Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 1.033
Filtrar
1.
PLoS Biol ; 17(10): e3000498, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31613879

RESUMO

During gastrulation, the pluripotent epiblast self-organizes into the 3 germ layers-endoderm, mesoderm and ectoderm, which eventually form the entire embryo. Decades of research in the mouse embryo have revealed that a signaling cascade involving the Bone Morphogenic Protein (BMP), WNT, and NODAL pathways is necessary for gastrulation. In vivo, WNT and NODAL ligands are expressed near the site of gastrulation in the posterior of the embryo, and knockout of these ligands leads to a failure to gastrulate. These data have led to the prevailing view that a signaling gradient in WNT and NODAL underlies patterning during gastrulation; however, the activities of these pathways in space and time have never been directly observed. In this study, we quantify BMP, WNT, and NODAL signaling dynamics in an in vitro model of human gastrulation. Our data suggest that BMP signaling initiates waves of WNT and NODAL signaling activity that move toward the colony center at a constant rate. Using a simple mathematical model, we show that this wave-like behavior is inconsistent with a reaction-diffusion-based Turing system, indicating that there is no stable signaling gradient of WNT/NODAL. Instead, the final signaling state is homogeneous, and spatial differences arise only from boundary effects. We further show that the durations of WNT and NODAL signaling control mesoderm differentiation, while the duration of BMP signaling controls differentiation of CDX2-positive extra-embryonic cells. The identity of these extra-embryonic cells has been controversial, and we use RNA sequencing (RNA-seq) to obtain their transcriptomes and show that they closely resemble human trophoblast cells in vivo. The domain of BMP signaling is identical to the domain of differentiation of these trophoblast-like cells; however, neither WNT nor NODAL forms a spatial pattern that maps directly to the mesodermal region, suggesting that mesoderm differentiation is controlled dynamically by the combinatorial effect of multiple signals. We synthesize our data into a mathematical model that accurately recapitulates signaling dynamics and predicts cell fate patterning upon chemical and physical perturbations. Taken together, our study shows that the dynamics of signaling events in the BMP, WNT, and NODAL cascade in the absence of a stable signaling gradient control fate patterning of human gastruloids.


Assuntos
Proteína Morfogenética Óssea 4/genética , Gastrulação/genética , Mesoderma/metabolismo , Proteína Nodal/genética , Transdução de Sinais , Proteínas Wnt/genética , Benzotiazóis/farmacologia , Padronização Corporal/efeitos dos fármacos , Padronização Corporal/genética , Proteína Morfogenética Óssea 4/metabolismo , Proteína Morfogenética Óssea 4/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Gástrula/citologia , Gástrula/efeitos dos fármacos , Gástrula/metabolismo , Gastrulação/efeitos dos fármacos , Regulação da Expressão Gênica , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Mesoderma/citologia , Mesoderma/efeitos dos fármacos , Modelos Biológicos , Modelos Estatísticos , Proteína Nodal/deficiência , Organoides/citologia , Organoides/efeitos dos fármacos , Organoides/metabolismo , Proteínas Wnt/metabolismo
2.
Int J Mol Sci ; 20(20)2019 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-31614900

RESUMO

Thrombin is an essential procoagulant and profibrotic mediator. However, its implication in tuberculous pleural effusion (TBPE) remains unknown. The effusion thrombin and plasminogen activator inhibitor-1 (PAI-1) levels were measured among transudative pleural effusion (TPE, n = 22) and TBPE (n = 24) patients. Pleural fibrosis, identified as radiological residual pleural thickening (RPT) and shadowing, was measured at 12-month follow-up. Moreover, in vivo and in vitro effects of thrombin on PAI-1 expression and mesothelial-mesenchymal transition (MMT) were assessed. We demonstrated the effusion thrombin levels were significantly higher in TBPE than TPE, especially greater in TBPE patients with RPT > 10mm than those without, and correlated positively with PAI-1 and pleural fibrosis area. In carbon black/bleomycin-treated mice, knockdown of protease-activated receptor-1 (PAR-1) markedly downregulated α-smooth muscle actin (α-SMA) and fibronectin, and attenuated pleural fibrosis. In pleural mesothelial cells (PMCs), thrombin concentration-dependently increased PAI-1, α-SMA, and collagen I expression. Specifically, Mycobacterium tuberculosis H37Ra (MTBRa) induced thrombin production by PMCs via upregulating tissue factor and prothrombin, and PAR-1 silencing considerably abrogated MTBRa-stimulated PAI-1 expression and MMT. Consistently, prothrombin/PAR-1 expression was evident in the pleural mesothelium of TBPE patients. Conclusively, thrombin upregulates PAI-1 and MMT and may contribute to tuberculous pleural fibrosis. Thrombin/PAR-1 inhibition may confer potential therapy for pleural fibrosis.


Assuntos
Inibidor 1 de Ativador de Plasminogênio/metabolismo , Pleura/patologia , Receptor PAR-1/metabolismo , Trombina/metabolismo , Tuberculose/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Modelos Animais de Doenças , Exsudatos e Transudatos/metabolismo , Feminino , Fibrose , Seguimentos , Humanos , Masculino , Mesoderma/efeitos dos fármacos , Mesoderma/crescimento & desenvolvimento , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Mycobacterium tuberculosis/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Derrame Pleural/metabolismo , Derrame Pleural/patologia , Transdução de Sinais , Tuberculose/patologia , Adulto Jovem
3.
Biomed Pharmacother ; 118: 109227, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31351433

RESUMO

Endothelial-to-mesenchymal transition (EndMT) is closely related to the pathogenesis of various diseases, including cardiac fibrosis. Transforming growth factor (TGF)-ß1 strongly induces EndMT, and sirtuin 1 (SIRT1) may play vital roles in TGF-ß/Smad pathway inhibition. This study aimed to determine whether SIRT1 activation inhibits EndMT, thereby attenuating cardiac fibrosis. Cardiac fibrosis was induced in C57BL/6 mice by subcutaneously injecting isoproterenol. SIRT1 was activated and then suppressed by intraperitoneally injecting resveratrol (RSV) and EX527, respectively. EndMT was induced by adding TGF-ß1 to H5V cells and measured by immunofluorescence and western blot. The role of SIRT1 in EndMT was determined by lentivirus-mediated overexpression of SIRT1. Interactions between SIRT1 and Smad2/3 in the TGF-ß/Smad2/3 pathway were examined by immunoprecipitation. SIRT1 activation upregulated CD31 and vascular endothelial-cadherin, and downregulated α-smooth muscle actin, fibroblast-specific protein 1, and vimentin. SIRT1 upregulated and EX527 inhibited TGF-ß receptor 1 (TGF-ßR1) and P-Smad2/3 expression, respectively. SIRT1 activation and overexpression by RSV/SRT2104 and lentivirus transfection, respectively, reduced TGF-ß1-induced EndMT. SIRT1 and Smad2/3 interaction was shown by immunoprecipitation in vivo and in vitro. TGF-ßR1 and P-Smad2/3 expression was downregulated and Smad2/3 nuclear translocation was inhibited. In conclusion, SIRT1 activated by RSV attenuated isoproterenol-induced cardiac fibrosis by regulating EndMT via the TGF-ß/Smad2/3 pathway.


Assuntos
Endotélio/patologia , Mesoderma/patologia , Miocárdio/patologia , Sirtuína 1/metabolismo , Animais , Cardiomegalia/tratamento farmacológico , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Colágeno/metabolismo , Regulação para Baixo/efeitos dos fármacos , Endotélio/efeitos dos fármacos , Fibrose , Isoproterenol , Masculino , Mesoderma/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Resveratrol/farmacologia , Resveratrol/uso terapêutico , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo
4.
Cells ; 8(6)2019 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-31207939

RESUMO

Hyperglycaemia causes endothelial dysfunction, which is the initial process in the development of diabetic vascular complications. Upon injury, endothelial cells undergo an endothelial-to-mesenchymal transition (EndMT), lose their specific marker, and gain mesenchymal phenotypes. This study investigated the effect of liraglutide, a glucagon-like peptide 1 (GLP-1) receptor agonist, on EndMT inhibition and neointima formation in diabetic mice induced by streptozotocin. The diabetic mice with a wire-induced vascular injury in the right carotid artery were treated with or without liraglutide for four weeks. The degree of neointima formation and re-endothelialisation was evaluated by histological assessments. Endothelial fate tracing revealed that endothelium-derived cells contribute to neointima formation through EndMT in vivo. In the diabetic mouse model, liraglutide attenuated wire injury-induced neointima formation and accelerated re-endothelialisation. In vitro, a high glucose condition (30 mmol/L) triggered morphological changes and mesenchymal marker expression in human umbilical vein endothelial cells (HUVECs), which were attenuated by liraglutide or Activin receptor-like 5 (ALK5) inhibitor SB431542. The inhibition of AMP-activated protein kinase (AMPK) signaling by Compound C diminished the liraglutide-mediated inhibitory effect on EndMT. Collectively, liraglutide was found to attenuate neointima formation in diabetic mice partially through EndMT inhibition, extending the potential therapeutic role of liraglutide.


Assuntos
Diabetes Mellitus Experimental/patologia , Endotélio/patologia , Liraglutida/farmacologia , Mesoderma/patologia , Neointima/patologia , Adenilato Quinase/metabolismo , Animais , Artérias/efeitos dos fármacos , Artérias/lesões , Artérias/patologia , Biomarcadores/metabolismo , Endotélio/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/antagonistas & inibidores , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Glucose/toxicidade , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Interleucina-1beta/farmacologia , Mesoderma/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Smad2/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Estreptozocina
5.
Endocrinology ; 160(8): 1868-1884, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31107524

RESUMO

Preterm birth is characterized by severe lung immaturity that is frequently treated antenatally or postnatally with the synthetic steroid betamethasone. The underlying cellular targets and pathways stimulated by betamethasone in the fetal lung are poorly defined. In this study, betamethasone was compared with corticosterone in steroid-treated primary cultures of fetal rat lung fibroblasts stimulated for 6 hours and analyzed by whole-cell transcriptome sequencing and glucocorticoid (GC) receptor (GR) chromatin immunoprecipitation sequencing (ChIP-Seq) analysis. Strikingly, betamethasone stimulated a much stronger transcriptional response compared with corticosterone for both induced and repressed genes. A total of 483 genes were significantly stimulated by betamethasone or corticosterone, with 476 stimulated by both steroids, indicating a strong overlap in regulation. Changes in mRNA levels were confirmed by quantitative PCR for eight induced and repressed target genes. Pathway analysis identified cell proliferation and cytoskeletal/cell matrix remodeling pathways as key processes regulated by both steroids. One target, transglutaminase 2 (Tgm2), was localized to fetal lung mesenchymal cells. Tgm2 mRNA and protein levels were strongly increased in fibroblasts by both steroids. Whole-genome GR ChIP-Seq analysis with betamethasone identified GC response element-binding sites close to the previously characterized GR target genes Per1, Dusp1, Fkbp5, and Sgk1 and near the genes identified by transcriptome sequencing encoding Crispld2, Tgm2, Hif3α, and Kdr, defining direct genomic induction of expression in fetal lung fibroblasts via the GR. These results demonstrate that betamethasone stimulates specific genes and cellular pathways controlling cell proliferation and extracellular matrix remodeling in lung mesenchymal fibroblasts, providing a basis for betamethasone's treatment efficacy in preterm birth.


Assuntos
Betametasona/farmacologia , Fibroblastos/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Mesoderma/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Corticosterona/farmacologia , Feminino , Perfilação da Expressão Gênica , Pulmão/citologia , Pulmão/metabolismo , Sistema de Sinalização das MAP Quinases , Ratos , Ratos Sprague-Dawley , Receptores de Glucocorticoides/fisiologia , Transdução de Sinais , Transglutaminases/análise
6.
Int J Mol Sci ; 20(8)2019 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-31010006

RESUMO

Vincristine is used in the clinical treatment of colon cancer, especially in patients diagnosed in the advanced phase of cancer development. Unfortunately, similar to other agents used during antitumor therapy, vincristine might induce chemoresistance. Studies of this process focus mainly on the analysis of the molecular mechanisms within cancer, usually ignoring the role of stromal cells. Our present findings confirm that vincristine stimulates the secretion of tumor growth factors class beta and interleukin-6 from cancer-associated fibroblasts as a result of paracrine stimulation by cancer cells. Based on alterations in morphology, modulation of capillary formation, and changes in endothelial and mesenchymal marker profile, our findings demonstrate that higher levels of tumor growth factor-ßs and interleukin-6 enhance cancer-associated fibroblast-like cell formation through endothelial-mesenchymal transition and that nonsteroidal anti-inflammatory drug treatment (aspirin and ibuprofen) is able to inhibit this phenomenon. The process appears to be regulated by the rate of microtubule polymerization, depending on ß-tubulin composition. While higher levels of tubulin-ß2 and tubulin-ß4 caused slowed polymerization and reduced the level of factors secreted to the extracellular matrix, tubulin-ß3 induced the opposite effect. We conclude that nonsteroidal anti-inflammatory drugs should be considered for use during vincristine monotherapy in the treatment of patients diagnosed with colorectal cancer.


Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Fibroblastos Associados a Câncer/patologia , Vincristina/farmacologia , Fibroblastos Associados a Câncer/efeitos dos fármacos , Fibroblastos Associados a Câncer/metabolismo , Comunicação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Transdiferenciação Celular/efeitos dos fármacos , Neoplasias do Colo/patologia , Meios de Cultivo Condicionados/farmacologia , Citocinas/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Endotélio/efeitos dos fármacos , Endotélio/patologia , Humanos , Mesoderma/efeitos dos fármacos , Mesoderma/patologia , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Polimerização , Tubulina (Proteína)/metabolismo
7.
Toxicol Lett ; 309: 51-58, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30946857

RESUMO

Arsenic trioxide (ATO) has been recommended as the first-line agent for the treatment of acute promyelocytic leukaemia (APL), due to its substantial anticancer effect. Numerous clinical reports have indicated that ATO is a developmental toxicant which can result in birth defects of human beings. But whether arsenic trioxide can lead to human cardiac developmental toxicity remains largely unknown. So the present study aims to explore the influence and mechanisms of ATO on human cardiac development by using a vitro cardiac differentiation model of human induced pluripotent stem cells (hiPSCs). Here we found that clinically achievable concentrations (0.1, 0.5 and 1 µM) of ATO resulted in a significant inhibition of proliferation during the whole process of cardiac differentiation of hiPSCs. Meanwhile, TUNEL assay revealed that ATO could cause cell apoptosis during cardiac differentiation in a concentration-dependent manner. Consistently, we found that ATO reduced the expressions of mesoderm markers Brachyury and EOMES, cardiac progenitor cell markers GATA-4, MESP-1 and TBX-5, and cardiac specific marker α-actinin in differentiated hiPSCs. Furthermore, ATO treatment had caused DNA damage which was shown in the upregulation of γH2AX, a sensitive marker for DNA double-strand breaks. Taken together, ATO blocked cardiomyocyte differentiation, induced apoptosis and cell growth arrest during cardiac differentiation of hiPSCs, which might be associated with DNA damage.


Assuntos
Trióxido de Arsênio/toxicidade , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Dano ao DNA , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Mesoderma/citologia , Mesoderma/efeitos dos fármacos , Miócitos Cardíacos/citologia
8.
Nano Lett ; 19(4): 2280-2290, 2019 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-30775927

RESUMO

Cancer cell invasion through physical barriers in the extracellular matrix (ECM) requires a complex synergy of traction force against the ECM, mechanosensitive feedback, and subsequent cytoskeletal rearrangement. PDMS microchannels were used to investigate the transition from mesenchymal to amoeboid invasion in cancer cells. Migration was faster in narrow 3 µm-wide channels than in wider 10 µm channels, even in the absence of cell-binding ECM proteins. Cells permeating narrow channels exhibited blebbing and had smooth leading edge profiles, suggesting an ECM-induced transition from mesenchymal invasion to amoeboid invasion. Live cell labeling revealed a mechanosensing period in which the cell attempts mesenchymal-based migration, reorganizes its cytoskeleton, and proceeds using an amoeboid phenotype. Rho/ROCK (amoeboid) and Rac (mesenchymal) pathway inhibition revealed that amoeboid invasion through confined environments relies on both pathways in a time- and ECM-dependent manner. This demonstrates that cancer cells can dynamically modify their invasion programming to navigate physically confining matrix conditions.


Assuntos
Citoesqueleto/efeitos dos fármacos , Mesoderma/efeitos dos fármacos , Invasividade Neoplásica/genética , Neoplasias/genética , Fenômenos Biomecânicos , Adesão Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Citoesqueleto/genética , Dimetilpolisiloxanos/química , Dimetilpolisiloxanos/farmacologia , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/genética , Humanos , Mesoderma/patologia , Invasividade Neoplásica/patologia , Neoplasias/patologia , Nylons/química , Nylons/farmacologia
9.
Life Sci ; 207: 442-450, 2018 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-29969608

RESUMO

AIMS: Endothelial-to-mesenchymal transition (EndoMT) was shown to lead to endothelial cell (EC) dysfunction in pulmonary arterial hypertension (PAH). Baicalein was reported to inhibit epithelial-to-mesenchymal transition (EMT), a biological process that has many regulatory pathways in common with EndoMT. Whether it can attenuate PAH by inhibiting EndoMT remains obscure. MAIN METHODS: PAH was induced by a single subcutaneous injection of MCT (60 mg/kg) in male Sprague Dawley rats. Two weeks after MCT administration, the rats in the treatment groups received baicalein orally (50 or 100 mg/kg/day) for an additional 2 weeks. Hemodynamic changes and right ventricular hypertrophy (RVH) were evaluated on day 28. Cardiopulmonary interstitial fibrosis was detected using Masson's trichrome, Picrosirius-red, and immunohistochemical staining. The reactivity of pulmonary arteries (PAs) was examined ex vivo. The protein expresson of EndoMT molecules, bone morphogenetic protein receptor 2 (BMPR2), and nuclear factor-κB (NF-κB) was examined to explore the mechanism of protective action of baicalein. KEY FINDINGS: Baicalein (50 and 100 mg/kg) significantly alleviated MCT-induced PAH and cardiopulmonary interstitial fibrosis. Furthermore, baicalein treatment enhanced PA responsiveness to acetylcholine (ACh) in PAH rats. The upregulation of EndoMT molecules (N-cadherin, vimentin, Snail, and Slug) strongly suggest that EndoMT participates in MCT-induced PAH, which was reversed by baicalein (50 and 100 mg/kg) treatment. Moreover, baicalein partially reversed MCT-induced reductions in BMPR2 and NF-κB activation in the PAs. SIGNIFICANCE: Baicalein attenuated MCT-induced PAH in rats by inhibiting EndoMT partially via the NF-κB-BMPR2 pathway. Thus, baicalein might be considered as a promising treatment option for PAH.


Assuntos
Endotélio/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Flavanonas/farmacologia , Hipertensão Pulmonar/tratamento farmacológico , Mesoderma/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Inibidores Enzimáticos/farmacologia , Hemodinâmica , Hipertensão Pulmonar/induzido quimicamente , Masculino , Monocrotalina , NF-kappa B/metabolismo , Ratos , Ratos Sprague-Dawley , Regulação para Cima
10.
Development ; 145(15)2018 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-29980563

RESUMO

The larval pharynx of the cephalochordate Branchiostoma (amphioxus) is asymmetrical. The mouth is on the left, and endostyle and gill slits are on the right. At the neurula, Nodal and Hedgehog (Hh) expression becomes restricted to the left. To dissect their respective roles in gill slit formation, we inhibited each pathway separately for 20 min at intervals during the neurula stage, before gill slits penetrate, and monitored the effects on morphology and expression of pharyngeal markers. The results pinpoint the short interval spanning the gastrula/neurula transition as the critical period for specification and positioning of future gill slits. Thus, reduced Nodal signaling shifts the gill slits ventrally, skews the pharyngeal domains of Hh, Pax1/9, Pax2/5/8, Six1/2 and IrxC towards the left, and reduces Hh and Tbx1/10 expression in endoderm and mesoderm, respectively. Nodal auto-regulates. Decreased Hh signaling does not affect gill slit positions or Hh or Nodal expression, but it does reduce the domain of Gli, the Hh target, in the pharyngeal endoderm. Thus, during the neurula stage, Nodal and Hh cooperate in gill slit development - Hh mediates gill slit formation and Nodal establishes their left-right position.


Assuntos
Padronização Corporal , Brânquias/metabolismo , Proteínas Hedgehog/metabolismo , Anfioxos/embriologia , Anfioxos/metabolismo , Proteína Nodal/metabolismo , Animais , Benzodioxóis/farmacologia , Padronização Corporal/efeitos dos fármacos , Padronização Corporal/genética , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Epistasia Genética/efeitos dos fármacos , Gástrula/efeitos dos fármacos , Gástrula/embriologia , Gástrula/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Brânquias/efeitos dos fármacos , Proteínas Hedgehog/genética , Imidazóis/farmacologia , Anfioxos/efeitos dos fármacos , Anfioxos/genética , Larva/efeitos dos fármacos , Larva/metabolismo , Mesoderma/efeitos dos fármacos , Mesoderma/embriologia , Mesoderma/metabolismo , Proteína Nodal/genética , Faringe/efeitos dos fármacos , Faringe/embriologia , Faringe/metabolismo , Piridinas/farmacologia , Alcaloides de Veratrum/farmacologia
11.
Mol Oncol ; 12(10): 1689-1705, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30051594

RESUMO

Ovarian cancer (OC) is one of the most intractable diseases, exhibiting tremendous molecular heterogeneity and lacking reliable methods for screening, resulting in late diagnosis and widespread peritoneal dissemination. Menopausal estrogen replacement therapy is a well-recognized risk factor for OC, but little is known about how estrogen might contribute to this disease at the cellular level. This study identifies chemokine receptor CXCR7/ACKR3 as an estrogen-responsive gene, whose expression is markedly enhanced by estrogen through direct recruitment of ERα and transcriptional active histone modifications in OC cells. The gene encoding CXCR7 chemokine ligand I-TAC/CXCL11 was also upregulated by estrogen, resulting in Ser-118 phosphorylation, activation, and recruitment of estrogen receptor ERα at the CXCR7 promoter locus for positive feedback regulation. Both CXCR7 and CXCL11, but not CXCR3 (also recognized to interact with CXCL11), were found to be significantly increased in stromal sections of microdissected tumors and positively correlated in mesenchymal subtype of OC. Estrogenic induction of mesenchymal markers SNAI1, SNAI2, and CDH2 expression, with a consequent increase in cancer cell migration, was shown to depend on CXCR7, indicating a key role for CXCR7 in mediating estrogen upregulation of mesenchymal markers to induce invasion of OC cells. These findings identify a feed-forward mechanism that sustains activation of the CXCR7/CXCL11 axis under ERα control to induce the epithelial-mesenchymal transition pathway and metastatic behavior of OC cells. Such interplay underlies the complex gene profile heterogeneity of OC that promotes changes in tumor microenvironment and metastatic acquisition.


Assuntos
Quimiocina CXCL11/metabolismo , Receptor alfa de Estrogênio/metabolismo , Retroalimentação Fisiológica , Neoplasias Ovarianas/metabolismo , Receptores CXCR/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Cromatina/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Estrogênios/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Loci Gênicos , Humanos , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Mesoderma/patologia , Neoplasias Ovarianas/patologia , Fosforilação/efeitos dos fármacos , Fosfosserina/metabolismo , Receptores CXCR/genética , Elementos de Resposta/genética , Transdução de Sinais/efeitos dos fármacos , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Transcrição Genética/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
12.
Cell Rep ; 23(6): 1651-1664, 2018 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-29742423

RESUMO

Cancer stem cells promote neoplastic growth, in part by deregulating asymmetric cell division and enhancing self-renewal. To uncover mechanisms and potential therapeutic targets in glioma stem cell (GSC) self-renewal, we performed a genetic suppressor screen for kinases to reverse the tumor phenotype of our Drosophila brain tumor model and identified dCdk5 as a critical regulator. CDK5, the human ortholog of dCdk5 (79% identity), is aberrantly activated in GBMs and tightly aligned with both chromosome 7 gains and stem cell markers affecting tumor-propagation. Our investigation revealed that pharmaceutical inhibition of CDK5 prevents GSC self-renewal in vitro and in xenografted tumors, at least partially by suppressing CREB1 activation independently of PKA/cAMP. Finally, our TCGA GBM data analysis revealed that CDK5, stem cell, and asymmetric cell division markers segregate within non-mesenchymal patient clusters, which may indicate preferential dependence on CDK5 signaling and sensitivity to its inhibition in this group.


Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Quinase 5 Dependente de Ciclina/antagonistas & inibidores , Proteínas de Drosophila/antagonistas & inibidores , Glioma/metabolismo , Glioma/patologia , Células-Tronco Neoplásicas/patologia , Transdução de Sinais , Animais , Divisão Celular Assimétrica/efeitos dos fármacos , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Proliferação de Células/efeitos dos fármacos , Autorrenovação Celular/efeitos dos fármacos , Quinase 5 Dependente de Ciclina/genética , Quinase 5 Dependente de Ciclina/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioma/genética , Mesoderma/efeitos dos fármacos , Mesoderma/patologia , Camundongos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Fosforilação/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Sci Rep ; 8(1): 6618, 2018 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-29700394

RESUMO

Bioactive lipids such as sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) regulate diverse processes including cell proliferation, differentiation, and migration. However, their roles in cardiac differentiation and cardiomyocyte proliferation have not been explored. Using a 96-well differentiation platform for generating human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) we found that S1P and LPA can independently enhance cardiomyocyte generation when administered at an early stage of differentiation. We showed that the combined S1P and LPA treatment of undifferentiated hiPSCs resulted in increased nuclear accumulation of ß-catenin, the canonical Wnt signaling pathway mediator, and synergized with CHIR99021, a glycogen synthase kinase 3 beta inhibitor, to enhance mesodermal induction and subsequent cardiac differentiation. At later stages of cardiac differentiation, the addition of S1P and LPA resulted in cell cycle initiation in hiPSC-CMs, an effect mediated through increased ERK signaling. Although the addition of S1P and LPA alone was insufficient to induce cell division, it was able to enhance ß-catenin-mediated hiPSC-CM proliferation. In summary, we demonstrated a developmental stage-specific effect of bioactive lipids to enhance hiPSC-CM differentiation and proliferation via modulating the effect of canonical Wnt/ß-catenin and ERK signaling. These findings may improve hiPSC-CM generation for cardiac disease modeling, precision medicine, and regenerative therapies.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Lipídeos/farmacologia , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Biomarcadores , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Perfilação da Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Mesoderma/citologia , Mesoderma/efeitos dos fármacos , Modelos Biológicos , Miócitos Cardíacos/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos
14.
Biochem Biophys Res Commun ; 501(4): 827-832, 2018 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-29654764

RESUMO

Recent study has showed that Ginsenoside Rg1, the mian active compound of Panax ginseng, could ameliorate oxidative stress and myocardial apoptosis in diabetes mellitus. However, the roles and mechanisms of Rg1 in proliferative diabetic retinopathy (PDR) are still unclear. In the present study, we aimed to investigate the effects of Rg1 on mesenchymal activation of high-glucose (HG) cultured müller cells. High glucose conditions up-regulate MMP-2, MMP-9 and down-regulate TIMP-2, and promote mesenchymal activation in Müller cells. And Rg1 inhibits the HG-induced mesenchymal activation and HG-increased MMP-2 and MMP-9 and HG-decreased TIMP-2 in Müller cells. HG up-regulates Zeb1 and lncRNA RP11-982M15.8, and down-regulates miR-2113, and Rg1 inhibits these effects of HG. Both inhibition of miR-2113 and over-expression of RP11-982M15.8 significantly restored the HG induced mesenchymal activasion. Taken together, our findings suggested that Rg1 inhibited HG-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway.


Assuntos
Ginsenosídeos/farmacologia , Glucose/toxicidade , Mesoderma/metabolismo , Mesoderma/patologia , MicroRNAs/genética , RNA Longo não Codificante/genética , Transdução de Sinais/efeitos dos fármacos , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Células Ependimogliais/efeitos dos fármacos , Células Ependimogliais/metabolismo , Fibrose , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz , Mesoderma/efeitos dos fármacos , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
15.
In Vitro Cell Dev Biol Anim ; 54(3): 231-240, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29435726

RESUMO

Thalidomide was once administered to pregnant women as a mild sedative; however, it was subsequently shown to be strongly teratogenic. Recently, there has been renewed interest in thalidomide because of its curative effects against intractable diseases. However, the teratogenicity of thalidomide is manifested in various ways and is still not fully understood. In the present study, we evaluated the effects of thalidomide on early mesodermal differentiation by examining the differentiation of human induced pluripotent stem cells (hiPSCs). The most common symptom of thalidomide teratogenicity is limb abnormality, which led us to hypothesize that thalidomide prevents early mesodermal differentiation. Therefore, mesodermal differentiation of hiPSCs was induced over a 6-d period. To induce early mesoderm differentiation, 1 d after seeding, the cells were incubated with the small molecule compound CHIR99021 for 3 d. Thalidomide exposure was initiated at the same time as CHIR99021 treatment. After 5 d of thalidomide exposure, the hiPSCs began expressing a mesodermal marker; however, the number of viable cells decreased significantly as compared to that of control cells. We observed that the proportion of apoptotic and dead cells increased on day 2; however, the proportion of dead cells on day 5 had decreased, suggesting that the cells were damaged by thalidomide during early mesodermal differentiation (days 0-2). Our findings may help elucidate the mechanism underlying thalidomide teratogenicity and bring us closer to the safe use of this drug.


Assuntos
Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/patologia , Mesoderma/patologia , Teratogênios/farmacologia , Talidomida/farmacologia , Células Cultivadas , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Mesoderma/efeitos dos fármacos
16.
Sci Rep ; 8(1): 2433, 2018 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-29402947

RESUMO

During the gastrulation stage in animal embryogenesis, the cells leading the axial mesoderm migrate toward the anterior side of the embryo, vigorously extending cell protrusions such as lamellipodia. It is thought that the leading cells sense gradients of chemoattractants emanating from the ectodermal cells and translate them to initiate and maintain the cell movements necessary for gastrulation. However, it is unclear how the extracellular information is converted to the intracellular chemical reactions that lead to motion. Here we demonstrated that intracellular Ca2+ levels in the protrusion-forming leading cells are markedly higher than those of the following cells and the axial mesoderm cells. We also showed that inhibiting the intracellular Ca2+ significantly retarded the gastrulation cell movements, while increasing the intracellular Ca2+ with an ionophore enhanced the migration. We further found that the ionophore treatment increased the active form of the small GTPase Rac1 in these cells. Our results suggest that transient intracellular Ca2+ signals play an essential role in the active cell migration during gastrulation.


Assuntos
Sinalização do Cálcio , Cálcio/metabolismo , Gastrulação/fisiologia , Mesoderma/metabolismo , Xenopus laevis/metabolismo , Animais , Ionóforos de Cálcio/farmacologia , Movimento Celular/efeitos dos fármacos , Quelantes/farmacologia , Ectoderma/citologia , Ectoderma/efeitos dos fármacos , Ectoderma/metabolismo , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Embrião não Mamífero , Gástrula/citologia , Gástrula/efeitos dos fármacos , Gástrula/metabolismo , Gastrulação/efeitos dos fármacos , Expressão Gênica , Ionomicina/farmacologia , Mesoderma/citologia , Mesoderma/efeitos dos fármacos , Pseudópodes/efeitos dos fármacos , Pseudópodes/metabolismo , Pseudópodes/ultraestrutura , Xenopus laevis/crescimento & desenvolvimento , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
17.
Mol Neurobiol ; 55(8): 6816-6833, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29349577

RESUMO

Glioblastoma (GBM) is the most common and aggressive malignant primary brain tumor and still lacks effective therapeutic strategies. It has already been shown that old drugs like sulfasalazine (SAS) and valproic acid (VPA) present antitumoral activities in glioma cell lines. SAS has also been associated with a decrease of intracellular glutathione (GSH) levels through a potent inhibition of xc- glutamate/cystine exchanger leading to an antioxidant deprotection. In the same way, VPA was recently identified as a histone deacetylase (HDAT) inhibitor capable of activating tumor suppression genes. As both drugs are widely used in clinical practice and their profile of adverse effects is well known, the aim of our study was to investigate the effects of the combined treatment with SAS and VPA in GBM cell lines. We observed that both drugs were able to reduce cell viability in a dose-dependent manner and the combined treatment potentiated these effects. Combined treatment also increased cell death and inhibited proliferation of GBM cells, while having no effect on human and rat cultured astrocytes. Also, we observed high protein expression of the catalytic subunit of xc- in all the examined GBM cell lines, and treatment with SAS blocked its activity and decreased intracellular GSH levels. Noteworthy, SAS but not VPA was also able to reduce the [14C]-ascorbate uptake. Together, these data indicate that SAS and VPA exhibit a substantial effect on GBM cell's death related to an intracellular oxidative response imbalance, making this combination of drugs a promising therapeutic strategy.


Assuntos
Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Espaço Intracelular/metabolismo , Sulfassalazina/farmacologia , Ácido Valproico/farmacologia , Sistema y+ de Transporte de Aminoácidos/metabolismo , Animais , Ácido Ascórbico/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Quimioterapia Combinada , Glutationa/metabolismo , Humanos , Mesoderma/efeitos dos fármacos , Mesoderma/patologia , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Neuroglia/patologia , Oxirredução , Ratos , Fatores de Tempo
18.
J Cell Mol Med ; 22(2): 808-822, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29063670

RESUMO

Emerging evidence indicates that irisin provides beneficial effects in diabetes. However, whether irisin influences the development of diabetic cardiomyopathy (DCM) remains unclear. Therefore, we investigated the potential role and mechanism of action of irisin in diabetes-induced myocardial dysfunction in mice. Type 1 diabetes was induced in mice by injecting streptozotocin, and the diabetic mice were administered recombinant r-irisin (low or high dose: 0.5 or 1.5 µg/g body weight/day, I.P.) or PBS for 16 weeks. Irisin treatment did not alter blood glucose levels in the diabetic mice. However, the results of echocardiographical and histopathological assays indicated that low-dose irisin treatment alleviated cardiac fibrosis and left ventricular function in the diabetic mice, whereas high-dose irisin failed to mitigate the ventricular function impairment and increased collagen deposition. The potential mechanism underlying the effect of low-dose irisin involved irisin-mediated inhibition of high glucose-induced endothelial-to-mesenchymal transition (EndMT); conversely, high-dose irisin treatment enhanced high glucose-induced MMP expression by stimulating MAPK (p38 and ERK) signalling and cardiac fibroblast proliferation and migration. Low-dose irisin alleviated DCM development by inhibiting high glucose-induced EndMT. By contrast, high-dose irisin disrupted normal MMP expression and induced cardiac fibroblast proliferation and migration, which results in excess collagen deposition. Thus, irisin can inhibit high glucose-induced EndMT and exert a dose-dependent bidirectional effect on DCM.


Assuntos
Cardiomiopatias Diabéticas/patologia , Fibronectinas/farmacologia , Glucose/toxicidade , Células Endoteliais da Veia Umbilical Humana/patologia , Mesoderma/patologia , Animais , Glicemia/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colágeno/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/fisiopatologia , Cardiomiopatias Diabéticas/sangue , Cardiomiopatias Diabéticas/fisiopatologia , Ativação Enzimática/efeitos dos fármacos , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Mesoderma/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , Miocárdio/patologia , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Estreptozocina , Fator de Crescimento Transformador beta/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
Nature ; 551(7679): 247-250, 2017 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-29088702

RESUMO

Acquired drug resistance prevents cancer therapies from achieving stable and complete responses. Emerging evidence implicates a key role for non-mutational drug resistance mechanisms underlying the survival of residual cancer 'persister' cells. The persister cell pool constitutes a reservoir from which drug-resistant tumours may emerge. Targeting persister cells therefore presents a therapeutic opportunity to impede tumour relapse. We previously found that cancer cells in a high mesenchymal therapy-resistant cell state are dependent on the lipid hydroperoxidase GPX4 for survival. Here we show that a similar therapy-resistant cell state underlies the behaviour of persister cells derived from a wide range of cancers and drug treatments. Consequently, we demonstrate that persister cells acquire a dependency on GPX4. Loss of GPX4 function results in selective persister cell ferroptotic death in vitro and prevents tumour relapse in mice. These findings suggest that targeting of GPX4 may represent a therapeutic strategy to prevent acquired drug resistance.


Assuntos
Apoptose/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Glutationa Peroxidase/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Animais , Antioxidantes/metabolismo , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Ferro/metabolismo , Masculino , Mesoderma/efeitos dos fármacos , Mesoderma/enzimologia , Mesoderma/patologia , Camundongos , Terapia de Alvo Molecular , Neoplasias/enzimologia , Recidiva , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Nucleic Acids Res ; 45(16): 9219-9228, 2017 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-28934500

RESUMO

Targeted differentiation of human induced pluripotent stem cells (hiPSCs) using only chemicals would have value-added clinical potential in the regeneration of complex cell types including cardiomyocytes. Despite the availability of several chemical inhibitors targeting proteins involved in signaling pathways, no bioactive synthetic DNA-binding inhibitors, targeting key cell fate-controlling genes such as SOX2, are yet available. Here, we demonstrate a novel DNA-based chemical approach to guide the differentiation of hiPSCs using pyrrole-imidazole polyamides (PIPs), which are sequence-selective DNA-binding synthetic molecules. Harnessing knowledge about key transcriptional changes during the induction of cardiomyocyte, we developed a DNA-binding inhibitor termed PIP-S2, targeting the 5'-CTTTGTT-3' and demonstrated that inhibition of SOX2-DNA interaction by PIP-S2 triggers the mesoderm induction in hiPSCs. Genome-wide gene expression analyses revealed that PIP-S2 induced mesoderm by targeted alterations in SOX2-associated gene regulatory networks. Also, employment of PIP-S2 along with a Wnt/ß-catenin inhibitor successfully generated spontaneously contracting cardiomyocytes, validating our concept that DNA-binding inhibitors could drive the directed differentiation of hiPSCs. Because PIPs can be fine-tuned to target specific DNA sequences, our DNA-based approach could be expanded to target and regulate key transcription factors specifically associated with desired cell types.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Mesoderma/citologia , Miócitos Cardíacos/citologia , Nylons/farmacologia , Pirróis/farmacologia , Fatores de Transcrição SOXB1/antagonistas & inibidores , Sequência de Bases , Sítios de Ligação , Linhagem Celular , Sequência Consenso , Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Miócitos Cardíacos/metabolismo , Nylons/química , Pirróis/química , Fatores de Transcrição SOXB1/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA