Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 7.255
Filtrar
1.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206850

RESUMO

Treating postoperative (PO) pain is a clinical challenge. Inadequate PO pain management can lead to worse outcomes, for example chronic post-surgical pain. Therefore, acquiring new information on the PO pain mechanism would increase the therapeutic options available. In this paper, we evaluated the role of a natural substance, epigallocatechin-3-gallate (EGCG), on pain and neuroinflammation induced by a surgical procedure in an animal model of PO pain. We performed an incision of the hind paw and EGCG was administered for five days. Mechanical allodynia, thermal hyperalgesia, and motor dysfunction were assessed 24 h, and three and five days after surgery. At the same time points, animals were sacrificed, and sera and lumbar spinal cord tissues were harvested for molecular analysis. EGCG administration significantly alleviated hyperalgesia and allodynia, and reduced motor disfunction. From the molecular point of view, EGCG reduced the activation of the WNT pathway, reducing WNT3a, cysteine-rich domain frizzled (FZ)1 and FZ8 expressions, and both cytosolic and nuclear ß-catenin expression, and the noncanonical ß-catenin-independent signaling pathways, reducing the activation of the NMDA receptor subtype NR2B (pNR2B), pPKC and cAMP response element-binding protein (pCREB) expressions at all time points. Additionally, EGCG reduced spinal astrocytes and microglia activation, cytokines overexpression and nuclear factor kappa-light-chain-enhancer of activated B cells (NFkB) pathway, downregulating inducible nitric oxide synthase (iNOS) activation, cyclooxygenase 2 (COX-2) expression, and prostaglandin E2 (PGE2) levels. Thus, EGCG administration managing the WNT/ß-catenin signaling pathways modulates PO pain related neurochemical and inflammatory alterations.


Assuntos
Analgésicos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Catequina/análogos & derivados , Dor Pós-Operatória/tratamento farmacológico , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Catequina/farmacologia , Catequina/uso terapêutico , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Masculino , NF-kappa B/genética , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Medula Espinal/efeitos dos fármacos , Medula Espinal/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , beta Catenina/genética , beta Catenina/metabolismo
2.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206401

RESUMO

Wingless-type MMTV integration site family, member 16 (wnt16), is a wnt ligand that participates in the regulation of vertebrate skeletal development. Studies have shown that wnt16 can regulate bone metabolism, but its molecular mechanism remains largely undefined. We obtained the wnt16-/- zebrafish model using the CRISPR-Cas9-mediated gene knockout screen with 11 bp deletion in wnt16, which led to the premature termination of amino acid translation and significantly reduced wnt16 expression, thus obtaining the wnt16-/- zebrafish model. The expression of wnt16 in bone-related parts was detected via in situ hybridization. The head, spine, and tail exhibited significant deformities, and the bone mineral density and trabecular bone decreased in wnt16-/- using light microscopy and micro-CT analysis. RNA sequencing was performed to explore the differentially expressed genes (DEGs). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis found that the down-regulated DEGs are mainly concentrated in mTOR, FoxO, and VEGF pathways. Protein-protein interaction (PPI) network analysis was performed with the detected DEGs. Eight down-regulated DEGs including akt1, bnip4, ptena, vegfaa, twsg1b, prkab1a, prkab1b, and pla2g4f.2 were validated by qRT-PCR and the results were consistent with the RNA-seq data. Overall, our work provides key insights into the influence of wnt16 gene on skeletal development.


Assuntos
Osso e Ossos/anormalidades , Anormalidades Musculoesqueléticas/genética , Anormalidades Musculoesqueléticas/metabolismo , Osteogênese/genética , Proteínas Wnt/deficiência , Proteínas de Peixe-Zebra/deficiência , Peixe-Zebra/genética , Animais , Animais Geneticamente Modificados , Biologia Computacional/métodos , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Técnicas de Inativação de Genes , Ontologia Genética , Anotação de Sequência Molecular , Anormalidades Musculoesqueléticas/diagnóstico , Fenótipo , Transcriptoma , Proteínas Wnt/química , Proteínas Wnt/metabolismo , Proteínas de Peixe-Zebra/química , Proteínas de Peixe-Zebra/metabolismo
3.
Nat Commun ; 12(1): 3464, 2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34103493

RESUMO

Right-sided (proximal) colorectal cancer (CRC) has a poor prognosis and a distinct mutational profile, characterized by oncogenic BRAF mutations and aberrations in mismatch repair and TGFß signalling. Here, we describe a mouse model of right-sided colon cancer driven by oncogenic BRAF and loss of epithelial TGFß-receptor signalling. The proximal colonic tumours that develop in this model exhibit a foetal-like progenitor phenotype (Ly6a/Sca1+) and, importantly, lack expression of Lgr5 and its associated intestinal stem cell signature. These features are recapitulated in human BRAF-mutant, right-sided CRCs and represent fundamental differences between left- and right-sided disease. Microbial-driven inflammation supports the initiation and progression of these tumours with foetal-like characteristics, consistent with their predilection for the microbe-rich right colon and their antibiotic sensitivity. While MAPK-pathway activating mutations drive this foetal-like signature via ERK-dependent activation of the transcriptional coactivator YAP, the same foetal-like transcriptional programs are also initiated by inflammation in a MAPK-independent manner. Importantly, in both contexts, epithelial TGFß-receptor signalling is instrumental in suppressing the tumorigenic potential of these foetal-like progenitor cells.


Assuntos
Carcinogênese/metabolismo , Neoplasias do Colo/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Carcinogênese/patologia , Diferenciação Celular , Sobrevivência Celular , Colo/patologia , Neoplasias do Colo/genética , Células Epiteliais/metabolismo , Feto/patologia , Inflamação/patologia , Estimativa de Kaplan-Meier , Sistema de Sinalização das MAP Quinases , Camundongos Endogâmicos C57BL , Mutação , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteínas Wnt/metabolismo , Via de Sinalização Wnt
4.
Life Sci ; 279: 119697, 2021 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-34102194

RESUMO

AIMS: Vitamin D and rosuvastatin are well-known drugs that mediate beneficial effects in treating type-2 diabetes (T2D) complications; however, their anti-neuropathic potential is debatable. Hence, our study investigates their neurotherapeutic potential and the possible underlying mechanisms using a T2D-associated neuropathy rat model. MAIN METHODS: Diabetic peripheral neuropathy (DPN) was induced with 8 weeks of administration of a high fat fructose diet followed by a single i.p. injection of streptozotocin (35 mg/kg). Six weeks later, DPN developed and rats were divided into five groups; viz., control, untreated DPN, DPN treated with vitamin D (cholecalciferol, 3500 IU/kg/week), DPN treated with rosuvastatin (10 mg/kg/day), or DPN treated with combination vitamin D and rosuvastatin. We determined their anti-neuropathic effects on small nerves (tail flick test); large nerves (electrophysiological and histological examination); neuronal inflammation (TNF-α and IL-18); apoptosis (caspase-3 activity and Bcl-2); mitochondrial function (NRF-1, TFAM, mtDNA, and ATP); and NICD1, Wnt-10α/ß-catenin, and TGF-ß/Smad-7 pathways. KEY FINDINGS: Two-month treatment with vitamin D and/or rosuvastatin regenerated neuronal function and architecture and abated neuronal inflammation and apoptosis. This was verified by the inhibition of the neuronal content of TNF-α, IL-18, and caspase-3 activity, while augmenting Bcl-2 content in the sciatic nerve. These treatments inhibited the protein expressions of NICD1, Wnt-10α, ß-catenin, and TGF-ß; increased the sciatic nerve content of Smad-7; and enhanced mitochondrial biogenesis and function. SIGNIFICANCE: Vitamin D and/or rosuvastatin alleviated diabetes-induced neuropathy by suppressing Notch1 and Wnt-10α/ß-catenin; modulating TGF-ß/Smad-7 signaling pathways; and enhancing mitochondrial function, which lessened neuronal degeneration, demyelination, and fibrosis.


Assuntos
Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Neuropatias Diabéticas/prevenção & controle , Regulação da Expressão Gênica/efeitos dos fármacos , Rosuvastatina Cálcica/farmacologia , Vitamina D/administração & dosagem , Animais , Anticolesterolemiantes/farmacologia , Neuropatias Diabéticas/etiologia , Neuropatias Diabéticas/metabolismo , Neuropatias Diabéticas/patologia , Quimioterapia Combinada , Masculino , Fator 1 Relacionado a NF-E2/genética , Fator 1 Relacionado a NF-E2/metabolismo , Ratos , Ratos Wistar , Receptor Notch1/genética , Receptor Notch1/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Vitaminas/administração & dosagem , Proteínas Wnt/genética , Proteínas Wnt/metabolismo
5.
FASEB J ; 35(7): e21683, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34118078

RESUMO

Glucocorticoids, widely prescribed for anti-inflammatory and immunosuppressive purposes, are the most common secondary cause for osteoporosis and related fractures. Current anti-resorptive and anabolic therapies are insufficient for treating glucocorticoid-induced osteoporosis due to contraindications or concerns of side effects. Glucocorticoids have been shown to disrupt Wnt signaling in osteoblast-lineage cells, but the efficacy for Wnt proteins to restore bone mass after glucocorticoid therapy has not been examined. Here by using two mouse genetic models wherein WNT7B expression is temporally activated by either tamoxifen or doxycycline in osteoblast-lineage cells, we show that WNT7B recovers bone mass following glucocorticoid-induced bone loss, thanks to increased osteoblast number and function. However, WNT7B overexpression in bone either before or after glucocorticoid treatments does not ameliorate the abnormal accumulation of body fat. The study demonstrates a potent bone anabolic function for WNT7B in countering glucocorticoid-induced bone loss.


Assuntos
Densidade Óssea , Glucocorticoides/toxicidade , Osteogênese , Osteoporose/prevenção & controle , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Wnt/metabolismo , Animais , Masculino , Camundongos , Osteoporose/induzido quimicamente , Osteoporose/patologia , Proteínas Proto-Oncogênicas/genética , Proteínas Wnt/genética
6.
Life Sci ; 278: 119573, 2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-33964297

RESUMO

AIMS: Myosin phosphatase targeting protein 1 (MYPT1) was identified to function as a tumor suppressor in several kinds of cancers, but its role and the molecular mechanisms in non-small cell lung cancer (NSCLC) remain undiscovered. Herein, we aimed to reveal MYPT1 expression pattern and role in NSCLC, and investigate the underlying mechanisms. MAIN METHODS: Sixty-eight paired NSCLC tissues and the adjacent normal tissues were included in this study. Western blotting and quantitative reverse transcription-polymerase chain (qPCR) technologies were applied for protein and RNA detection. CCK-8, colony formation, flow cytometry, wound healing, transwell chambers coated with Matrigel and in vivo experiments were applied to detect cell viability, colony formation, apoptosis, migration, invasiveness and tumorigenesis, respectively. KEY FINDINGS: MYPT1 expressed at a lower level in NSCLC tissues as compared with the adjacent normal tissues, which predicted advanced clinic process and poor prognosis. Overexpression of MYPT1 resulted in obvious inhibitions in cell viability, colony formation, migration, invasiveness and tumorigenesis, and induced cell apoptotic rates, as well as decreased the expression levels of ß-catenin and TCF4. Besides, overexpression of ß-catenin weakened the above roles of MYPT1. In addition, the luciferase gene reporter assay verified that MYPT1 was a target of miR-19b-3p. Further experiments showed that miR-19b-3p promoted cell viability, invasiveness and migration and repressed cell apoptosis by targeting MYPT1. SIGNIFICANCE: In conclusion, this study demonstrates that MYPT1, regulated by miR-19b-3p, inhibits the progression of NSCLC via inhibiting the activation of wnt/ß-catenin signaling.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , MicroRNAs/genética , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Via de Sinalização Wnt , Células A549 , Apoptose , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Progressão da Doença , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , MicroRNAs/metabolismo , Invasividade Neoplásica , Pneumonectomia , RNA Longo não Codificante/genética , Reprodutibilidade dos Testes , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
7.
Cancer Med ; 10(10): 3332-3345, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33934523

RESUMO

Mining databases and data obtained from assays on human specimens had shown that Fzd7 is closely associated with Wnt7b, that Fzd7/Wnt7b expression is upregulated in pancreatic cancer tissues compared with normal tissues, and its expression is negatively correlated with survival. Fzd7/Wnt7b knockdown in Capan-2 and Panc-1 cells reduced the proliferative capacity of pancreatic cancer stem cells (PCSCs), reduced drug resistance, decreased the percentage of CD24+ CD44+ subset of cells and the levels of ABCG2, inhibited cell-sphere formation, and reduced gemcitabine (GEM) resistance. In contrast, Fzd7/Wnt7b overexpression increased the percentage of the CD24+ CD44+ subset of cells, and increased the levels of ABCG2 detected in cell spheroids. The gem-resistant cells exhibited higher levels of Fzd7/Wnt7b expression, an increased percentage of CD24+ CD44+ cells, and higher levels of ABCG2 compared with the parental cells. Taken together, Fzd7/Wnt7b knockdown can reduce PDAC cell stemness and chemoresistance by reducing the percentage of CSCs. Mechanistically, Fzd7 binds with Wnt7b and modulates the levels of ß-catenin, and they may exert their role via modulation of the canonical Wnt pathway.


Assuntos
Resistencia a Medicamentos Antineoplásicos/fisiologia , Receptores Frizzled/metabolismo , Células-Tronco Neoplásicas/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas Wnt/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Antígeno CD24/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Receptores de Hialuronatos/metabolismo , Masculino , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/efeitos dos fármacos , Neoplasias Pancreáticas/tratamento farmacológico , Regulação para Cima/fisiologia , beta Catenina/metabolismo
8.
Bone ; 150: 116006, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34000432

RESUMO

Recently, the accumulation of senescent cells (SnCs) within joints was found to promote osteoarthritis (OA) progression. Our previous study found that Wnt proteins, especially Wnt10a, have marked effects on cellular senescence and joint health. However, the effect of WNT10A on SnCs in OA joints remains unknown. In this study, we confirmed that the synovium was the first and most marked site of SnC accumulation in the OA joint, and synovial resident mesenchymal stem cells (SMSCs) seemed to be the main source of these SnCs. In synovium samples from OA patients, WNT10A level inversely correlated with the extent of SnCs accumulation. Therefore, we further explored the possible regulatory role and mechanism of WNT10A in intraarticular senescent SMSCs. In brief, we confirmed that WNT10A could specifically clear these senescent OA-SMSCs in vitro experiments and naturally occurring OA models via proapoptotic effects. Mechanistically, WNT10A activated noncanonical Wnt/calcium signaling in senescent OA-SMSCs, which in turn induced histone deacetylase 5 (HDAC5) phosphorylation and nuclear export via its downstream Ca2+/calmodulin-dependent protein kinase II (CaMKII) to regulate cell fate. The regulation of this pathway significantly improved the regenerative microenvironment of OA, exhibiting its potential as a novel clinical disease-modifying OA drugs (DMOADs) target.


Assuntos
Células-Tronco Mesenquimais , Osteoartrite , Apoptose , Cálcio/metabolismo , Histona Desacetilases/metabolismo , Humanos , Osteoartrite/metabolismo , Fosforilação , Membrana Sinovial/metabolismo , Proteínas Wnt/metabolismo
9.
Int J Mol Sci ; 22(7)2021 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-33800649

RESUMO

The transforming growth factor-ß (TGF-ß) family of cytokines performs a multifunctional signaling, which is integrated and coordinated in a signaling network that involves other pathways, such as Wintless, Forkhead box-O (FOXO) and Hedgehog and regulates pivotal functions related to cell fate in all tissues. In the hematopoietic system, TGF-ß signaling controls a wide spectrum of biological processes, from immune system homeostasis to the quiescence and self-renewal of hematopoietic stem cells (HSCs). Recently an important role in post-transcription regulation has been attributed to two type of ncRNAs: microRNAs and pseudogenes. Ciona robusta, due to its philogenetic position close to vertebrates, is an excellent model to investigate mechanisms of post-transcriptional regulation evolutionarily highly conserved in immune homeostasis. The combined use of NGS and bioinformatic analyses suggests that in the pharynx, the hematopoietic organ of Ciona robusta, the Tgf-ß, Wnt, Hedgehog and FoxO pathways are involved in tissue homeostasis, as they are in human. Furthermore, ceRNA network interactions and 3'UTR elements analyses of Tgf-ß, Wnt, Hedgehog and FoxO pathways genes suggest that different miRNAs conserved (cin-let-7d, cin-mir-92c, cin-mir-153), species-specific (cin-mir-4187, cin-mir-4011a, cin-mir-4056, cin-mir-4150, cin-mir-4189, cin-mir-4053, cin-mir-4016, cin-mir-4075), pseudogenes (ENSCING00000011392, ENSCING00000018651, ENSCING00000007698) and mRNA 3'UTR elements are involved in post-transcriptional regulation in an integrated way in C. robusta.


Assuntos
Ciona/metabolismo , Proteína Forkhead Box O1/metabolismo , Regulação da Expressão Gênica , Fator de Crescimento Transformador beta/metabolismo , Proteínas Wnt/metabolismo , Regiões 3' não Traduzidas , Animais , Linhagem da Célula , Biologia Computacional , Proteínas Hedgehog/metabolismo , Hematopoese , Sequenciamento de Nucleotídeos em Larga Escala , Homeostase , Sistema Imunitário , MicroRNAs/metabolismo , Faringe/metabolismo , Mapeamento de Interação de Proteínas , RNA-Seq
10.
J Med Chem ; 64(8): 4257-4288, 2021 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-33822624

RESUMO

Canonical WNT signaling is an important developmental pathway that has attracted increased attention for anticancer drug discovery. From the production and secretion of WNT ligands, their binding to membrane receptors, and the ß-catenin destruction complex to the expansive ß-catenin transcriptional complex, multiple components have been investigated as drug targets to modulate WNT signaling. Significant progress in developing WNT inhibitors such as porcupine inhibitors, tankyrase inhibitors, ß-catenin/coactivators, protein-protein interaction inhibitors, casein kinase modulators, DVL inhibitors, and dCTPP1 inhibitors has been made, with several candidates (e.g., LGK-974, PRI-724, and ETC-159) in human clinical trials. Herein we summarize recent progress in the drug discovery and development of small-molecule inhibitors targeting the canonical WNT pathway, focusing on their specific target proteins, in vitro and in vivo activities, physicochemical properties, and therapeutic potential. The relevant opportunities and challenges toward maintaining the balance between efficacy and toxicity in effectively targeting this pathway are also highlighted.


Assuntos
Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas Wnt/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Sítios de Ligação , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Humanos , Simulação de Dinâmica Molecular , Neoplasias/tratamento farmacológico , Peptídeos/química , Peptídeos/metabolismo , Mapas de Interação de Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/uso terapêutico , Fatores de Transcrição TCF/química , Fatores de Transcrição TCF/metabolismo , Tanquirases/antagonistas & inibidores , Tanquirases/metabolismo , Proteínas Wnt/química , beta Catenina/química , beta Catenina/metabolismo
11.
Biomolecules ; 11(2)2021 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-33672636

RESUMO

The prevalence of patients suffering from mental disorders is substantially increasing in recent years and represents a major burden to society. The underlying causes and neuronal circuits affected are complex and difficult to unravel. Frequent disorders such as depression, schizophrenia, autism, and bipolar disorder share links to the habenular neural circuit. This conserved neurotransmitter system relays cognitive information between different brain areas steering behaviors ranging from fear and anxiety to reward, sleep, and social behaviors. Advances in the field using the zebrafish model organism have uncovered major genetic mechanisms underlying the formation of the habenular neural circuit. Some of the identified genes involved in regulating Wnt/beta-catenin signaling have previously been suggested as risk genes of human mental disorders. Hence, these studies on habenular genetics contribute to a better understanding of brain diseases. We are here summarizing how the gained knowledge on the mechanisms underlying habenular neural circuit development can be used to introduce defined manipulations into the system to study the functional behavioral consequences. We further give an overview of existing behavior assays to address phenotypes related to mental disorders and critically discuss the power but also the limits of the zebrafish model for identifying suitable targets to develop therapies.


Assuntos
Habenula/fisiologia , Transtornos Mentais/genética , Peixe-Zebra/genética , Animais , Ansiedade , Axônios/metabolismo , Comportamento Animal , Depressão/tratamento farmacológico , Depressão/metabolismo , Depressão/fisiopatologia , Modelos Animais de Doenças , Habenula/metabolismo , Transtornos Mentais/metabolismo , Mutação , Rede Nervosa , Neurogênese , Neurônios/metabolismo , Neurotransmissores , Fenótipo , Comportamento Social , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , Peixe-Zebra/fisiologia , Proteínas de Peixe-Zebra/genética
12.
PLoS Biol ; 19(3): e3001111, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33657096

RESUMO

Development of the Drosophila wing-a paradigm of organ development-is governed by 2 morphogens, Decapentaplegic (Dpp, a BMP) and Wingless (Wg, a Wnt). Both proteins are produced by defined subpopulations of cells and spread outwards, forming gradients that control gene expression and cell pattern as a function of concentration. They also control growth, but how is unknown. Most studies have focused on Dpp and yielded disparate models in which cells throughout the wing grow at similar rates in response to the grade or temporal change in Dpp concentration or to the different amounts of Dpp "equalized" by molecular or mechanical feedbacks. In contrast, a model for Wg posits that growth is governed by a progressive expansion in morphogen range, via a mechanism in which a minimum threshold of Wg sustains the growth of cells within the wing and recruits surrounding "pre-wing" cells to grow and enter the wing. This mechanism depends on the capacity of Wg to fuel the autoregulation of vestigial (vg)-the selector gene that specifies the wing state-both to sustain vg expression in wing cells and by a feed-forward (FF) circuit of Fat (Ft)/Dachsous (Ds) protocadherin signaling to induce vg expression in neighboring pre-wing cells. Here, we have subjected Dpp to the same experimental tests used to elucidate the Wg model and find that it behaves indistinguishably. Hence, we posit that both morphogens act together, via a common mechanism, to control wing growth as a function of morphogen range.


Assuntos
Proteínas de Drosophila/genética , Asas de Animais/crescimento & desenvolvimento , Proteína Wnt1/genética , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Homeostase , Morfogênese , Proteínas Nucleares/metabolismo , Transdução de Sinais , Asas de Animais/metabolismo , Proteínas Wnt/metabolismo
13.
Int J Mol Sci ; 22(5)2021 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-33668092

RESUMO

The mechanistic target of rapamycin (mTOR) and wingless-related integration site (Wnt) signal transduction networks are evolutionarily conserved mammalian growth and cellular development networks. Most cells express many of the proteins in both pathways, and this review will briefly describe only the key proteins and their intra- and extracellular crosstalk. These complex interactions will be discussed in relation to cancer development, drug resistance, and stem cell exhaustion. This review will also highlight the tumor-suppressive tuberous sclerosis complex (TSC) mutated, mTOR-hyperactive lung disease of women, lymphangioleiomyomatosis (LAM). We will summarize recent advances in the targeting of these pathways by monotherapy or combination therapy, as well as future potential treatments.


Assuntos
Linfangioleiomiomatose/fisiopatologia , Terapia de Alvo Molecular , Serina-Treonina Quinases TOR/metabolismo , Proteínas Wnt/metabolismo , Animais , Humanos , Linfangioleiomiomatose/tratamento farmacológico , Linfangioleiomiomatose/metabolismo
14.
Int J Mol Sci ; 22(4)2021 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-33670029

RESUMO

Hair follicle morphogenesis is heavily dependent on reciprocal, sequential, and epithelial-mesenchymal interaction (EMI) between epidermal stem cells and the specialized cells of the underlying mesenchyme, which aggregate to form the dermal condensate (DC) and will later become the dermal papilla (DP). Similar models were developed with a co-culture of keratinocytes and DP cells. Previous studies have demonstrated that co-culture with keratinocytes maintains the in vivo characteristics of the DP. However, it is often challenging to develop three-dimensional (3D) DP and keratinocyte co-culture models for long term in vitro studies, due to the poor intercellular adherence between keratinocytes. Keratinocytes exhibit exfoliative behavior, and the integrity of the DP and keratinocyte co-cultured spheroids cannot be maintained over prolonged culture. Short durations of culture are unable to sufficiently allow the differentiation and re-programming of the keratinocytes into hair follicular fate by the DP. In this study, we explored a microgel array approach fabricated with two different hydrogel systems. Using poly (ethylene glycol) diacrylate (PEGDA) and gelatin methacrylate (GelMA), we compare their effects on maintaining the integrity of the cultures and their expression of important genes responsible for hair follicle morphogenesis, namely Wnt10A, Wnt10B, and Shh, over prolonged duration. We discovered that low attachment surfaces such as PEGDA result in the exfoliation of keratinocytes and were not suitable for long-term culture. GelMA, on the hand, was able to sustain the integrity of co-cultures and showed higher expression of the morphogens overtime.


Assuntos
Derme/citologia , Queratinócitos/citologia , Microgéis/química , Polietilenoglicóis/farmacologia , Adesão Celular/efeitos dos fármacos , Agregação Celular/efeitos dos fármacos , Linhagem Celular , Técnicas de Cocultura , Proteínas de Fluorescência Verde/metabolismo , Células HaCaT/citologia , Células HaCaT/efeitos dos fármacos , Humanos , Hidrogéis/farmacologia , Proteínas Luminescentes/metabolismo , Esferoides Celulares/citologia , Esferoides Celulares/efeitos dos fármacos , Proteínas Wnt/metabolismo
15.
J Vis Exp ; (168)2021 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-33645570

RESUMO

Generating patient-specific cardiomyocytes from a single blood draw has attracted tremendous interest in precision medicine on cardiovascular disease. Cardiac differentiation from human induced pluripotent stem cells (iPSCs) is modulated by defined signaling pathways that are essential for embryonic heart development. Numerous cardiac differentiation methods on 2-D and 3-D platforms have been developed with various efficiencies and cardiomyocyte yield. This has puzzled investigators outside the field as the variety of these methods can be difficult to follow. Here we present a comprehensive protocol that elaborates robust generation and expansion of patient-specific cardiomyocytes from peripheral blood mononuclear cells (PBMCs). We first describe a high-efficiency iPSC reprogramming protocol from a patient's blood sample using non-integration Sendai virus vectors. We then detail a small molecule-mediated monolayer differentiation method that can robustly produce beating cardiomyocytes from most human iPSC lines. In addition, a scalable cardiomyocyte expansion protocol is introduced using a small molecule (CHIR99021) that could rapidly expand patient-derived cardiomyocytes for industrial- and clinical-grade applications. At the end, detailed protocols for molecular identification and electrophysiological characterization of these iPSC-CMs are depicted. We expect this protocol to be pragmatic for beginners with limited knowledge on cardiovascular development and stem cell biology.


Assuntos
Técnicas de Cultura de Células/métodos , Leucócitos Mononucleares/citologia , Miócitos Cardíacos/citologia , Diferenciação Celular , Proliferação de Células , Reprogramação Celular , Citometria de Fluxo , Imunofluorescência , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Técnicas de Patch-Clamp , Proteínas Wnt/metabolismo
16.
BMC Cancer ; 21(1): 180, 2021 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-33607955

RESUMO

BACKGROUND: Aberrant activation of the Wnt/ß-catenin signaling pathway is one of the most frequent abnormalities in human cancer, including colorectal cancer (CRC). Previous studies revealed pivotal functions of WNT family members in colorectal cancer, as well as their prognostic values. Nevertheless, the prognostic role and mechanisms underlying WNT7b in colorectal cancer development remains unclear. METHODS: In this study, WNT7b expression was measured by immunohistochemical staining of 100 cases of surgically resected human colorectal cancerous tissues as well as matched adjacent normal tissues constructed as tissue microarrays. In vitro studies, we attempted to substantiate the WNT7b expressional pattern previously found in immunohistochemistry staining. We used the colorectal cancer cell-line HCT116 and normal colorectal cell-line FHC for immunofluorescence staining and nuclear/cytoplasmic separated western blotting. We measured epithelial-mesenchymal transition (EMT) markers and migration capacity of HCT116 in the context of WNT7b knocked-down using short interfering RNA. Finally, clinical and prognostic values of WNT7b activation levels were examined. RESULTS: WNT7b was expressed in the nucleus in adjacent normal tissues. In CRC tissues, nuclear expression of WNT7b was similar; however, membrane and cytoplasmic expression was strikingly enhanced. Consistently, in vitro analysis confirmed the same expression pattern of WNT7b. Compared with FHC cells, HCT116 cells displayed higher levels of WNT7b membrane and cytoplasmic enrichment, as well as higher migration capacity with a sensitized EMT process. Either partial knockdown of WNT7b or blockade of the Wnt/ß-catenin signaling pathway reversed EMT process and inhibited the migration of HCT116 cells. Finally, elevated secretion levels of WNT7b were significantly associated with lymphatic and remote metastasis and predicted worse prognosis in the CRC cohort. CONCLUSION: In summary, we demonstrated that the activation of WNT7b autocrine probably contributes to CRC metastasis by triggering EMT process through the Wnt/ß-catenin signaling pathway. High levels of WNT7b autocrine secretion predicts poor outcome in patients with CRC. This molecule is a promising candidate for clinical CRC treatments.


Assuntos
Neoplasias Colorretais/metabolismo , Proteínas Wnt/metabolismo , Via de Sinalização Wnt , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Neoplasias Colorretais/patologia , Neoplasias Colorretais/cirurgia , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Prognóstico , Taxa de Sobrevida
17.
Int J Mol Sci ; 22(4)2021 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-33557112

RESUMO

Breast cancer (BCa) is one of the leading health problems among women. Although significant achievements have led to advanced therapeutic success with targeted therapy options, more efforts are required for different subtypes of tumors and according to genomic, transcriptomic, and proteomic alterations. This study underlines the role of microRNA-21 (miR-21) in metastatic MDA-MB-231 breast cancer cells. Following the knockout of miR-21 from MDA-MB-231 cells, which have the highest miR-21 expression levels compared to MCF-7 and SK-BR-3 BCa cells, a decrease in epithelial-mesenchymal transition (EMT) via downregulation of mesenchymal markers was observed. Wnt-11 was a critical target for miR-21, and the Wnt-11 related signaling axis was altered in the stable miR-21 knockout cells. miR-21 expression was associated with a significant increase in mesenchymal markers in MDA-MB-231 BCa cells. Furthermore, the release of extracellular vesicles (EVs) was significantly reduced in the miR-21 KO cells, alongside a significant reduction in relative miR-21 export in EV cargo, compared with control cells. We conclude that miR-21 is a leading factor involved in mesenchymal transition in MDA-MB-231 BCa. Future therapeutic strategies could focus on its role in the treatment of metastatic breast cancer.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Biomarcadores Tumorais , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Feminino , Edição de Genes , Técnicas de Inativação de Genes , Humanos , Prognóstico , Interferência de RNA , Proteínas Wnt/metabolismo
18.
Cancer Sci ; 112(5): 1695-1706, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33605517

RESUMO

Wnt/ß-catenin signaling is indispensable for many biological processes, including embryonic development, cell cycle, inflammation, and carcinogenesis. Aberrant activation of the Wnt/ß-catenin signaling can promote tumorigenicity and enhance metastatic potential in hepatocellular carcinoma (HCC). Targeting this pathway is a new opportunity for precise medicine for HCC. However, inhibiting Wnt/ß-catenin signaling alone is unlikely to significantly improve HCC patient outcome due to the lack of specific inhibitors and the complexity of this pathway. Combination with other therapies will be an important next step in improving the efficacy of Wnt/ß-catenin signaling inhibitors. Protein kinases play a key and evolutionarily conserved role in the Wnt/ß-catenin signaling and have become one of the most important drug targets in cancer. Targeting Wnt/ß-catenin signaling and its regulatory kinase together will be a promising HCC management strategy. In this review, we summarize the kinases that modulate the Wnt/ß-catenin signaling in HCC and briefly discuss their molecular mechanisms. Furthermore, we list some small molecules that target the kinases and may inhibit Wnt/ß-catenin signaling, to offer new perspectives for preclinical and clinical HCC studies.


Assuntos
Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas Quinases/metabolismo , Via de Sinalização Wnt/fisiologia , beta Catenina/antagonistas & inibidores , Complexo de Sinalização da Axina/metabolismo , Proteína Quinase CDC2/metabolismo , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/terapia , Terapia Combinada/métodos , Creatina Quinase/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Receptores ErbB/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/terapia , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Medicina de Precisão , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Proteínas Wnt/antagonistas & inibidores , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Quinases Ativadas por p21/metabolismo , Quinases da Família src/metabolismo
19.
J Orthop Surg Res ; 16(1): 71, 2021 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-33472642

RESUMO

BACKGROUND: Bone neoplasms present poor prognosis due to recurrence and metastasis. Although the role microRNAs (miRNAs) in inhibiting growth and metastasis of bone neoplasms has been investigated, the underlying potential molecular mechanisms mediated by miRNA-128 (miR-218) for the invasiveness of bone neoplasms cells are still not completely understood. The purpose of this study was to identify the regulatory mechanisms of miR-218 in bone neoplasms cells. METHODS: Western blotting, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), Counting Kit-8 assay, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, luciferase activity assay immunofluorescence, and immunohistochemistry were used to analyze the regulatory effects of miR-218 on bone neoplasms cells. RESULTS: Here, the results showed that transfection of miR-128 suppressed bone neoplasms cells proliferation, migration, and invasion. Genetic knockdown of miR-128 in bone neoplasms cells suppressed the activation of the Wnt/ß-catenin and epithelial-mesenchymal transition (EMT) signaling pathways. Activation of Wnt or EMT blocked miR-128-inhibited cells proliferation and migration in bone neoplasms cells. Exogenously introduced miR-128 markedly inhibited tumor regeneration in bone neoplasms xenograft models. CONCLUSIONS: These results define a tumor-regulated function for miR-128 in bone neoplasms by down-regulation of the Wnt/ß-catenin and EMT signal pathways, which provided a potential target for bone neoplasms gene therapy.


Assuntos
Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Movimento Celular/genética , Proliferação de Células/genética , MicroRNAs/fisiologia , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Neoplasias Ósseas/terapia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Regulação para Baixo/genética , Feminino , Humanos , Masculino , Camundongos Nus , MicroRNAs/uso terapêutico , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Transplante de Neoplasias
20.
J Ethnopharmacol ; 270: 113807, 2021 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-33450290

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: There is substantial experimental evidence to support the view that Ginkgo biloba L. (Ginkgoaceae), a traditional Chinese medicine known to treating stroke, has a protective effect on the central nervous system and significantly improves the cognitive dysfunction caused by disease, including alzheimer disease (AD), vascular dementia, and diabetic encephalopathy. Although a number of studies have reported that ginkgolide B (GB), a diterpenoid lactone compound extracted from Ginkgo biloba leaves, has neuroprotective effects, very little research has been performed to explore its potential pharmacological mechanism on astrocytes under abnormal glutamate (Glu) metabolism in the pathological environment of AD. AIM OF THE STUDY: We investigated the protective effect and mechanism of GB on Glu-induced astrocytes injury. METHODS: Astrocytes were randomly divided into the control group, Glu group, GB group, and GB + IWP-4 group.The CCK-8 assay was used to determine relative cell viability in vitro. Furthermore, RNA sequencing (RNA-seq) was performed to assess the preventive effects of GB in the Glu-induced astrocyte model and reverse transcription quantitative polymerase chain reaction (RT-qPCR) was used to validate the possible molecular mechanisms. The effects of GB on the Glu transporter and Glu-induced apoptosis of astrocytes were studied by RT-qPCR and western blot. RESULTS: GB attenuated Glu-induced apoptosis in a concentration-dependent manner, while the Wnt inhibitor IWP-4 reversed the protective effect of GB on astrocytes. The RNA-seq results revealed 4,032 differential gene expression profiles; 3,491 genes were up-regulated, and 543 genes were down-regulated in the GB group compared with the Glu group. Differentially expressed genes involved in a variety of signaling pathways, including the Hippo and Wnt pathways have been associated with the development and progression of AD. RT-qPCR was used to validate 14 key genes, and the results were consistent with the RNA-seq data. IWP-4 inhibited the regulation of GB, disturbed the apoptosis protective effect on astrocytes, and promoted Glu transporter gene and protein expression caused by Glu. CONCLUSION: Our findings demonstrate that GB may play a protective role in Glu-induced astrocyte injury by regulating the Hippo and Wnt pathways. GB was closely associated with the Wnt pathway by promoting expression of the Glu transporter and inhibiting Glu-induced injury in astrocytes.


Assuntos
Astrócitos/efeitos dos fármacos , Ginkgolídeos/farmacologia , Lactonas/farmacologia , Fármacos Neuroprotetores/farmacologia , Animais , Apoptose/efeitos dos fármacos , Astrócitos/citologia , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Ácido Glutâmico/toxicidade , Cultura Primária de Células , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Análise de Sequência de RNA/métodos , Transdução de Sinais/efeitos dos fármacos , Proteínas Wnt/antagonistas & inibidores , Proteínas Wnt/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...