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1.
Gastroenterology ; 157(5): 1413-1428.e11, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31352001

RESUMO

BACKGROUND & AIMS: Obesity is a risk factor for pancreatic cancer. In mice, a high-fat diet (HFD) and expression of oncogenic KRAS lead to development of invasive pancreatic ductal adenocarcinoma (PDAC) by unknown mechanisms. We investigated how oncogenic KRAS regulates the expression of fibroblast growth factor 21, FGF21, a metabolic regulator that prevents obesity, and the effects of recombinant human FGF21 (rhFGF21) on pancreatic tumorigenesis. METHODS: We performed immunohistochemical analyses of FGF21 levels in human pancreatic tissue arrays, comprising 59 PDAC specimens and 45 nontumor tissues. We also studied mice with tamoxifen-inducible expression of oncogenic KRAS in acinar cells (KrasG12D/+ mice) and fElasCreERT mice (controls). KrasG12D/+ mice were placed on an HFD or regular chow diet (control) and given injections of rhFGF21 or vehicle; pancreata were collected and analyzed by histology, immunoblots, quantitative polymerase chain reaction, and immunohistochemistry. We measured markers of inflammation in the pancreas, liver, and adipose tissue. Activity of RAS was measured based on the amount of bound guanosine triphosphate. RESULTS: Pancreatic tissues of mice expressed high levels of FGF21 compared with liver tissues. FGF21 and its receptor proteins were expressed by acinar cells. Acinar cells that expressed KrasG12D/+ had significantly lower expression of Fgf21 messenger RNA compared with acinar cells from control mice, partly due to down-regulation of PPARG expression-a transcription factor that activates Fgf21 transcription. Pancreata from KrasG12D/+ mice on a control diet and given injections of rhFGF21 had reduced pancreatic inflammation, infiltration by immune cells, and acinar-to-ductal metaplasia compared with mice given injections of vehicle. HFD-fed KrasG12D/+ mice given injections of vehicle accumulated abdominal fat, developed extensive inflammation, pancreatic cysts, and high-grade pancreatic intraepithelial neoplasias (PanINs); half the mice developed PDAC with liver metastases. HFD-fed KrasG12D/+ mice given injections of rhFGF21 had reduced accumulation of abdominal fat and pancreatic triglycerides, fewer pancreatic cysts, reduced systemic and pancreatic markers of inflammation, fewer PanINs, and longer survival-only approximately 12% of the mice developed PDACs, and none of the mice had metastases. Pancreata from HFD-fed KrasG12D/+ mice given injections of rhFGF21 had lower levels of active RAS than from mice given vehicle. CONCLUSIONS: Normal acinar cells from mice and humans express high levels of FGF21. In mice, acinar expression of oncogenic KRAS significantly reduces FGF21 expression. When these mice are placed on an HFD, they develop extensive inflammation, pancreatic cysts, PanINs, and PDACs, which are reduced by injection of FGF21. FGF21 also reduces the guanosine triphosphate binding capacity of RAS. FGF21 might be used in the prevention or treatment of pancreatic cancer.


Assuntos
Células Acinares/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Transformação Celular Neoplásica/metabolismo , Dieta Hiperlipídica , Fatores de Crescimento de Fibroblastos/metabolismo , Neoplasias Intraductais Pancreáticas/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Células Acinares/patologia , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/prevenção & controle , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Regulação para Baixo , Fatores de Crescimento de Fibroblastos/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Transgênicos , Mutação , PPAR gama/genética , PPAR gama/metabolismo , Cisto Pancreático/genética , Cisto Pancreático/metabolismo , Cisto Pancreático/patologia , Neoplasias Intraductais Pancreáticas/genética , Neoplasias Intraductais Pancreáticas/patologia , Neoplasias Intraductais Pancreáticas/prevenção & controle , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/prevenção & controle , Pancreatite/genética , Pancreatite/metabolismo , Pancreatite/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
2.
Cell Commun Signal ; 17(1): 19, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30819189

RESUMO

Oncogenic KRAS plays a vital role in controlling tumor metabolism by enhancing aerobic glycolysis. Obesity driven by chronic consumption of high-fat diet (HFD) is a major risk factor for oncogenic KRAS-mediated pancreatic ductal adenocarcinoma (PDAC). However, the role of HFD in KRAS-mediated metabolic reprogramming has been obscure. Here, by using genetically engineered mouse models expressing an endogenous level of KRASG12D in pancreatic acinar cells, we demonstrate that hyperactivation of KRASG12D by obesogenic HFD, as compared to carbohydrate-rich diet, is responsible for enhanced aerobic glycolysis that associates with critical pathogenic responses in the path towards PDAC. Ablation of Cox-2 attenuates KRAS hyperactivation leading to the reversal of both aggravated aerobic glycolysis and high-grade dysplasia under HFD challenge. Our data highlight a pivotal role of the cooperative interaction between obesity-ensuing HFD and oncogenic KRAS in driving the heightened aerobic glycolysis during pancreatic tumorigenesis and suggest that in addition to directly targeting KRAS and aerobic glycolysis pathway, strategies to target the upstream of KRAS hyperactivation may bear important therapeutic value.


Assuntos
Dieta Hiperlipídica , Glicólise , Obesidade/metabolismo , Oncogenes , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Aerobiose , Animais , Ciclo-Oxigenase 2/metabolismo , Carboidratos da Dieta , Camundongos , Modelos Biológicos , Obesidade/patologia , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia
3.
Trends Pharmacol Sci ; 40(2): 142-153, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30616873

RESUMO

Endocrine fibroblast growth factors (eFGFs) control pathways that are crucial for maintaining metabolic homeostasis of lipids, glucose, energy, bile acids, and minerals. Unlike the heparin-binding paracrine FGFs, eFGFs require a unique Klotho family protein to form a productive triad complex, but the structural and mechanistical details of this complex have remained obscure since the beginning of the eFGF field. However, recent breakthroughs in resolving the 3D structures of eFGF signaling complexes have now unveiled the atomic details of multivalent interactions among eFGF, FGFR, and Klotho. We provide here a timely review on the architecture and the structure-function relationships of these complexes, and highlight how the structural knowledge opens a new door to structure-based drug design against a repertoire of eFGF-associated metabolic diseases.


Assuntos
Fatores de Crescimento de Fibroblastos/química , Fatores de Crescimento de Fibroblastos/metabolismo , Doenças Metabólicas/tratamento farmacológico , Doenças Metabólicas/metabolismo , Animais , Desenho de Drogas , Sistema Endócrino/metabolismo , Glucuronidase/química , Glucuronidase/metabolismo , Humanos , Receptores de Fatores de Crescimento de Fibroblastos/química , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Relação Estrutura-Atividade
4.
Am J Physiol Gastrointest Liver Physiol ; 316(1): G179-G186, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30431318

RESUMO

Replacement of the exocrine parenchyma by fibrous tissue is a main characteristic of chronic pancreatitis. Understanding the mechanisms of pancreatic fibrogenesis is critical for the development of preventive and therapeutic interventions. Cyclooxygenase-2 (COX-2), a rate-limiting enzyme for prostaglandin synthesis, is expressed in patients with chronic pancreatitis. However, it is unknown whether COX-2 can cause chronic pancreatitis. To investigate the roles of pancreatic acinar COX-2 in fibrogenesis and the development of chronic pancreatitis, COX-2 was ectopically expressed specifically in pancreatic acinar cells in transgenic mice. Histopathological changes and expression levels of several profibrogenic factors related to chronic pancreatitis were evaluated. COX-2 was expressed in the pancreas of the transgenic mice, as detected by Western blot analysis. Immunohistochemical staining showed COX-2 was specifically expressed in pancreatic acinar cells. COX-2 expression led to progressive changes in the pancreas, including pancreas megaly, persistent inflammation, collagen deposition, and acinar-to-ductal metaplasia. Quantitative RT-PCR and immunostaining showed that profibrogenic factors were upregulated and pancreatic stellate cells were activated in the COX-2 transgenic mice. Expression of COX-2 in pancreatic acinar cells is sufficient to induce chronic pancreatitis. Targeting this pathway may be valuable in the prevention of chronic pancreatitis. NEW & NOTEWORTHY COX-2 expression is observed in pancreatic tissues of human chronic pancreatitis. In this study, we showed that COX-2 expression caused the development of chronic pancreatitis in transgenic mice, supporting the idea that COX-2 inhibition may be an effective preventive and therapeutic strategy.


Assuntos
Células Acinares/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Pancreatite Crônica/metabolismo , Animais , Transformação Celular Neoplásica/metabolismo , Inflamação/metabolismo , Camundongos Transgênicos , Pâncreas/metabolismo , Pâncreas Exócrino/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Células Estreladas do Pâncreas/metabolismo
5.
Methods Mol Biol ; 1882: 207-219, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30378057

RESUMO

Pancreatic cancer is a highly lethal disease and is projected to become the second leading cause of cancer-related death by 2020. Among the different subtypes, pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic cancer. The genetic landscape of PDAC shows nearly ubiquitous mutations of KRAS. However, expression of KRAS somatic mutants alone is insufficient to drive PDAC. Redox deregulation may contribute significantly to KRAS-mediated PDAC. Thus, measurement of cellular reactive oxygen species (ROS) levels is essential to determine how oxidative stress affects mutant KRAS and modulates intracellular signaling pathways leading to the change of cellular functions and the development of PDAC. Here we describe the protocol for comparative measurement of several key forms of ROS, including intracellular and mitochondrial levels of superoxide as well as extracellular H2O2 and general cellular ROS, with oxidation-sensitive fluorescent probes using flow cytometry in pancreatic cancer cells or mutant KRAS transformed cells.


Assuntos
Carcinoma Ductal Pancreático/patologia , Citometria de Fluxo/métodos , Neoplasias Pancreáticas/patologia , Espécies Reativas de Oxigênio/análise , Carcinoma Ductal Pancreático/genética , Técnicas de Cultura de Células/instrumentação , Técnicas de Cultura de Células/métodos , Linhagem Celular Tumoral , Citometria de Fluxo/instrumentação , Corantes Fluorescentes/química , Humanos , Mitocôndrias/patologia , Mutação , Estresse Oxidativo/genética , Pâncreas/patologia , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/genética
6.
Trends Biochem Sci ; 43(8): 563-566, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29895507

RESUMO

Metabolic homeostasis is critical to cellular and organismal health. The newly revealed crystal structures of the endocrine factors FGF21 and FGF23, in association with the glycosidase coreceptor Klotho and transmembrane tyrosine kinase FGFR, set a platform for structure-based novel drug design against common metabolic disorders.


Assuntos
Doenças Metabólicas , Transdução de Sinais , Comunicação Celular , Homeostase , Humanos
9.
Cytokine Growth Factor Rev ; 38: 59-65, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28887067

RESUMO

FGF21 is a master regulator of homeostasis of local and systemic lipid, glucose and energy metabolism. Since its discovery a decade ago, significant progress has been made in understanding the basic molecular, cellular and physiological mechanisms underlying its metabolic roles, and characterizing its beneficial pharmacological activities and possible pathological roles in obesity, diabetes, dyslipidemia, fatty liver disease and their collateral complications and tissue damage. Under basal or normal conditions, FGF21 appears to play a dispensable role in metabolism. However, in response to a variety of cellular and metabolic stress, FGF21 is significantly upregulated to serve as a potent catabolic factor leading to the clearance of excessive lipids and glucose, and therefore, antagonizes metabolic and energy imbalance in a negative fashion. Furthermore, FGF21 treatment ameliorates tissue damage resulted from the harmful effects of metabolic abnormalities, which often ensue an oxidative, pro-inflammatory, inflammatory and/or immune stress state, the so-called metaflammation. Most notably, studies focusing on the liver, pancreas, cardio-vasculature and kidney have revealed its significant protective effects against the structural and functional damages induced by the obese, diabetic or other abnormal metabolic conditions. In this review, we will summarize the current progress on the roles of FGF21 against metaflammation and metabolic tissue damage.


Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Estresse Fisiológico/fisiologia , Animais , Humanos , Inflamação/metabolismo , Doenças Metabólicas/metabolismo
11.
Endocrinology ; 157(12): 4754-4769, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27690692

RESUMO

The antiobese and antidiabetic fibroblast growth factor 21 (FGF21) regulates lipid metabolism and energy homeostasis by targeting the ßKlotho-FGFR1 (fibroblast growth factor receptor 1) binary complex in adipose tissue adipocytes. Because lipid droplet is the organelle responsible for storing lipid energy in adipocytes, it is the plausible target of FGF21 action. However, the impact of the FGF21-ßKlotho-FGFR1 signaling pathway on the functions of the lipid droplet is not clearly understood. Using our mouse models of adipocyte-specific FGFR1 ablation and hepatic overexpression of FGF21 with diet-induced obesity established previously, we analyzed the alterations of the pathways involved in energy and substrate metabolism that is attributable to the dynamic functions of the lipid droplet. In addition to the previous reports showing that FGFR1 deficiency abrogated lipolysis, fatty acid oxidation, and energy expenditure promoted by the elevated FGF21 signal, we observed that the deficiency up-regulated the biosynthesis and remodeling of membrane phospholipids that are important for the biogenesis and expansion of the droplet, whereas the enhanced FGF21 signal constrained the biosynthesis of phospholipids. As a result, the loss of adipose FGFR1 led to a sustained droplet expansion and endoplasmic reticulum (ER) stress, whereas the enhanced FGF21 signal suppressed them in obesogenesis. These new findings reveal that the FGF21-ßKlotho-FGFR1 signaling axis plays roles in maintaining phospholipid homeostasis and the dynamic functions of the lipid droplet, whereas protecting against ER stress, and suggest a potential link of phospholipid biosynthesis, lipid droplet dynamics, ER stress, and energy homeostasis in adipose tissue coordinated by this signaling axis.


Assuntos
Estresse do Retículo Endoplasmático/fisiologia , Fatores de Crescimento de Fibroblastos/metabolismo , Homeostase/fisiologia , Gotículas Lipídicas/metabolismo , Obesidade/metabolismo , Fosfolipídeos/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Adipócitos/metabolismo , Tecido Adiposo/metabolismo , Animais , Metabolismo Energético/fisiologia , Fatores de Crescimento de Fibroblastos/genética , Metabolismo dos Lipídeos/fisiologia , Fígado/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Transdução de Sinais/fisiologia
12.
Int J Biol Sci ; 12(3): 338-46, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26929740

RESUMO

The genetic landscape of pancreatic cancer shows nearly ubiquitous mutations of K-RAS. However, oncogenic K-Ras(mt) alone is not sufficient to lead to pancreatic ductal adenocarcinoma (PDAC) in either human or in genetically modified adult mouse models. Many stimulants, such as high fat diet, CCK, LPS, PGE2 and others, have physiological effects at low concentrations that are mediated in part through modest increases in K-Ras activity. However, at high concentrations, they induce inflammation that, in the presence of oncogenic K-Ras expression, substantially accelerates PDAC formation. The mechanism involves increased activity of oncogenic K-Ras(mt). Unlike what has been proposed in the standard paradigm for the role of Ras in oncogenesis, oncogenic K-Ras(mt) is now known to not be constitutively active. Rather, it can be activated by standard mechanisms similar to wild-type K-Ras, but its activity is sustained for a prolonged period. Furthermore, if the level of K-Ras activity exceeds a threshold at which it begins to generate its own activators, then a feed-forward loop is formed between K-Ras activity and inflammation and pathological processes including oncogenesis are initiated. Oncogenic K-Ras(mt) activation, a key event in PDAC initiation and development, is subject to complex regulatory mechanisms. Reagents which inhibit inflammation, such as the Cox2 inhibitor celecoxib, block the feed-forward loop and prevent induction of PDAC in models with endogenous oncogenic K-Ras(mt). Increased understanding of the role of activating and inhibitory mechanisms on oncogenic K-Ras(mt) activity is of paramount importance for the development of preventive and therapeutic strategies to fight against this lethal disease.


Assuntos
Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Animais , Humanos , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/genética
13.
Mol Cell ; 61(4): 520-534, 2016 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-26853146

RESUMO

Altered energy metabolism is a cancer hallmark as malignant cells tailor their metabolic pathways to meet their energy requirements. Glucose and glutamine are the major nutrients that fuel cellular metabolism, and the pathways utilizing these nutrients are often altered in cancer. Here, we show that the long ncRNA CCAT2, located at the 8q24 amplicon on cancer risk-associated rs6983267 SNP, regulates cancer metabolism in vitro and in vivo in an allele-specific manner by binding the Cleavage Factor I (CFIm) complex with distinct affinities for the two subunits (CFIm25 and CFIm68). The CCAT2 interaction with the CFIm complex fine-tunes the alternative splicing of Glutaminase (GLS) by selecting the poly(A) site in intron 14 of the precursor mRNA. These findings uncover a complex, allele-specific regulatory mechanism of cancer metabolism orchestrated by the two alleles of a long ncRNA.


Assuntos
Glutaminase/genética , Neoplasias/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo , Alelos , Processamento Alternativo , Metabolismo Energético , Células HCT116 , Humanos , Neoplasias/genética , Precursores de RNA/química , Precursores de RNA/metabolismo , RNA Mensageiro/metabolismo
15.
Stem Cell Res Ther ; 6: 198, 2015 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-26472041

RESUMO

INTRODUCTION: Cancer stem cells (CSCs) possess characteristics associated with normal stem cells, specifically the abilities to renew themselves and to give rise to all cell types (differentiation). It is assumed that induction of differentiation in CSCs would reduce their ability to form tumors. What triggers CSC differentiation and the role of "differentiation" in tumorigenesis remain elusive. METHODS: Glioma stem cell (GSC) lines and subcutaneous as well as orthotopic xenografts established from fresh surgical specimens of glioblastoma multiforme were used. RESULTS: Exposure of GSCs to serum activates mitochondrial respiration and causes an increase in mitochondrial reactive oxygen species (ROS) as well as oxidative stress responses, leading to the appearance of differentiation morphology and a deceased expression of CSC markers. Chemical perturbation of the mitochondrial electron transport chain causes ROS increase and further downregulation of stem cell markers, while antioxidant N-acetyl-cysteine reduces ROS and suppresses the differentiation of GSCs. Surprisingly, the serum-induced differentiated GSCs exhibit greater ability to form tumor in both orthotopic and subcutaneous xenograft models, which can be suppressed by N-acetyl-cysteine. Mitochondrial ROS from the serum-stimulated cells triggered the activation of nuclear factor-kappa-B (NFκB) pathway, which is a potential mechanism for the promotion of tumorigenesis. CONCLUSION: This study suggests that ROS generated from active mitochondrial respiration in the presence of serum is critical in CSCs activation, which promotes tumor development in vivo.


Assuntos
Glioma/metabolismo , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ativação Metabólica , Animais , Antioxidantes/farmacologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Diferenciação Celular , Linhagem Celular Tumoral , Meios de Cultura , Transporte de Elétrons , Glioblastoma/metabolismo , Glioma/patologia , Humanos , Camundongos , Camundongos Nus , Camundongos SCID , Mitocôndrias/efeitos dos fármacos , NF-kappa B/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Estresse Oxidativo , Transdução de Sinais
16.
Int J Mol Sci ; 16(10): 24011-31, 2015 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-26473837

RESUMO

Most anti-angiogenic therapies currently being evaluated target the vascular endothelial growth factor (VEGF) pathway; however, the tumor vasculature can acquire resistance to VEGF-targeted therapy by shifting to other angiogenesis mechanisms. Therefore, other therapeutic agents that block non-VEGF angiogenic pathways need to be evaluated. Here, we identified ferulic acid as a novel fibroblast growth factor receptor 1 (FGFR1) inhibitor and a novel agent with potential anti-angiogenic and anti-cancer activities. Ferulic acid demonstrated inhibition of endothelial cell proliferation, migration and tube formation in response to basic fibroblast growth factor 1 (FGF1). In ex vivo and in vivo angiogenesis assays, ferulic acid suppressed FGF1-induced microvessel sprouting of rat aortic rings and angiogenesis. To understand the underlying molecular basis, we examined the effects of ferulic acid on different molecular components and found that ferulic acid suppressed FGF1-triggered activation of FGFR1 and phosphatidyl inositol 3-kinase (PI3K)-protein kinase B (Akt) signaling. Moreover, ferulic acid directly inhibited proliferation and blocked the PI3K-Akt pathway in melanoma cell. In vivo, using a melanoma xenograft model, ferulic acid showed growth-inhibitory activity associated with inhibition of angiogenesis. Taken together, our results indicate that ferulic acid targets the FGFR1-mediated PI3K-Akt signaling pathway, leading to the suppression of melanoma growth and angiogenesis.


Assuntos
Inibidores da Angiogênese/farmacologia , Antineoplásicos/farmacologia , Ácidos Cumáricos/farmacologia , Fator 1 de Crescimento de Fibroblastos/farmacologia , Melanoma/patologia , Neovascularização Patológica/patologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Embrião de Galinha , Ativação Enzimática/efeitos dos fármacos , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Melanoma/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinase/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-Dawley , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
17.
BMC Gastroenterol ; 13: 67, 2013 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-23590285

RESUMO

BACKGROUND: FGF21 is a promising intervention therapy for metabolic diseases as fatty liver, obesity and diabetes. Recent results suggest that FGF21 is highly expressed in hepatocytes under metabolic stress caused by starvation, hepatosteatosis, obesity and diabetes. Hepatic FGF21 elicits metabolic benefits by targeting adipocytes of the peripheral adipose tissue through the transmembrane FGFR1-KLB complex. Ablation of adipose FGFR1 resulted in increased hepatosteatosis under starvation conditions and abrogation of the anti-obesogenic action of FGF21. These results indicate that FGF21 may be a stress responsive hepatokine that targets adipocytes and adipose tissue for alleviating the damaging effects of stress on the liver. However, it is unclear whether hepatic induction of FGF21 is limited to only metabolic stress, or to a more general hepatic stress resulting from liver pathogenesis and injury. METHODS: In this survey-based study, we examine the nature of hepatic FGF21 activation in liver tissues and tissue sections from several mouse liver disease models and human patients, by quantitative PCR, immunohistochemistry, protein chemistry, and reporter and CHIP assays. The liver diseases include genetic and chemical-induced HCC, liver injury and regeneration, cirrhosis, and other types of liver diseases. RESULTS: We found that mouse FGF21 is induced in response to chemical (DEN treatment) and genetic-induced hepatocarcinogenesis (disruptions in LKB1, p53, MST1/2, SAV1 and PTEN). It is also induced in response to loss of liver mass due to partial hepatectomy followed by regeneration. The induction of FGF21 expression is potentially under the control of stress responsive transcription factors p53 and STAT3. Serum FGF21 levels correlate with FGF21 expression in hepatocytes. In patients with hepatitis, fatty degeneration, cirrhosis and liver tumors, FGF21 levels in hepatocytes or phenotypically normal hepatocytes are invariably elevated compared to normal health subjects. CONCLUSION: FGF21 is an inducible hepatokine and could be a biomarker for normal hepatocyte function. Activation of its expression is a response of functional hepatocytes to a broad spectrum of pathological changes that impose both cellular and metabolic stress on the liver. Taken together with our recent data, we suggest that hepatic FGF21 is a general stress responsive factor that targets adipose tissue for normalizing local and systemic metabolic parameters while alleviating the overload and damaging effects imposed by the pathogenic stress on the liver. This study therefore provides a rationale for clinical biomarker studies in humans.


Assuntos
Carcinoma Hepatocelular/metabolismo , Transformação Celular Neoplásica/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Cirrose Hepática/metabolismo , Neoplasias Hepáticas/metabolismo , Animais , Carcinoma Hepatocelular/induzido quimicamente , Transformação Celular Neoplásica/genética , Dietilnitrosamina , Modelos Animais de Doenças , Fatores de Crescimento de Fibroblastos/genética , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Fígado/patologia , Fígado/cirurgia , Neoplasias Hepáticas/induzido quimicamente , Masculino , Proteínas de Membrana/genética , Camundongos , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/metabolismo , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Fator de Transcrição STAT3/metabolismo , Estresse Fisiológico , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
18.
Free Radic Biol Med ; 58: 1-13, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23376231

RESUMO

Tumor microenvironment plays a major role in cancer development. Understanding how the stroma affects epithelial transformation will provide a basis for new preventive strategies. Recent evidence suggests that oxidative stress in stroma may play a role in cancer progression, and loss of p53 function in the stromal cells was associated with poor prognosis and high tumor recurrence. However, the underlying mechanisms remain poorly understood. Here, we investigated the role of p53 loss in fibroblasts in epithelial transformation and the mechanistic involvement of reactive species. Using 3D organotypic culture and other assays, we report that the stroma containing p53-deficient fibroblasts could induce the nontumorigenic epithelial cells of oral and ovarian tissue origins to become invasive through reactive nitrogen species (RNS)-mediated release of the cytokine ICAM1. The p53-deficient fibroblasts have increased RNS production and accumulation of oxidative DNA-damage products associated with specific upregulation of endothelial nitric oxide synthase (eNOS). Suppression of RNS production by siRNA of eNOS or the antioxidant NAC reduced ICAM1 expression and prevented the stroma-mediated epithelial invasion. Our study uncovers the novel mechanism by which redox alteration associated with loss of p53 in stromal fibroblasts functions as a key inducer of epithelial transformation and invasion via RNS-mediated ICAM1 signaling. Thus, the modulation of redox signaling in the microenvironment may serve as a new approach to preventing epithelial transformation and suppressing cancer invasion.


Assuntos
Transformação Celular Neoplásica/genética , Células Epiteliais/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Proteína Supressora de Tumor p53 , Dano ao DNA/genética , Células Epiteliais/citologia , Feminino , Humanos , Invasividade Neoplásica/genética , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Ovário/metabolismo , Ovário/patologia , Oxirredução , RNA Interferente Pequeno , Espécies Reativas de Nitrogênio/metabolismo , Transdução de Sinais , Células Estromais/citologia , Células Estromais/metabolismo , Microambiente Tumoral , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
19.
Clin Endocrinol (Oxf) ; 78(6): 920-5, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23252994

RESUMO

BACKGROUND: Familial neurohypophyseal diabetes insipidus, an autosomal dominant disorder, is mostly caused by mutations in the genes that encode AVP or its intracellular binding protein, neurophysin-II. The mutations lead to aberrant preprohormone processing and progressive destruction of AVP-secreting cells, which gradually manifests a progressive polyuria and polydipsia during early childhood, and a disorder of water homeostasis. OBJECTIVE: We characterized the clinical and biochemical features, and sequenced the AVP neurophysin-II(AVP-NPII) gene of the affected individuals with autosomal dominant neurohypophyseal diabetes insipidus(ADNDI)to determine whether this disease was genetically determined. PATIENTS AND METHODS: We obtained the histories of eight affected and four unaffected family individuals. The diagnosis of ADNDI was established using a water deprivation test and exogenous AVP administration. For molecular analysis, genomic DNA was extracted and the AVP-NPII gene was amplified using polymerase chain reaction and sequenced. RESULTS: The eight affected individuals showed different spectra of age of onsets (7-15 years) and urine volumes (132-253 ml/kg/24 h). All affected individuals responded to vasopressin administration, with a resolution of symptoms and an increase in urine osmolality by more than 50%. The characteristic hyperintense signal in the posterior pituitary on T1-weighted magnetic resonance imaging was absent in six family members and present in one. Sequencing analysis revealed a missense heterozygous mutation 1516G > T (Gly17Val) in exon 2 of the AVP-NPII gene among the ADNDI individuals. CONCLUSIONS: We identified a missense mutation in the AVP-NPII gene and the same mutation showed different spectra of age of onsets and urine volumes in a new Chinese family with ADNDI. The mutation may provide a molecular basis for understanding the characteristics of NPII and add to our knowledge of the pathogenesis of ADNDI, which would allow the presymptomatic diagnosis of asymptomatic subjects.


Assuntos
Arginina Vasopressina/genética , Arginina Vasopressina/metabolismo , Diabetes Insípido Neurogênico/genética , Neurofisinas/genética , Adolescente , Idade de Início , Grupo com Ancestrais do Continente Asiático/genética , Criança , Feminino , Humanos , Capacidade de Concentração Renal , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Concentração Osmolar , Linhagem , Solução Salina Hipertônica , Privação de Água
20.
Protein Cell ; 3(12): 950-61, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23011846

RESUMO

Heparinase III (HepIII) is a 73-kDa polysaccharide lyase (PL) that degrades the heparan sulfate (HS) polysaccharides at sulfate-rare regions, which are important co-factors for a vast array of functional distinct proteins including the well-characterized antithrombin and the FGF/FGFR signal transduction system. It functions in cleaving metazoan heparan sulfate (HS) and providing carbon, nitrogen and sulfate sources for host microorganisms. It has long been used to deduce the structure of HS and heparin motifs; however, the structure of its own is unknown. Here we report the crystal structure of the HepIII from Bacteroides thetaiotaomicron at a resolution of 1.6 Å. The overall architecture of HepIII belongs to the (α/α)5 toroid subclass with an N-terminal toroid-like domain and a C-terminal ß-sandwich domain. Analysis of this high-resolution structure allows us to identify a potential HS substrate binding site in a tunnel between the two domains. A tetrasaccharide substrate bound model suggests an elimination mechanism in the HS degradation. Asn260 and His464 neutralize the carboxylic group, whereas Tyr314 serves both as a general base in C-5 proton abstraction, and a general acid in a proton donation to reconstitute the terminal hydroxyl group, respectively. The structure of HepIII and the proposed reaction model provide a molecular basis for its potential practical utilization and the mechanism of its eliminative degradation for HS polysaccarides.


Assuntos
Heparitina Sulfato/metabolismo , Polissacarídeo-Liase/química , Polissacarídeo-Liase/metabolismo , Sequência de Aminoácidos , Bacteroides/enzimologia , Domínio Catalítico , Cristalografia por Raios X , Cinética , Modelos Moleculares , Dados de Sequência Molecular , Especificidade por Substrato
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