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1.
J Cell Sci ; 134(19)2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34477203

RESUMO

Both tumor cell-intrinsic signals and tumor cell-extrinsic signals from cells within the tumor microenvironment influence tumor cell dissemination and metastasis. The fibrillar collagen receptor tyrosine kinase (RTK) discoidin domain receptor 2 (DDR2) is essential for breast cancer metastasis in mouse models, and high expression of DDR2 in tumor and tumor stromal cells is strongly associated with poorer clinical outcomes. DDR2 tyrosine kinase activity has been hypothesized to be required for the metastatic activity of DDR2; however, inhibition of DDR2 tyrosine kinase activity, along with that of other RTKs, has failed to provide clinically relevant responses in metastatic patients. Here, we show that tyrosine kinase activity-independent action of DDR2 in tumor cells can support Matrigel invasion and in vivo metastasis. Paracrine actions of DDR2 in tumor cells and cancer-associated fibroblasts (CAFs) also support tumor invasion, migration and lung colonization in vivo. These data suggest that tyrosine kinase-independent functions of DDR2 could explain failures of tyrosine kinase inhibitor treatment in metastatic breast cancer patients and highlight the need for alternative therapeutic strategies that inhibit both tyrosine kinase-dependent and -independent actions of RTKs in the treatment of breast cancer. This article has an associated First Person interview with the first author of the paper.


Assuntos
Neoplasias da Mama , Fibroblastos Associados a Câncer , Receptor com Domínio Discoidina 2 , Animais , Neoplasias da Mama/genética , Fibroblastos Associados a Câncer/metabolismo , Movimento Celular , Receptor com Domínio Discoidina 2/genética , Receptor com Domínio Discoidina 2/metabolismo , Feminino , Fibroblastos/metabolismo , Humanos , Camundongos , Fosforilação , Microambiente Tumoral
2.
Dev Biol ; 458(2): 153-163, 2020 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-31697938

RESUMO

To investigate the role of adipose tissue in reproductive function and mammary gland development and function, we have examined lipodystrophic (LD) mice. LD mice of both sexes are sterile, but fertility can be restored with leptin injections. Mammary glands from lipodystrophic mice were rudimentary and lacked terminal end buds. Leptin-injected LD mice were able to become pregnant, showed normal pregnancy-associated glandular proliferation despite a smaller glandular area, were able to produce a small amount of milk that had grossly normal content of milk proteins and neutral lipids, but could not sustain pups to weaning. In order to separate the individual requirements for 1) adipokines such as leptin, 2) estradiol, and 3) physical epithelial-adipocyte interactions, we performed a series of experiments with both lipodystrophic mice and ob (obese mice with a mutation in the lep gene encoding the adipokine leptin) mice that received either estradiol treatment or preadipocyte transplant. The resulting fat pad did not rescue the defect in mammary gland development in lipodystrophic mice. The defect also could not be rescued with estradiol pellets. Ob/ob mice, like LD mice, lack leptin and estradiol, but retain adipose tissue. Ob mice have defective mammary gland development. However, in striking contrast to what was observed in lipodystrophic mice, reconstitution of a WT fat pad in ob mice rescued the defect in mammary gland development. Estradiol treatment did not rescue mammary gland development in ob mice. Therefore direct interaction between mammary gland epithelia and adipocytes is a requirement for full invasion and expansion of the gland, but is not required for glandular proliferation during pregnancy and milk production.


Assuntos
Tecido Adiposo/metabolismo , Células Epiteliais/metabolismo , Glândulas Mamárias Animais/metabolismo , Adipócitos/metabolismo , Tecido Adiposo/fisiologia , Animais , Estradiol/farmacologia , Feminino , Fertilidade , Lactação , Leptina/metabolismo , Leptina/farmacologia , Lipodistrofia/metabolismo , Lipodistrofia/patologia , Masculino , Glândulas Mamárias Animais/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Embrionárias Murinas , Obesidade , Transdução de Sinais
3.
Proc Natl Acad Sci U S A ; 115(5): E992-E1001, 2018 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-29339479

RESUMO

Bone metastasis involves dynamic interplay between tumor cells and the local stromal environment. In bones, local hypoxia and activation of the hypoxia-inducible factor (HIF)-1α in osteoblasts are essential to maintain skeletal homeostasis. However, the role of osteoblast-specific HIF signaling in cancer metastasis is unknown. Here, we show that osteoprogenitor cells (OPCs) are located in hypoxic niches in the bone marrow and that activation of HIF signaling in these cells increases bone mass and favors breast cancer metastasis to bone locally. Remarkably, HIF signaling in osteoblast-lineage cells also promotes breast cancer growth and dissemination remotely, in the lungs and in other tissues distant from bones. Mechanistically, we found that activation of HIF signaling in OPCs increases blood levels of the chemokine C-X-C motif ligand 12 (CXCL12), which leads to a systemic increase of breast cancer cell proliferation and dissemination through direct activation of the CXCR4 receptor. Hence, our data reveal a previously unrecognized role of the hypoxic osteogenic niche in promoting tumorigenesis beyond the local bone microenvironment. They also support the concept that the skeleton is an important regulator of the systemic tumor environment.


Assuntos
Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Osteoblastos/metabolismo , Alelos , Motivos de Aminoácidos , Animais , Neoplasias Ósseas/secundário , Osso e Ossos/metabolismo , Linhagem da Célula , Quimiocina CXCL12/sangue , Progressão da Doença , Feminino , Proteínas de Fluorescência Verde/metabolismo , Hipóxia , Ligantes , Camundongos , Camundongos Transgênicos , Metástase Neoplásica , Osteoclastos/metabolismo , Transdução de Sinais
4.
BMC Cancer ; 20(1): 542, 2020 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-32522170

RESUMO

BACKGROUND: Novel biomarkers are required to discern between breast tumors that should be targeted for treatment from those that would never become clinically apparent and/or life threatening for patients. Moreover, therapeutics that specifically target breast cancer (BC) cells with tumor-initiating capacity to prevent recurrence are an unmet need. We investigated the clinical importance of LGR5 in BC and ductal carcinoma in situ (DCIS) to explore LGR5 as a biomarker and a therapeutic target. METHODS: We stained BC (n = 401) and DCIS (n = 119) tissue microarrays with an antibody against LGR5. We examined an LGR5 knockdown ER- cell line that was orthotopically transplanted and used for in vitro colony assays. We also determined the tumor-initiating role of Lgr5 in lineage-tracing experiments. Lastly, we transplanted ER- patient-derived xenografts into mice that were subsequently treated with a LGR5 antibody drug conjugate (anti-LGR5-ADC). RESULTS: LGR5 expression correlated with small tumor size, lower grade, lymph node negativity, and ER-positivity. ER+ patients with LGR5high tumors rarely had recurrence, while high-grade ER- patients with LGR5high expression recurred and died due to BC more often. Intriguingly, all the DCIS patients who later died of BC had LGR5-positive tumors. Colony assays and xenograft experiments substantiated a role for LGR5 in ER- tumor initiation and subsequent growth, which was further validated by lineage-tracing experiments in ER- /triple-negative BC mouse models. Importantly, by utilizing LGR5high patient-derived xenografts, we showed that anti-LGR5-ADC should be considered as a therapeutic for high-grade ER- BC. CONCLUSION: LGR5 has distinct roles in ER- vs. ER+ BC with potential clinical applicability as a biomarker to identify patients in need of therapy and could serve as a therapeutic target for high-grade ER- BC.


Assuntos
Biomarcadores Tumorais/análise , Neoplasias da Mama/química , Carcinoma Intraductal não Infiltrante/química , Receptores Acoplados a Proteínas G/análise , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Biomarcadores Tumorais/imunologia , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Carcinoma Intraductal não Infiltrante/diagnóstico , Carcinoma Intraductal não Infiltrante/mortalidade , Carcinoma Intraductal não Infiltrante/patologia , Linhagem Celular Tumoral , Feminino , Xenoenxertos , Humanos , Camundongos , Pessoa de Meia-Idade , Prognóstico , RNA Neoplásico/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real , Receptor ErbB-2/análise , Receptores Acoplados a Proteínas G/imunologia , Análise Serial de Tecidos/métodos
5.
Proc Natl Acad Sci U S A ; 109(39): E2595-604, 2012 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-22923691

RESUMO

Breast cancer progression involves genetic changes and changes in the extracellular matrix (ECM). To test the importance of the ECM in tumor cell dissemination, we cultured epithelium from primary human breast carcinomas in different ECM gels. We used basement membrane gels to model the normal microenvironment and collagen I to model the stromal ECM. In basement membrane gels, malignant epithelium either was indolent or grew collectively, without protrusions. In collagen I, epithelium from the same tumor invaded with protrusions and disseminated cells. Importantly, collagen I induced a similar initial response of protrusions and dissemination in both normal and malignant mammary epithelium. However, dissemination of normal cells into collagen I was transient and ceased as laminin 111 localized to the basal surface, whereas dissemination of carcinoma cells was sustained throughout culture, and laminin 111 was not detected. Despite the large impact of ECM on migration strategy, transcriptome analysis of our 3D cultures revealed few ECM-dependent changes in RNA expression. However, we observed many differences between normal and malignant epithelium, including reduced expression of cell-adhesion genes in tumors. Therefore, we tested whether deletion of an adhesion gene could induce sustained dissemination of nontransformed cells into collagen I. We found that deletion of P-cadherin was sufficient for sustained dissemination, but exclusively into collagen I. Our data reveal that metastatic tumors preferentially disseminate in specific ECM microenvironments. Furthermore, these data suggest that breaks in the basement membrane could induce invasion and dissemination via the resulting direct contact between cancer cells and collagen I.


Assuntos
Neoplasias da Mama , Movimento Celular , Regulação Neoplásica da Expressão Gênica , Neoplasias Mamárias Animais , Microambiente Tumoral , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Feminino , Humanos , Neoplasias Mamárias Animais/genética , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Camundongos , Invasividade Neoplásica
6.
Infect Immun ; 77(7): 2840-8, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19398546

RESUMO

"Cluster 9" family lipoproteins function as ligand-binding subunits of ABC-type transporters in maintaining transition metal homeostasis and have been implicated in the virulence of several bacteria. While these proteins share high similarity, the specific metal that they recognize and whether their role in virulence directly involves metal homeostasis cannot be reliably predicted. We examined the cluster 9 protein Lsp of Streptococcus pyogenes and found that specific deletion of lsp produced mutants highly attenuated in a murine model of soft tissue infection. Under standard in vitro conditions, growth of the Lsp(-) mutant was indistinguishable from that of the wild type, but growth was defective under zinc-limited conditions. The growth defect could be complemented by plasmids expressing wild-type Lsp but not Lsp engineered to lack its putative lipidation residue. Furthermore, Zn(2+) but not Mn(2+) rescued Lsp(-) growth, implicating Zn(2+) as the physiological ligand for Lsp. Mutation of residues in the putative Zn(2+)-binding pocket generated variants both hypo- and hyper-resistant to zinc starvation, and both mutant classes displayed attenuated virulence. Together, these data suggest that Lsp is a ligand-binding component of an ABC-type zinc permease and that perturbation of zinc homeostasis inhibits the ability of S. pyogenes to cause disease in a zinc-limited host milieu.


Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Streptococcus pyogenes/metabolismo , Streptococcus pyogenes/patogenicidade , Fatores de Virulência/fisiologia , Zinco/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Sítios de Ligação , Deleção de Genes , Teste de Complementação Genética , Manganês/metabolismo , Camundongos , Mutagênese Sítio-Dirigida , Mutação de Sentido Incorreto , Infecções dos Tecidos Moles/microbiologia , Infecções dos Tecidos Moles/patologia , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/patologia , Streptococcus pyogenes/crescimento & desenvolvimento , Virulência , Fatores de Virulência/genética
7.
Cancer Res ; 79(8): 1899-1912, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30862718

RESUMO

Collective cell migration is an adaptive, coordinated interactive process involving cell-cell and cell-extracellular matrix (ECM) microenvironmental interactions. A critical aspect of collective migration is the sensing and establishment of directional movement. It has been proposed that a subgroup of cells known as leader cells localize at the front edge of a collectively migrating cluster and are responsible for directing migration. However, it is unknown how and when leader cells arrive at the front edge and what environmental cues dictate leader cell development and behavior. Here, we addressed these questions by combining a microfluidic device design that mimics multiple tumor microenvironmental cues concurrently with biologically relevant primary, heterogeneous tumor cell organoids. Prior to migration, breast tumor leader cells (K14+) were present throughout a tumor organoid and migrated (polarized) to the leading edge in response to biochemical and biomechanical cues. Impairment of either CXCR4 (biochemical responsive) or the collagen receptor DDR2 (biomechanical responsive) abrogated polarization of leader cells and directed collective migration. This work demonstrates that K14+ leader cells utilize both chemical and mechanical cues from the microenvironment to polarize to the leading edge of collectively migrating tumors. SIGNIFICANCE: These findings demonstrate that pre-existing, randomly distributed leader cells within primary tumor organoids use CXCR4 and DDR2 to polarize to the leading edge and direct migration.


Assuntos
Movimento Celular , Receptor com Domínio Discoidina 2/fisiologia , Queratina-14/metabolismo , Neoplasias Mamárias Experimentais/patologia , Organoides/patologia , Receptores CXCR4/metabolismo , Animais , Comunicação Celular , Diferenciação Celular , Matriz Extracelular , Feminino , Humanos , Queratina-14/genética , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Transgênicos , Organoides/metabolismo , Receptores CXCR4/genética , Transdução de Sinais , Microambiente Tumoral
8.
Elife ; 82019 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-31144616

RESUMO

Biomechanical changes in the tumor microenvironment influence tumor progression and metastases. Collagen content and fiber organization within the tumor stroma are major contributors to biomechanical changes (e., tumor stiffness) and correlated with tumor aggressiveness and outcome. What signals and in what cells control collagen organization within the tumors, and how, is not fully understood. We show in mouse breast tumors that the action of the collagen receptor DDR2 in CAFs controls tumor stiffness by reorganizing collagen fibers specifically at the tumor-stromal boundary. These changes were associated with lung metastases. The action of DDR2 in mouse and human CAFs, and tumors in vivo, was found to influence mechanotransduction by controlling full collagen-binding integrin activation via Rap1-mediated Talin1 and Kindlin2 recruitment. The action of DDR2 in tumor CAFs is thus critical for remodeling collagen fibers at the tumor-stromal boundary to generate a physically permissive tumor microenvironment for tumor cell invasion and metastases.


Assuntos
Neoplasias da Mama/patologia , Neoplasias da Mama/fisiopatologia , Fibroblastos Associados a Câncer/metabolismo , Receptor com Domínio Discoidina 2/metabolismo , Integrinas/metabolismo , Metástase Neoplásica/fisiopatologia , Animais , Colágeno/metabolismo , Modelos Animais de Doenças , Humanos , Camundongos , Microambiente Tumoral
9.
Oncogenesis ; 7(3): 32, 2018 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-29593211

RESUMO

The EMT inducer SNAIL1 regulates breast cancer metastasis and its expression in human primary breast tumor predicts for poor outcomes. During tumor progression SNAIL1 has multiple effects in tumor cells that can impact metastasis. An inflammatory tumor microenvironment also impacts metastasis and recently SNAIL1 has been implicated as modulating the secretion of cytokines that can influence the tumor immune infiltrate. Using a spontaneous genetic model of breast cancer metastasis and syngeneic orthotopic transplant experiments we show that the action of SNAIL1 in primary breast tumor cells is required for breast tumor growth and metastasis. It does so, in part, by regulating production of GM-CSF, IL1α, IL-6, and TNFα by breast cancer cells. The SNAIL1-dependent tumor cell secretome modulates the primary tumor-associated macrophage (TAM) polarization. GM-CSF alone modulates TAM polarization and impacts breast cancer metastasis in vivo. This study highlights another role for breast tumor SNAIL1 in cancer progression to metastasis-modulation of the immune microenvironment of primary breast tumors.

10.
Biochim Biophys Acta ; 1761(11): 1391-9, 2006 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17081801

RESUMO

As saprophytes or disease causing microorganisms, fungi acquire nutrients from dead organic material or living host organisms. Lipids as structural components of cell membranes and storage compartments play an important role as energy-rich food source. In recent years, it also has become clear that lipids have a wide range of bioactive properties including signal transduction and cell to cell communication. Thus, it is not surprising that fungi possess a broad range of hydrolytic enzymes that attack neutral lipids and phospholipids. Especially during infection of a mammalian host, phospholipase A(2) (PLA(2)) enzymes released by fungi could play important roles not only for nutrient acquisition and tissue invasion, but for intricate modulation of the host's immune response. Sequencing of fungal genomes has revealed a wide range of genes encoding PLA(2) activities in fungi. We are just beginning to become aware of the significance these enzymes could have for the fungal cells and their interaction with the host.


Assuntos
Proteínas Fúngicas/metabolismo , Fungos/enzimologia , Lisofosfolipase/metabolismo , Micoses/enzimologia , Fosfolipases A/metabolismo , Transdução de Sinais , Animais , Proteínas Fúngicas/genética , Proteínas Fúngicas/imunologia , Fungos/genética , Fungos/imunologia , Genoma Fúngico/imunologia , Fosfolipases A2 do Grupo IV , Humanos , Lisofosfolipase/genética , Lisofosfolipase/imunologia , Micoses/genética , Micoses/imunologia , Fosfolipases A/genética , Fosfolipases A/imunologia , Fosfolipases A2 , Fosfolipídeos/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia
11.
Cell Rep ; 15(11): 2510-23, 2016 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-27264173

RESUMO

High levels of collagen deposition in human and mouse breast tumors are associated with poor outcome due to increased local invasion and distant metastases. Using a genetic approach, we show that, in mice, the action of the fibrillar collagen receptor discoidin domain receptor 2 (DDR2) in both tumor and tumor-stromal cells is critical for breast cancer metastasis yet does not affect primary tumor growth. In tumor cells, DDR2 in basal epithelial cells regulates the collective invasion of tumor organoids. In stromal cancer-associated fibroblasts (CAFs), DDR2 is critical for extracellular matrix production and the organization of collagen fibers. The action of DDR2 in CAFs also enhances tumor cell collective invasion through a pathway distinct from the tumor-cell-intrinsic function of DDR2. This work identifies DDR2 as a potential therapeutic target that controls breast cancer metastases through its action in both tumor cells and tumor-stromal cells at the primary tumor site.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/patologia , Receptor com Domínio Discoidina 2/metabolismo , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Alelos , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Matriz Extracelular/metabolismo , Feminino , Deleção de Genes , Humanos , Queratina-14/metabolismo , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Invasividade Neoplásica , Organoides/patologia , Células Estromais/patologia , Microambiente Tumoral
12.
Nat Cell Biol ; 15(2): 201-13, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23354167

RESUMO

Despite advances in our understanding of breast cancer, patients with metastatic disease have poor prognoses. GATA3 is a transcription factor that specifies and maintains mammary luminal epithelial cell fate, and its expression is lost in breast cancer, correlating with a worse prognosis in human patients. Here, we show that GATA3 promotes differentiation, suppresses metastasis and alters the tumour microenvironment in breast cancer by inducing microRNA-29b (miR-29b) expression. Accordingly, miR-29b is enriched in luminal breast cancers and loss of miR-29b, even in GATA3-expressing cells, increases metastasis and promotes a mesenchymal phenotype. Mechanistically, miR-29b inhibits metastasis by targeting a network of pro-metastatic regulators involved in angiogenesis, collagen remodelling and proteolysis, including VEGFA, ANGPTL4, PDGF, LOX and MMP9, and targeting ITGA6, ITGB1 and TGFB, thereby indirectly affecting differentiation and epithelial plasticity. The discovery that a GATA3-miR-29b axis regulates the tumour microenvironment and inhibits metastasis opens up possibilities for therapeutic intervention in breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Movimento Celular , Fator de Transcrição GATA3/metabolismo , MicroRNAs/metabolismo , Microambiente Tumoral , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/secundário , Diferenciação Celular , Linhagem Celular Tumoral , Feminino , Fator de Transcrição GATA3/genética , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Genótipo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Fenótipo , Transfecção , Regulação para Cima
13.
Cell Rep ; 3(1): 70-8, 2013 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-23352663

RESUMO

Mammary epithelial stem cells are vital to tissue expansion and remodeling during various phases of postnatal mammary development. Basal mammary epithelial cells are enriched in Wnt-responsive cells and can reconstitute cleared mammary fat pads upon transplantation into mice. Lgr5 is a Wnt-regulated target gene and was identified as a major stem cell marker in the small intestine, colon, stomach, and hair follicle, as well as in kidney nephrons. Here, we demonstrate the outstanding regenerative potential of a rare population of Lgr5-expressing (Lgr5(+)) mammary epithelial cells (MECs). We found that Lgr5(+) cells reside within the basal population, are superior to other basal cells in regenerating functional mammary glands (MGs), are exceptionally efficient in reconstituting MGs from single cells, and exhibit regenerative capacity in serial transplantations. Loss-of-function and depletion experiments of Lgr5(+) cells from transplanted MECs or from pubertal MGs revealed that these cells are not only sufficient but also necessary for postnatal mammary organogenesis.


Assuntos
Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Organogênese , Receptores Acoplados a Proteínas G/metabolismo , Animais , Animais Recém-Nascidos , Antígeno CD24/metabolismo , Toxina Diftérica/farmacologia , Feminino , Proteínas de Fluorescência Verde/metabolismo , Queratina-14/metabolismo , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/transplante , Camundongos , Camundongos Endogâmicos C57BL , Regeneração/efeitos dos fármacos , Maturidade Sexual/efeitos dos fármacos , Análise de Célula Única , Tamoxifeno/farmacologia
14.
Cancer Res ; 70(6): 2224-34, 2010 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-20215503

RESUMO

Prostate cancer is the leading form of cancer in men. Prostate tumors often contain neuroendocrine differentiation, which correlates with androgen-independent progression and poor prognosis. Matrix metalloproteinases (MMP), a family of enzymes that remodel the microenvironment, are associated with tumorigenesis and metastasis. To evaluate MMPs during metastatic prostatic neuroendocrine cancer development, we used transgenic mice expressing SV40 large T antigen in their prostatic neuroendocrine cells, under the control of transcriptional regulatory elements from the mouse cryptdin-2 gene (CR2-TAg). These mice have a stereotypical pattern of tumorigenesis and metastasis. MMP-2, MMP-7, and MMP-9 activities increased concurrently with the transition to invasive metastatic carcinoma, but they were expressed in different prostatic cell types: stromal, luminal epithelium, and macrophages, respectively. CR2-TAg mice treated with AG3340/Prinomastat, an MMP inhibitor that blocks activity of MMP-2, MMP-9, MMP-13, and MMP-14, had reduced tumor burden. CR2-TAg animals were crossed to mice homozygous for null alleles of MMP-2, MMP-7, or MMP-9 genes. At 24 weeks CR2-TAg; MMP-2(-/-) mice showed reduced tumor burden, prolonged survival, decreased lung metastasis, and decreased blood vessel density, whereas deficiencies in MMP-7 or MMP-9 did not influence tumor growth or survival. Mice deficient for MMP-7 had reduced endothelial area coverage and decreased vessel size, and mice lacking MMP-9 had increased numbers of invasive foci and increased perivascular invasion, as well as decreased tumor blood vessel size. Together, these results suggest distinct contributions by MMPs to the progression of aggressive prostate tumor and to helping tumors cleverly find alternative routes to malignant progression.


Assuntos
Metaloproteinases da Matriz/metabolismo , Tumores Neuroendócrinos/enzimologia , Tumores Neuroendócrinos/patologia , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/patologia , Animais , Antineoplásicos/farmacologia , Apoptose/fisiologia , Processos de Crescimento Celular/fisiologia , Progressão da Doença , Inibidores Enzimáticos/farmacologia , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/secundário , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 7 da Matriz/genética , Metaloproteinase 7 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz , Metaloproteinases da Matriz/genética , Camundongos , Camundongos Transgênicos , Metástase Neoplásica , Neovascularização Patológica/enzimologia , Neovascularização Patológica/patologia , Tumores Neuroendócrinos/irrigação sanguínea , Compostos Orgânicos/farmacologia , Neoplasias da Próstata/irrigação sanguínea , Células Estromais/enzimologia , Células Estromais/patologia
15.
Dev Cell ; 14(4): 570-81, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18410732

RESUMO

Epithelial organs are built through the movement of groups of interconnected cells. We observed cells in elongating mammary ducts reorganize into a multilayered epithelium, migrate collectively, and rearrange dynamically, all without forming leading cellular extensions. Duct initiation required proliferation, Rac, and myosin light-chain kinase, whereas repolarization to a bilayer depended on Rho kinase. We observed that branching morphogenesis results from the active motility of both luminal and myoepithelial cells. Luminal epithelial cells advanced collectively, whereas myoepithelial cells appeared to restrain elongating ducts. Significantly, we observed that normal epithelium and neoplastic hyperplasias are organized similarly, suggesting common mechanisms of epithelial growth.


Assuntos
Movimento Celular/fisiologia , Células Epiteliais/fisiologia , Glândulas Mamárias Animais , Morfogênese/fisiologia , Actinas/metabolismo , Animais , Biomarcadores/metabolismo , Polaridade Celular , Proliferação de Células , Extensões da Superfície Celular/metabolismo , Extensões da Superfície Celular/ultraestrutura , Células Cultivadas , Progressão da Doença , Inibidores Enzimáticos/metabolismo , Células Epiteliais/citologia , Epitélio/anatomia & histologia , Epitélio/fisiologia , Feminino , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/crescimento & desenvolvimento , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Camundongos , Camundongos Transgênicos , Quinase de Cadeia Leve de Miosina/antagonistas & inibidores , Quinase de Cadeia Leve de Miosina/metabolismo , Organoides/citologia , Organoides/fisiologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais/fisiologia , Proteínas rac1 de Ligação ao GTP/antagonistas & inibidores , Proteínas rac1 de Ligação ao GTP/metabolismo , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/metabolismo
16.
Mol Microbiol ; 63(4): 1185-96, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17238923

RESUMO

In Streptococcus pyogenes, mutation of the peroxide sensor PerR results in avirulence despite producing hyper-resistance to peroxide stress. To understand the basis of this effect, global transcription profiling was conducted revealing one highly downregulated gene (czcD), and five highly upregulated genes in the mutant. Of the latter, only pmtA contained a binding site for PerR, while phtY, phtD, lsp and rpsN2 harboured an AdcR motif and were regulated by AdcR, a repressor of an ABC-type metal transporter. Furthermore, only the PerR-regulated pmtA (PerR-regulated metal transporter A), a putative metal transporter, contributed to resistance against peroxide stress, while AdcR and the other AdcR-regulated genes did not. However, overexpression of pmtA resulted in upregulation of several AdcR-regulated genes, suggesting that the AdcR regulon is sensitive to PerR regulation of metal homeostasis. Finally, examination of S. pyogenes following murine subcutaneous infection revealed that while pmtA was not upregulated in a late infection, the AdcR-regulated genes were. Taken together, these data suggest that PerR has a greater impact on the transcriptome than can be predicted by its binding sites and that pmtA functions to link metal homeostasis and oxidative stress responses.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Transporte/metabolismo , Metais/metabolismo , Estresse Oxidativo , Proteínas Repressoras/metabolismo , Streptococcus pyogenes/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteínas de Bactérias/genética , Sítios de Ligação , Proteínas de Transporte/genética , Regulação Bacteriana da Expressão Gênica , Homeostase , Peróxido de Hidrogênio/farmacologia , Camundongos , Mutação , Regulon , Proteínas Repressoras/genética , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/microbiologia , Streptococcus pyogenes/efeitos dos fármacos , Streptococcus pyogenes/genética , Fatores de Transcrição/genética , Zinco/farmacologia
17.
Int J Med Microbiol ; 296(6): 405-20, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16759910

RESUMO

Phospholipases are critical for modification and redistribution of lipid substrates, membrane remodeling and microbial virulence. Among the many different classes of phospholipases, fungal phospholipase B (Plb) proteins show the broadest range of substrate specificity and hydrolytic activity, hydrolyzing acyl ester bonds in phospholipids and lysophospholipids and further catalyzing lysophospholipase-transacylase reactions. The genome of the opportunistic fungal pathogen Candida albicans encodes a PLB multigene family with five putative members; we present the first characterization of this group of potential virulence determinants. CaPLB5, the third member of this multigene family characterized herein is a putative secretory protein with a predicted GPI-anchor attachment site. Real-time RT-PCR gene expression analysis of CaPLB5 and the additional CaPLB gene family members revealed that filamentous growth and physiologically relevant environmental conditions are associated with increased PLB gene activity. The phenotypes expressed by null mutant and revertant strains of CaPLB5 indicate that this lipid hydrolase plays an important role for cell-associated phospholipase A(2) activity and in vivo organ colonization.


Assuntos
Candida albicans/enzimologia , Candida albicans/genética , Lisofosfolipase/genética , Fosfolipases A/genética , Virulência/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Candida albicans/patogenicidade , Candidíase/etiologia , Perfilação da Expressão Gênica , Camundongos , Dados de Sequência Molecular , Família Multigênica/genética , Fenótipo , Fosfolipases A2
18.
Mol Microbiol ; 55(1): 221-34, 2005 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-15612930

RESUMO

Prior studies have shown that the catalase-deficient pathogen Streptococcus pyogenes (group A streptococcus) has a robust ability to resist oxidative stress that partially involves the transcriptional regulator PerR. However, the extent of the PerR regulon and the contribution of the members of this regulon to virulence are unknown. In this study, DNase I footprinting revealed that PerR binds specifically to a single site upstream of the promoter for the gene encoding alkyl hydroperoxide reductase (ahpC). However, analyses of transcript abundance revealed that while ahpC is regulated in response to growth phase, its regulation is independent of PerR. Instead, PerR regulates transcription of a divergent gene cluster that encodes a putative cold shock protein. The gene encoding the Dps-like peroxide resistance protein MrgA was repressed by PerR, consistent with the presence of a PerR binding site in its promoter. Phenotypic analyses of PerR-, AhpC- and MrgA- mutants revealed that while AhpC is not essential for resistance to challenge with hydrogen peroxide in vitro, AhpC does contribute to scavenging of endogenous hydrogen peroxide and is required for virulence in a murine model of infection. In contrast, a MrgA- mutant was hypersensitive to challenge with peroxide in vitro, but was fully virulent in all animal models tested. Finally, a PerR- mutant was hyper-resistant to peroxide, yet was highly attenuated for virulence in all murine models. These data demonstrate that while a mutant's capacity to resist peroxide stress did not directly correlate with its ability to cause disease, the appropriate regulation of the peroxide stress response is critical for virulence.


Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Farmacorresistência Bacteriana/genética , Regulação Bacteriana da Expressão Gênica , Peróxidos/farmacologia , Regulon , Proteínas Repressoras/genética , Streptococcus pyogenes/genética , Fatores de Transcrição/genética , Animais , Proteínas de Bactérias/fisiologia , Sequência de Bases , Pegada de DNA , DNA Bacteriano , Farmacorresistência Bacteriana/fisiologia , Camundongos , Dados de Sequência Molecular , Mutação , Oxidantes/farmacologia , Estresse Oxidativo , Peroxidases/genética , Peroxidases/metabolismo , Peroxirredoxinas , Regiões Promotoras Genéticas , Ligação Proteica , Proteínas Repressoras/fisiologia , Infecções Estreptocócicas/microbiologia , Streptococcus pyogenes/efeitos dos fármacos , Streptococcus pyogenes/patogenicidade , Fatores de Transcrição/fisiologia , Virulência/genética
19.
Infect Immun ; 72(1): 408-13, 2004 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-14688122

RESUMO

Glutathione peroxidases are widespread among eukaryotic organisms and function as a major defense against hydrogen peroxide and organic peroxides. However, glutathione peroxidases are not well studied among prokaryotic organisms and have not previously been shown to promote bacterial virulence. Recently, a gene with homology to glutathione peroxidase was shown to contribute to the antioxidant defenses of Streptococcus pyogenes (group A streptococcus). Since this bacterium causes numerous suppurative diseases that require it to thrive in highly inflamed tissue, it was of interest to determine if glutathione peroxidase is important for virulence. In this study, we report that GpoA glutathione peroxidase is the major glutathione peroxidase in S. pyogenes and is essential for S. pyogenes pathogenesis in several murine models that mimic different aspects of streptococcal suppurative disease. In contrast, glutathione peroxidase is not essential for virulence in a zebrafish model of streptococcal myositis, a disease characterized by the absence of an inflammatory cell infiltrate. Taken together, these data suggest that S. pyogenes requires glutathione peroxidase to adapt to oxidative stress that accompanies an inflammatory response, and the data provide the first demonstration of a role for glutathione peroxidase in bacterial virulence. The fact that genes encoding putative glutathione peroxidases are found in the genomes of many pathogenic bacterial species suggests that glutathione peroxidase may have a general role in bacterial pathogenesis.


Assuntos
Glutationa Peroxidase/metabolismo , Infecções Estreptocócicas/fisiopatologia , Streptococcus pyogenes/enzimologia , Streptococcus pyogenes/patogenicidade , Animais , Modelos Animais de Doenças , Feminino , Glutationa Peroxidase/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Índice de Gravidade de Doença , Pele/microbiologia , Infecções Estreptocócicas/microbiologia , Virulência , Peixe-Zebra
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