RESUMO
Microbial communities in lakes can profoundly impact biogeochemical processes through their individual activities and collective interactions. However, the complexity of these communities poses challenges, particularly for studying rare organisms such as Candidate Phyla Radiation bacteria (CPR) and enigmatic entities such as aster-like nanoparticles (ALNs). Here, a reactor was inoculated with water from Lake Fargette, France, and maintained under dark conditions at 4°C for 31 months and enriched for ALNs, diverse Planctomycetes, and CPR bacteria. We reconstructed draft genomes and predicted metabolic traits for 12 diverse Planctomycetes and 9 CPR bacteria, some of which are likely representatives of undescribed families or genera. One CPR genome representing the little-studied lineage "Candidatus Peribacter" was curated to completion (1.239 Mbp) and unexpectedly encodes the full gluconeogenesis pathway. Metatranscriptomic data indicate that some planctomycetes and CPR bacteria were active under the culture conditions, accounting for â¼30% and â¼1% of RNA reads mapping to the genome set, respectively. We also reconstructed genomes and obtained transmission electron microscope images for numerous viruses, including one with a >300-kbp genome and several predicted to infect Planctomycetes. Together, our analyses suggest that freshwater Planctomycetes are central players in a subsystem that includes ALNs, symbiotic CPR bacteria, and viruses. IMPORTANCE Laboratory incubations of natural microbial communities can aid in the study of member organisms and their networks of interaction. This is particularly important for understudied lineages for which key elements of basic biology are still emerging. Using genomics and microscopy, we found that members of the bacterial lineage Planctomycetes may be central players in a subset of a freshwater lake microbiome that includes other bacteria, archaea, viruses, and mysterious entities, called aster-like nanoparticles (ALNs), whose origin is unknown. Our results help constrain the possible origins of ALNs and provide insight into possible interactions within a complex lake ecosystem.
Assuntos
Microbiota , Planctomicetos , Humanos , Lagos/microbiologia , Metagenômica , Filogenia , Bactérias , Genômica , Água/metabolismoRESUMO
Obesity, a recognized risk factor for breast cancer in postmenopausal women, is associated with higher mortality rates regardless of menopausal status, which could in part be explained by therapeutic escape. Indeed, adipose microenvironment has been described to influence the efficiency of chemo- and hormonal therapies. Residual cancer stem cells could also have a key role in this process. To understand the mechanisms involved in the reduced efficacy of hormonal therapy on breast cancer cells in the presence of adipose secretome, human adipose stem cells (hMAD cell line) differentiated into mature adipocytes were co-cultured with mammary breast cancer cells and treated with hormonal therapies (tamoxifen, fulvestrant). Proliferation and apoptosis were measured (fluorescence test, impedancemetry, cytometry) and the gene expression profile was evaluated. Cancer stem cells were isolated from mammospheres made from MCF-7. The impact of chemo- and hormonal therapies and leptin was evaluated in this population. hMAD-differentiated mature adipocytes and their secretions were able to increase mammary cancer cell proliferation and to suppress the antiproliferative effect of tamoxifen, confirming previous data and validating our model. Apoptosis and cell cycle did not seem to be involved in this process. The evaluation of gene expression profiles suggested that STAT3 could be a possible target. On the contrary, leptin did not seem to be involved. The study of isolated cancer stem cells revealed that their proliferation was stimulated in the presence of anticancer therapies (tamoxifen, fulvestrant, doxorubicine) and leptin. Our study confirmed the role of adipocytes and their secretome, but above all, the role of communication between adipose and cancer cells in interfering with the efficiency of hormonal therapy. Among the pathophysiological mechanisms involved, leptin does not seem to interfere with the estrogenic pathway but seems to promote the proliferation of cancer stem cells.
Assuntos
Adipócitos/metabolismo , Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Fulvestranto/farmacologia , Leptina/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Tamoxifeno/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Comunicação Celular , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Microambiente TumoralRESUMO
BACKGROUND: Obesity is a well-known risk factor of breast cancer in post-menopausal women that also correlates with a diminished therapeutic response. The influence of adipocytes and their secretome, i.e. adipokines, on the efficacy of hormone therapy has yet to be elucidated. METHODS: We investigated, ex vivo, whether mature adipocytes, differentiated from adipose stem cells of normal-weight (MA20) or obese (MA30) women, and their secretions, were able to counteract the effects of tamoxifen (Tx) which is known to decrease neoplastic cell proliferation. RESULTS: In a tridimensional model and in a model of co-culture, the anti-proliferative effect of Tx on MCF-7 cancer cells was counteracted by MA30. These two models highlighted two different specific gene expression profiles for genes encoding cytokines or involved in angiogenesis based on the adipocyte microenvironment and the treatment. Thus it notably showed altered expression of genes such as TNFα that correlated with IL-6. In addition, leptin, IL-6 and TNFα, at concentrations reflecting plasma concentrations in obese patients, decreased the anti-proliferative efficacy of 4-hydroxytamoxifen (a major active metabolite of Tx). CONCLUSIONS: These findings bring insights on adipocytes and mammary cancer cell interactions in Tx therapy, particularly in overweight/obese people. Indeed, patient' adipokine status would give valuable information for developing individual strategies and avoid resistance to treatment.
Assuntos
Adipócitos/patologia , Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Obesidade/patologia , Tamoxifeno/farmacologia , Linhagem Celular , Feminino , Expressão Gênica , Humanos , Leptina/metabolismo , Células MCF-7 , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Breast cancer is correlated with a higher risk of metastasis in obese postmenopausal women. Adipokines, whose plasma concentrations are modulated in obese subjects and adipocytes surround mammary cells, suggesting that adipocyte secretome affect mammary tumorogenesis. We hypothesize that mature adipocyte secretions from obese women conditioned or not by breast neoplasic cells, increase changes on the angiogenesis stages. Supernatants of human mature adipocytes, differentiated from stem cells of either adipose tissue of normal weight (MA20) or obese (MA30) women or obtained from co-cultures between MA20 and MA30 and breast cancer cell line MCF-7, were collected. The impact of these supernatants was investigated on proliferation, migration, and tube formation by endothelial cells (HUVEC). MA20 and MA30 showed a preservation of their "metabolic memory" (increase of Leptin, ObR, VEGF, CYP19A1, and a decrease of Adiponectin expression in MA30 compared to MA20). Supernatants from obese-adipocytes increased HUVEC proliferation, migration, and sprouting like with supernatants obtained from co-cultures of MA/MCF-7 regardless the women's BMI. Additional analyses such as the use of neutralizing antibodies, analysis of supernatants (Milliplex®) and variations in gene expression (qRT-PCR), strongly suggest an implication of IL-6, or a synergistic action among adipokines, probably associated with that of VEGF or IL-6. As a conclusion, supernatants from co-cultures of MA30 and MCF-7 cells increase proliferation, migration, and sprouting of HUVEC cells. These results provide insights into the interaction between adipocytes and epithelial cancer cells, particularly in case of obesity. The identification of synergistic action of adipokines would therefore be a great interest in developing preventive strategies. J. Cell. Physiol. 232: 1808-1816, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Adipócitos/patologia , Neoplasias da Mama/irrigação sanguínea , Neoplasias da Mama/patologia , Neovascularização Patológica/patologia , Obesidade/patologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Tecido Adiposo/patologia , Anticorpos Neutralizantes/farmacologia , Índice de Massa Corporal , Peso Corporal/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Células MCF-7 , Receptores para Leptina/genética , Receptores para Leptina/metabolismo , Coloração e Rotulagem , Frações Subcelulares/efeitos dos fármacos , Frações Subcelulares/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Obesity is a risk factor for breast cancer in postmenopausal women. Leptin, a hormone excessively produced during obesity, is suggested to be involved in breast cancer. The aim of the study was to investigate procarcinogenic potential of leptin by evaluating influence of leptin on cell proliferation, cell cycle, apoptosis, and signaling on numerous breast cells lines, including 184B5 normal cells, MCF10A fibrocystic cells and MCF-7, MDA-MB-231, and T47D cancer cells. Expressions of leptin and Ob-R were analyzed using qRT-PCR and immunohistochemistry, proliferation using fluorimetric resazurin reduction test and xCELLigence system, apoptosis and cell cycle by flow cytometry, and effect of leptin on different signalling pathways using qRT-PCR and Western blot. Cells were exposed to increasing concentrations of leptin. All cell lines expressed mRNA and protein of leptin and Ob-R. Leptin stimulated proliferation of all cell lines except for 184B5 and MDA-MB-231 cells. Leptin inhibited apoptosis but didn't alter proportion of cells within cell cycle in MCF7 cells. Leptin induced overexpression of leptin, Ob-R, estrogen receptor, and aromatase mRNA in MCF-7 and T47D cells. Autoregulation induced by leptin, relationship with estrogen pathway, and proliferative and antiapoptic activity in breast cancer cells may explain that obesity-associated hyperleptinemia may be a breast cancer risk factor.
Assuntos
Neoplasias da Mama/sangue , Proliferação de Células/efeitos dos fármacos , Leptina/sangue , Obesidade/sangue , Apoptose/efeitos dos fármacos , Neoplasias da Mama/etiologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Doença da Mama Fibrocística/sangue , Doença da Mama Fibrocística/etiologia , Humanos , Imuno-Histoquímica , Leptina/genética , Células MCF-7 , Obesidade/complicações , Receptores para Leptina/sangue , Receptores para Leptina/genética , Transdução de Sinais/efeitos dos fármacosRESUMO
Photodynamic therapy (PDT) and vascular-disrupting agents (VDA) each have their advantages in the treatment of solid tumors, but also present drawbacks. In PDT, hypoxia at the center of the tumor limits conversion of molecular oxygen into singlet oxygen, while VDAs are deficient at affecting the rim of the tumor. A phthalocyanine-chalcone conjugate combining the VDA properties of chalcones with the PDT properties of phthalocyanines was designed to address these deficiencies. Its vascular targeting, photophysical, photochemical, photodynamic activities are reported herein.
Assuntos
Chalcona/química , Indóis/química , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Chalcona/farmacologia , Humanos , Indóis/farmacologia , Isoindóis , Estrutura Molecular , Fármacos Fotossensibilizantes/síntese química , Oxigênio Singlete/metabolismoRESUMO
Breast cancer has become the most common cancer among women in industrialized countries. Obesity is well established as a risk factor, in particular owing to the attendant secretion of the entities called adipokines; there is growing evidence for a role of cells and factors present in the mammary tumor microenvironment such as fibroblasts, preadipocytes, adipocytes and their secretions. To study how the microenvironment influences breast cancer growth, we developed a novel tridimensional adipose model epithelialized with normal human keratinocytes or with breast cancer cell lines. These mimicked a breast tumor in contact with an adipose microenvironment and allowed monitoring of the interactions between the cells. Leptin and adiponectin, two major adipokines, and their respective receptors, ObRt and AdipoR1, were expressed in the model, but not the second adiponectin receptor, AdipoR2. The differentiation of preadipocytes into adipocytes was greater when they were in contact with the breast cancer cell lines. The contact of breast cancer cell lines with the microenvironment completely modified their transcriptional programs by increasing the expression of genes involved in cell proliferation (cyclinD1, MAPK), angiogenesis (MMP9, VEGF) and hormonal pathways (ESR1, IL6). This tridimensional adipose model provides new insights into the interactions between breast cancer cells and their adipose microenvironment, and provides a tool to develop new drugs for the treatment of both cancer and obesity.
Assuntos
Tecido Adiposo/patologia , Neoplasias da Mama/patologia , Modelos Biológicos , Microambiente Tumoral , Adipocinas/metabolismo , Neoplasias da Mama/genética , Diferenciação Celular , Linhagem Celular Tumoral , Criança , Células Epiteliais/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Queratinócitos/patologia , Receptores de Adipocina/metabolismo , Transcrição GênicaRESUMO
Obesity is now considered as a risk factor for breast cancer in postmenopausal women. Adipokine levels are modulated in obesity, and may play a role in carcinogenesis. Moreover, obesity increases risk of cancer mortality. Here, we hypothesized that this increase could be due to a modification in angiogenesis, capital event in the development of metastases, and/or in effectiveness of cancer treatments. To test these assumptions, following a same experimental design and simultaneously the effects of leptin and adiponectin on angiogenesis were investigated, and the impact of hyperleptinemia on anticancer drug effectiveness was measured in physiological and obesity situations. Focusing on angiogenesis, the proliferation of endothelial cells (HUVEC), which expressed leptin and adiponectin receptors, was stimulated by leptin and inhibited by adiponectin. Both adipokines globally reduced apoptosis and caspase activity. Leptin increased migration whereas adiponectin decreased migration, and leptin enhanced the area of the tubes formed by HUVEC cells while adiponectin inhibited their formation. MCF7 and MDA-MB-231 cells treated with leptin secreted more VEGF than untreated cells, whereas adiponectin treatment inhibited VEGF secretion. Finally, MCF7 cells pre-treated with leptin were more invasive than untreated cells. This effect was not reproduced in MDA-MB-231 cells. In the MCF7 breast cancer cell line, leptin could induce cell proliferation and reduced the efficacy of all breast cancer therapies (tamoxifen, 5-fluorouracil, taxol and vinblastin). These results suggest that, in obesity situation, leptin- in contrast to adiponectin - may promote tumor invasion and angiogenesis, leading to metastases 'apparition, and reduce treatment efficacy, which could explain the increased risk of cancer mortality in cases of overweight. The evidence suggests adipokines influence breast cancer issue and could play a significant role, especially in obese patients for which hyperleptinemia, hypoadiponectinemia and increased metastatic potential are described.
Assuntos
Adiponectina/metabolismo , Antineoplásicos/farmacologia , Neoplasias da Mama , Leptina/metabolismo , Proteínas de Neoplasias/metabolismo , Neovascularização Patológica/metabolismo , Obesidade/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adiponectina/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Leptina/farmacologia , Invasividade Neoplásica , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Obesidade/patologia , Receptores para Leptina/metabolismoRESUMO
Zinc-α2-glycoprotein (ZAG) is a new adipokine whose gene expression is downregulated in obese patients. We recently reported ZAG expression in breast tumor or healthy breast tissue and detected this expression at high levels in ductal carcinoma and in normal epithelial adjacent tissue but not in normal tissue of healthy women. In the present study, we used two human breast tumor cell lines (MCF-7 and MDA-MB231) and one fibrocystic breast cell line (MCF10a) to examine whether recombinant ZAG has an effect on proliferative/apoptotic response in breast cancer cell lines. ZAG seemed to exert a proliferative effect on breast cancer cell proliferation [+11 to 27% in MCF-7 with (ZAG) = 5-20 µg/ml; +13% in MDA-MB-231 with (ZAG) = 5 µg/ml] and, on the contrary, an anti-proliferative effect in the fibrocystic breast cell line [-5 to -8% in MCF-10a with (ZAG) = 5-10 µg/ml]. ZAG was able to modulate gene and protein expression involved in the apoptotic response. However, further studies are required to fully elucidate the effects of ZAG on the proliferation of mammary cells.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteínas de Plasma Seminal/genética , Proteínas de Plasma Seminal/metabolismo , Apoptose/genética , Neoplasias da Mama/patologia , Processos de Crescimento Celular/genética , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Expressão Gênica , Humanos , Células MCF-7 , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Glicoproteína Zn-alfa-2RESUMO
Few data are available regarding the role of human skin dendritic cells (DCs) in driving T-cell responses. In this study we analyzed the relative capacity of Langerhans cells (LCs) and dermal CD14(-)CD1c(+) DCs (DDCs) to trigger naive CD4(+) T-cell proliferation and differentiation. DC subsets were purified after a 2-day migration from epidermis and dermis of the same skin sample. Migratory LCs showed far more activated phenotype than CD1c(+)DDCs and distinct expression of new molecules of the B7 family; when compared with LCs, CD1c(+)DDCs showed higher PD-L1 and lower inducible co-stimulator ligand (ICOS-L) expression. As expected, CD1c(+)DDCs showed lower allostimulatory property than LCs, a process that was partly reversed by anti-PD-L1 mAb. LCs were significantly more efficient than CD1c(+)DDCs at inducing allogeneic naive CD4(+) T cells to secrete both T helper cell 1 (Th1; IFN-gamma and tumor necrosis factor-alpha ) and Th2 (IL-4 and IL-5) cytokines. Moreover, anti-PD-L1 mAb increased the production of IFN-gamma by both LC- and CD1c(+)DDC-stimulated T cells. Globally, these results argue for a preponderant role of human LCs in inducing naive CD4(+) T-cell priming. Low expression of co-stimulatory molecules together with high expression of PD-L1 might limit the efficiency of CD1c(+)DDCs at inducing naive CD4(+) T-cell proliferation and secretion of cytokines.