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1.
Methods Mol Biol ; 2619: 273-292, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36662477

RESUMO

MicroRNAs are small noncoding RNAs that regulate gene expression at the posttranscriptional level. Proteoglycans are glycoproteins characterized by covalent attachment of a glycosaminoglycan chain, which have been identified as regulatory targets of microRNAs in a physiological and pathophysiological context. We present a strategy and detailed methods for the functional analysis of microRNA regulation of proteoglycans using human cancer cells as an application example. The experimental setup includes in silico microRNA target prediction, transfection of cancer cells with microRNA precursors, validation of target regulation by qPCR, flow cytometry and luciferase reporter assays, and an example for functional analysis and phenotype confirmation by complementation analysis.


Assuntos
MicroRNAs , Neoplasias , Humanos , MicroRNAs/metabolismo , Proteoglicanas/genética , Proteoglicanas/metabolismo , Transfecção , Luciferases/metabolismo
2.
Arch Med Res ; 53(8): 826-839, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36411172

RESUMO

BACKGROUND AND AIM: Radiation resistance represents a major challenge in the treatment of breast cancer. As heparan sulfate (HS) chains are known to contribute to tumorigenesis, we aimed to investigate the interplay between HS degradation and radiation response in triple-negative breast cancer (TNBC) cells. METHODS: HS chains were degraded in vitro as TNBC cells MDA-MB-231 and HCC1806 were treated with heparinase I and III. Subsequently, radioresistance was determined via colony formation assay after doses of 2, 4 and 6 Gy. Cell cycle profile, stem cell characteristics, expression of HS, activation of beta integrins, and apoptosis were determined by flow cytometry. Additionally, cell motility was analyzed via wound-healing assays, and expression and activation of FAK, CDK-6, Src, and Erk1/2 were quantified by western blot pre- and post-irradiation. Finally, the expression of cytokines was analyzed using a cytokine array. RESULTS: Radiation promoted cell cycle changes, while heparinase treatment induced apoptosis in both cell lines. Colony formation assays showed significantly increased radio-resistance for both cell lines after degradation of HS. Cell migration was similarly upregulated after degradation of HS compared to controls. This effect was even more prominent after irradiation. Interestingly, FAK, a marker of radioresistance, was significantly activated in the heparinase-treated group. Additionally, we found Src to be dysregulated in MDA-MB-231 cells. Finally, we observed differential secretion of GRO, CXCL1, IGFBP1, IL8, Angiogenin, and Osteoprotegerin after HS degradation and radiotherapy. CONCLUSION: Our results suggest an influence of HS chains on the development of radioresistance in TNBC.


Assuntos
Neoplasias de Mama Triplo Negativas , Humanos , Neoplasias de Mama Triplo Negativas/radioterapia , Neoplasias de Mama Triplo Negativas/metabolismo , Heparitina Sulfato/metabolismo , Apoptose , Movimento Celular , Linhagem Celular Tumoral
3.
Int J Mol Sci ; 23(10)2022 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-35628603

RESUMO

Syndecans are transmembrane heparan sulfate proteoglycans that integrate signaling at the cell surface. By interacting with cytokines, signaling receptors, proteases, and extracellular matrix proteins, syndecans regulate cell proliferation, metastasis, angiogenesis, and inflammation. We analyzed public gene expression datasets to evaluate the dysregulation and potential prognostic impact of Syndecan-3 in ovarian cancer. Moreover, we performed functional in vitro analysis in syndecan-3-siRNA-treated SKOV3 and CAOV3 ovarian cancer cells. In silico analysis of public gene array datasets revealed that syndecan-3 mRNA expression was significantly increased 5.8-fold in ovarian cancer tissues (n = 744) and 3.4-fold in metastases (n = 44) compared with control tissue (n = 46), as independently confirmed in an RNAseq dataset on ovarian serous cystadenocarcinoma tissue (n = 374, controls: n = 133, 3.5-fold increase tumor vs. normal). Syndecan-3 siRNA knockdown impaired 3D spheroid growth and colony formation as stemness-related readouts in SKOV3 and CAOV3 cells. In SKOV3, but not in CAOV3 cells, syndecan-3 depletion reduced cell viability both under basal conditions and under chemotherapy with cisplatin, or cisplatin and paclitaxel. While analysis of the SIOVDB database did not reveal differences in Syndecan-3 expression between patients, sensitive, resistant or refractory to chemotherapy, KM Plotter analysis of 1435 ovarian cancer patients revealed that high syndecan-3 expression was associated with reduced survival in patients treated with taxol and platin. At the molecular level, a reduction in Stat3 activation and changes in the expression of Wnt and notch signaling constituents were observed. Our study suggests that up-regulation of syndecan-3 promotes the pathogenesis of ovarian cancer by modulating stemness-associated pathways.


Assuntos
Neoplasias Ovarianas , Sindecana-3 , Carcinoma Epitelial do Ovário/tratamento farmacológico , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/metabolismo , Linhagem Celular Tumoral , Cisplatino/farmacologia , Feminino , Humanos , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Sindecana-3/genética , Sindecana-3/metabolismo
4.
IUBMB Life ; 74(10): 955-968, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35587107

RESUMO

Ductal carcinoma in situ (DCIS) is a form of breast cancer that is restricted to the lactiferous ducts and has not yet invaded the surrounding breast tissue. Dysregulation of the transmembrane heparan sulphate proteoglycan Syndecan-1 (Sdc-1) plays a role in tumour progression of invasive breast cancer (IBC). In DCIS, Sdc-1, c-Met and E-cadherin are part of a proangiogenic expression signature. In this study, we employed a siRNA knockdown approach in the DCIS model cell line MCF10A DCIS.com to investigate a potential connection between Sdc-1 and epithelial mesenchymal transition (EMT), proteolysis and the Rho kinase pathway. Analysis of gene expression data of the TNMplot.com database revealed that Sdc-1 expression was higher in primary breast tumours compared to metastases. The impact of Sdc-1-depletion on the cellular phenotype was investigated in a Matrigel-based three-dimensional cell culture model. Sdc-1 depletion resulted in the formation of larger spheroids and the formation of invasive protrusions. Application of matrix metalloproteinase (MMP) and Rho kinase inhibitors could block the Sdc-1-induced phenotype. qPCR analysis of Sdc-1-depleted cells in two-dimensional culture revealed upregulated expression of the EMT-markers CDH1, FN-1, CLDN1, the proteolysis markers MMP3, and MMP9, and HPSE, while MMP2, VIM and ROCK-2 were downregulated. Immunocytochemistry confirmed upregulation of MMP9 and fibronectin, the latter being particular prominent after ROCK inhibition. STRING analysis confirmed an interaction of the investigated gene products at the protein level. Our results suggest that diminished Sdc-1 expression plays a role in DCIS progression to IBC through deregulation of proteolytic factors and a partial EMT.


Assuntos
Carcinoma Ductal de Mama , Carcinoma Intraductal não Infiltrante , Sindecana-1 , Caderinas/genética , Caderinas/metabolismo , Carcinoma Ductal de Mama/patologia , Carcinoma Intraductal não Infiltrante/genética , Carcinoma Intraductal não Infiltrante/metabolismo , Carcinoma Intraductal não Infiltrante/patologia , Fibronectinas , Humanos , Metaloproteinase 2 da Matriz , Metaloproteinase 3 da Matriz , Metaloproteinase 9 da Matriz/metabolismo , RNA Interferente Pequeno , Sindecana-1/genética , Quinases Associadas a rho/genética
5.
J Cell Biochem ; 122(10): 1491-1505, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34180077

RESUMO

Syndecan-4, a predicted target of the microRNA miR-140-3p, plays an important role in multiple steps of tumor progression and is the second most abundant heparan sulfate proteoglycan produced by breast carcinoma cell lines. To investigate the potential functional relationship of miR-140-3p and syndecan-4, MDA-MB-231, SKBR3, and MCF-7 breast cancer (BC) cells were transiently transfected with pre-miR-140-3p, syndecan-4 small interfering RNAJ, or control reagents, respectively. Altered cell behavior was monitored by adhesion, migration, and invasion chamber assays. Moreover, the prognostic value of syndecan-4 was assessed by Kaplan-Maier Plotter analysis of gene expression data from tumor samples of 4929 patients. High expression of syndecan-4 was associated with better relapse-free survival in the whole collective of BC patients, but correlated with a worse survival in the subgroup of estrogen receptor negative and estrogen/progesterone-receptor negative patients. miR-140-3p expression was associated with improved survival irrespective of hormone receptor status. miR-140-3p overexpression induced posttranscriptional downregulation of syndecan-4, as demonstrated by quantitative real-time PCR (qPCR), flow cytometry, and luciferase assays, resulting in decreased BC cell migration and matrigel invasiveness. Furthermore, miR-140-3p overexpression and syndecan-4 silencing increased the adhesion of BC to fibronectin and laminin. qPCR analysis demonstrated that syndecan-4 silencing leads to altered gene expression of adhesion-related molecules, such as fibronectin and focal adhesion kinase, as well as in the gene expression of the proinvasive factors matrix metalloproteinase 2 and heparanase (also known as HPSE). We conclude that syndecan-4 is a novel target of miR-140-3p that regulates BC cell invasiveness and cell-matrix interactions in the tumor microenvironment.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Matriz Extracelular/patologia , MicroRNAs/genética , Sindecana-4/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Adesão Celular , Movimento Celular , Proliferação de Células , Matriz Extracelular/metabolismo , Feminino , Humanos , Invasividade Neoplásica , Prognóstico , Taxa de Sobrevida , Sindecana-4/genética , Células Tumorais Cultivadas
6.
Int J Mol Sci ; 22(11)2021 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-34070901

RESUMO

Glycosaminoglycans (GAGs) and proteoglycans (PGs) are major components of the glycocalyx. The secreted GAG and CD44 ligand hyaluronic acid (HA), and the cell surface PG syndecan-1 (Sdc-1) modulate the expression and activity of cytokines, chemokines, growth factors, and adhesion molecules, acting as critical regulators of tumor cell behavior. Here, we studied the effect of Sdc-1 siRNA depletion and HA treatment on hallmark processes of cancer in breast cancer cell lines of different levels of aggressiveness. We analyzed HA synthesis, and parameters relevant to tumor progression, including the stem cell phenotype, Wnt signaling constituents, cell cycle progression and apoptosis, and angiogenic markers in luminal MCF-7 and triple-negative MDA-MB-231 cells. Sdc-1 knockdown enhanced HAS-2 synthesis and HA binding in MCF-7, but not in MDA-MB-231 cells. Sdc-1-depleted MDA-MB-231 cells showed a reduced CD24-/CD44+ population. Furthermore, Sdc-1 depletion was associated with survival signals in both cell lines, affecting cell cycle progression and apoptosis evasion. These changes were linked to the altered expression of KLF4, MSI2, and miR-10b and differential changes in Erk, Akt, and PTEN signaling. We conclude that Sdc-1 knockdown differentially affects HA metabolism in luminal and triple-negative breast cancer model cell lines and impacts the stem phenotype, cell survival, and angiogenic factors.


Assuntos
Regulação Neoplásica da Expressão Gênica , Glicocálix/metabolismo , Ácido Hialurônico/metabolismo , Sindecana-1/genética , Neoplasias de Mama Triplo Negativas/genética , Via de Sinalização Wnt/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Antígeno CD24/genética , Antígeno CD24/metabolismo , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Bases de Dados Factuais , Feminino , Glicocálix/química , Glicocálix/efeitos dos fármacos , Humanos , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/metabolismo , Hialuronan Sintases/genética , Hialuronan Sintases/metabolismo , Ácido Hialurônico/farmacologia , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Células MCF-7 , MicroRNAs/genética , MicroRNAs/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Ligação Proteica , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Análise de Sobrevida , Sindecana-1/antagonistas & inibidores , Sindecana-1/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/mortalidade , Neoplasias de Mama Triplo Negativas/patologia
7.
J Cell Biochem ; 122(5): 577-597, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33417295

RESUMO

Breast cancer continues to be a serious public health problem. The role of the hedgehog pathway in normal development of the mammary gland as well as in carcinogenesis and progression of breast cancer is the subject of intense investigation, revealing functional interactions with cell surface heparan sulfate. Nevertheless, its influence on breast cancer prognosis, and its relation to specific sulfation motifs in heparan sulfate have only been poorly studied in large patient cohorts. Using the public database KMplotter that includes gene expression and survival data of 3951 patients, we found that the higher expression of SHH, HHAT, PTCH1, GLI1, GLI2, and GLI3 positively influences breast cancer prognosis. Stratifying patients according to the expression of hormone receptors, histological grade, lymph node metastasis, and systemic therapy, we observed that GLI1, GLI2, and GLI3 expression, as well as co-expression of SHH and ELP1 were associated with worse relapse-free survival in patients with HER2-positive tumors. Moreover, GLI1 expression in progesterone receptor-negative tumors and GLI3 expression in grade 3 tumors correlated with poor prognosis. SHH, in a panel of cell lines representing different breast cancer subtypes, and HHAT, PTCH1, GLI1, GLI2, and GLI3 were mostly expressed in cell lines classified as HER2-positive and basal-like. Expression of SHH, HHAT, GLI2, and GLI3 was differentially affected by overexpression of the heparan sulfate sulfotransferases HS2ST1 and HS3ST2 in vitro. Although high HS2ST1 expression was associated with poor prognosis in KMplotter analysis, high levels of HS3ST2 were associated with a good prognosis, except for ER-positive breast cancer. We suggest the GLI transcription factors as possible markers for the diagnosis, treatment, and prognosis of breast cancer especially in HER2-positive tumors, but also in progesterone receptor-negative and grade-3 tumors. The pathway interaction and prognostic impact of specific heparan sulfate sulfotransferases provide novel perspectives regarding a therapeutical targeting of the hedgehog pathway in breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proteínas Hedgehog/metabolismo , Feminino , Heparitina Sulfato , Humanos , Proteínas do Tecido Nervoso/metabolismo , Receptor Patched-1/metabolismo , Prognóstico , Fatores de Elongação da Transcrição/metabolismo , Proteína GLI1 em Dedos de Zinco/metabolismo , Proteína Gli2 com Dedos de Zinco/metabolismo , Proteína Gli3 com Dedos de Zinco/metabolismo
8.
Sci Rep ; 10(1): 2262, 2020 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-32042016

RESUMO

In several carcinomas, the SET Domain Containing 3, Actin Histidine Methyltransferase (SETD3) is associated with oncogenesis. However, there is little knowledge about the role of SETD3 in the progression and prognosis of breast cancer. In this study, we first analyzed the prognostic value of SETD3 in breast cancer patients using the database of the public Kaplan-Meier plotter. Moreover, in vitro assays were performed to assess the role of SETD3 in the viability and capacity of invasion of human breast cancer cell lines. We observed that the high expression of SETD3 was associated with better relapse-free survival (RFS) of the whole collective of 3,951 patients, of Estrogen Receptor-positive, and of Luminal A-type breast cancer patients. However, in patients lacking expression of estrogen-, progesterone- and HER2-receptor, and those affected by a p53-mutation, SETD3 was associated with poor RFS. In vitro analysis showed that SETD3 siRNA depletion affects the viability of triple-negative cells as well as the cytoskeletal function and capacity of invasion of highly invasive MDA-MB-231 cells. Interestingly, SETD3 regulates the expression of other genes associated with cancer such as ß-actin, FOXM1, FBXW7, Fascin, eNOS, and MMP-2. Our study suggests that SETD3 expression can act as a subtype-specific biomarker for breast cancer progression and prognosis.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Histona Metiltransferases/metabolismo , Recidiva Local de Neoplasia/diagnóstico , Biomarcadores Tumorais/genética , Mama/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Metilação de DNA , Progressão da Doença , Intervalo Livre de Doença , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Histona Metiltransferases/genética , Humanos , Estimativa de Kaplan-Meier , Mutação , Invasividade Neoplásica/genética , Recidiva Local de Neoplasia/epidemiologia , Prognóstico , RNA Interferente Pequeno/metabolismo , Receptor ErbB-2/metabolismo , Receptores de Estrogênio , Receptores de Progesterona/metabolismo , Proteína Supressora de Tumor p53/genética
9.
J Cancer ; 10(21): 5191-5211, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31602271

RESUMO

Inflammation is a well-known driver of carcinogenesis and cancer progression, often attributed to the tumor microenvironment. However, tumor cells themselves are capable of secreting a variety of inflammatory molecules, leading to the activation of specific signaling pathways that promote tumor progression. The NF-κB signaling pathway is one of the most important connections between inflammation and tumorigenesis. NF-κB is a superfamily of transcription factors that plays an important role in several types of hematological and solid tumors, including breast cancer. However, the role of the NF-κB pathway in the survival of breast cancer patients is poorly studied. In this study, we analyzed and related the expression of both canonical and alternative NF-κB pathways and selected target genes with the relapse-free and overall survival of breast cancer patients. We used the public database Kaplan-Meier plotter (KMplot) which includes gene expression data and survival information of 3951 breast cancer patients. We found that the expression of IKKα was associated with poor relapse-free survival in patients with ER-positive tumors. Moreover, the expression of IL-8 and MMP-1 was associated with poor relapse-free and overall survival. In contrast, expression of IKKß, p50, and p65 from the canonical pathway, and NIK and RELB from the alternative pathway correlated with better relapse-free survival also when the patients were classified by their hormonal and nodal status. Our study suggests that the expression of genes of the canonical and alternative NF-κB pathways is ultimately critical for tumor persistence. Understanding the communication between both pathways would help to find better therapeutic and prophylactic targets to prevent breast cancer progression and relapse.

10.
Oncotarget ; 9(42): 26679-26700, 2018 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-29928478

RESUMO

Breast cancer is a complex disease exhibiting extensive inter- and intra-tumor heterogeneity. Inflammation is a well-known driver of cancer progression, often attributed to immune cells infiltrating the tumor stroma. However, tumor cells themselves are capable to secrete a variety of inflammatory molecules, of which we understand very little about their role in intra-clonal communication. We recently reported the capacity of triple negative cell lines to induce a cancer stem cell (CSC)-like phenotype and invasion properties into luminal cells, a mechanism mediated by pro-inflammatory cytokines that up-regulated the CXCL12/CXCR4/CXCR7 chemokine signaling axis. We performed transcriptional array analyses of CSCs-associated genes and cancer-inflammatory cell crosstalk genes and built regulatory networks with the data collected. We found a specific molecular signature segregating with the induced-invasive/stemness phenotype. Regulatory network analysis pointed out to an NFκB transcriptional signature, active in aggressive triple negative cells and in induced-invasive/CSC-like luminal cells. In agreement, NFκB inhibition abolished the induction of the stemness/invasive features. These data support an NFκB dependent mechanism of intra-clonal communication responsible for tumor cell plasticity leading the acquisition of cancer aggressive features. Understanding the communication between different tumor clones would help to find better therapeutic and prophylactic targets to prevent BrC progression and relapse.

11.
J Vis Exp ; (131)2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29364225

RESUMO

Embedded in the extracellular matrix (ECM), normal and neoplastic epithelial cells intimately communicate with hematopoietic and non-hematopoietic cells, thus greatly influencing normal tissue homeostasis and disease outcome. In breast cancer, tumor-associated macrophages (TAMs) play a critical role in disease progression, metastasis, and recurrence; therefore, understanding the mechanisms of monocyte chemoattraction to the tumor microenvironment and their interactions with tumor cells is important to control the disease. Here, we provide a detailed description of a three-dimensional (3D) co-culture system of human breast cancer (BrC) cells and human monocytes. BrC cells produced high basal levels of regulated on-activation, normal T-cell expressed and secreted (RANTES), monocyte chemoattractant protein-1 (MCP-1), and granulocyte-macrophage colony-stimulating factor (G-CSF), while in co-culture with monocytes, pro-inflammatory cytokines Interleukin (IL)-1 beta (IL-1ß) and IL-8 were enriched together with matrix metalloproteinases (MMP)-1, MMP-2, and MMP-10. This tumor stroma microenvironment promoted resistance to anoikis in MCF-10A 3D acini-like structures, chemoattraction of monocytes, and invasion of aggressive BrC cells. The protocols presented here provide an affordable alternative to study intra-tumor communication and are an example of the great potential that in vitro 3D cell systems provide to interrogate specific features of tumor biology related to tumor aggression.


Assuntos
Neoplasias da Mama/patologia , Técnicas de Cultura de Células/métodos , Matriz Extracelular/patologia , Monócitos/citologia , Neoplasias da Mama/metabolismo , Comunicação Celular/fisiologia , Linhagem Celular Tumoral , Matriz Extracelular/metabolismo , Feminino , Humanos , Células MCF-7 , Monócitos/metabolismo , Microambiente Tumoral
12.
Int J Oncol ; 51(5): 1482-1496, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29048610

RESUMO

Breast cancer (BrC) is a major public health problem worldwide. The intra-tumoral heterogeneity and tumor cell plasticity importantly contribute to disease progression and treatment failure. However, the dynamic interactions between different tumor clones, as well as their contribution to tumor aggressiveness are still poorly understood. In this study, we provide evidence of a lateral transmission of aggressive features between aggressive and non-aggressive tumor cells, consisting of gain of expression of cancer stem cell markers, increased expression of CXCL12 receptors CXCR4 and CXCR7 and increased invasiveness in response to CXCL12, which correlated with high levels of secretion of pro-inflammatory mediators G-CSF, GM-CSF, MCP-1, IL-8 and metalloproteinases 1 and 2 by the aggressive cells. Noteworthy, we found no evidence of a TGF-ß participation in the inducible-invasive phenotype. Altogether, our results provide evidence of communication between tumor cells with different potentials for aggressiveness, which could influence intra-tumoral population dynamics promoting the emergence of clones with novel functions. Understanding these interactions will provide better targets for diagnosis, prognosis and therapeutic strategies.


Assuntos
Neoplasias da Mama/genética , Evolução Clonal/genética , Heterogeneidade Genética , Invasividade Neoplásica/genética , Biomarcadores Tumorais/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Linhagem da Célula/genética , Movimento Celular , Proliferação de Células/genética , Quimiocina CXCL12/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica/patologia , Receptores CXCR/genética , Receptores CXCR4/genética , Transdução de Sinais , Fator de Crescimento Transformador beta/genética
14.
Front Immunol ; 8: 205, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28337194

RESUMO

Breast cancer remains the first cancer-related cause of death in women worldwide, particularly in developing countries in which most cases are diagnosed in late stages. Although most cancer studies are based in the genetic or epigenetic changes of the tumor cells, immune cells within the tumor stroma often cooperate with cancer progression. Particularly, monocytes are attracted to the tumor primary site in which they are differentiated into tumor-associated macrophages that facilitate tumor cell invasion and metastasis. In this study, we used three-dimensional cultures to form acini-like structures to analyze the inflammatory secretion profile of tumor cells individually or in co-culture with monocytes. Breast cancer cell lines and primary isolates from eight Mexican patients with breast cancer were used. We found high levels of RANTES/CCL5, MCP-1/CCL2, and G-CSF in the breast cancer individual cultures, supporting an important recruitment capacity of monocytes, but also of neutrophils. The co-cultures of the tumor cells and monocytes were significantly enriched with the potent pro-inflammatory cytokines interleukin (IL)-1ß and IL-8, known to support malignant progression. We also found that the interaction of tumor cells with monocytes promoted high levels of matrix metalloproteinases (MMP)-1, MMP-2, and MMP-10. Our study supports that a key event for malignant progression is the recruitment of different immune cell populations, which help to sustain and enhance a chronic inflammatory microenvironment that highly favors tumor malignancy.

15.
J Immunol Res ; 2016: 6031486, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27376091

RESUMO

Macrophages facilitate breast cancer progression. Macrophages were initially classified as M1 or M2 based on their distinct metabolic programs and then expanded to include antitumoral (M1) and protumoral (M2) activities. However, it is still uncertain what markers define the pro- and antitumoral phenotypes and what conditions lead to their formation. In this study, monocytic cell lines and primary monocytes were subjected to commonly reported protocols of M1/M2 polarization and conditions known to engage monocytes into protumoral functions. The results showed that only IDO enzyme and CD86 M1 markers were upregulated correlating with M1 polarization. TNF-α, CCR7, IL-10, arginase I, CD36, and CD163 were expressed indistinguishably from M1 or M2 polarization. Similarly, protumoral engaging resulted in upregulation of both M1 and M2 markers, with conditioned media from the most aggressive breast cancer cell line promoting the greatest changes. In spite of the mixed phenotype, M1-polarized macrophages exhibited the highest expression/secretion of inflammatory mediators, many of which have previously been associated with breast cancer aggressiveness. These data argue that although the existence of protumoral macrophages is unquestionable, their associated phenotypes and the precise conditions driving their formation are still unclear, and those conditions may need both M1 and M2 stimuli.


Assuntos
Diferenciação Celular , Macrófagos/fisiologia , Monócitos/fisiologia , Arginase/genética , Antígeno B7-2/genética , Ligante CD30/genética , Antígenos CD36/genética , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Células Cultivadas , Citocinas/genética , Feminino , Citometria de Fluxo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Interleucina-10/genética , Macrófagos/classificação , Macrófagos/imunologia , Fenótipo , Receptores CCR7/genética , Fator de Necrose Tumoral alfa/genética , Células U937 , Regulação para Cima
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