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1.
Magn Reson Imaging ; 75: 156-161, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33130057

RESUMO

PURPOSE: Spectrally selective fat saturation (FatSat) sequence is commonly used to suppress signal from adipose tissue. Conventional SINC-shaped pulses are sensitive to B0 off-resonance and B1+ offset. Uniform fat saturation with large spatial coverage is especially challenging for the body and breast MRI. The aim of this study is to develop spectrally selective FatSat pulses that offer more immunity to B0/B1+ field inhomogeneities than SINC pulses and evaluate them in bilateral breast imaging at 3 T. MATERIALS AND METHODS: Optimized composite pulses (OCP) were designed based on the optimal control theory with robustness to a targeted B0/ B1+ conditions. OCP pulses also allows flexible flip angles to meet different requirements. Comparisons with the vendor-provided SINC pulses were conducted by numerical simulation and in vivo scans using a 3D T1-weighted (T1w) gradient-echo (GRE) sequence with coverage of the whole-breast. RESULTS: Simulation revealed that OCP pulses yielded almost half of the transition band and much less sensitivity to B1+ inhomogeneity compared to SINC pulses with B0 off-resonance within ±200 Hz and B1+ scale error within ±0.3 (P < 0.001). Across five normal subjects, OCP FatSat pulses produced 25-41% lower residual fat signals (P < 0.05) with 27-36% less spatial variation (P < 0.05) than SINC. CONCLUSION: In contrast to conventional SINC-shaped pulses, the newly designed OCP FatSat pulses mitigated challenges of wide range of B0/ B1+ field inhomogeneities and achieved more uniform fat suppression in bilateral breast T1w imaging at 3 T.


Assuntos
Tecido Adiposo/diagnóstico por imagem , Mama/citologia , Mama/diagnóstico por imagem , Imageamento Tridimensional , Imagem por Ressonância Magnética , Simulação por Computador , Feminino , Humanos , Imagens de Fantasmas
2.
Int J Mol Sci ; 21(24)2020 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-33322711

RESUMO

Circulating tumor cells (CTCs) and the immune infiltration of tumors are closely related to clinical outcomes. This study aimed to verify the influence of stromal lymphocyte infiltration and the immune context of tumor microenvironment on the hematogenous spread and prognosis of 282 chemotherapy naïve primary BC patients. To detect the presence of mesenchymal CTCs, RNA extracted from CD45-depleted peripheral blood was interrogated for the expression of mesenchymal gene transcripts. Tumor-infiltrating lymphocytes (TILs) were detected in the stromal areas by immunohistochemistry, using CD3, CD8, and CD45RO antibodies. The concentrations of 51 plasma cytokines were measured by multiplex bead arrays. TILs infiltration in mesenchymal CTC-positive patients significantly decreased their progression-free survival (HR = 4.88, 95% CI 2.30-10.37, p < 0.001 for CD3high; HR = 6.17, 95% CI 2.75-13.80, p < 0.001 for CD8high; HR = 6.93, 95% CI 2.86-16.81, p < 0.001 for CD45ROhigh). Moreover, the combination of elevated plasma concentrations of transforming growth factor beta-3 (cut-off 662 pg/mL), decreased monocyte chemotactic protein-3 (cut-off 52.5 pg/mL) and interleukin-15 (cut-off 17.1 pg/mL) significantly increased the risk of disease recurrence (HR = 4.838, 95% CI 2.048-11.427, p < 0.001). Our results suggest a strong impact of the immune tumor microenvironment on BC progression, especially through influencing the dissemination and survival of more aggressive, mesenchymal CTC subtypes.


Assuntos
Neoplasias da Mama/imunologia , Neoplasias da Mama/mortalidade , Citocinas/sangue , Linfócitos do Interstício Tumoral/imunologia , Microambiente Tumoral/imunologia , Mama/citologia , Mama/imunologia , Mama/patologia , Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Complexo CD3/metabolismo , Antígenos CD8/metabolismo , Quimiocina CCL7/sangue , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Interleucina-15/sangue , Antígenos Comuns de Leucócito/metabolismo , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Células Neoplásicas Circulantes/patologia , Prognóstico , Fatores de Risco , Células Estromais/imunologia , Células Estromais/metabolismo , Fator de Crescimento Transformador beta3/sangue
3.
Nat Commun ; 11(1): 4977, 2020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-33020483

RESUMO

Although thousands of breast cancer cells disseminate and home to bone marrow until primary surgery, usually less than a handful will succeed in establishing manifest metastases months to years later. To identify signals that support survival or outgrowth in patients, we profile rare bone marrow-derived disseminated cancer cells (DCCs) long before manifestation of metastasis and identify IL6/PI3K-signaling as candidate pathway for DCC activation. Surprisingly, and similar to mammary epithelial cells, DCCs lack membranous IL6 receptor expression and mechanistic dissection reveals IL6 trans-signaling to regulate a stem-like state of mammary epithelial cells via gp130. Responsiveness to IL6 trans-signals is found to be niche-dependent as bone marrow stromal and endosteal cells down-regulate gp130 in premalignant mammary epithelial cells as opposed to vascular niche cells. PIK3CA activation renders cells independent from IL6 trans-signaling. Consistent with a bottleneck function of microenvironmental DCC control, we find PIK3CA mutations highly associated with late-stage metastatic cells while being extremely rare in early DCCs. Our data suggest that the initial steps of metastasis formation are often not cancer cell-autonomous, but also depend on microenvironmental signals.


Assuntos
Interleucina-6/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Transdução de Sinais , Medula Óssea/patologia , Mama/citologia , Neoplasias da Mama/patologia , Classe I de Fosfatidilinositol 3-Quinases/genética , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Receptor gp130 de Citocina/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Humanos , Interleucina-6/genética , Mutação , Metástase Neoplásica/genética , Receptores de Interleucina-6/deficiência , Receptores de Interleucina-6/metabolismo , Células Estromais/metabolismo , Microambiente Tumoral
4.
Adv Exp Med Biol ; 1252: 27-32, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32816259

RESUMO

Breast tissue reveals some physiologic changes during pregnancy and lactation due to hormonal alterations. Whole range of breast diseases including inflammatory, benign and malignant neoplasms can be seen in pregnancy but due to concurrent physiologic changes, may lead to diagnostic challenges. This chapter reviews sampling methods and histologic features of common benign breast lesions in pregnancy and lactation periods.


Assuntos
Doenças Mamárias/patologia , Mama/citologia , Mama/patologia , Lactação/fisiologia , Complicações na Gravidez , Gravidez/fisiologia , Neoplasias da Mama/patologia , Feminino , Humanos
5.
Cancer Causes Control ; 31(9): 827-837, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32476101

RESUMO

PURPOSE: We investigated the associations of aspirin and other non-steroid anti-inflammatory drugs with mammographic breast density (MBD) and their interactions in relation to breast cancer risk. METHODS: This study included 3,675 cancer-free women within the Nurses' Health Study (NHS) and Nurses' Health Study II (NHSII) cohorts. Percent breast density (PD), absolute dense area (DA), and non-dense area (NDA) were measured from digitized film mammograms using a computer-assisted thresholding technique; all measures were square root-transformed. Information on medication use was collected in 1980 (NHS) and 1989 (NHSII) and updated biennially. Medication use was defined as none, past or current; average cumulative dose and frequency were calculated for all past or current users from all bi-annual questionnaires preceding the mammogram date. We used generalized linear regression to quantify associations of medications with MBD. Two-way interactions were examined in logistic regression models. RESULTS: In multivariate analysis, none of the anti-inflammatory medications were associated with PD, DA, and NDA. We found no interactions of any of the medications with PD with respect to breast cancer risk (all p-interactions > 0.05). However, some of the aspirin variables appeared to have positive associations with breast cancer risk limited only to women with PD 10-24% (past aspirin OR 1.56, 95% CI 1.03-2.35; current aspirin with < 5 years of use OR 1.82, 95% CI 1.01-3.28; current aspirin with ≥ 5 years of use OR 1.89, 95% CI 1.26-2.82). CONCLUSIONS: Aspirin and NSAIDs are not associated with breast density measures. We found no interactions of aspirin with MBD in relation to breast cancer risk.


Assuntos
Anti-Inflamatórios não Esteroides/administração & dosagem , Aspirina/administração & dosagem , Densidade da Mama/efeitos dos fármacos , Neoplasias da Mama/epidemiologia , Mama/diagnóstico por imagem , Adulto , Mama/citologia , Mama/efeitos dos fármacos , Mama/patologia , Neoplasias da Mama/diagnóstico por imagem , Estudos de Coortes , Feminino , Humanos , Modelos Lineares , Mamografia , Pessoa de Meia-Idade , Análise Multivariada , Risco , Inquéritos e Questionários , Estados Unidos/epidemiologia
6.
J Surg Oncol ; 121(8): 1181-1190, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32167588

RESUMO

BACKGROUND AND OBJECTIVES: To investigate the performance of texture analysis based on enhancement kinetic parametric maps derived from breast dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in discriminating benign from malignant tumors. METHODS: A total of 192 cases confirmed by histopathology were retrospectively selected from our Picture Archiving and Communication System, including 93 benign and 99 malignant tumors. Lesion areas were delineated semi-automatically, and six kinetic parametric maps reflecting the perfusion information were generated, namely the maximum slope of increase (MSI), slope of signal intensity (SIslope ), initial percentage of peak enhancement (Einitial ), percentage of peak enhancement (Epeak ), early signal enhancement ratio (ESER), and second enhancement percentage (SEP) maps. A total of 286 texture features were extracted from those quantitative parametric maps. The Student t test or Mann-Whitney U test was used to select features that were statistically significantly different between the benign and malignant groups. A support vector machine was employed with a leave-one-out cross-validation method to establish the classification model. Classification performance was evaluated according to the receiver operating characteristic (ROC) theory. RESULTS: The areas under ROC curves (AUCs) indicating the diagnostic performance were 0.925 for MSI, 0.854 for SIslope , 0.756 for Einitial , 0.923 for Epeak , 0.871 for ESER and 0.881 for SEP. Significant differences in AUCs were found between Einitial vs MSI, Einitial vs Epeak and Einitial vs SEP (P < .05). There were no significant differences in other pairwise comparisons. CONCLUSION: Texture analysis of the kinetic parametric maps derived from breast DCE-MRI can contribute to the discrimination between malignant and benign lesions. It can be considered as a supplementary tool for breast diagnosis.


Assuntos
Neoplasias da Mama/diagnóstico por imagem , Mama/diagnóstico por imagem , Adulto , Mama/citologia , Mama/patologia , Neoplasias da Mama/patologia , Meios de Contraste , Feminino , Humanos , Interpretação de Imagem Assistida por Computador/métodos , Imagem por Ressonância Magnética/métodos , Pessoa de Meia-Idade , Estudos Retrospectivos , Adulto Jovem
7.
Mol Cell Biol ; 40(9)2020 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-32015100

RESUMO

During aging, cellular plasticity and senescence play important roles in tissue regeneration and the pathogenesis of different diseases, including cancer. We have recently shown that senescent breast luminal cells can activate their adjacent stromal fibroblasts. In the present report, we present clear evidence that these senescence-related active fibroblasts can dedifferentiate proliferating primary human luminal cells to multipotent stem cells in an interleukin-8 (IL-8)-dependent manner. This was confirmed using recombinant IL-8, while the truncated protein was not active. This IL-8-related dedifferentiation of luminal cells was mediated through the STAT3-dependent downregulation of p16INK4A and the microRNA miR-141. Importantly, these in vitro-generated mammary stem cells exhibited high molecular and cellular similarities to human mammary stem cells. They have also shown a long-term mammary gland-reconstituting ability and the capacity to produce milk postdelivery. Thereby, these IL-8-generated mammary stem cells could be of great value for autologous cell therapy procedures and also for biomedical research as well as drug development.


Assuntos
Mama/citologia , Mama/metabolismo , Interleucina-8/metabolismo , Células-Tronco Multipotentes/citologia , Células-Tronco Multipotentes/metabolismo , Desdiferenciação Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Senescência Celular/fisiologia , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Transição Epitelial-Mesenquimal , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Interleucina-6/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Fator de Transcrição STAT3/metabolismo , Células Estromais/citologia , Células Estromais/metabolismo
8.
PLoS One ; 15(1): e0228226, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32004334

RESUMO

OBJECTIVE: Exposure to toxic metals such as mercury has been proposed to be a risk factor for the development of breast cancer since some metals can promote genetic mutations and epigenetic changes. We sought to find what toxic metals are present in normal breast tissue and in the tumours of women who had mastectomies for invasive ductal breast carcinoma. MATERIALS AND METHODS: Formalin-fixed paraffin-embedded blocks from mastectomies for breast carcinoma were examined from 50 women aged 34-69 years. Paraffin blocks selected for elemental analysis were from breast tissue not involved by carcinoma and from the carcinoma itself. Seven micrometer-thick sections were stained with autometallography to demonstrate the presence of mercury, and subjected to laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) to confirm the presence of mercury and to detect other toxic metals. RESULTS: Autometallography-detected mercury was seen in intraductal secretions and some luminal epithelial cells of normal breast lobules in 26 (55%) of the 47 samples where lobules were present, and in 10 (23%) of carcinomas from the 44 samples where carcinoma was present. In eight samples ductal carcinoma in situ was present and one of these contained mercury. LA-ICP-MS confirmed the presence of mercury in samples that stained with autometallography, and detected lead, iron, nickel, aluminium, chromium and cadmium in some samples. CONCLUSIONS: Mercury was present in normal breast lobules in more than half of mastectomy samples that contained an invasive carcinoma, and in a smaller proportion of carcinomas and ductal carcinomas in situ. Other toxic metals that may interact synergistically with mercury could be detected in some samples. These findings do not provide direct evidence that toxic metals such as mercury play a role in the pathogenesis of breast cancer, but suggest that future molecular biological investigations on the role of toxic metals in breast cancer are warranted.


Assuntos
Mama/diagnóstico por imagem , Mama/metabolismo , Carcinoma Ductal de Mama/diagnóstico por imagem , Carcinoma Ductal de Mama/metabolismo , Poluentes Ambientais/metabolismo , Mercúrio/metabolismo , Imagem Molecular , Adulto , Idoso , Mama/citologia , Mama/patologia , Carcinoma Ductal de Mama/patologia , Feminino , Humanos , Pessoa de Meia-Idade
9.
Int J Mol Sci ; 21(3)2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-32019271

RESUMO

Brg1 (Brahma-related gene 1) is one of two mutually exclusive ATPases that can act as the catalytic subunit of mammalian SWI/SNF (mSWI/SfigureNF) chromatin remodeling enzymes that facilitate utilization of the DNA in eukaryotic cells. Brg1 is a phospho-protein, and its activity is regulated by specific kinases and phosphatases. Previously, we showed that Brg1 interacts with and is phosphorylated by casein kinase 2 (CK2) in a manner that regulates myoblast proliferation. Here, we use biochemical and cell and molecular biology approaches to demonstrate that the Brg1-CK2 interaction occurred during mitosis in embryonic mouse somites and in primary myoblasts derived from satellite cells isolated from mouse skeletal muscle tissue. The interaction of CK2 with Brg1 and the incorporation of a number of other subunits into the mSWI/SNF enzyme complex were independent of CK2 enzymatic activity. CK2-mediated hyperphosphorylation of Brg1 was observed in mitotic cells derived from multiple cell types and organisms, suggesting functional conservation across tissues and species. The mitotically hyperphosphorylated form of Brg1 was localized with soluble chromatin, demonstrating that CK2-mediated phosphorylation of Brg1 is associated with specific partitioning of Brg1 within subcellular compartments. Thus, CK2 acts as a mitotic kinase that regulates Brg1 phosphorylation and subcellular localization.


Assuntos
Mama/metabolismo , Caseína Quinase II/metabolismo , DNA Helicases/metabolismo , Células Epiteliais/metabolismo , Mitose , Mioblastos/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Mama/citologia , Montagem e Desmontagem da Cromatina , DNA Helicases/genética , Células Epiteliais/citologia , Feminino , Humanos , Camundongos , Mioblastos/citologia , Proteínas Nucleares/genética , Fosforilação , Fatores de Transcrição/genética
10.
Am J Case Rep ; 21: e919856, 2020 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-31941879

RESUMO

BACKGROUND Pseudoangiomatous stromal hyperplasia (PASH) is an uncommon benign breast lesion. CASE REPORT PASH is reported in a young female in treatment for neurological diseases with multi-drug therapy (clonazepam, valproate and risperidone). Her menstrual cycles are irregular, and she reached menarche very late. CONCLUSIONS The higher PASH prevalence in premenopausal woman (the majority of whom are actively taking oral contraceptive pills), in 24% to 47% of men with gynecomastia and during pregnancy supports a hormonal etiology; the interaction between clonazepam, valproate, risperidone and progesterone could increase the level of progesterone that could stimulate PASH growth.


Assuntos
Angiomatose/diagnóstico , Doenças Mamárias/diagnóstico , Mama/patologia , Hiperplasia/diagnóstico , Células Estromais/patologia , Adulto , Angiomatose/etiologia , Angiomatose/cirurgia , Biópsia com Agulha de Grande Calibre , Mama/citologia , Doenças Mamárias/etiologia , Doenças Mamárias/cirurgia , Interações Medicamentosas , Feminino , Humanos , Hiperplasia/etiologia , Hiperplasia/cirurgia , Mastectomia Segmentar , Progesterona/metabolismo
11.
Breast Cancer Res Treat ; 179(3): 565-575, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31720917

RESUMO

PURPOSE: Tumor microenvironment, including inflammatory cells, adipocytes and extracellular matrix constituents such as hyaluronan (HA), impacts on cancer progression. Systemic metabolism also influences tumor growth e.g. obesity and type 2 diabetes (T2D) are risk factors for breast cancer. Here, in 262 breast cancer cases, we explored the combined impacts on survival of M2-like tumor associated macrophages (TAMs), the abundance of breast fat visualized as low density in mammograms, and tumor HA, and their associations with T2D. METHODS: Mammographic densities were assessed visually from the diagnostic images and dichotomized into very low density (VLD, density ≤ 10%, "fatty breast") and mixed density (MID, density > 10%). The amounts of TAMs (CD163+ and CD68+) and tumor HA were determined by immunohistochemistry. The data of T2D was collected from the patient records. Statistical differences between the parameters were calculated with Chi square or Mann-Whitney test and survival analyses with Cox's model. RESULTS: A combination of fatty breasts (VLD), abundance of M2-like TAMs (CD163+) and tumor HA associated with poor survival, as survival was 88-89% in the absence of these factors but only 40-47% when all three factors were present (p < 0.001). Also, an association between T2D and fatty breasts was found (p < 0.01). Furthermore, tumors in fatty breasts contained more frequently high levels of M2-like TAMs than tumors in MID breasts (p = 0.01). CONCLUSIONS: Our results demonstrate a dramatic effect of the tumor microenvironment on breast cancer progression. We hypothesize that T2D as well as obesity increase the fat content of the breasts, subsequently enhancing local pro-tumoral inflammation.


Assuntos
Tecido Adiposo/fisiologia , Densidade da Mama/fisiologia , Neoplasias da Mama/patologia , Ácido Hialurônico/metabolismo , Macrófagos/imunologia , Microambiente Tumoral/fisiologia , Adipócitos/fisiologia , Tecido Adiposo/citologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Mama/citologia , Mama/patologia , Neoplasias da Mama/mortalidade , Diabetes Mellitus Tipo 2/patologia , Intervalo Livre de Doença , Feminino , Humanos , Pessoa de Meia-Idade , Obesidade/patologia , Estudos Retrospectivos , Análise de Sobrevida
12.
Cancer Res ; 80(5): 1064-1077, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31862778

RESUMO

miR-223 is an anti-inflammatory miRNA that in cancer acts either as an oncosuppressor or oncopromoter, in a context-dependent manner. In breast cancer, we demonstrated that it dampens the activation of the EGF pathway. However, little is known on the role of miR-223 during breast cancer onset and progression. miR-223 expression was decreased in breast cancer of luminal and HER2 subtypes and inversely correlated with patients' prognosis. In normal luminal mammary epithelial cells, miR-223 acted cell autonomously in the control of their growth and morphology in three-dimensional context. In the MMTV-Δ16HER2 transgenic mouse model, oncogene transformation resulted in a timely abrogation of miR-223 expression, likely due to activation of E2F1, a known repressor of miR-223 transcription. Accordingly, treatment with CDK4/6 inhibitors, which eventually results in restraining E2F1 activity, restored miR-223 expression and miR-223 ablation induced luminal breast cancer resistance to CDK4/6 inhibition, both in vitro and in vivo. Notably, miR-223 expression was lost in microdissected ductal carcinoma in situ (DCIS) from patients with luminal and HER2-positive breast cancer. Altogether, these results identify downmodulation of miR-223 as an early step in luminal breast cancer onset and suggest that it could be used to identify aggressive DCIS and predict the response to targeted therapy. SIGNIFICANCE: miR-223 may represent a predictive biomarker of response to CDK4/6 inhibitors and its loss could identify DCIS lesions that are likely to progress into invasive breast cancer.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/genética , Carcinoma Intraductal não Infiltrante/genética , Transformação Celular Neoplásica/genética , MicroRNAs/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antineoplásicos/uso terapêutico , Mama/citologia , Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Carcinoma Intraductal não Infiltrante/tratamento farmacológico , Carcinoma Intraductal não Infiltrante/mortalidade , Carcinoma Intraductal não Infiltrante/patologia , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Modelos Animais de Doenças , Progressão da Doença , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Fator de Transcrição E2F1/metabolismo , Células Epiteliais , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/patologia , Camundongos Knockout , MicroRNAs/genética , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Prognóstico , Piridinas/farmacologia , Piridinas/uso terapêutico , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo
13.
Cancer Res ; 80(2): 204-218, 2020 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-31676574

RESUMO

Dissemination is an essential early step in metastasis but its molecular basis remains incompletely understood. To define the essential targetable effectors of this process, we developed a 3D mammary epithelial culture model, in which dissemination is induced by overexpression of the transcription factor Twist1. Transcriptomic analysis and ChIP-PCR together demonstrated that protein kinase D1 (Prkd1) is a direct transcriptional target of Twist1 and is not expressed in the normal mammary epithelium. Pharmacologic and genetic inhibition of Prkd1 in the Twist1-induced dissemination model demonstrated that Prkd1 was required for cells to initiate extracellular matrix (ECM)-directed protrusions, release from the epithelium, and migrate through the ECM. Antibody-based protein profiling revealed that Prkd1 induced broad phosphorylation changes, including an inactivating phosphorylation of ß-catenin and two microtubule depolymerizing phosphorylations of Tau, potentially explaining the release of cell-cell contacts and persistent activation of Prkd1. In patients with breast cancer, TWIST1 and PRKD1 expression correlated with metastatic recurrence, particularly in basal breast cancer. Prkd1 knockdown was sufficient to block dissemination of both murine and human mammary tumor organoids. Finally, Prkd1 knockdown in vivo blocked primary tumor invasion and distant metastasis in a mouse model of basal breast cancer. Collectively, these data identify Prkd1 as a novel and targetable signaling node downstream of Twist1 that is required for epithelial invasion and dissemination. SIGNIFICANCE: Twist1 is a known regulator of metastatic cell behaviors but not directly targetable. This study provides a molecular explanation for how Twist1-induced dissemination works and demonstrates that it can be targeted. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/2/204/F1.large.jpg.


Assuntos
Neoplasias da Mama/genética , Neoplasias Mamárias Experimentais/genética , Proteínas Nucleares/metabolismo , Proteína Quinase C/genética , Proteína 1 Relacionada a Twist/metabolismo , Animais , Mama/citologia , Mama/patologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Conjuntos de Dados como Assunto , Células Epiteliais/citologia , Células Epiteliais/patologia , Transição Epitelial-Mesenquimal/genética , Epitélio/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Mamárias Experimentais/patologia , Camundongos , Microtúbulos/metabolismo , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Cultura Primária de Células , RNA-Seq , Transdução de Sinais/genética
14.
Rev Bras Ginecol Obstet ; 41(12): 703-709, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31856289

RESUMO

OBJECTIVE: To investigate the action of testosterone (T), isolated or associated with estradiol benzoate (EB), on the proliferation markers and apoptosis of breasts of ovariectomized rats. METHODS: A total of 48 castrated female Wistar rats were divided into 6 groups, and each of them were submitted to one of the following treatments for 5 weeks: 1) control; 2) EB 50 mcg/day + T 50 mcg/day; 3) T 50mcg/day; 4) EB 50 mcg + T 300 mcg/day; 5) T 300 mcg/day; and 6) EB 50 mcg/day. After the treatment, the mammary tissue was submitted to a histological analysis and immunoexpression evaluation of proliferation markers (proliferating cell nuclear antigen, PCNA) and apoptosis (caspase-3). RESULTS: There was a statistically significant difference among the groups regarding microcalcifications and secretory activity, with higher prevalence in the groups treated with EB. There was no difference among the groups regarding atrophy, but a higher prevalence of atrophy was found in the groups that received T versus those that received EB + T. There was a difference among the groups regarding the PCNA (p = 0.028), with higher expression in the group submitted to EB + T 300 mcg/day. Regarding caspase-3, there was no difference among the groups; however, in the group submitted to EB + T 300 mcg/day, the expression was higher than in the isolated T group. CONCLUSION: Isolated T did not have a proliferative effect on the mammary tissue, contrary to EB. Testosterone in combination with EB may or may not decrease the proliferation, depending on the dose of T.


Assuntos
Apoptose/efeitos dos fármacos , Mama/citologia , Proliferação de Células/efeitos dos fármacos , Testosterona/farmacologia , Animais , Biomarcadores/análise , Mama/patologia , Calcinose/patologia , Caspase 3/análise , Estradiol/análogos & derivados , Estradiol/farmacologia , Feminino , Ovariectomia , Antígeno Nuclear de Célula em Proliferação/análise , Ratos Wistar
15.
Biofabrication ; 12(1): 015023, 2019 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-31665714

RESUMO

Breast tissue engineering is a promising alternative to standard treatments for breast defects. Although there is a consensus that the mechanical property of the scaffold should best match the reconstructed tissue, the simulation of the soft and elastic tactility of native breast tissues using conventional materials and architecture design requires further study. Previous research has shown that the crystal microstructure-like design can drastically alter the mechanical properties of the constructed scaffolds. In this study, we designed and additive manufactured four kinds of breast scaffolds using polyurethane and termed their architectures as N5S4, N9S8, N7S6 and N4S6. The basic unit cell of each scaffold was similar to a lattice structure from the isometric crystal system. The scaffolds possessed identical porosity but different mechanical properties in which the compressive modulus of the softest scaffolds (N5S4) were similar to that of native breast tissue. When applied in the construction of tissue-engineered breast combining with delayed fat injection technique in nude rat models, the soft scaffolds(N5S4) performed better compared to its stiff counterpart (N4S6), as higher adipose survival, vascularization and milder fibrosis could be observed in N5S4 scaffolds . Lastly, using finite element analysis, we further investigated the influence of the unit cell architectures on the mechanical properties of the scaffolds and simulated the deformation as well as stress distribution patterns of the implanted scaffolds in detail. Thus, a crystal lattice-like architecture design was introduced to tune the mechanical properties of the scaffolds and match the requirements for tissue engineering applications.


Assuntos
Mama/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Bioimpressão , Mama/citologia , Feminino , Porosidade , Impressão Tridimensional , Ratos , Ratos Nus , Engenharia Tecidual/instrumentação
16.
J Vis Exp ; (151)2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31609342

RESUMO

In modern pathology, optical microscopy plays an important role in disease diagnosis by revealing microscopic structures of clinical specimens. However, the fundamental physical diffraction limit prevents interrogation of nanoscale anatomy and subtle pathological changes when using conventional optical imaging approaches. Here, we describe a simple and inexpensive protocol, called expansion pathology (ExPath), for nanoscale optical imaging of common types of clinical primary tissue specimens, including both fixed-frozen or formalin-fixed paraffin embedded (FFPE) tissue sections. This method circumvents the optical diffraction limit by chemically transforming the tissue samples into tissue-hydrogel hybrid and physically expanding them isotropically across multiple scales in pure water. Due to expansion, previously unresolvable molecules are separated and thus can be observed using a conventional optical microscope.


Assuntos
Imageamento Tridimensional , Nanopartículas/química , Fixação de Tecidos , Mama/citologia , Feminino , Formaldeído/química , Humanos , Rim/citologia , Inclusão em Parafina
17.
J Physiol Pharmacol ; 70(4)2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31642813

RESUMO

The tissue renin-angiotensin system (RAS) plays an important role in the development and progression of many diseases. It has been confirmed that angiotensin II (ANG II) participates in the proliferation and angiogenesis of breast cancer. Moreover, some RAS dysregulations in cancer have been observed. Recent studies on the role of two opposite axes of angiotensinogen metabolism - ACE (angiotensin-converting enzyme)/ANGII/AT1R (angiotensin receptor type 1) and ACE-2/ANG 1-7/MAS (mitochondrial assembly) - indicate their importance in tumor growth and invasion, but studies describing the metabolic pathways in breast cancer and the role of newer angiotensins, such as ANG 1-12, remain lacking. In this study, the metabolism of angiotensinogen fragments in three breast cancer lines, namely, MDA-MB-231, MCF-7, and T-47D, compared with normal breast tissue cells (PCS-600) was estimated. Incubation of the cancer cells with angiotensinogen resulted in the prevalent formation of ANG 1-7. A difference in the ability to form ANG II was observed between cell lines. In normal breast cells, the strong predominance of the ACE-2/ANG 1-7/MAS pathway was detected. In cancer cells, differences in angiotensinogen metabolism depending on cancer line were observed; the prevalence of the ACE/ANG II/AT1R pathway was shown. Expressions of the RAS component were dysregulated in cancer cells and differed between cell lines. In conclusion, the ability of breast cancer cells to produce numerous angiotensin peptide metabolites was demonstrated. The metabolism of angiotensinogen differed between various types of breast cancer cells. The obtained results indicate the greater importance of the classical pathway - ACE/ANG II/AT1R - in breast cancer cells. The production of ANG 1-12 seems to be marginal in breast tissue, but a tendency for the higher formation of this peptide in cancer cells was observed. The production of ANG 1-7 was significantly lower in cancer cells, whereas the expression of MAS receptor was higher than that in the control. This finding suggests that substances with MAS receptor agonist activity could be useful in the treatment of breast cancer, but this requires further investigations.


Assuntos
Angiotensina I/metabolismo , Angiotensinogênio/metabolismo , Neoplasias da Mama/metabolismo , Fragmentos de Peptídeos/metabolismo , Mama/citologia , Mama/metabolismo , Linhagem Celular , Feminino , Humanos , Proteínas Proto-Oncogênicas/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo
18.
Cell Mol Biol Lett ; 24: 57, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31660059

RESUMO

Background: Glucose-regulated protein 78 (GRP78) is a member of the HSP70 protein family and a key endoplasmic reticulum chaperone. It has been revealed to play important roles both in the maturation, folding and transport of proteins and in cellproliferation. However, its involvement in milk biosynthesis or the proliferation of bovine primary mammary epithelial cells (BMECs) has yet to be established. Methods: The expressions of GRP78 in BMECs stimulated with methionine, leucine, estrogen and prolactin were determined using western blotting and immunofluorescence assays. To explore the function of GRP78 in BMECs, the protein was overexpressed or knocked down, respectively using an overexpression vector or an siRNA mixture transfected into cells cultured in vitro. Flow cytometry was used to analyze cell proliferation and cell activity. The contents of lactose and triglyceride (TG) secreted from the treated BMECs were measured using lactose and TG assay kits, respectively. Western blotting analysis was used to measure the ß-casein content and the protein levels of the signaling molecules known to be involved in milk biosynthesis and cell proliferation. Results: GRP78overexpression significantly stimulated milk protein and milk fat synthesis, enhanced cell proliferation, positively regulated the phosphorylation of mammalian target of rapamycin (mTOR), and increased the amount of protein of cyclinD1andsterol regulatory element-binding protein 1c (SREBP-1c). GRP78 knockdown after siRNA transfection had the opposite effects. We further found that GRP78 was located in the cytoplasm of BMECs, and that stimulating methionine, leucine, estrogen and prolactin expression led to a significant increase in the protein expression of GRP78 in BMECs. Conclusions: These data reveal that GRP78 is an important regulator of milk biosynthesis and the proliferation of BMECs through the mTOR signaling pathway.


Assuntos
Bovinos/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Leite/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Animais , Mama/citologia , Mama/metabolismo , Proliferação de Células , Células Cultivadas , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino
19.
Nano Lett ; 19(10): 7526-7533, 2019 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-31487192

RESUMO

Amplitude, duration, and frequency of activation of the extracellular-signal-regulated kinase (ERK) pathway code distinct information to instruct cells to migrate, proliferate, or differentiate. Synchronized frequency control of ERK activation would provide a powerful approach to regulate cell behaviors. Here we demonstrated modulation of ERK activities using alternative current (AC) electric fields (EFs) applied through high-k dielectric passivated microelectrodes. Both the amplitude and frequency of ERK activation can be precisely synchronized and modulated. ERK activation in our system is independent of Faradaic currents and electroporation, thus excluding mechanisms of changes in pH, reactive oxygen species, and other electrochemical reaction. Further experiments pinpointed a mechanism of phosphorylation site of epidermal growth factor (EGF) receptor to activate the EGFR-ERK pathway, and independent of EGF. AC EFs thus provide a powerful platform for practical and precise control of EGFR-ERK pathway.


Assuntos
Mama/citologia , Estimulação Elétrica/instrumentação , Células Epiteliais/enzimologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Mama/enzimologia , Linhagem Celular , Ativação Enzimática , Células Epiteliais/citologia , Desenho de Equipamento , Feminino , Humanos , Microeletrodos
20.
Nat Commun ; 10(1): 3840, 2019 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477698

RESUMO

Resistant tumours are thought to arise from the action of Darwinian selection on genetically heterogenous cancer cell populations. However, simple clonal selection is inadequate to describe the late relapses often characterising luminal breast cancers treated with endocrine therapy (ET), suggesting a more complex interplay between genetic and non-genetic factors. Here, we dissect the contributions of clonal genetic diversity and transcriptional plasticity during the early and late phases of ET at single-cell resolution. Using single-cell RNA-sequencing and imaging we disentangle the transcriptional variability of plastic cells and define a rare subpopulation of pre-adapted (PA) cells which undergoes further transcriptomic reprogramming and copy number changes to acquire full resistance. We find evidence for sub-clonal expression of a PA signature in primary tumours and for dominant expression in clustered circulating tumour cells. We propose a multi-step model for ET resistance development and advocate the use of stage-specific biomarkers.


Assuntos
Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Antineoplásicos Hormonais/uso terapêutico , Mama/citologia , Mama/patologia , Neoplasias da Mama/sangue , Neoplasias da Mama/genética , Plasticidade Celular/efeitos dos fármacos , Plasticidade Celular/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Microscopia Intravital , Células MCF-7 , Aprendizado de Máquina , Mutação , Células Neoplásicas Circulantes/efeitos dos fármacos , RNA-Seq , Análise de Célula Única , Esferoides Celulares
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