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1.
Cell Commun Signal ; 16(1): 36, 2018 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-29966518

RESUMO

BACKGROUND: Chemotherapeutic efficacy can be improved by targeting the structure and function of the extracellular matrix (ECM) in the carcinomal stroma. This can be accomplished by e.g. inhibiting TGF-ß1 and -ß3 or treating with Imatinib, which results in scarcer collagen fibril structure in xenografted human KAT-4/HT29 (KAT-4) colon adenocarcinoma. METHODS: The potential role of αVß6 integrin-mediated activation of latent TGF-ß was studied in cultured KAT-4 and Capan-2 human ductal pancreatic carcinoma cells as well as in xenograft carcinoma generated by these cells. The monoclonal αVß6 integrin-specific monoclonal antibody 3G9 was used to inhibit the αVß6 integrin activity. RESULTS: Both KAT-4 and Capan-2 cells expressed the αVß6 integrin but only KAT-4 cells could utilize this integrin to activate latent TGF-ß in vitro. Only when Capan-2 cells were co-cultured with human F99 fibroblasts was the integrin activation mechanism triggered, suggesting a more complex, fibroblast-dependent, activation pathway. In nude mice, a 10-day treatment with 3G9 reduced collagen fibril thickness and interstitial fluid pressure in KAT-4 but not in the more desmoplastic Capan-2 tumors that, to achieve a similar effect, required a prolonged 3G9 treatment. In contrast, a 10-day direct inhibition of TGF-ß1 and -ß3 reduced collagen fibril thickness in both tumor models. CONCLUSION: Our data demonstrate that the αVß6-directed activation of latent TGF-ß plays a pivotal role in modulating the stromal collagen network in carcinoma, but that the sensitivity to αVß6 inhibition depends on the simultaneous presence of alternative paths for latent TGF-ß activation and the extent of desmoplasia.


Assuntos
Antígenos de Neoplasias/imunologia , Colágeno/química , Integrinas/imunologia , Animais , Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Colágeno/metabolismo , Líquido Extracelular/metabolismo , Feminino , Perfilação da Expressão Gênica , Humanos , Integrinas/metabolismo , Camundongos , Pressão , Fator de Crescimento Transformador beta/metabolismo
2.
Mol Cancer Ther ; 15(10): 2455-2464, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27474147

RESUMO

A typical obstacle to cancer therapy is the limited distribution of low molecular weight anticancer drugs within the carcinoma tissue. In experimental carcinoma, imatinib (STI571) increases efficacy of synchronized chemotherapy, reduces tumor interstitial fluid pressure, and increases interstitial fluid volume. STI571 also increases the water-perfusable fraction in metastases from human colorectal adenocarcinomas. Because the mechanism(s) behind these effects have not been fully elucidated, we investigated the hypothesis that STI571 alters specific properties of the stromal extracellular matrix. We analyzed STI571-treated human colorectal KAT-4/HT-29 experimental carcinomas, known to have a well-developed stromal compartment, for solute exchange and glycosaminoglycan content, as well as collagen content, structure, and synthesis. MRI of STI571-treated KAT-4/HT-29 experimental carcinomas showed a significantly increased efficacy in dynamic exchanges of solutes between tumor interstitium and blood. This effect was paralleled by a distinct change of the stromal collagen network architecture, manifested by a decreased average collagen fibril diameter, and increased collagen turnover. The glycosaminoglycan content was unchanged. Furthermore, the apparent effects on the stromal cellular composition were limited to a reduction in an NG2-positive stromal cell population. The current data support the hypothesis that the collagen network architecture influences the dynamic exchanges of solutes between blood and carcinoma tissue. It is conceivable that STI571 reprograms distinct nonvascular stromal cells to produce a looser extracellular matrix, ultimately improving transport characteristics for traditional chemotherapeutic agents. Mol Cancer Ther; 15(10); 2455-64. ©2016 AACR.


Assuntos
Antineoplásicos/farmacologia , Carcinoma/metabolismo , Colágeno/metabolismo , Líquido Extracelular/metabolismo , Mesilato de Imatinib/farmacologia , Agregados Proteicos , Inibidores de Proteínas Quinases/farmacologia , Animais , Carcinoma/tratamento farmacológico , Carcinoma/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Camundongos , Pericitos/efeitos dos fármacos , Pericitos/metabolismo , Células Estromais/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Exp Dermatol ; 25(7): 558-61, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-26997256

RESUMO

Excessive production of collagen is the hallmark of fatal diseases of fibrosis such as systemic sclerosis. Overexpression of the proteoglycan fibromodulin (FMOD) has been associated with improved wound healing and scarless repair. In this study, we have investigated the consequences of FMOD deficiency on the development of experimental skin fibrosis. Using immunohistochemistry, we identified FMOD in both human and murine fibrotic skin. In the bleomycin model of skin fibrosis, FMOD(-/-) mice developed skin fibrosis to a similar degree compared to FMOD(+/+) mice. Analysis of skin ultrastructure using transmission electron microscopy revealed a significant reduction in collagen fibril diameter in FMOD(-/-) but not FMOD(+/+) mice following fibrosis. We conclude that the impact of FMOD deficiency on the development of experimental skin fibrosis is limited.


Assuntos
Fibromodulina/deficiência , Escleroderma Sistêmico/metabolismo , Adolescente , Animais , Bleomicina , Estudos de Casos e Controles , Criança , Modelos Animais de Doenças , Fibrose , Humanos , Camundongos Endogâmicos C57BL , Escleroderma Sistêmico/etiologia , Pele/metabolismo , Pele/patologia , Pele/ultraestrutura
4.
Birth Defects Res A Clin Mol Teratol ; 100(9): 712-20, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25186462

RESUMO

BACKGROUND: Dermatan sulfate (DS) is a highly sulfated polysaccharide with a variety of biological functions in extracellular matrix organization and processes such as tumorigenesis and wound healing. A distinct feature of DS is the presence of iduronic acid, produced by the two enzymes, DS-epimerase 1 and 2, which are encoded by Dse and Dsel, respectively. METHODS: We have previously shown that Dse knockout (KO) mice in a mixed C57BL/6-129/SvJ background have an altered collagen matrix structure in skin. In the current work we studied Dse KO mice in a pure NFR genetic background. RESULTS: Dse KO embryos and newborns had kinked tails and histological staining revealed significantly thicker epidermal layers in Dse KO mice when compared with heterozygote (Het) or wild-type (WT) littermates. Immunochemical analysis of the epidermal layers in newborn pups showed increased expression of keratin 5 in the basal layer and keratin 1 in the spinous layer. In addition, we observed an abdominal wall defect with herniated intestines in 16% of the Dse KO embryos. Other, less frequent, developmental defects were exencephaly and spina bifida. CONCLUSION: We conclude that the combination of defective collagen structure in the dermis and imbalanced keratinocyte maturation could be responsible for the observed developmental defects in Dse KO mice. In addition, we propose that Dse KO mice could be used as a model in pathogenetic studies of human fetal abdominal wall defects.


Assuntos
Parede Abdominal/anormalidades , Carboidratos Epimerases/genética , Derme/metabolismo , Hérnia Abdominal/genética , Queratinócitos/metabolismo , Animais , Carboidratos Epimerases/deficiência , Dermatan Sulfato/metabolismo , Derme/patologia , Modelos Animais de Doenças , Embrião de Mamíferos , Expressão Gênica , Hérnia Abdominal/complicações , Hérnia Abdominal/patologia , Humanos , Queratina-1/genética , Queratina-1/metabolismo , Queratina-15/genética , Queratina-15/metabolismo , Queratinócitos/patologia , Camundongos , Camundongos Knockout , Defeitos do Tubo Neural/complicações , Defeitos do Tubo Neural/genética , Defeitos do Tubo Neural/patologia , Disrafismo Espinal/complicações , Disrafismo Espinal/genética , Disrafismo Espinal/patologia
5.
Biochem J ; 462(1): 113-23, 2014 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-24840544

RESUMO

The functional significance of fibrin deposits typically seen in inflammatory lesions, carcinomas and in healing wounds is not fully understood. In the present study, we demonstrate that fibrinogen/fibrin specifically bound to native Col I (collagen type I) and used the Col I fibre network as a base to provide a functional interface matrix that connects cells to the Col I fibres through αVß3 integrins. This allowed murine myoblast C2C12 cells to contract the collagenous composite gel via αVß3 integrin. We show that fibrinogen specifically bound to immobilized native Col I at the site known to bind matrix metalloproteinase-1, discoidin domain receptor-2 and fibronectin, and that binding had no effect on Col I fibrillation. A specific competitive inhibitor blocking the Col-I-binding site for fibrinogen abolished the organization of fibrin into discernable fibrils, as well as the C2C12-mediated contraction of Col I gels. Our data show that fibrin can function as a linkage protein between Col I fibres and cells, and suggest that fibrin at inflammatory sites indirectly connects αVß3 integrins to Col I fibres and thereby promotes cell-mediated contraction of collagenous tissue structures.


Assuntos
Colágeno Tipo I/metabolismo , Fibrina/metabolismo , Integrina alfaVbeta3/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Matriz Extracelular/metabolismo , Géis , Camundongos , Trombina/farmacologia
6.
PLoS One ; 8(12): e84148, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24358335

RESUMO

The NADPH oxidase 2 (NOX2) complex is a professional producer of reactive oxygen species (ROS) and is mainly expressed in phagocytes. While the activity of the NOX2 complex is essential for immunity against pathogens and protection against autoimmunity, its role in the development of malignant tumors remains unclear. We compared wild type and Ncf1 (m1J) mutated mice, which lack functional NOX2 complex, in four different tumor models. Ncf1 (m1J) mutated mice developed significantly smaller tumors in two melanoma models in which B16 melanoma cells expressing a hematopoietic growth factor FLT3L or luciferase reporter were used. Ncf1 (m1J) mutated mice developed significantly fewer Lewis Lung Carcinoma (LLC) tumors, but the tumors that did develop, grew at a pace that was similar to the wild type mice. In the spontaneously arising prostate carcinoma model (TRAMP), tumor growth was not affected. The lack of ROS-mediated protection against tumor growth was associated with increased production of immunity-associated cytokines. A significant increase in Th2 associated cytokines was observed in the LLC model. Our present data show that ROS regulate rejection of the antigenic B16-luc and LLC tumors, whereas the data do not support a role for ROS in growth of intrinsically generated tumors.


Assuntos
Carcinoma/genética , Carcinoma/patologia , Melanoma/genética , Melanoma/patologia , NADPH Oxidases/genética , Neoplasias/genética , Neoplasias/patologia , Animais , Carcinoma/metabolismo , Carcinoma/mortalidade , Carcinoma Pulmonar de Lewis , Modelos Animais de Doenças , Inflamação/genética , Inflamação/metabolismo , Melanoma/metabolismo , Melanoma/mortalidade , Melanoma Experimental , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Mutação , NADPH Oxidases/deficiência , NADPH Oxidases/metabolismo , Neoplasias/metabolismo , Neoplasias/mortalidade , Espécies Reativas de Oxigênio/imunologia , Carga Tumoral/genética
7.
Am J Pathol ; 182(6): 2132-46, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23570836

RESUMO

Fibrosis is a deleterious consequence of chronic inflammation in a number of human pathologies ultimately leading to organ dysfunction and failure. Two growth factors that are important in blood vessel physiology and tissue fibrosis, platelet-derived growth factor (PDGF)-B and transforming growth factor (TGF)-ß1, were investigated. Adenoviral vectors were used to induce transient overexpression of these growth factors in mouse skin. Changes in tissue structure and protein and mRNA expressions were investigated. Both PDGF-B and TGF-ß1 could initiate but neither could sustain angiogenesis. Instead, vascular regression was observed. Overexpression of both TGF-ß1 and PDGF-B led to a marked macrophage influx and an expansion of the connective tissue cell population. Over time, this effect was sustained in mice treated with TGF-ß1, whereas it was partially reversible in mice treated with PDGF-B. On the basis of structure and expression of phenotypical markers, the emerging connective tissue cell population may originate from microvascular pericytes. TGF-ß1 induced expansion of connective tissue cells with a myofibroblast phenotype, whereas PDGF-B induced a fibroblast phenotype negative for α-smooth muscle actin. TGF-ß1 and PDGF-B overexpressions mediated distinct effects on mRNA transcript levels of fibrillar procollagens, their modifying enzymes, small leucin-rich repeat proteoglycans, and matricellular proteins affecting both the composition and the quantity of the extracellular matrix. This study offers new insight into the effects of PDGF-B and TGF-ß1 on the vasculature and connective tissue in vivo.


Assuntos
Células do Tecido Conjuntivo/citologia , Pericitos/citologia , Proteínas Proto-Oncogênicas c-sis/fisiologia , Pele/citologia , Fator de Crescimento Transformador beta1/fisiologia , Adenoviridae/genética , Animais , Endotélio Vascular/citologia , Matriz Extracelular/fisiologia , Proteínas da Matriz Extracelular/metabolismo , Fibromodulina , Regulação da Expressão Gênica/fisiologia , Vetores Genéticos , Macrófagos/fisiologia , Camundongos , Camundongos Nus , Microvasos/citologia , Neovascularização Fisiológica/fisiologia , Pericitos/metabolismo , Fenótipo , Proteoglicanas/metabolismo , RNA Mensageiro/genética , Pele/irrigação sanguínea , Pele/metabolismo
8.
FEBS J ; 280(10): 2431-46, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23441919

RESUMO

The presence of iduronic acid in chondroitin/dermatan sulfate changes the properties of the polysaccharides because it generates a more flexible chain with increased binding potentials. Iduronic acid in chondroitin/dermatan sulfate influences multiple cellular properties, such as migration, proliferation, differentiation, angiogenesis and the regulation of cytokine/growth factor activities. Under pathological conditions such as wound healing, inflammation and cancer, iduronic acid has diverse regulatory functions. Iduronic acid is formed by two epimerases (i.e. dermatan sulfate epimerase 1 and 2) that have different tissue distribution and properties. The role of iduronic acid in chondroitin/dermatan sulfate is highlighted by the vast changes in connective tissue features in patients with a new type of Ehler-Danlos syndrome: adducted thumb-clubfoot syndrome. Future research aims to understand the roles of the two epimerases and their interplay with the sulfotransferases involved in chondroitin sulfate/dermatan sulfate biosynthesis. Furthermore, a better definition of chondroitin/dermatan sulfate functions using different knockout models is needed. In this review, we focus on the two enzymes responsible for iduronic acid formation, as well as the role of iduronic acid in health and disease.


Assuntos
Antígenos de Neoplasias/metabolismo , Carboidratos Epimerases/metabolismo , Sulfatos de Condroitina/metabolismo , Proteínas de Ligação a DNA/metabolismo , Dermatan Sulfato/metabolismo , Ácido Idurônico/metabolismo , Proteínas de Neoplasias/metabolismo , Motivos de Aminoácidos , Animais , Antígenos de Neoplasias/genética , Carboidratos Epimerases/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Movimento Celular , Proteínas de Ligação a DNA/genética , Dermatan Sulfato/biossíntese , Síndrome de Ehlers-Danlos/patologia , Matriz Extracelular/metabolismo , Anormalidades do Olho , Deformidades Congênitas do Pé/patologia , Deformidades Congênitas da Mão/patologia , Humanos , Instabilidade Articular/congênito , Conformação Molecular , Proteínas de Neoplasias/genética , Anormalidades da Pele , Células-Tronco/metabolismo , Sulfotransferases/genética , Sulfotransferases/metabolismo , Polegar/anormalidades , Polegar/patologia
9.
Arterioscler Thromb Vasc Biol ; 33(2): 354-61, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23202368

RESUMO

OBJECTIVE: The aim of this study was to analyze how an altered collagen structure affects development of atherosclerotic plaques. METHODS AND RESULTS: Fibromodulin-null mice develop an abnormal collagen fibril structure. In apolipoprotein E (ApoE)-null and ApoE/fibromodulin-null mice, a shear stress-modifying carotid artery cast induced formation of atherosclerotic plaques of different phenotypes; inflammatory in low-shear stress regions and fibrous in oscillatory shear stress regions. Electron microscopy showed that collagen fibrils were thicker and more heterogeneous in oscillatory shear stress lesions from ApoE/fibromodulin-null mice. Low-shear stress lesions were smaller in ApoE/fibromodulin-null mice and contained less lipids. Total plaque burden in aortas stained en face with Oil Red O, as well as lipid accumulation in aortic root lesions, was also decreased in ApoE/fibromodulin-null mice. In addition, lipid accumulation in RAW264.7 macrophages cultured on fibromodulin-deficient extracellular matrix was decreased, whereas levels of interleukin-6 and -10 were increased. Our results show that an abnormal plaque collagen fibril structure can influence atherosclerotic plaque development. CONCLUSIONS: The present findings suggest a more complex role for collagen in plaque stability than previously anticipated, in that it may promote lipid-accumulation and inflammation at the same time as it provides mechanical stability.


Assuntos
Aorta/metabolismo , Doenças da Aorta/metabolismo , Apolipoproteínas E/deficiência , Aterosclerose/metabolismo , Artérias Carótidas/metabolismo , Doenças das Artérias Carótidas/metabolismo , Proteínas da Matriz Extracelular/deficiência , Lipoproteínas LDL/metabolismo , Proteoglicanas/deficiência , Animais , Aorta/imunologia , Aorta/fisiopatologia , Aorta/ultraestrutura , Doenças da Aorta/genética , Doenças da Aorta/imunologia , Doenças da Aorta/patologia , Doenças da Aorta/fisiopatologia , Doenças da Aorta/prevenção & controle , Apolipoproteínas E/genética , Aterosclerose/genética , Aterosclerose/imunologia , Aterosclerose/patologia , Aterosclerose/fisiopatologia , Aterosclerose/prevenção & controle , Artérias Carótidas/imunologia , Artérias Carótidas/fisiopatologia , Artérias Carótidas/ultraestrutura , Doenças das Artérias Carótidas/genética , Doenças das Artérias Carótidas/imunologia , Doenças das Artérias Carótidas/patologia , Doenças das Artérias Carótidas/fisiopatologia , Doenças das Artérias Carótidas/prevenção & controle , Linhagem Celular , Proliferação de Células , Modelos Animais de Doenças , Regulação para Baixo , Proteínas da Matriz Extracelular/genética , Colágenos Fibrilares/metabolismo , Colágenos Fibrilares/ultraestrutura , Fibromodulina , Genótipo , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Eletrônica de Transmissão , Fenótipo , Placa Aterosclerótica , Proteoglicanas/genética , Fluxo Sanguíneo Regional , Estresse Mecânico
10.
PLoS One ; 7(3): e34082, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22479530

RESUMO

Stroma properties affect carcinoma physiology and direct malignant cell development. Here we present data showing that α(V)ß(3) expressed by stromal cells is involved in the control of interstitial fluid pressure (IFP), extracellular volume (ECV) and collagen scaffold architecture in experimental murine carcinoma. IFP was elevated and ECV lowered in syngeneic CT26 colon and LM3 mammary carcinomas grown in integrin ß(3)-deficient compared to wild-type BALB/c mice. Integrin ß(3)-deficiency had no effect on carcinoma growth rate or on vascular morphology and function. Analyses by electron microscopy of carcinomas from integrin ß(3)-deficient mice revealed a coarser and denser collagen network compared to carcinomas in wild-type littermates. Collagen fibers were built from heterogeneous and thicker collagen fibrils in carcinomas from integrin ß(3)-deficient mice. The fibrotic extracellular matrix (ECM) did not correlate with increased macrophage infiltration in integrin ß(3)-deficient mice bearing CT26 tumors, indicating that the fibrotic phenotype was not mediated by increased inflammation. In conclusion, we report that integrin ß(3)-deficiency in tumor stroma led to an elevated IFP and lowered ECV that correlated with a more fibrotic ECM, underlining the role of the collagen network for carcinoma physiology.


Assuntos
Carcinoma/metabolismo , Fibrose/patologia , Regulação Neoplásica da Expressão Gênica , Integrina beta3/genética , Animais , Linhagem Celular Tumoral , Colágeno/metabolismo , Líquido Extracelular , Feminino , Hidroxiprolina/metabolismo , Integrina alfaVbeta3/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Neoplasias , Fenótipo , Pressão
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