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1.
Int J Biochem Cell Biol ; 94: 71-78, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29203233

RESUMO

Notch signaling plays an essential role in the proliferation, differentiation and cell fate determination of various tissues, including the developing pancreas. One regulator of the Notch pathway is GDE2 (or GDPD5), a transmembrane ecto-phosphodiesterase that cleaves GPI-anchored proteins at the plasma membrane, including a Notch ligand regulator. Here we report that Gdpd5-knockdown in zebrafish embryos leads to developmental defects, particularly, impaired motility and reduced pancreas differentiation, as shown by decreased expression of insulin and other pancreatic markers. Exogenous expression of human GDE2, but not catalytically dead GDE2, similarly leads to developmental defects. Human GDE2 restores insulin expression in Gdpd5a-depleted zebrafish embryos. Importantly, zebrafish Gdpd5 orthologues localize to the plasma membrane where they show catalytic activity against GPI-anchored GPC6. Thus, our data reveal functional conservation between zebrafish Gdpd5 and human GDE2, and suggest that strict regulation of GDE2 expression and catalytic activity is critical for correct embryonic patterning. In particular, our data uncover a role for GDE2 in regulating pancreas differentiation.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Organogênese , Pâncreas/metabolismo , Diester Fosfórico Hidrolases/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Animais , Embrião não Mamífero/anormalidades , Embrião não Mamífero/diagnóstico por imagem , Embrião não Mamífero/metabolismo , Técnicas de Silenciamento de Genes , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Morfolinos/metabolismo , Pâncreas/diagnóstico por imagem , Pâncreas/embriologia , Fragmentos de Peptídeos/antagonistas & inibidores , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Diester Fosfórico Hidrolases/química , Diester Fosfórico Hidrolases/genética , Filogenia , Domínios Proteicos , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Homologia de Sequência de Aminoácidos , Peixe-Zebra , Proteínas de Peixe-Zebra/antagonistas & inibidores , Proteínas de Peixe-Zebra/química , Proteínas de Peixe-Zebra/genética
2.
Sci Rep ; 7(1): 7327, 2017 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-28779098

RESUMO

Autophagy is an evolutionarily conserved process that degrades cellular components to restore energy homeostasis under limited nutrient conditions. How this starvation-induced autophagy is regulated at the whole-body level is not fully understood. Here, we show that the tumor suppressor Lkb1, which activates the key energy sensor AMPK, also regulates starvation-induced autophagy at the organismal level. Lkb1-deficient zebrafish larvae fail to activate autophagy in response to nutrient restriction upon yolk termination, shown by reduced levels of the autophagy-activating proteins Atg5, Lc3-II and Becn1, and aberrant accumulation of the cargo receptor and autophagy substrate p62. We demonstrate that the autophagy defect in lkb1 mutants can be partially rescued by inhibiting mTOR signaling but not by inhibiting the PI3K pathway. Interestingly, mTOR-independent activation of autophagy restores degradation of the aberrantly accumulated p62 in lkb1 mutants and prolongs their survival. Our data uncover a novel critical role for Lkb1 in regulating starvation-induced autophagy at the organismal level, providing mechanistic insight into metabolic adaptation during development.


Assuntos
Autofagia , Proteínas Serina-Treonina Quinases/metabolismo , Inanição , Estresse Fisiológico , Proteínas Supressoras de Tumor/metabolismo , Animais , Autofagia/genética , Biomarcadores , Imunofluorescência , Imuno-Histoquímica , Larva , Mutação , Proteínas Serina-Treonina Quinases/genética , Estresse Fisiológico/genética , Serina-Treonina Quinases TOR/metabolismo , Proteínas Supressoras de Tumor/genética , Peixe-Zebra
3.
Clin Cancer Res ; 18(14): 3961-3971, 2012 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-22573349

RESUMO

PURPOSE: To investigate the angiogenic changes in primary tumor tissue of renal cell carcinoma (RCC) patients treated with VEGF-targeted therapy. EXPERIMENTAL DESIGN: Phase II trials of VEGF pathway-targeted therapy given before cytoreductive surgery were carried out with metastatic RCC patients with the primary tumor in situ to investigate the necessity of nephrectomy. Primary tumor tissues were obtained and assessed for angiogenesis parameters. Results were compared with similar analyses on untreated tumors. RESULTS: Sunitinib or bevacizumab pretreatment resulted in a significant reduction of microvessel density in the primary tumor. Also, an increase in vascular pericyte coverage was found in sunitinib-pretreated tumors, consistent with efficient angiogenesis inhibition. Expression of several key regulators of angiogenesis was found to be suppressed in pretreated tissues, among which VEGFR-1 and VEGFR-2, angiopoietin-1 and angiopoietin-2 and platelet-derived growth factor-B. In addition, apoptosis in tumor and endothelial cells was induced. Interestingly, in sunitinib-pretreated tissues a dramatic increase of the number of proliferating endothelial cells was observed, which was not the case in bevacizumab-pretreated tumors. A positive correlation with the interval between halting the therapy and surgery was found, suggesting a compensatory angiogenic response caused by the discontinuation of sunitinib treatment. CONCLUSION: This study describes, for the first time, the angiostatic response in human primary renal cancers at the tissue level upon treatment with VEGF-targeted therapy. Discontinuation of treatment with tyrosine kinase inhibitors leads to accelerated endothelial cell proliferation. The results of this study contribute important data to the ongoing discussion on the discontinuation of treatment with kinase inhibitors.


Assuntos
Anticorpos Monoclonais Humanizados/administração & dosagem , Carcinoma de Células Renais/tratamento farmacológico , Indóis/administração & dosagem , Neovascularização Patológica/tratamento farmacológico , Pirróis/administração & dosagem , Adulto , Idoso , Apoptose/efeitos dos fármacos , Bevacizumab , Carcinoma de Células Renais/complicações , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/secundário , Ensaios Clínicos Fase II como Assunto , Feminino , Humanos , Masculino , Microvasos/efeitos dos fármacos , Pessoa de Meia-Idade , Neovascularização Patológica/complicações , Estudos Retrospectivos , Sunitinibe
4.
J Cell Mol Med ; 16(9): 2035-48, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22128761

RESUMO

Prolactin is best known as the polypeptide anterior pituitary hormone, which regulates the development of the mammary gland. However, it became clear over the last decade that prolactin contributes to a broad range of pathologies, including breast cancer. Prolactin is also involved in angiogenesis via the release of pro-angiogenic factors by leukocytes and epithelial cells. However, whether prolactin also influences endothelial cells, and whether there are functional consequences of prolactin-induced signalling in the perspective of angiogenesis, remains so far elusive. In the present study, we show that prolactin induces phosphorylation of ERK1/2 and STAT5 and induces tube formation of endothelial cells on Matrigel. These effects are blocked by a specific prolactin receptor antagonist, del1-9-G129R-hPRL. Moreover, in an in vivo model of the chorioallantoic membrane of the chicken embryo, prolactin enhances vessel density and the tortuosity of the vasculature and pillar formation, which are hallmarks of intussusceptive angiogenesis. Interestingly, while prolactin has only little effect on endothelial cell proliferation, it markedly stimulates endothelial cell migration. Again, migration was reverted by del1-9-G129R-hPRL, indicating a direct effect of prolactin on its receptor. Immunohistochemistry and spectral imaging revealed that the prolactin receptor is present in the microvasculature of human breast carcinoma tissue. Altogether, these results suggest that prolactin may directly stimulate angiogenesis, which could be one of the mechanisms by which prolactin contributes to breast cancer progression, thereby providing a potential tool for intervention.


Assuntos
Células Endoteliais/patologia , Neovascularização Patológica/patologia , Prolactina/efeitos adversos , Transdução de Sinais/efeitos dos fármacos , Indutores da Angiogênese/efeitos adversos , Animais , Neoplasias da Mama/patologia , Linhagem Celular , Embrião de Galinha , Colágeno/metabolismo , Combinação de Medicamentos , Células Endoteliais/metabolismo , Feminino , Imuno-Histoquímica , Laminina/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Fosforilação , Proteoglicanas/metabolismo , Receptores da Prolactina/antagonistas & inibidores , Receptores da Prolactina/metabolismo , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo
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