulation of endothelial proliferation and sprouting is essential for the effective inhibition of angiogenesis in cancer or its induction in cardiovascular disease. The current view is that an increase in growth factor concentration, and the resulting mitogenic activity, increases both endothelial proliferation and sprouting. Here, we modulate mitogenic stimuli in different vascular contexts by interfering with the function of the VEGF and Notch signalling pathways at high spatiotemporal resolution in vivo. Contrary to the prevailing view, our results indicate that high mitogenic stimulation induced by VEGF, or Notch inhibition, arrests the proliferation of angiogenic vessels. This is due to the existence of a bell-shaped dose-response to VEGF and MAPK activity that is counteracted by Notch and p21, determining whether endothelial cells sprout, proliferate, or become quiescent. The identified mechanism should be considered to achieve optimal therapeutic modulation of angiogenesis.


Assuntos
Endotélio Vascular/efeitos dos fármacos , Mitógenos/farmacologia , Neovascularização Patológica/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Endotélio Vascular/patologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Knockout , Neovascularização Patológica/patologia , Receptores Notch/antagonistas & inibidores , Receptores Notch/metabolismo , Retina , Vasos Retinianos , Transdução de Sinais/genética , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismo
11.
Nat Commun ; 10(1): 2071, 2019 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-31061501

RESUMO

Translation and transcription are frequently dysregulated in cancer. These two processes are generally regulated by distinct sets of factors. The CBFB gene, which encodes a transcription factor, has recently emerged as a highly mutated driver in a variety of human cancers including breast cancer. Here we report a noncanonical role of CBFB in translation regulation. RNA immunoprecipitation followed by deep sequencing (RIP-seq) reveals that cytoplasmic CBFB binds to hundreds of transcripts and regulates their translation. CBFB binds to mRNAs via hnRNPK and enhances translation through eIF4B, a general translation initiation factor. Interestingly, the RUNX1 mRNA, which encodes the transcriptional partner of CBFB, is bound and translationally regulated by CBFB. Furthermore, nuclear CBFB/RUNX1 complex transcriptionally represses the oncogenic NOTCH signaling pathway in breast cancer. Thus, our data reveal an unexpected function of CBFB in translation regulation and propose that breast cancer cells evade translation and transcription surveillance simultaneously through downregulating CBFB.


Assuntos
Neoplasias da Mama/genética , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade beta de Fator de Ligação ao Core/metabolismo , Regulação Neoplásica da Expressão Gênica , Ribonucleoproteínas Nucleares Heterogêneas Grupo K/metabolismo , Animais , Mama/patologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Regulação para Baixo , Fatores de Iniciação em Eucariotos/metabolismo , Feminino , Células HEK293 , Humanos , Camundongos , Camundongos Nus , RNA Mensageiro/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais/genética , Análise Serial de Tecidos , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Methods Mol Biol ; 1981: 203-236, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31016657

RESUMO

Cholangiopathies are an important group of liver diseases affecting the biliary system, and the purpose of this review is to describe how diseases in the biliary system can be studied in mouse models. A particular focus is placed on mouse models for Alagille syndrome, a cholangiopathy with a strong genetic link to dysfunctional Notch signaling. Recently, a number of different genetic mouse models based on various manipulations of the Notch signaling pathway have been generated to study Alagille syndrome, and we discuss the resulting phenotypes, and possible causes for the phenotypic heterogeneity among the various models. In the final section, we provide a more general discussion on how well mouse models can be expected to mimic human liver disease, as well as an outlook toward the need for new technologies that can help us to gain new insights from mouse models for liver disease.


Assuntos
Doenças dos Ductos Biliares/metabolismo , Modelos Animais de Doenças , Animais , Camundongos , Receptores Notch/metabolismo , Transdução de Sinais
13.
Int J Mol Sci ; 20(8)2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-31018488

RESUMO

Histopathological findings of oral neoplasm cell differentiation and metaplasia suggest that tumor cells induce their own dedifferentiation and re-differentiation and may lead to the formation of tumor-specific histological features. Notch signaling is involved in the maintenance of tissue stem cell nature and regulation of differentiation and is responsible for the cytological regulation of cell fate, morphogenesis, and/or development. In our previous study, immunohistochemistry was used to examine Notch expression using cases of odontogenic tumors and pleomorphic adenoma as oral neoplasms. According to our results, Notch signaling was specifically associated with tumor cell differentiation and metaplastic cells of developmental tissues. Notch signaling was involved in the differentiation of the ductal epithelial cells of salivary gland tumors and ameloblast-like cells of odontogenic tumors. However, Notch signaling was also involved in squamous metaplasia, irrespective of the type of developmental tissue. In odontogenic tumors, Notch signaling was involved in epithelial-mesenchymal interactions and may be related to tumor development and tumorigenesis. This signaling may also be associated with the malignant transformation of ameloblastomas. Overall, Notch signaling appears to play a major role in the formation of the characteristic cellular composition and histological features of oral neoplasms, and this involvement has been reviewed here.


Assuntos
Adenoma Pleomorfo/patologia , Transformação Celular Neoplásica/patologia , Neoplasias Bucais/patologia , Mixoma/patologia , Tumores Odontogênicos/patologia , Receptores Notch/metabolismo , Transdução de Sinais , Adenoma Pleomorfo/metabolismo , Ameloblastoma/metabolismo , Ameloblastoma/patologia , Animais , Diferenciação Celular , Transformação Celular Neoplásica/metabolismo , Humanos , Neoplasias Bucais/metabolismo , Mixoma/metabolismo , Tumores Odontogênicos/metabolismo
14.
PLoS Genet ; 15(4): e1008058, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30933982

RESUMO

In the skin and gill epidermis of fish, ionocytes develop alongside keratinocytes and maintain body fluid ionic homeostasis that is essential for adaptation to environmental fluctuations. It is known that ionocyte progenitors in zebrafish embryos are specified from p63+ epidermal stem cells through a patterning process involving DeltaC (Dlc)-Notch-mediated lateral inhibition, which selects scattered dlc+ cells into the ionocyte progenitor fate. However, mechanisms by which the ionocyte progenitor population is modulated remain unclear. Krüppel-like factor 4 (Klf4) transcription factor was previously implicated in the terminal differentiation of mammalian skin epidermis and is known for its bifunctional regulation of cell proliferation in a tissue context-dependent manner. Here, we report novel roles for zebrafish Klf4 in the ventral ectoderm during embryonic skin development. We found that Klf4 was expressed in p63+ epidermal stem cells of the ventral ectoderm from 90% epiboly onward. Knockdown or knockout of klf4 expression reduced the proliferation rate of p63+ stem cells, resulting in decreased numbers of p63+ stem cells, dlc-p63+ keratinocyte progenitors and dlc+ p63+ ionocyte progenitor cells. These reductions subsequently led to diminished keratinocyte and ionocyte densities and resulted from upregulation of the well-known cell cycle regulators, p53 and cdkn1a/p21. Moreover, mutation analyses of the KLF motif in the dlc promoter, combined with VP16-klf4 or engrailed-klf4 mRNA overexpression analyses, showed that Klf4 can bind the dlc promoter and modulate lateral inhibition by directly repressing dlc expression. This idea was further supported by observing the lateral inhibition outcomes in klf4-overexpressing or knockdown embryos. Overall, our experiments delineate novel roles for zebrafish Klf4 in regulating the ionocyte progenitor population throughout early stem cell stage to initiation of terminal differentiation, which is dependent on Dlc-Notch-mediated lateral inhibition.


Assuntos
Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Células Epidérmicas/citologia , Células Epidérmicas/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Padronização Corporal , Diferenciação Celular , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Ectoderma/citologia , Ectoderma/embriologia , Ectoderma/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Brânquias/citologia , Brânquias/embriologia , Brânquias/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Transporte de Íons , Fatores de Transcrição Kruppel-Like/deficiência , Fatores de Transcrição Kruppel-Like/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Regiões Promotoras Genéticas , Receptores Notch/genética , Receptores Notch/metabolismo , Transativadores/genética , Transativadores/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/deficiência , Proteínas de Peixe-Zebra/genética
15.
Nat Commun ; 10(1): 1839, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015398

RESUMO

Hematopoietic stem and progenitor cells (HSPCs) are capable of producing all mature blood lineages, as well as maintaining the self-renewal ability throughout life. The hairy-like organelle, cilium, is present in most types of vertebrate cells, and plays important roles in various biological processes. However, it is unclear whether and how cilia regulate HSPC development in vertebrates. Here, we show that cilia-specific genes, involved in primary cilia formation and function, are required for HSPC development, especially in hemogenic endothelium (HE) specification in zebrafish embryos. Blocking primary cilia formation or function by genetic or chemical manipulations impairs HSPC development. Mechanistically, we uncover that primary cilia in endothelial cells transduce Notch signal to the earliest HE for proper HSPC specification during embryogenesis. Altogether, our findings reveal a pivotal role of endothelial primary cilia in HSPC development, and may shed lights into in vitro directed differentiation of HSPCs.


Assuntos
Cílios/metabolismo , Células-Tronco Hematopoéticas/fisiologia , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia , Proteínas de Peixe-Zebra/metabolismo , Animais , Animais Geneticamente Modificados , Cílios/genética , Embrião não Mamífero , Desenvolvimento Embrionário/fisiologia , Hemangioblastos/citologia , Hemangioblastos/metabolismo , Hematopoese/fisiologia , Modelos Animais , Peixe-Zebra/fisiologia
16.
Cell Mol Gastroenterol Hepatol ; 7(3): 533-554, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30827941

RESUMO

BACKGROUND & AIMS: Loss of leucine-rich repeat-containing G-protein-coupled receptor 5-positive crypt base columnar cells provides permissive conditions for different facultative stem cell populations to dedifferentiate and repopulate the stem cell compartment. In this study, we used a defensin α4-Cre recombinase (Defa4Cre) line to define the potential of Paneth cells to dedifferentiate and contribute to intestinal stem cell (ISC) maintenance during normal homeostasis and after intestinal injury. METHODS: Small intestine and enteroids from Defa4Cre;Rosa26 tandem dimer Tomato (tdTomato), a red fluoresent protein, (or Rosa26 Enhanced Yellow Fluorescent Protein (EYFP)) reporter, Notch gain-of-function (Defa4Cre;Rosa26 Notch Intracellular Domain (NICD)-ires-nuclear Green Fluorescent Protein (nGFP) and Defa4Cre;Rosa26reverse tetracycline transactivator-ires Enhanced Green Fluorescent Protein (EGFP);TetONICD), A Disintegrin and Metalloproteinase domain-containing protein 10 (ADAM10) loss-of-function (Defa4Cre;ADAM10flox/flox), and Adenomatous polyposis coli (APC) inactivation (Defa4Cre;APCflox/flox) mice were analyzed. Doxorubicin treatment was used as an acute intestinal injury model. Lineage tracing, proliferation, and differentiation were assessed in vitro and in vivo. RESULTS: Defa4Cre-expressing cells are fated to become mature Paneth cells and do not contribute to ISC maintenance during normal homeostasis in vivo. However, spontaneous lineage tracing was observed in enteroids, and fluorescent-activated cell sorter-sorted Defa4Cre-marked cells showed clonogenic enteroid growth. Notch activation in Defa4Cre-expressing cells caused dedifferentiation to multipotent ISCs in vivo and was required for adenoma formation. ADAM10 deletion had no significant effect on crypt homeostasis. However, after acute doxorubicin-induced injury, Defa4Cre-expressing cells contributed to regeneration in an ADAM10-Notch-dependent manner. CONCLUSIONS: Our studies have shown that Defa4Cre-expressing Paneth cells possess cellular plasticity, can dedifferentiate into multipotent stem cells upon Notch activation, and can contribute to intestinal regeneration in an acute injury model.


Assuntos
Plasticidade Celular , Integrases/metabolismo , Intestinos/lesões , Intestinos/patologia , Celulas de Paneth/metabolismo , Receptores Notch/metabolismo , alfa-Defensinas/metabolismo , Proteína ADAM10/metabolismo , Adenoma/patologia , Proteína da Polipose Adenomatosa do Colo/metabolismo , Alelos , Animais , Desdiferenciação Celular , Linhagem da Célula , Células Clonais , Doxorrubicina , Deleção de Genes , Homeostase , Hiperplasia , Camundongos , Mitose , Células-Tronco Multipotentes/metabolismo , Organoides/crescimento & desenvolvimento , Organoides/patologia , Regeneração
17.
Genes Dev ; 33(9-10): 498-510, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30842215

RESUMO

Developmental signal transduction pathways act diversely, with context-dependent roles across systems and disease types. Glioblastomas (GBMs), which are the poorest prognosis primary brain cancers, strongly resemble developmental systems, but these growth processes have not been exploited therapeutically, likely in part due to the extreme cellular and genetic heterogeneity observed in these tumors. The role of Wnt/ßcatenin signaling in GBM stem cell (GSC) renewal and fate decisions remains controversial. Here, we report context-specific actions of Wnt/ßcatenin signaling in directing cellular fate specification and renewal. A subset of primary GBM-derived stem cells requires Wnt proteins for self-renewal, and this subset specifically relies on Wnt/ßcatenin signaling for enhanced tumor burden in xenograft models. In an orthotopic Wnt reporter model, Wnthi GBM cells (which exhibit high levels of ßcatenin signaling) are a faster-cycling, highly self-renewing stem cell pool. In contrast, Wntlo cells (with low levels of signaling) are slower cycling and have decreased self-renewing potential. Dual inhibition of Wnt/ßcatenin and Notch signaling in GSCs that express high levels of the proneural transcription factor ASCL1 leads to robust neuronal differentiation and inhibits clonogenic potential. Our work identifies new contexts for Wnt modulation for targeting stem cell differentiation and self-renewal in GBM heterogeneity, which deserve further exploration therapeutically.


Assuntos
Diferenciação Celular/genética , Células-Tronco Neoplásicas/citologia , Transdução de Sinais , Linhagem Celular Tumoral , Autorrenovação Celular/genética , Regulação Neoplásica da Expressão Gênica/genética , Glioblastoma/fisiopatologia , Humanos , Receptores Notch/genética , Receptores Notch/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo
18.
Genes Dev ; 33(9-10): 524-535, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30862660

RESUMO

The balance between proliferation and differentiation of muscle stem cells is tightly controlled, ensuring the maintenance of a cellular pool needed for muscle growth and repair. We demonstrate here that the transcriptional regulator Hes1 controls the balance between proliferation and differentiation of activated muscle stem cells in both developing and regenerating muscle. We observed that Hes1 is expressed in an oscillatory manner in activated stem cells where it drives the oscillatory expression of MyoD. MyoD expression oscillates in activated muscle stem cells from postnatal and adult muscle under various conditions: when the stem cells are dispersed in culture, when they remain associated with single muscle fibers, or when they reside in muscle biopsies. Unstable MyoD oscillations and long periods of sustained MyoD expression are observed in differentiating cells. Ablation of the Hes1 oscillator in stem cells interfered with stable MyoD oscillations and led to prolonged periods of sustained MyoD expression, resulting in increased differentiation propensity. This interfered with the maintenance of activated muscle stem cells, and impaired muscle growth and repair. We conclude that oscillatory MyoD expression allows the cells to remain in an undifferentiated and proliferative state and is required for amplification of the activated stem cell pool.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento/genética , Proteína MyoD/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Fatores de Transcrição HES-1/metabolismo , Animais , Células Cultivadas , Camundongos , Proteína MyoD/genética , Receptores Notch/metabolismo , Transdução de Sinais , Fatores de Transcrição HES-1/genética
19.
Artif Cells Nanomed Biotechnol ; 47(1): 833-843, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30862190

RESUMO

We aimed to explore the mediating role of Notch signal in macrophage polarization. This signal was knocked out from macrophages of Lyz2 cre and RBP-J flox mice. Bone marrow-derived macrophages (BMDMs) were isolated and polarized. The expressions of polarization markers in BMDMs 24 h after transfection were detected by qPCR. After Notch knockout, the expressions of M1 markers decreased but those of M2 markers increased significantly. MiR-125a/miR-99b and Spaca6 were highly and lowly expressed upon M1 and M2 polarizations, respectively. The expressions of experimental group were significantly lower than those of control group. Overexpression of miR-125a significantly promoted the expressions of M1 markers, whereas inhibited those of M2 markers. NO release in the culture supernatant of miR-125a overexpression group significantly exceeded that of control group. Transfection with miR-125a inhibitor significantly down-regulated IL-12 expression but up-regulated MR expression in BMDMs. The supernatant secreted by M1 macrophages significantly facilitated BS524 cell apoptosis, which was enhanced after miR-125a overexpression. The TNF-α expression of miR-99b overexpression group increased whereas that of MR decreased significantly. The miR-125a/miR-99b cluster contained an RBP-J specific recognition site in the first intron of initial transcript. The Notch signalling pathway promoted macrophage polarization into M1 phenotype by up-regulating miR-125a/miR-99b expression.


Assuntos
Regulação da Expressão Gênica , Macrófagos/imunologia , MicroRNAs/genética , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Biomarcadores/metabolismo , Linhagem Celular Tumoral , Técnicas de Inativação de Genes , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Ativação de Macrófagos/genética , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , MicroRNAs/antagonistas & inibidores , Fenótipo , Receptores Notch/genética , Proteínas de Plasma Seminal/genética
20.
Front Biosci (Landmark Ed) ; 24: 947-960, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30844722

RESUMO

Enhancer of zeste homolog 2 (EZH2), a catalytic component of polycomb repressive complex 2 (PRC2), epigenetically regulates chromatin structure and gene expression through trimethylation at histone H3K27 and recruitment of DNA methyltransferases for gene silencing. Despite extensive studies of the role of EZH2 in cancer progression and malignancy, increasing evidences suggest that EZH2 plays a critical role in stem cells renewal, maintenance, and differentiation into specific cell lineages. Here, we reviewed the update information regarding how EZH2 contributes to stem cell maintenance and cell lineage determination (including gonadogenesis, neurogenesis, myogenesis, osteogenesis, hematopoiesis, lymphopoiesis, adipogenesis, epidermal differentiation and hepatogenesis), and the regulation of EZH2 by phosphorylation and different signaling pathways.


Assuntos
Linhagem da Célula , Proteína Potenciadora do Homólogo 2 de Zeste/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Transdução de Sinais , Animais , Feminino , Hematopoese , Humanos , Linfopoese , Masculino , Camundongos , Desenvolvimento Muscular , Neurogênese , Oogênese , Osteogênese , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Notch/metabolismo , Fator de Transcrição STAT3/metabolismo , Espermatogênese , Células-Tronco/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA