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1.
Arch Pharm Res ; 43(12): 1311-1324, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33245516

RESUMO

Chronic inflammation in adipose tissue is the hallmark of obesity and a major risk factor for the development of obesity-induced insulin resistance. NLRP3 inflammasome regulates the maturation and secretion of pro-inflammatory cytokines, such as IL-1ß and IL-18, and was recently discovered to be involved in obesity-related metabolic diseases. Fibroblast growth factors (FGFs) such as FGF1, FGF10, and FGF21 are adipokines that regulate adipocyte development and metabolism, but reports on the effect of other FGFs on adipocytes are lacking. In the present study, the novel role of FGF2 in NLRP3 inflammasome activation was elucidated. Our results showed that FGF2 levels were increased during adipocyte differentiation and in the adipose tissue of high-fat diet (HFD)-induced obese mice. Recombinant FGF2 treatment upregulated inflammasome markers such as NLRP3, which was further exaggerated by TNF-ɑ treatment. Interestingly, ß-Klotho, a co-receptor of FGF21, was significantly decreased by FGF2 treatment. Results from mice confirmed the positive correlation between FGF2 and NLRP3 expression in epididymal and subcutaneous adipose tissue, while exercise training effectively reversed HFD-induced NLRP3 expression as well as FGF2 levels in both adipose depots. Our results suggest that FGF2 is an adipokine that may exacerbate the inflammatory response in adipocytes through NLRP3 inflammasome activation.


Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Inflamassomos/metabolismo , Inflamação/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Obesidade/metabolismo , Gordura Subcutânea/efeitos dos fármacos , Células 3T3-L1 , Adipócitos/imunologia , Adipócitos/metabolismo , Animais , Modelos Animais de Doenças , Fator 2 de Crescimento de Fibroblastos/genética , Fator 2 de Crescimento de Fibroblastos/metabolismo , Inflamação/genética , Inflamação/imunologia , Proteínas Klotho , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/genética , Obesidade/imunologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais , Gordura Subcutânea/imunologia , Gordura Subcutânea/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
2.
J Biol Chem ; 293(10): 3758-3769, 2018 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-29363574

RESUMO

Investigating stimulation of endogenous wound healing in corneal endothelial cells (CECs) may help address the global shortage of donor corneas by decreasing the number of transplants performed for blindness because of endothelial dysfunction. We previously reported that IL-1ß stimulation leads to fibroblast growth factor (FGF2) expression, enhancing migration and proliferation of mammalian CECs. However, FGF2 also promotes the endothelial-mesenchymal transition, which can lead to retrocorneal membrane formation and blindness. This prompted us to investigate downstream FGF2 signaling targets that could be manipulated to prevent retrocorneal membrane formation. FGF2 stimulation altered cell morphology and induced expression of mesenchymal transition marker genes such as snail family transcriptional repressor 1 (SNAI1), SNAI2, zinc finger E-box-binding homeobox 1 (ZEB1), and ZEB2 This, in turn, induced expression of fibronectin, vimentin, and type I collagen, and suppressed E-cadherin in CECs in vitro and ex vivo siRNA-mediated SNAI1 knockdown revealed that SNAI1 induces ZEB1 expression, in turn inducing expression of type I collagen, the major component of retrocorneal membranes, and of cyclin-dependent kinase 2 (CDK2) and cyclin E1, promoting cell proliferation. siRNA-mediated knockdown of SNAI1 or ZEB1, but not of CDK2, inhibited FGF2-dependent expression of fibronectin, vimentin, and type I collagen and of suppression of E-cadherin expression. We conclude that SNAI1 is a key regulator of FGF2-dependent mesenchymal transition in human ex vivo corneal endothelium, with ZEB1 regulating type I collagen expression and CDK2 regulating cell proliferation. These results suggest that SNAI1 promotes fibrosis and cell proliferation in human corneal endothelium through ZEB1 and CDK2.


Assuntos
Quinase 2 Dependente de Ciclina/metabolismo , Endotélio Corneano/metabolismo , Proteínas do Olho/metabolismo , Regulação da Expressão Gênica , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Fatores de Transcrição da Família Snail/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Biomarcadores/metabolismo , Movimento Celular , Proliferação de Células , Forma Celular , Transdiferenciação Celular , Células Cultivadas , Colágeno Tipo I/agonistas , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Cadeia alfa 1 do Colágeno Tipo I , Quinase 2 Dependente de Ciclina/antagonistas & inibidores , Quinase 2 Dependente de Ciclina/química , Quinase 2 Dependente de Ciclina/genética , Endotélio Corneano/citologia , Endotélio Corneano/patologia , Ativação Enzimática , Proteínas do Olho/agonistas , Proteínas do Olho/antagonistas & inibidores , Proteínas do Olho/genética , Humanos , Interferência de RNA , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Fatores de Transcrição da Família Snail/antagonistas & inibidores , Fatores de Transcrição da Família Snail/genética , Cicatrização , Homeobox 2 de Ligação a E-box com Dedos de Zinco/agonistas , Homeobox 2 de Ligação a E-box com Dedos de Zinco/genética , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/agonistas , Homeobox 1 de Ligação a E-box em Dedo de Zinco/antagonistas & inibidores , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
3.
Am J Physiol Lung Cell Mol Physiol ; 308(10): L1014-24, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25820524

RESUMO

Fibroblast growth factors (Fgfs) mediate organ repair. Lung epithelial cell overexpression of Fgf10 postbleomycin injury is both protective and therapeutic, characterized by increased survival and attenuated fibrosis. Exogenous administration of FGF7 (palifermin) also showed prophylactic survival benefits in mice. The role of endogenous Fgfr2b ligands on bleomycin-induced lung fibrosis is still elusive. This study reports the expression of endogenous Fgfr2b ligands, receptors, and signaling targets in wild-type mice following bleomycin lung injury. In addition, the impact of attenuating endogenous Fgfr2b-ligands following bleomycin-induced fibrosis was tested by using a doxycycline (dox)-based inducible, soluble, dominant-negative form of the Fgfr2b receptor. Double-transgenic (DTG) Rosa26(rtTA/+);tet(O)solFgfr2b mice were validated for the expression and activity of soluble Fgfr2b (failure to regenerate maxillary incisors, attenuated recombinant FGF7 signal in the lung). As previously reported, no defects in lung morphometry were detected in DTG (+dox) mice exposed from postnatal days (PN) 1 through PN105. Female single-transgenic (STG) and DTG mice were subjected to various levels of bleomycin injury (1.0, 2.0, and 3.0 U/kg). Fgfr2b ligands were attenuated either throughout injury (days 0-11; days 0-28) or during later stages (days 6-28 and 14-28). No significant changes in survival, weight, lung function, confluent areas of fibrosis, or hydroxyproline deposition were detected in DTG mice. These results indicate that endogenous Fgfr2b ligands do not significantly protect against bleomycin injury, nor do they expedite the resolution of bleomycin-induced lung injury in mice.


Assuntos
Antibióticos Antineoplásicos/efeitos adversos , Bleomicina/efeitos adversos , Fator 7 de Crescimento de Fibroblastos/farmacologia , Fibrose Pulmonar , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Antibióticos Antineoplásicos/farmacologia , Bleomicina/farmacologia , Feminino , Camundongos , Camundongos Transgênicos , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Transdução de Sinais/genética
4.
Biol Reprod ; 89(4): 81, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23946540

RESUMO

The prostate gland is unique in that it undergoes rapid regression following castration but regenerates completely once androgens are replaced. Residual ductal components play an important role in the regeneration of a fully functional prostate. In this study, to examine how androgen status affects prostate structure and components, we conducted histopathological studies of the involuted and regenerated mouse dorsolateral prostate (DLP). In the castrated mouse DLP, the number of luminal epithelial cells decreased in a time-dependent manner. On Day 14 postandrogen replacement, the number of luminal epithelial cells was completely restored to the baseline level. In contrast, the number of basal epithelial cells gradually increased in the castrated mouse prostate. The Ki67-labeling index of prostate basal epithelial cells was significantly increased after castration. The number of basal epithelial cells decreased to baseline after androgen replacement. After castration, mRNA expression levels of specific growth factors, such as Fgf2, Fgf7, Hgf, Tgfa, and Tgfb, were relatively abundant in whole mouse DLPs. In organ culture experiments, basal epithelial proliferation was recapitulated in the absence of dihydrotestosterone (DHT). The proliferation of basal epithelial cells in the absence of DHT was suppressed by treatment with an FGF receptor inhibitor (PD173074). Moreover, FGF2 treatment directly stimulated the proliferation of basal epithelial cells. Taken together, these data indicated that the FGF2-FGF receptor signal cascade in the prostate gland may be one of the pathways stimulating the proliferation of basal epithelial cells in the absence of androgens.


Assuntos
Castração/efeitos adversos , Células Epiteliais/fisiologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Próstata/fisiologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Regeneração , Transdução de Sinais , Androgênios/farmacologia , Androgênios/uso terapêutico , Animais , Membrana Basal/citologia , Membrana Basal/efeitos dos fármacos , Membrana Basal/fisiologia , Proliferação de Células/efeitos dos fármacos , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/antagonistas & inibidores , Fator 2 de Crescimento de Fibroblastos/genética , Fator 7 de Crescimento de Fibroblastos/antagonistas & inibidores , Fator 7 de Crescimento de Fibroblastos/genética , Fator 7 de Crescimento de Fibroblastos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Fator de Crescimento de Hepatócito/antagonistas & inibidores , Fator de Crescimento de Hepatócito/genética , Fator de Crescimento de Hepatócito/metabolismo , Terapia de Reposição Hormonal , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Técnicas de Cultura de Órgãos , Próstata/citologia , Próstata/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Proteínas Recombinantes/metabolismo , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fatores de Crescimento Transformadores/antagonistas & inibidores , Fatores de Crescimento Transformadores/genética , Fatores de Crescimento Transformadores/metabolismo
5.
J Dermatol Sci ; 54(2): 106-13, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19250802

RESUMO

BACKGROUND: Exposure to ultraviolet (UV) radiation causes a complex cellular response, mostly mediated by the production of reactive oxygen species (ROS), which can be counteracted by exogenous treatments and endogenous mechanisms with anti-oxidant and scavenger properties. Keratinocyte growth factor (KGF/FGF7), a member of the fibroblast growth factor family, promotes epithelial growth and differentiation and is involved in cell survival after oxidant injuries. OBJECTIVE: We analyzed the role of KGF in the control of intracellular ROS production and oxidative stress after UVB exposure on KGF receptor (KGFR) transfected cells and human immortalized and primary keratinocytes. METHODS: We assessed the intracellular ROS production measuring the intensity of the oxidation-sensitive fluorescent probe 2',7'-dichlorofluorescein diacetate (DCFH-DA) by confocal microscopy, as well as the catalase activity by spectrophotometric assay. Moreover, morphological and biochemical analysis of actin cytoskeleton reorganization was evaluated as a further marker of oxidative damage. RESULTS: Our data show that KGF significantly reduces intracellular ROS generation in response to UVB, preserves the decrease of catalase activity and prevents actin cytoskeleton rearrangement. CONCLUSION: Our results provide a further evidence that KGF may be crucial for an efficient skin photoprotection, demonstrating a direct role for KGF in the reduction of intracellular ROS content following UVB exposure.


Assuntos
Actinas/metabolismo , Catalase/metabolismo , Fator 7 de Crescimento de Fibroblastos/fisiologia , Queratinócitos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Actinas/efeitos dos fármacos , Actinas/efeitos da radiação , Animais , Catalase/efeitos dos fármacos , Catalase/efeitos da radiação , Linhagem Celular , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Citoesqueleto/efeitos da radiação , Regulação para Baixo/efeitos dos fármacos , Fator 7 de Crescimento de Fibroblastos/farmacologia , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/efeitos da radiação , Camundongos , Células NIH 3T3 , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Transfecção , Raios Ultravioleta
6.
J Clin Immunol ; 28(5): 600-15, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18592360

RESUMO

INTRODUCTION: Treating recipient mice with palifermin (recombinant human keratinocyte growth factor) prevents the development of acute, lethal, graft-versus-host disease (GVHD). This is due, at least in part, to the ability of palifermin to protect epithelial cells from injury. Using the C57BL/6-->(C57BL/6 x DBA/2)F(1)-hybrid model, we previously showed that the protective effect of palifermin was also associated with redirection of the cytokine profile from Th1 to Th2. DISCUSSION: To study this immunoregulatory effect more directly, we induced acute GVH reactions in which we treated the donors rather than the recipients with palifermin. The recipient mice were protected from GVHD-associated morbidity, and their cytokine profile was predominantly Th2. The palifermin-treated donor mice alone showed a similar Th2 cytokine profile, and we observed elevated levels of thymic stromal lymphopoietin mRNA in the thymus. We further demonstrated that treating the donor mice with palifermin protects against GVHD-associated morbidity, even if the donors are deficient in Valpha14i natural killer T cells. Our findings clearly show that palifermin mediates immunoregulatory effects in addition to its cytoprotective effects and that both are likely to be involved in the mechanism through which palifermin provides protection from acute murine GVHD.


Assuntos
Fator 7 de Crescimento de Fibroblastos/administração & dosagem , Doença Enxerto-Hospedeiro/prevenção & controle , Proteínas Recombinantes/administração & dosagem , Células Th1/metabolismo , Células Th2/metabolismo , Doença Aguda , Animais , Linhagem Celular Transformada , Citocinas/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Doença Enxerto-Hospedeiro/imunologia , Transplante de Células-Tronco Hematopoéticas , Injeções Subcutâneas , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Knockout , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Células Th1/imunologia , Células Th1/patologia , Células Th2/imunologia , Células Th2/patologia
7.
J Biol Chem ; 283(9): 5287-95, 2008 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-18171671

RESUMO

It has been reported recently that bone marrow stromal cells (BMSCs) are able to differentiate into various neural cells both in vivo and in vitro (Egusa, H., Schweizer, F. E., Wang, C. C., Matsuka, Y., and Nishimura, I. (2005) J. Biol. Chem. 280, 23691-23697). However, the underlying mechanisms remain largely unknown. In this report, we have demonstrated that basic fibroblast growth factor (bFGF) alone effectively induces mouse BMSC neuronal differentiation. These differentiated neuronal cells exhibit characteristic electrophysiological properties and elevated levels of the neuronal differentiation marker, growth-associated protein-43 (GAP-43). To explore possible signaling pathways, we first analyzed the expression of various FGF receptors in mouse BMSCs. FGF receptor-1, -2, and -3 were detected, but only FGFR-1 was shown to be activated by bFGF. Small interfering RNA knock down of FGFR-1 in BMSCs significantly inhibited neuronal differentiation. Moreover, we have shown that the mitogen-activated protein kinase (ERK1/2) is persistently activated and blockage of ERK activity with the ERK-specific inhibitor U0126 prevents neuronal differentiation. It appears that activation of ERK cascade and neuronal differentiation of BMSCs induced by bFGF are independent of Ras activity but require functions of phospholipase C-gamma pathway. Lastly, we examined the role of the immediate-early transcription factors AP-1 and NF-kappaB and have found that phospholipase C-gamma-dependent c-Jun and ERK-dependent c-fos, but not the NF-kappaB, are strongly activated by bFGF, which in turn regulates the neuronal differentiation of BMSCs.


Assuntos
Células da Medula Óssea/metabolismo , Diferenciação Celular/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Neurônios/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Fator de Transcrição AP-1/metabolismo , Animais , Células da Medula Óssea/citologia , Butadienos/farmacologia , Diferenciação Celular/fisiologia , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Proteína GAP-43/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , NF-kappa B/metabolismo , Neurônios/citologia , Nitrilas/farmacologia , Proteína Oncogênica p21(ras)/antagonistas & inibidores , Proteína Oncogênica p21(ras)/metabolismo , Fosfolipase C gama/antagonistas & inibidores , Fosfolipase C gama/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/metabolismo , Células Estromais/citologia , Células Estromais/metabolismo
8.
Cytokine ; 38(1): 22-31, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17537644

RESUMO

AIM: A growing number of mutations mapped in the receptor gene for fibroblast growth factor have been implicated in several cranial development disorders including the Apert and Crouzon syndromes. The present paper investigated cellular mechanisms underlying Apert phenotype, by analyzing the effects of FGF2 in primary cultures of Apert periosteal fibroblasts carrying the FGFR2 Pro253Arg mutation. RESULTS: FGF2 administration significantly decreased extracellular matrix production in mutant cells by stimulating degradative enzymatic activities. Gene expression analysis revealed that decorin and biglycan, two proteoglycans involved in collagen fibrillogenesis, were more expressed in mutant cells and down-regulated by FGF2. FGF2 receptor binding showed little differences in high affinity receptor counts between mutant and wild-type cells, while we showed for the first time that low affinity receptors are significantly fewer in mutant cells. Differences were found in Crouzon syndrome, where both high and low affinity receptor counts were up-regulated. CONCLUSIONS: The different mutation and low affinity receptor regulation in mutant receptors support the hypothesis that the impact on the activity of the ligand-receptor complex could allow distinct modes of FGF2 activation in Apert and Crouzon syndromes, which interfere with the FGFR2 signalling cascade.


Assuntos
Acrocefalossindactilia/genética , Disostose Craniofacial/genética , Matriz Extracelular/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Periósteo/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Acrocefalossindactilia/metabolismo , Adolescente , Arginina/química , Arginina/genética , Contagem de Células , Colágeno Tipo I/metabolismo , Disostose Craniofacial/metabolismo , Análise Mutacional de DNA , Fator 2 de Crescimento de Fibroblastos/farmacologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibronectinas/metabolismo , Glicosídeo Hidrolases/metabolismo , Humanos , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/metabolismo , Mutação , Peptídeo Hidrolases/metabolismo , Periósteo/citologia , Periósteo/efeitos dos fármacos , Fenótipo , Prolina/química , Prolina/genética , Proteoglicanas/genética , Proteoglicanas/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética
9.
Mol Endocrinol ; 21(4): 987-1000, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17284663

RESUMO

Two related receptor tyrosine kinases (RTKs), fibroblast growth factor receptor 1 and 2 (FGFR1 and FGFR2), exert distinct effects during carcinogenesis. To examine FGFR1 and FGFR2 signaling in polarized epithelia, we have developed an in vitro three-dimensional HC11 mouse mammary epithelial cell culture model combined with a chemically inducible FGFR (iFGFR) dimerization system. Although activation of both RTKs led to reinitiation of cell proliferation and loss of cell polarity, only iFGFR1 activation induced cell survival and epithelial to mesenchymal transition. In contrast, iFGFR2 activation induced cell apoptosis even in the cells in direct contact with the extracellular matrix. Activation of iFGFR2, but not iFGFR1, led to rapid receptor down-regulation and transient activation of downstream signaling, which were partially rescued by Cbl small interfering RNA knockdown or the proteasome inhibitor lactacystin. Importantly, inhibition of proteasome activity in iFGFR2-activated structures led to epithelial to mesenchymal transition and invasive phenotypes resembling those observed after iFGFR1 activation. These studies demonstrate, for the first time, that the duration of downstream signaling determines the distinct phenotypes mediated by very homologous RTKs in three-dimensional cultures.


Assuntos
Transformação Celular Neoplásica/metabolismo , Glândulas Mamárias Animais/enzimologia , Neoplasias Mamárias Animais/enzimologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/fisiologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/fisiologia , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacologia , Animais , Apoptose , Técnicas de Cultura de Células , Sobrevivência Celular , Transformação Celular Neoplásica/genética , Células Cultivadas , Inibidores de Cisteína Proteinase/farmacologia , Epitélio/patologia , Glândulas Mamárias Animais/patologia , Mesoderma/patologia , Camundongos , Camundongos Transgênicos , Fenótipo , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Proto-Oncogênicas c-cbl/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-cbl/genética , RNA Interferente Pequeno/farmacologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética
10.
J Periodontal Res ; 41(4): 340-9, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16827730

RESUMO

BACKGROUND AND OBJECTIVE: Regulation of epithelial cell behavior associated with periodontitis is not well elucidated but many responses will ultimately be regulated by growth factor receptors. Using a rat experimental periodontitis model, protein and gene expression of select growth factor receptors in junctional and pocket epithelium were examined. MATERIAL AND METHODS: Periodontal disease was induced by daily topical application of lipopolysaccharide using an established protocol. Animals were killed at time 0 (control), and at 2 and 8 wk. Frozen tissue samples were collected from the right palatal gingival soft tissue, and the left periodontal tissues were decalcified and embedded in paraffin. Laser microdissection and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to quantify keratinocyte growth factor receptor (KGFR), hepatocyte growth factor receptor (HGFR), epidermal growth factor receptor (EGFR) and fibroblast growth factor receptor 1 (FGFR1) gene expression, and in situ RT-PCR localized these increases to specific epithelial cells. Receptor protein expression was examined immunohistochemically. In cell culture, induction of HGFR and KGFR protein expression by serum, lipopolysaccharide and pro-inflammatory cytokines were examined using flow cytometry. RESULTS: Eight-week tissue samples exhibited histological changes consistent with periodontitis. KGFR and HGFR gene and protein expression were significantly induced at the 8 wk time point. KGFR expression was significantly up-regulated in basal and parabasal pocket epithelial cells, but HGFR was up-regulated throughout the pocket epithelium. In cell culture serum, lipopolysaccharide and pro-inflammatory cytokines, interleukin-1beta and tumour necrosis factor-alpha significantly induced KGFR protein receptor expression, but HGFR expression was only induced by serum. CONCLUSION: KGFR and HGFR are highly up-regulated in this model of periodontal disease and may play a significant role in regulating the proliferation and migration of pocket epithelium.


Assuntos
Inserção Epitelial/metabolismo , Bolsa Periodontal/metabolismo , Receptores Proteína Tirosina Quinases/biossíntese , Animais , Inserção Epitelial/citologia , Células Epiteliais/metabolismo , Citometria de Fluxo , Regulação Enzimológica da Expressão Gênica , Imuno-Histoquímica , Interleucina-1/metabolismo , Lipopolissacarídeos/metabolismo , Masculino , Proteínas Proto-Oncogênicas c-met/biossíntese , Ratos , Ratos Wistar , Receptores Proteína Tirosina Quinases/análise , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/biossíntese , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Suínos , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima
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