RESUMO
BACKGROUND: When Wnt binds to the N-terminal of Frizzled, a conformational change occurs in the C-terminal of Frizzled, which binds to Dishevelled1 (Dvl1), a Wnt signaling component protein. When Dvl1 binds to the C-terminal of Frizzled, the concentration of ß-catenin increases and it enters the nucleus to transmit cell proliferation signals. CXXC-type zinc finger protein 5 (CXXC5) binds to the Frizzled binding site of Dvl1 and interferes with Dvl1-Frizzled binding. Therefore, blocking CXXC5-Dvl1 binding may induce Wnt signal transduction. MATERIALS AND METHODS: We used WD-aptamer, a DNA aptamer that specifically binds to Dvl1 and interferes with CXXC5-Dvl1 interaction. We confirmed the penetration of WD-aptamer into human hair follicle dermal papilla cells (HFDPCs) and measured ß-catenin expression following treatment with WD-aptamer in HFDPCs, wherein Wnt signaling was activated by Wnt3a. In addition, MTT assay was performed to investigate the effect of WD-aptamer on cell proliferation. RESULTS: WD-aptamer penetrated the cell, affected Wnt signaling, and increased ß-catenin expression, which plays an important role in signaling. Additionally, WD-aptamer induced HFDPC proliferation. CONCLUSION: CXXC5-associated negative feedback of Wnt/ß-catenin signaling can be regulated by interfering with CXXC5-Dvl1 interaction.
Assuntos
Aptâmeros de Nucleotídeos , Via de Sinalização Wnt , Humanos , Via de Sinalização Wnt/fisiologia , Folículo Piloso/metabolismo , beta Catenina/metabolismo , beta Catenina/farmacologia , Aptâmeros de Nucleotídeos/farmacologia , Aptâmeros de Nucleotídeos/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/farmacologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/farmacologiaRESUMO
OBJECTIVE: To investigate the mechanism by which fibroblasts with high WNT2b expression causes intestinal mucosa barrier disruption and promote the progression of inflammatory bowel disease (IBD). METHODS: Caco-2 cells were treated with 20% fibroblast conditioned medium or co-cultured with fibroblasts highly expressing WNT2b, with the cells without treatment with the conditioned medium and cells co-cultured with wild-type fibroblasts as the control groups. The changes in barrier permeability of Caco-2 cells were assessed by measuring transmembrane resistance and Lucifer Yellow permeability. In Caco-2 cells co-cultured with WNT2b-overexpressing or control intestinal fibroblasts, nuclear entry of ß-catenin was detected with immunofluorescence assay, and the expressions of tight junction proteins ZO-1 and E-cadherin were detected with Western blotting. In a C57 mouse model of dextran sulfate sodium (DSS)-induced IBD-like enteritis, the therapeutic effect of intraperitoneal injection of salinomycin (5 mg/kg, an inhibitor of WNT/ß-catenin signaling pathway) was evaluated by observing the changes in intestinal inflammation and detecting the expressions of tight junction proteins. RESULTS: In the coculture system, WNT2b overexpression in the fibroblasts significantly promoted nuclear entry of ß-catenin (P < 0.01) and decreased the expressions of tight junction proteins in Caco-2 cells; knockdown of FZD4 expression in Caco-2 cells obviously reversed this effect. In DSS-treated mice, salinomycin treatment significantly reduced intestinal inflammation and increased the expressions of tight junction proteins in the intestinal mucosa. CONCLUSION: Intestinal fibroblasts overexpressing WNT2b causes impairment of intestinal mucosal barrier function and can be a potential target for treatment of IBD.
Assuntos
Doenças Inflamatórias Intestinais , beta Catenina , Humanos , Camundongos , Animais , Células CACO-2 , beta Catenina/metabolismo , Meios de Cultivo Condicionados/farmacologia , Junções Íntimas/metabolismo , Mucosa Intestinal , Proteínas de Junções Íntimas/metabolismo , Inflamação/metabolismo , Fibroblastos/metabolismo , Camundongos Endogâmicos C57BL , Glicoproteínas/metabolismo , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , Receptores Frizzled/metabolismoRESUMO
BACKGROUND: The anti-osteosarcoma effects of hydrocortisone and thiram, an inhibitor of type 2 11ß-hydroxysteroid dehydrogenase (11HSD2), have not been reported. The purpose of this study was to investigate the effects of hydrocortisone alone or the combination of hydrocortisone with thiram on osteosarcoma and the molecular mechanism, and determine whether they can be as new therapeutic agents for osteosarcoma. METHODS: Normal bone cells and osteosarcoma cells were treated with hydrocortisone or thiram alone or in combination. The cell proliferation, migration, cell cycle and apoptosis were detected by using CCK8 assay, wound healing assay, and flow cytometry, respectively. An osteosarcoma mouse model was established. The effect of drugs on osteosarcoma in vivo was assessed by measuring tumor volume. Transcriptome sequencing, bioinformatics analysis, RT-qPCR, Western blotting (WB), enzymelinked immunosorbent assay (ELISA) and siRNA transfection were performed to determine the molecular mechanisms. RESULTS: Hydrocortisone inhibited the proliferation and migration, and induced apoptosis and cell cycle arrest of osteosarcoma cells in vitro. Hydrocortisone also reduced the volume of osteosarcoma in mice in vivo. Mechanistically, hydrocortisone decreased the levels of Wnt/ß-catenin pathway-associated proteins, and induced the expression of glucocorticoid receptor α (GCR), CCAAT enhancer-binding protein ß (C/EBP-beta) and 11HSD2, resulting in a hydrocortisone resistance loop. Thiram inhibited the activity of the 11HSD2 enzyme, the combination of thiram and hydrocortisone further enhanced the inhibition of osteosarcoma through Wnt/ß-catenin pathway. CONCLUSIONS: Hydrocortisone inhibits osteosarcoma through the Wnt/ß-catenin pathway. Thiram inhibits 11HSD2 enzyme activity, reducing hydrocortisone inactivation and promoting the effect of hydrocortisone through the same pathway.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Animais , Camundongos , Apoptose , beta Catenina/metabolismo , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Hidrocortisona/farmacologia , Hidrocortisona/uso terapêutico , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Osteossarcoma/metabolismo , Tiram/farmacologia , Tiram/uso terapêutico , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , 11-beta-Hidroxiesteroide Desidrogenase Tipo 2RESUMO
Alzheimer disease (AD) is the most common, irreversible and progressive form of dementia for which the exact pathology and cause are still not clear. At present, we are only confined to symptomatic treatment, and the lack of disease-modifying therapeutics is worrisome. Alteration of Wnt signaling has been linked to metabolic diseases as well as AD. The crosstalk between Canonical Wnt signaling and insulin signaling pathway has been widely studied and accepted from several clinical and preclinical studies that have proven the beneficial effect of antidiabetic medications in the case of memory and cognition loss. This structure-based in silico study was focused on exploring the link between the currently available FDA approved antidiabetic drugs and the Wnt signaling pathway. The library of antidiabetics was obtained from drug bank and was screened for their binding affinity with protein (PDB ID: 3S2K) LRP6, a coreceptor of the Wnt signaling pathway using GLIDE module of Schrodinger. The top molecules, with higher docking score, binding energy and stable interactions, were subjected to energy-based calculation using MMGBSA, followed by a molecular dynamics-based simulation study. Drugs of class α-glucosidase inhibitors and peroxisome proliferator-activated receptors (PPARs) agonists were found to have a strong affinity towards LRP6 proteins, highlighting the possibility of the modulation of Wnt signaling by antidiabetics as one of the possible mechanisms for use in AD. However, further experimental based in vitro and in vivo studies are warranted for verification and support.Communicated by Ramaswamy H. Sarma.
Assuntos
Doença de Alzheimer , Via de Sinalização Wnt , Humanos , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/química , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , Reposicionamento de Medicamentos , Hipoglicemiantes/farmacologiaRESUMO
Our study was aimed to investigate the effects of lgals3a (Gal-3 encoding gene) on the development of zebrafish embryo and its underlying mechanisms. Morpholino (MO) technology was used to inhibit the expression of zebrafish lgals3a, and the effect of lgals3a gene knockdown on zebrafish embryo development and the number of monocyte macrophages was observed. Effect of lgals3a-e3i3-MO on apoptosis of zebrafish was detected by acridine orange staining. In addition, the mRNA expression levels of Wnt/ß-catenin signaling pathway-related genes were detected by RT-qPCR. Compared with control-MO group, the zebrafish embryos injected with lgals3a-e3i3-MO had obvious defects in the head, eyes, and tail, and pericardial edema. Lgals3a-e3i3-MO significantly reduced the number of mononuclear macrophages in zebrafish embryos compared with the control-MO group. The results of acridine orange staining showed that compared with the control-MO group, lgals3a-e3i3-MO promoted cardiomyocyte apoptosis in zebrafish. Furthermore, lgals3a-e3i3-MO significantly up-regulated the expression of dkk1b, wnt9a, lrp5, fzd7a, ß-catenin, Gsk-3ß, mycn, myca in the Wnt/ß-catenin pathway, and decreased the expression of lef1. These results indicate that lgals3a-e3i3-MO inhibits zebrafish embryo development, reduces the number of mononuclear macrophages, activates Wnt/ß-catenin signaling pathway and promotes cardiomyocyte apoptosis.
Assuntos
Peixe-Zebra , beta Catenina , Laranja de Acridina/metabolismo , Laranja de Acridina/farmacologia , Animais , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Glicogênio Sintase Quinase 3 beta/farmacologia , Receptores de Superfície Celular , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , Via de Sinalização Wnt/genética , Proteínas de Peixe-Zebra/genética , beta Catenina/genética , beta Catenina/metabolismoRESUMO
Accumulating evidence has shown that Wnt signaling is deeply involved in male reproductive physiology, and malfunction of the signal path can cause pathological changes in genital organs and sperm cells. These abnormalities are diverse in manifestation and have been constantly found in the knockout models of Wnt studies. Nevertheless, most of the research solely focused on a certain factor in the Wnt pathway, and there are few reports on the overall relation between Wnt signals and male reproductive physiology. In our review, Wnt findings relating to the reproductive system were sought and summarized in terms of Wnt ligands, Wnt receptors, Wnt intracellular signals and Wnt regulators. By sorting out and integrating relevant functions, as well as underlining the controversies among different reports, our review aims to offer an overview of Wnt signaling in male reproductive physiology and pathology for further mechanistic studies.
Assuntos
Reprodução/fisiologia , Proteínas Wnt/farmacologia , Via de Sinalização Wnt/fisiologia , Animais , Humanos , Infertilidade Masculina/metabolismo , Masculino , Receptores Wnt/fisiologiaRESUMO
Medulloblastoma (MB) is defined by four molecular subgroups (Wnt, Shh, Group 3, Group 4) with Wnt MB having the most favorable prognosis. Since prior reports have illustrated the antitumorigenic role of Wnt activation in Shh MB, we aimed to assess the effects of activated canonical Wnt signaling in Group 3 and 4 MBs. By using primary patient-derived MB brain tumor-initiating cell (BTIC) lines, we characterize differences in the tumor-initiating capacity of Wnt, Group 3, and Group 4 MB. With single cell RNA-seq technology, we demonstrate the presence of rare Wnt-active cells in non-Wnt MBs, which functionally retain the impaired tumorigenic potential of Wnt MB. In treating MB xenografts with a Wnt agonist, we provide a rational therapeutic option in which the protective effects of Wnt-driven MBs may be augmented in Group 3 and 4 MB and thereby support emerging data for a context-dependent tumor suppressive role for Wnt/ß-catenin signaling.
Assuntos
Neoplasias Cerebelares/terapia , Meduloblastoma/terapia , Proteínas Wnt/farmacologia , Proteínas Wnt/uso terapêutico , Animais , Carcinogênese , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Cerebelares/patologia , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Meduloblastoma/genética , Meduloblastoma/patologia , Camundongos , Células-Tronco , Proteínas Wnt/genética , Via de Sinalização Wnt , beta Catenina/uso terapêuticoRESUMO
The multiple specialized cell types of the hematopoietic system originate from differentiation of hematopoietic stem cells and progenitors (HSPC), which can generate both lymphoid and myeloid lineages. The myeloid lineage is preferentially maintained during ageing, but the mechanisms that contribute to this process are incompletely understood. Here, we studied the roles of Wnt5a and Wnt5b, ligands that have previously been linked to hematopoietic stem cell ageing and that are abundantly expressed by both hematopoietic progenitors and bone-marrow derived niche cells. Whereas Wnt5a had no major effects on primitive cell differentiation, Wnt5b had profound and divergent effects on cytokine-induced myeloid differentiation. Remarkably, while IL-3-mediated myeloid differentiation was largely repressed by Wnt5b, GM-CSF-induced myeloid differentiation was augmented. Furthermore, in the presence of IL-3, Wnt5b enhanced HSPC self-renewal, whereas in the presence of GM-CSF, Wnt5b accelerated differentiation, leading to progenitor cell exhaustion. Our results highlight discrepancies between IL-3 and GM-CSF, and reveal novel effects of Wnt5b on the hematopoietic system.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Interleucina-3/farmacologia , Células Mieloides/citologia , Proteínas Wnt/farmacologia , Animais , Camundongos Endogâmicos C57BL , Células Mieloides/efeitos dos fármacos , Células Mieloides/metabolismo , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacosRESUMO
Type 2 diabetes mellitus (T2DM) is an important reason to cause chronic wound healing or even amputation to patients. A common characteristic of T2DM is the presence of hyperglycemia. Autophagy, a cellular pathway which related to protein and organelle degradation, is relevant to many types of cellular homeostasis change and human diseases including diabetes. However, the relationship between high glucose and autophagy in keratinocytes remains unclear. Previously in our research, wnt7a was proved to accelerate healing process by promoting angiogenesis and ameliorating local inflammation, while little is known about its role in epithelial cells, namely keratinocytes. We hypothesized that reduced expression of wnt7a may contribute to the increment of autophagy. We then compared the expression of autophagic markers such as nlrp3, LC3A/B and bip by high glucose-cultured HaCaT cells, with or without wnt7a treatment. Then we examined the expression levels of TNF-α, TLR-4 and p62 to assess inflammatory statement. High glucose induced a significant increasement in the expression of both autophagic markers and inflammatory markers and these elevated protein levels were reversed after the use of wnt7a. Therefore, these results showed that wnt7a regulates overwhelmed autophagy and inflammation promoted under high glucose condition.
Assuntos
Autofagia/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Epiderme/metabolismo , Hiperglicemia/metabolismo , Inflamação/metabolismo , Queratinócitos/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Wnt/metabolismo , Animais , Autofagia/efeitos dos fármacos , Linhagem Celular , Glucose/farmacologia , Humanos , Queratinócitos/efeitos dos fármacos , Proteínas Associadas aos Microtúbulos/efeitos dos fármacos , Proteínas Associadas aos Microtúbulos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos , Ratos Sprague-Dawley , Receptor 4 Toll-Like/efeitos dos fármacos , Receptor 4 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Wnt/farmacologiaRESUMO
BACKGROUND AND OBJECTIVE: Obesity is highly complicated by hypertension and hyperglycemia. In particular, it has been proposed that obesity-related hypertension is caused by adipocyte-derived factors that are recognized as undetermined proteins secreted from adipocytes. Adipocyte-derived factors have been known to be related to aldosterone secretion in the adrenal gland. So far, Wnt proteins, CTRP-1, VLDL, LDL, HDL and leptin have been demonstrated to stimulate aldosterone secretion. In contrast, it has not yet been clarified whether adipocyte-derived factors also affect adrenal cortisol secretion. METHODS AND RESULTS: In the present study, we investigated the effect of adipocyte-derived factors on cortisol synthase gene CYP11B1 mRNA expression in vitro study using adrenocortical carcinoma H295R cells and mouse fibroblast 3T3-L1cells. Interestingly, adipocyte-derived factors were demonstrated to have the ability to stimulate CYP11B1 mRNA expression. CONCLUSION: Since CYP11B1 is well known as a limiting enzyme of cortisol synthesis, our study suggests that adipocyte-derived factors may stimulate cortisol secretion, as well as aldosterone secretion. Taken together, adipocyte-derived factors may be the cause of metabolic syndrome due to their stimulating effects on aldosterone/cortisol secretion. Therefore, the innovation of novel drugs against them may possibly be a new approach against metabolic syndrome.
Assuntos
Adipócitos/química , Córtex Suprarrenal/efeitos dos fármacos , Citocromo P-450 CYP11B2/biossíntese , Esteroide 11-beta-Hidroxilase/biossíntese , Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/metabolismo , Neoplasias do Córtex Suprarrenal/patologia , Animais , Carcinoma/metabolismo , Carcinoma/patologia , Linhagem Celular , Linhagem Celular Tumoral , Citocromo P-450 CYP11B2/genética , Fibroblastos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hidrocortisona/metabolismo , Leptina/metabolismo , Leptina/farmacologia , Lipoproteínas LDL/metabolismo , Lipoproteínas LDL/farmacologia , Camundongos , Proteínas/genética , Proteínas/fisiologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Esteroide 11-beta-Hidroxilase/genética , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologia , Zona Fasciculada/efeitos dos fármacos , Zona Fasciculada/metabolismoRESUMO
Necrotizing enterocolitis (NEC) is a devastating neonatal disease characterized by acute intestinal injury. Intestinal stem cell (ISC) renewal is required for gut regeneration in response to acute injury. The Wnt/ß-catenin pathway is essential for intestinal renewal and ISC maintenance. We found that ISC expression, Wnt activity and intestinal regeneration were all decreased in both mice with experimental NEC and in infants with acute active NEC. Moreover, intestinal organoids derived from NEC-injured intestine of both mice and humans failed to maintain proliferation and presented more differentiation. Administration of Wnt7b reversed these changes and promoted growth of intestinal organoids. Additionally, administration of exogenous Wnt7b rescued intestinal injury, restored ISC, and reestablished intestinal epithelial homeostasis in mice with NEC. Our findings demonstrate that during NEC, Wnt/ß-catenin signaling is decreased, ISC activity is impaired, and intestinal regeneration is defective. Administration of Wnt resulted in the maintenance of intestinal epithelial homeostasis and avoidance of NEC intestinal injury.
Assuntos
Enterocolite Necrosante/fisiopatologia , Intestinos/fisiopatologia , Regeneração/fisiologia , Via de Sinalização Wnt , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Enterocolite Necrosante/genética , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Intestinos/efeitos dos fármacos , Intestinos/patologia , Camundongos Endogâmicos C57BL , Modelos Biológicos , Organoides/efeitos dos fármacos , Organoides/metabolismo , Proteínas Proto-Oncogênicas/administração & dosagem , Proteínas Proto-Oncogênicas/farmacologia , Regeneração/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Análise de Sobrevida , Proteínas Wnt/administração & dosagem , Proteínas Wnt/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Via de Sinalização Wnt/genéticaRESUMO
KEY POINTS: Wnt ligands belonging to both canonical and non-canonical Wnt pathways regulate membrane potential signifying a very early event in the signal transduction. Wnts activate K+ currents by elevating intracellular Ca2+ and trigger Ca2+ release from intracellular stores. Control of potential by Wnt ligands has significant implications for gene transcription and opens up a novel avenue to interfere with this critical pathway. ABSTRACT: The Wnt signalling network determines gene transcription with free intracellular Ca2+ ( Cai2+ ) and ß-catenin as major intracellular signal transducers. Despite its critical importance during development and disease, many of the basic mechanisms of Wnt signal activation remain unclear. Here we show by single cell recording and simultaneous Cai2+ imaging in mammalian prostate cancer cells that an early step in the signal cascade is direct action on the cell membrane potential. We show that Wnt ligands 5A, 9B and 10B rapidly hyperpolarized the cells by activating K+ current by Ca2+ release from intracellular stores. Medium-throughput multi-well recordings showed responses to Wnts at concentrations of 2 nm. We identify a putative target for early events as a TRPM channel. Wnts thus act as ligands for ion channel activation in mammalian cells and membrane potential is an early indicator of control of transcription.
Assuntos
Sinalização do Cálcio , Potenciais da Membrana , Via de Sinalização Wnt , Membrana Celular/efeitos dos fármacos , Membrana Celular/fisiologia , Humanos , Células MCF-7 , Canais de Potássio/metabolismo , Canais de Cátion TRPM/metabolismo , Proteínas Wnt/metabolismo , Proteínas Wnt/farmacologiaRESUMO
Mutations and inadequate methylation profiles of CITED2 are associated with human congenital heart disease (CHD). In mouse, Cited2 is necessary for embryogenesis, particularly for heart development, and its depletion in embryonic stem cells (ESC) impairs cardiac differentiation. We have now determined that Cited2 depletion in ESC affects the expression of transcription factors and cardiopoietic genes involved in early mesoderm and cardiac specification. Interestingly, the supplementation of the secretome prepared from ESC overexpressing CITED2, during the onset of differentiation, rescued the cardiogenic defects of Cited2-depleted ESC. In addition, we demonstrate that the proteins WNT5A and WNT11 held the potential for rescue. We also validated the zebrafish as a model to investigate cited2 function during development. Indeed, the microinjection of morpholinos targeting cited2 transcripts caused developmental defects recapitulating those of mice knockout models, including the increased propensity for cardiac defects and severe death rate. Importantly, the co-injection of anti-cited2 morpholinos with either CITED2 or WNT5A and WNT11 recombinant proteins corrected the developmental defects of Cited2-morphants. This study argues that defects caused by the dysfunction of Cited2 at early stages of development, including heart anomalies, may be remediable by supplementation of exogenous molecules, offering the opportunity to develop novel therapeutic strategies aiming to prevent CHD.
Assuntos
Cardiopatias Congênitas/metabolismo , Células-Tronco Embrionárias Murinas/metabolismo , Proteínas Repressoras/metabolismo , Transativadores/metabolismo , Proteínas Wnt/farmacologia , Proteína Wnt-5a/farmacologia , Peixe-Zebra/embriologia , Animais , Diferenciação Celular/genética , Linhagem Celular , Modelos Animais de Doenças , Feminino , Cardiopatias Congênitas/prevenção & controle , Masculino , Camundongos , Camundongos Knockout , Morfolinos/administração & dosagem , Morfolinos/farmacologia , Proteínas Recombinantes/farmacologia , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Transativadores/antagonistas & inibidores , Transativadores/genética , TransfecçãoRESUMO
Over the past 15 years, many studies have revealed that Wnt signaling has a strong impact on hematopoietic stem cell fate. After a controversy over the interpretation of some results, the current understanding is that an appropriate degree of canonical Wnt signaling induces hematopoietic stem cell self-renewal and that noncanonical Wnt signaling keeps the quiescence. It is also likely that the balance between canonical and noncanonical Wnt pathways regulates the stress response and aging of hematopoietic stem cells.
Assuntos
Células-Tronco Hematopoéticas/efeitos dos fármacos , Proteínas Wnt/farmacologia , Via de Sinalização Wnt , Diferenciação Celular , Senescência Celular , Células-Tronco Hematopoéticas/fisiologia , HumanosRESUMO
Class III Semaphorin (Sema) secreted ligands are known to repel neurites expressing Neuropilin (Nrp) and/or Plexin (Plxn) receptors. There is, however, a growing body of literature supporting that Sema signaling also has alternative roles in development such as synaptogenesis, boundary formation, and vasculogenesis. To evaluate these options during inner ear development, we used in situ hybridization or immunohistochemistry to map the expression of Sema3D, Sema3F, Nrp1, Nrp2, and PlxnA1 in the chicken (Gallus gallus) inner ear from embryonic day (E)5-E10. The resulting expression patterns in either the otic epithelium or its surrounding mesenchyme suggest that Sema signaling could be involved in each of the varied functions reported for other tissues. Sema3D expression flanking the sensory tissue in vestibular organs suggests that it may repel Nrp2- and PlxnA1-expressing neurites of the vestibular ganglion away from nonsensory epithelia, thus channeling them into the sensory domains at E5-E8. Expression of Sema signaling genes in the sensory hair cells of both the auditory and vestibular organs on E8-E10 may implicate Sema signaling in synaptogenesis. In the nonsensory regions of the cochlea, Sema3D in the future tegmentum vasculosum opposes Nrp1 and PlxnA1 in the future cuboidal cells; the abutment of ligand and receptors in adjacent domains may enforce or maintain the boundary between them. In the mesenchyme, Nrp1 colocalized with capillary-rich tissue. Sema3D immediately flanks this Nrp1-expressing tissue, suggesting a role in endothelial cell migration towards the inner ear. In summary, Sema signaling may play multiple roles in the developing inner ear.
Assuntos
Galinhas/metabolismo , Orelha Interna/metabolismo , Neuropilina-1/biossíntese , Semaforinas/biossíntese , Animais , Movimento Celular , Embrião de Galinha , Orelha Interna/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Neurogênese , Neuropilina-1/genética , Órgão Espiral/embriologia , Órgão Espiral/metabolismo , Semaforinas/genética , Transdução de Sinais , Organismos Livres de Patógenos Específicos , Gânglio Espiral da Cóclea/embriologia , Gânglio Espiral da Cóclea/metabolismo , Proteínas Wnt/farmacologia , Proteínas Wnt/fisiologiaRESUMO
Wnt signaling regulates brain development and synapse maturation; however, the precise molecular mechanism remains elusive. Here, we report that Wnt-7a stimulates dendritic spine morphogenesis in the hippocampus via glycogen synthase kinase-3 ß (GSK-3ß) inhibition, triggering ß-catenin/T cell factor/lymphoid enhancer factor (TCF/LEF)-dependent gene transcription and promoting postsynaptic density-95 (PSD-95) protein expression. In addition, wild-type mice treated with an inhibitor of ß-catenin/TCF/LEF-mediated transcription showed a reduction in spatial memory acquisition accompanied by a reduction in PSD-95 and decreases in spine density measured by Golgi staining, suggesting that PSD-95 is a novel Wnt target gene. Together, our data strongly demonstrate that Wnt-dependent target gene transcription is essential to hippocampal synaptic plasticity.
Assuntos
Espinhas Dendríticas/metabolismo , Proteína 4 Homóloga a Disks-Large/metabolismo , Plasticidade Neuronal/fisiologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Wnt/farmacologia , Animais , Espinhas Dendríticas/efeitos dos fármacos , Glicogênio Sintase Quinase 3 beta/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Memória/efeitos dos fármacos , Memória/fisiologia , Camundongos , Plasticidade Neuronal/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , beta Catenina/metabolismoRESUMO
BACKGROUND: Alterations in maternal environment can sometimes affect embryonic development in a sexually-dimorphic manner. The objective was to determine whether preimplantation bovine embryos respond to three maternally-derived cell signaling molecules in a sex-dependent manner. RESULTS: Actions of three embryokines known to increase competence of bovine embryos to develop to the blastocyst stage, insulin-like growth factor 1 (IGF1), activin A, and WNT member 7A (WNT7A), were evaluated for actions on embryos produced in vitro with X- or Y- sorted semen from the same bull. Each embryokine was tested in embryos produced by in vitro fertilization of groups of oocytes with either pooled sperm from two bulls or with sperm from individual bulls. Embryos were treated with IGF1, activin A, or WNT7A on day 5 of culture. All three embryokines increased the proportion of cleaved zygotes that developed to the blastocyst stage and the effect was similar for female and male embryos. As an additional test of sexual dimorphism, effects of IGF1 on blastocyst expression of a total of 127 genes were determined by RT-qPCR using the Fluidigm Delta Gene assay. Expression of 18 genes was affected by sex, expression of 4 genes was affected by IGF1 and expression of 3 genes was affected by the IGF1 by sex interaction. CONCLUSION: Sex did not alter how IGF1, activin A or WNT7A altered developmental competence to the blastocyst stage. Thus, sex-dependent differences in regulation of developmental competence of embryos by maternal regulatory signals is not a general phenomenon. The fact that sex altered how IGF1 regulates gene expression is indicative that there could be sexual dimorphism in embryokine regulation of some aspects of embryonic function other than developmental potential to become a blastocyst.
Assuntos
Blastocisto/efeitos dos fármacos , Subunidades beta de Inibinas/farmacologia , Fator de Crescimento Insulin-Like I/farmacologia , Caracteres Sexuais , Proteínas Wnt/farmacologia , Animais , Bovinos , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , MasculinoRESUMO
Recently several studies demonstrated a role for the Wnt pathway in lymphocyte development and self-renewal of hematopoietic stem cells (HSCs). B-1 cells constitute a separate lineage of B lymphocytes, originating during fetal hematopoiesis, expressing lymphoid and myeloid markers and possessing self-renewal ability, similar to early hematopoietic progenitors and HSCs. A plethora of studies have shown an important role for the evolutionary conserved Wnt pathway in the biology of HSCs and T lymphocyte development. Our previous data demonstrated abundant expression of Wnt pathway components by B-1 cells, including Wnt ligands and receptors. Here we report that the canonical Wnt pathway is activated in B-1 cell precursors, but not in mature B-1 cells. However, both B-1 precursors and B-1 cells are able to respond to Wnt ligands in vitro. Canonical Wnt activity promotes proliferation of B-1 cells, while non-canonical Wnt signals induce the expansion of B-1 precursors. Interestingly, using a co-culture system with OP9 cells, Wnt3a stimulus supported the generation of B-1a cells. Taking together, these results indicate that B-1 cells and their progenitors are differentially responsive to Wnt ligands, and that the balance of activation of canonical and non-canonical Wnt signaling may regulate the maintenance and differentiation of different B-1 cell subsets.
Assuntos
Linfócitos B/citologia , Linfócitos B/metabolismo , Células-Tronco/metabolismo , Via de Sinalização Wnt , Animais , Linfócitos B/efeitos dos fármacos , Antígenos CD5/metabolismo , Proliferação de Células/efeitos dos fármacos , Feminino , Interleucina-7/farmacologia , Ligantes , Camundongos Endogâmicos C57BL , Receptores de Interleucina-7/metabolismo , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Proteínas Wnt/farmacologia , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
Autografts tend to be unreliable in older patients. Some of these age-related skeletal changes appear to be attributable to a decline in endogenous WNT signaling. We used a functional in vivo transplantation assay to demonstrate that the bone-forming capacity of an autograft can be traced back to a Wnt-responsive cell population associated with the mineralized bone matrix fraction of a bone graft. Micro-CT imaging, flow cytometry and quantitative analyses demonstrate that this mineralized fraction declines with age, along with a waning in endogenous Wnt signaling; together these factors contribute to the age-related deterioration in autograft efficacy. Using a lipid formulation to stabilize the hydrophobic WNT3A protein, we demonstrate that osteogenic capacity can be restored by incubating the bone graft ex vivo with WNT3A. Compared to control bone grafts, WNT-treated bone grafts give rise to three times more bone. These preclinical results establish a pivotal role for WNT signaling in the age-related decline of autologous bone grafting efficacy, and demonstrate a means to restore that efficacy via local, transient amplification of endogenous Wnt signaling.
Assuntos
Regeneração Óssea/efeitos dos fármacos , Transplante Ósseo , Osteogênese/efeitos dos fármacos , Proteínas Wnt/farmacologia , Fatores Etários , Animais , Densidade Óssea , Calcificação Fisiológica , Modelos Animais de Doenças , Matriz Extracelular , Camundongos , Camundongos Knockout , Transplante Autólogo , Via de Sinalização WntRESUMO
Human perivascular stem/stromal cells (hPSC) are a multipotent mesenchymogenic stromal cell population defined by their perivascular locale. Recent studies have demonstrated the high potential for clinical translation of this fluorescence-activated cell sorting (FACS)-derived cell population for autologous bone tissue engineering. However, the mechanisms underlying the osteogenic differentiation of PSC are incompletely understood. The current study investigates the roles of canonical and noncanonical Wnt signaling in the osteogenic and adipogenic differentiation of PSC. Results showed that both canonical and noncanonical Wnt signaling activity transiently increased during PSC osteogenic differentiation in vitro. Sustained WNT3A treatment significantly decreased PSC osteogenic differentiation. Conversely, sustained treatment with Wnt family member 16 (WNT16), a mixed canonical and noncanonical ligand, increased osteogenic differentiation in a c-Jun N-terminal kinase (JNK) pathway-dependent manner. Conversely, WNT16 knockdown significantly diminished PSC osteogenic differentiation. Finally, WNT16 but not WNT3A increased the adipogenic differentiation of PSC. These results indicate the importance of regulation of canonical and noncanonical Wnt signaling for PSC fate and differentiation. Moreover, these data suggest that WNT16 plays a functional and necessary role in PSC osteogenesis.