RESUMO
BACKGROUND & AIMS: Despite recent progress in identifying aberrant genetic and epigenetic alterations in esophageal squamous cell carcinoma (ESCC), the mechanism of ESCC initiation remains unknown. METHODS: Using CRISPR/Cas 9-based genetic ablation, we targeted 9 genes (TP53, CDKN2A, NOTCH1, NOTCH3, KMT2D, KMT2C, FAT1, FAT4, and AJUBA) in murine esophageal organoids. Transcriptomic phenotypes of organoids and chemokine released by organoids were analyzed by single-cell RNA sequencing. Tumorigenicity and immune evasion of organoids were monitored by allograft transplantation. Human ESCC single-cell RNA sequencing data sets were analyzed to classify patients and find subsets relevant to organoid models and immune evasion. RESULTS: We established 32 genetically engineered esophageal organoids and identified key genetic determinants that drive ESCC initiation. A single-cell transcriptomic analysis uncovered that Trp53, Cdkn2a, and Notch1 (PCN) triple-knockout induces neoplastic features of ESCC by generating cell lineage heterogeneity and high cell plasticity. PCN knockout also generates an immunosuppressive niche enriched with exhausted T cells and M2 macrophages via the CCL2-CCR2 axis. Mechanistically, CDKN2A inactivation transactivates CCL2 via nuclear factor-κB. Moreover, comparative single-cell transcriptomic analyses stratified patients with ESCC and identified a specific subtype recapitulating the PCN-type ESCC signatures, including the high expression of CCL2 and CD274/PD-L1. CONCLUSIONS: Our study unveils that loss of TP53, CDKN2A, and NOTCH1 induces esophageal neoplasia and immune evasion for ESCC initiation and proposes the CCL2 blockade as a viable option for targeting PCN-type ESCC.
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Carcinoma de Células Escamosas , Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , Humanos , Animais , Camundongos , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/patologia , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Evasão da Resposta Imune/genética , Mutação , Proteínas com Domínio LIM/genéticaRESUMO
Over this century, coral reefs will run the gauntlet of climate change, as marine heatwaves (MHWs) become more intense and frequent, and ocean acidification (OA) progresses. However, we still lack a quantitative assessment of how, and to what degree, OA will moderate the responses of corals to MHWs as they intensify throughout this century. Here, we first projected future MHW intensities for tropical regions under three future greenhouse gas emissions scenario (representative concentration pathways, RCP2.6, RCP4.5 and RCP8.5) for the near-term (2021-2040), mid-century (2041-2060) and late-century (2081-2100). We then combined these MHW intensity projections with a global data set of 1,788 experiments to assess coral attribute performance and survival under the three emissions scenarios for the near-term, mid-century and late-century in the presence and absence of OA. Although warming and OA had predominately additive impacts on the coral responses, the contribution of OA in affecting most coral attributes was minor relative to the dominant role of intensifying MHWs. However, the addition of OA led to greater decreases in photosynthesis and survival under intermediate and unrestricted emissions scenario for the mid- and late-century than if intensifying MHWs were considered as the only driver. These results show that role of OA in modulating coral responses to intensifying MHWs depended on the focal coral attribute and extremity of the scenario examined. Specifically, intensifying MHWs and OA will cause increasing instances of coral bleaching and substantial declines in coral productivity, calcification and survival within the next two decades under the low and intermediate emissions scenario. These projections suggest that corals must rapidly adapt or acclimatize to projected ocean conditions to persist, which is far more likely under a low emissions scenario and with increasing efforts to manage reefs to enhance resilience.
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Antozoários , Animais , Antozoários/fisiologia , Mudança Climática , Recifes de Corais , Concentração de Íons de Hidrogênio , Oceanos e Mares , Água do MarRESUMO
The inflammatory response after traumatic brain injury (TBI) can lead to significant secondary brain injury and chronic inflammation within the central nervous system. Cell therapies, including mesenchymal stromal cells (MSC), have led to improvements in animal models of TBI and are under investigation in human trials. One potential mechanism for the therapeutic potential of MSC is their ability to augment the endogenous response of immune suppressive regulatory T cells (Treg). We have recently shown that infusion of human cord blood Treg decreased chronic microgliosis after TBI and altered the systemic immune response in a rodent model. These cells likely use both overlapping and distinct mechanisms to modulate the immune system; therefore, combining Treg and MSC as a combination therapy may confer therapeutic benefit over either monotherapy. However, investigation of Treg + MSC combination therapy in TBI is lacking. In this study, we compared the ability MSC + Treg combination therapy, as well as MSC and Treg monotherapies, to inhibit the neuroinflammatory response to TBI in vivo and in vitro. Treg + MSC combination therapy demonstrated increased potency to reduce the neuro- and peripheral inflammatory response compared to monotherapy; furthermore, the timing of infusion proved to be a significant variable in the efficacy of both MSC monotherapy and Treg + MSC combination therapy in vivo and in vitro.
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Lesões Encefálicas Traumáticas/terapia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/imunologia , Linfócitos T Reguladores/imunologia , Animais , Lesões Encefálicas Traumáticas/imunologia , Terapia Combinada/métodos , Modelos Animais de Doenças , Imunidade , Inflamação/terapia , Transplante de Células-Tronco Mesenquimais/métodos , Ratos Sprague-DawleyRESUMO
Coastal wetlands trap plastics from terrestrial and marine sources, but the stocks of plastics and their impacts on coastal wetlands are poorly known. We evaluated the stocks, fate, and biological and biogeochemical effects of plastics in coastal wetlands with plastic abundance data from 112 studies. The representative abundance of plastics that occurs in coastal wetland sediments and is ingested by marine animals reaches 156.7 and 98.3 items kg-1, respectively, 200 times higher than that (0.43 items kg-1) in the water column. Plastics are more abundant in mangrove forests and tidal marshes than in tidal flats and seagrass meadows. The variation in plastic abundance is related to climatic and geographic zones, seasons, and population density or plastic waste management. The abundance of plastics ingested by pelagic and demersal fish increases with fish length and dry weight. The dominant characteristics of plastics ingested by marine animals are correlated with those found in coastal wetland sediments. Microplastics exert negative effects on biota abundance and mangrove survival but positive effects on sediment nutrients, leaf drop, and carbon emission. We highlight that plastic pollution is widespread in coastal wetlands and actions are urged to include microplastics in ecosystem health and degradation assessment.
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Microplásticos , Poluentes Químicos da Água , Animais , Ecossistema , Monitoramento Ambiental , Sedimentos Geológicos , Plásticos , Poluentes Químicos da Água/análise , Áreas AlagadasRESUMO
Our understanding of the response of reef-building corals to changes in their physical environment is largely based on laboratory experiments, analysis of long-term field data, and model projections. Experimental data provide unique insights into how organisms respond to variation of environmental drivers. However, an assessment of how well experimental conditions cover the breadth of environmental conditions and variability where corals live successfully is missing. Here, we compiled and analyzed a globally distributed dataset of in-situ seasonal and diurnal variability of key environmental drivers (temperature, pCO2 , and O2 ) critical for the growth and livelihood of reef-building corals. Using a meta-analysis approach, we compared the variability of environmental conditions assayed in coral experimental studies to current and projected conditions in their natural habitats. We found that annual temperature profiles projected for the end of the 21st century were characterized by distributional shifts in temperatures with warmer winters and longer warm periods in the summer, not just peak temperatures. Furthermore, short-term hourly fluctuations of temperature and pCO2 may regularly expose corals to conditions beyond the projected average increases for the end of the 21st century. Coral reef sites varied in the degree of coupling between temperature, pCO2 , and dissolved O2 , which warrants site-specific, differentiated experimental approaches depending on the local hydrography and influence of biological processes on the carbonate system and O2 availability. Our analysis highlights that a large portion of the natural environmental variability at short and long timescales is underexplored in experimental designs, which may provide a path to extend our understanding on the response of corals to global climate change.
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Antozoários , Animais , Mudança Climática , Recifes de Corais , Oceanos e Mares , TemperaturaRESUMO
Characterization of the color of the plastic is often included in studies on plastic pollution. However, the comparability and relevance of this information is limited by methodology or observer subjectivity. Based on the analysis of thousands of floating plastic fragments from a global collection, here we propose a systematic semiautomatic method to analyze colors by using a reference palette of 120 Pantone colors. The most abundant colors were white and transparent/translucent (47%), yellow and brown (26%), and blue-like (9%). The white color increased in the smallest pieces (<5 mm) and far from coastal sources (>500 km). Both fragmentation and discolouration of ocean plastics may occur because of longer exposure time to sunlight in nature. In addition, yellow items peaked at around 1 cm and brown colors at around 1 mm, supporting the notion that yellowing precedes tanning in the aging process, which is paralleled by fragmentation. Apart from the effects of the weathering, our results suggest a second-order modulation of the color distributions of marine microplastics by the selective action of visual predators. The present work provides methodological tools and a wide empirical background to further the interpretation and applicability of the color information on ocean plastics.
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Plásticos , Poluentes Químicos da Água , Cor , Monitoramento Ambiental , Oceanos e Mares , Poluentes Químicos da Água/análiseRESUMO
The adipocyte-derived hormone leptin plays a critical role in the central transmission of energy balance to modulate reproductive function. However, the neurocircuitry underlying this interaction remains elusive, in part due to incomplete knowledge of first-order leptin-responsive neurons. To address this gap, we explored the contribution of predominantly inhibitory (GABAergic) neurons versus excitatory (glutamatergic) neurons in the female mouse by selective ablation of the leptin receptor in each neuronal population: Vgat-Cre;Lepr(lox/lox) and Vglut2-Cre;Lepr(lox/lox) mice, respectively. Female Vgat-Cre;Lepr(lox/lox) but not Vglut2-Cre;Lepr(lox/lox) mice were obese. Vgat-Cre;Lepr(lox/lox) mice had delayed or absent vaginal opening, persistent diestrus, and atrophic reproductive tracts with absent corpora lutea. In contrast, Vglut2-Cre;Lepr(lox/lox) females exhibited reproductive maturation and function comparable to Lepr(lox/lox) control mice. Intracerebroventricular administration of kisspeptin-10 to Vgat-Cre;Lepr(lox/lox) female mice elicited robust gonadotropin responses, suggesting normal gonadotropin-releasing hormone neuronal and gonadotrope function. However, adult ovariectomized Vgat-Cre;Lepr(lox/lox) mice displayed significantly reduced levels of Kiss1 (but not Tac2) mRNA in the arcuate nucleus, and a reduced compensatory luteinizing hormone increase compared with control animals. Estradiol replacement after ovariectomy inhibited gonadotropin release to a similar extent in both groups. These animals also exhibited a compromised positive feedback response to sex steroids, as shown by significantly lower Kiss1 mRNA levels in the AVPV, compared with Lepr(lox/lox) mice. We conclude that leptin-responsive GABAergic neurons, but not glutamatergic neurons, act as metabolic sensors to regulate fertility, at least in part through modulatory effects on kisspeptin neurons.
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Fertilidade/fisiologia , Neurônios GABAérgicos/metabolismo , Kisspeptinas/metabolismo , Leptina/metabolismo , Receptores para Leptina/metabolismo , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Núcleo Arqueado do Hipotálamo/metabolismo , Estradiol/farmacologia , Feminino , Fertilidade/efeitos dos fármacos , Neurônios GABAérgicos/efeitos dos fármacos , Kisspeptinas/genética , Kisspeptinas/farmacologia , Hormônio Luteinizante/sangue , Camundongos , Camundongos Knockout , Receptores para Leptina/genética , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Functional hypothalamic amenorrhea is a reversible form of gonadotropin-releasing hormone (GnRH) deficiency commonly triggered by stressors such as excessive exercise, nutritional deficits, or psychological distress. Women vary in their susceptibility to inhibition of the reproductive axis by such stressors, but it is unknown whether this variability reflects a genetic predisposition to hypothalamic amenorrhea. We hypothesized that mutations in genes involved in idiopathic hypogonadotropic hypogonadism, a congenital form of GnRH deficiency, are associated with hypothalamic amenorrhea. METHODS: We analyzed the coding sequence of genes associated with idiopathic hypogonadotropic hypogonadism in 55 women with hypothalamic amenorrhea and performed in vitro studies of the identified mutations. RESULTS: Six heterozygous mutations were identified in 7 of the 55 patients with hypothalamic amenorrhea: two variants in the fibroblast growth factor receptor 1 gene FGFR1 (G260E and R756H), two in the prokineticin receptor 2 gene PROKR2 (R85H and L173R), one in the GnRH receptor gene GNRHR (R262Q), and one in the Kallmann syndrome 1 sequence gene KAL1 (V371I). No mutations were found in a cohort of 422 controls with normal menstrual cycles. In vitro studies showed that FGFR1 G260E, FGFR1 R756H, and PROKR2 R85H are loss-of-function mutations, as has been previously shown for PROKR2 L173R and GNRHR R262Q. CONCLUSIONS: Rare variants in genes associated with idiopathic hypogonadotropic hypogonadism are found in women with hypothalamic amenorrhea, suggesting that these mutations may contribute to the variable susceptibility of women to the functional changes in GnRH secretion that characterize hypothalamic amenorrhea. Our observations provide evidence for the role of rare variants in common multifactorial disease. (Funded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development and others; ClinicalTrials.gov number, NCT00494169.).
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Amenorreia/genética , Hormônio Liberador de Gonadotropina/deficiência , Doenças Hipotalâmicas/genética , Mutação , Amenorreia/etiologia , Proteínas da Matriz Extracelular/genética , Feminino , Expressão Gênica , Predisposição Genética para Doença , Hormônio Liberador de Gonadotropina/genética , Humanos , Hipogonadismo/genética , Doenças Hipotalâmicas/complicações , Hormônio Luteinizante/metabolismo , Proteínas do Tecido Nervoso/genética , Precursores de Proteínas/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptores Acoplados a Proteínas G/genética , Receptores LHRH/genética , Receptores de Peptídeos/genética , Análise de Sequência de DNARESUMO
For an organ to maintain correct architecture and function, its diverse cellular components must coordinate their size and shape. Although cell-intrinsic mechanisms driving homotypic cell-cell coordination are known, it is unclear how cell shape is regulated across heterotypic cells. We find that epithelial cells maintain the shape of neighboring sense-organ glia-neuron units in adult Caenorhabditis elegans (C. elegans). Hsp co-chaperone UNC-23/BAG2 prevents epithelial cell shape from deforming, and its loss causes head epithelia to stretch aberrantly during animal movement. In the sense-organ glia, amphid sheath (AMsh), this causes progressive fibroblast growth factor receptor (FGFR)-dependent disruption of the glial apical cytoskeleton. Resultant glial cell shape alteration causes concomitant shape change in glia-associated neuron endings. Epithelial UNC-23 maintenance of glia-neuron shape is specific both spatially, within a defined anatomical zone, and temporally, in a developmentally critical period. As all molecular components uncovered are broadly conserved across central and peripheral nervous systems, we posit that epithelia may similarly regulate glia-neuron architecture cross-species.
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Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Neuroglia , Neurônios , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Forma Celular , Citoesqueleto/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/citologia , Neuroglia/metabolismo , Neurônios/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/genética , Estresse MecânicoRESUMO
Despite the promising outcomes of immune checkpoint inhibitors (ICIs), resistance to ICI presents a new challenge. Therefore, selecting patients for specific ICI applications is crucial for maximizing therapeutic efficacy. Herein, we curated 69 human esophageal squamous cell cancer (ESCC) patients' tumor microenvironment (TME) single-cell transcriptomic datasets to subtype ESCC. Integrative analyses of the cellular network and transcriptional signatures of T cells and myeloid cells define distinct ESCC subtypes characterized by T cell exhaustion, and interleukin (IL) and interferon (IFN) signaling. Furthermore, this approach classifies ESCC patients into ICI responders and non-responders, as validated by whole tumor transcriptomes and liquid biopsy-based single-cell transcriptomes of anti-PD-1 ICI responders and non-responders. Our study stratifies ESCC patients based on TME transcriptional network, providing novel insights into tumor niche remodeling and potentially predicting ICI responses in ESCC patients.
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Background: Esophageal organoids from a variety of pathologies including cancer are grown in Advanced Dulbecco's Modified Eagle Medium-Nutrient Mixture F12 (hereafter ADF). However, the currently available ADF-based formulations are suboptimal for normal human esophageal organoids, limiting the ability to compare normal esophageal organoids with those representing a given disease state. Methods: We have utilized immortalized normal human esophageal epithelial cell (keratinocyte) lines EPC1 and EPC2 and endoscopic normal esophageal biopsies to generate three-dimensional (3D) organoids. To optimize ADF-based medium, we evaluated the requirement of exogenous epidermal growth factor (EGF) and inhibition of transforming growth factor-(TGF)-ß receptor-mediated signaling, both key regulators of proliferation of human esophageal keratinocytes. We have modeled human esophageal epithelial pathology by stimulating esophageal 3D organoids with interleukin (IL)-13, an inflammatory cytokine, or UAB30, a novel pharmacological activator of retinoic acid signaling. Results: The formation of normal human esophageal 3D organoids was limited by excessive EGF and intrinsic TGFß receptor-mediated signaling. In optimized HOME0, normal human esophageal organoid formation was improved, whereas IL-13 and UAB30 induced epithelial changes reminiscent of basal cell hyperplasia, a common histopathologic feature in broad esophageal disease conditions including eosinophilic esophagitis. Conclusions: HOME0 allows modeling of the homeostatic differentiation gradient and perturbation of the human esophageal epithelium while permitting a comparison of organoids from mice and other organs grown in ADF-based media.
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The abundance of anthropogenic debris dispersed in the environment is exponentially growing, raising concerns about marine life and human exposure to microplastics. Microfibers are the most abundant microplastic type in the environment. However, recent research suggests that most microfibers dispersed in the environment are not made of synthetic polymers. In this work, we systematically tested this assumption by determining the man-made or natural origin of microfibers found in different environments, including surface waters, sediments at depths >5000 m and highly sensitive habitats like mangroves and seagrass, and treated water using stimulated Raman scattering (SRS) microscopy. Our findings show that ¾th of analyzed microfibers are of natural origin. One plastic fiber is estimated per every 50 L in surface seawater, every 5 L in desalinated drinking water, every 3 g in deep sea sediments and every 27 g in coastal sediments. Synthetic fibers were significantly larger in surface seawaters compared to organic fibers due to higher resistance to solar radiation. These results emphasize the necessity of using spectroscopical methods to assess the origin of environmental microfibers to accurately estimate the abundance of synthetic materials in the environment.
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Microplásticos , Poluentes Químicos da Água , Humanos , Poluentes Químicos da Água/análise , Plásticos , Monitoramento Ambiental/métodos , Água do Mar , Sedimentos Geológicos/químicaRESUMO
Tumors originating from the head and neck represent diverse histologies and are comprised of several cell types, including malignant cells, cancer-associated fibroblasts, endothelial cells, and immune cells. In this protocol, we describe a step-by-step approach for the dissociation of fresh human head and neck tumor specimens, followed by isolation of viable single cells using fluorescence-activated cell sorting. Our protocol facilitates the effective downstream use of techniques, including single-cell RNA sequencing and generation of three-dimensional patient-derived organoids. For complete details on the use and execution of this protocol, please refer to Puram et al. (2017)1 and Parikh et al. (2022).2.
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The microscopic species colonizing the human body, collectively referred to as the microbiome, play a crucial role in the maintenance of tissue homeostasis, immunity, and the development of disease. There is evidence to suggest associations between alterations in the microbiome and the development of head and neck squamous cell carcinomas (HNSCC). The use of two-dimensional (2D) modeling systems has made significant strides in uncovering the role of microbes in carcinogenesis; however, direct mechanistic links remain in their infancy. Patient-derived three-dimensional (3D) HNSCC organoid and organotypic models have recently been described. Compared to 2D models, 3D organoid culture systems effectively capture the genetic and epigenetic features of parent tissue in a patient-specific manner and may offer a more nuanced understanding of the role of host-microbe responses in carcinogenesis. This review provides a topical literature review assessing the current state of the field investigating the role of the microbiome in HNSCC; including in vivo and in vitro modeling methods that may be used to characterize microbiome-epithelial interactions.
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Despite the promising outcomes of immune checkpoint blockade (ICB), resistance to ICB presents a new challenge. Therefore, selecting patients for specific ICB applications is crucial for maximizing therapeutic efficacy. Herein we curated 69 human esophageal squamous cell cancer (ESCC) patients' tumor microenvironment (TME) single-cell transcriptomic datasets to subtype ESCC. Integrative analyses of the cellular network transcriptional signatures of T cells, myeloid cells, and fibroblasts define distinct ESCC subtypes characterized by T cell exhaustion, Interferon (IFN) a/b signaling, TIGIT enrichment, and specific marker genes. Furthermore, this approach classifies ESCC patients into ICB responders and non-responders, as validated by liquid biopsy single-cell transcriptomics. Our study stratifies ESCC patients based on TME transcriptional network, providing novel insights into tumor niche remodeling and predicting ICB responses in ESCC patients.
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Esophageal squamous cell carcinoma (ESCC) is prevalent worldwide, accounting for 90% of all esophageal cancer cases each year, and is the deadliest of all human squamous cell carcinomas. Despite recent progress in defining the molecular changes accompanying ESCC initiation and development, patient prognosis remains poor. The functional annotation of these molecular changes is the necessary next step and requires models that both capture the molecular features of ESCC and can be readily and inexpensively manipulated for functional annotation. Mice treated with the tobacco smoke mimetic 4-nitroquinoline 1-oxide (4NQO) predictably form ESCC and esophageal preneoplasia. Of note, 4NQO lesions also arise in the oral cavity, most commonly in the tongue, as well as the forestomach, which all share the stratified squamous epithelium. However, these mice cannot be simply manipulated for functional hypothesis testing, as generating isogenic mouse models is time- and resource-intensive. Herein, we overcome this limitation by generating single cell-derived three-dimensional (3D) organoids from mice treated with 4NQO to characterize murine ESCC or preneoplastic cells ex vivo. These organoids capture the salient features of ESCC and esophageal preneoplasia, can be cheaply and quickly leveraged to form isogenic models, and can be utilized for syngeneic transplantation experiments. We demonstrate how to generate 3D organoids from normal, preneoplastic, and SCC murine esophageal tissue and maintain and cryopreserve these organoids. The applications of these versatile organoids are broad and include the utilization of genetically engineered mice and further characterization by flow cytometry or immunohistochemistry, the generation of isogeneic organoid lines using CRISPR technologies, and drug screening or syngeneic transplantation. We believe that the widespread adoption of the techniques demonstrated in this protocol will accelerate progress in this field to combat the severe burden of ESCC.
Assuntos
Carcinoma de Células Escamosas , Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Humanos , Camundongos , Animais , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço , Organoides/metabolismo , Linhagem Celular Tumoral , Proliferação de CélulasRESUMO
BACKGROUND: Cyclooxygenase-2 (COX-2) expression in primary breast cancer predicts tumor cell dissemination to bone marrow, which is a risk factor for recurrence and distant metastasis. "Stem-like" phenotype may be important in cancer metastasis. METHODS: To investigate the role of COX-2 protein in breast cancer stem-like cells, we analyzed it by co-immunofluorescence in tumorospheres derived from the MCF7 estrogen receptor-positive breast cancer cell line. To evaluate COX-2 function we utilized a COX-2 inhibitor in a clonogenicity assay performed with tumorospheres-derived cells. RESULTS: We detected rare cells in tumorospheres (one cell per tumorosphere) with very high COX-2 expression (COX-2(high)). COX-2 transfected MCF7 cells were able to generate long-term tumorospheres culture, even though transfection efficiency was only one in a million cells. We detected expression of OCT4 in some COX-2(high) cells, supporting the hypothesis that these cells could be cancer stem-like cells. It is important that COX-2(high) cells showed less expression of Ki-67 than did neighboring cells, indicating that COX-2(high) cells may be progenitors of tumorospheres. Celecoxib inhibited the growth of tumorosphere cultures and the ability of tumorosphere-derived cells to form colonies in vitro, indicating an active role of COX-2 in these processes. However, 2 µM celecoxib failed to eradicate tumorosphere-initiating cells. Finally, we detected rare COX-2(high) cells among SUM149 inflammatory breast cancer cells growing on plastic in serum-containing medium; the SUM149 cell line produces a very high level of COX-2 protein. CONCLUSION: Our results support a role for COX-2 in stem-like breast cancer cells and suggest a mechanism behind a role for COX-2 in disseminated tumor cells, which are known to exhibit characteristic biomarkers and functional properties of stem-like cells.
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Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclo-Oxigenase 2/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Neoplasias da Mama/fisiopatologia , Celecoxib , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Inibidores de Ciclo-Oxigenase 2/farmacologia , Feminino , Humanos , Metástase Neoplásica/fisiopatologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Fator 3 de Transcrição de Octâmero/metabolismo , Pirazóis/farmacologia , Receptores de Estrogênio/metabolismo , Sulfonamidas/farmacologiaRESUMO
Anthropogenic litter density and composition data were obtained by conducting aerial surveys on 44 beaches along the Saudi Arabian Coast of the Red Sea [1]. The aerial surveys were completed with commercial drones of the DJI Phantom suite flown at a 10 m altitude. The stills have a resolution of less than 0.5 cm pixels-1, hence, litter objects of few centimetres like bottle caps are easily detectable in the drone images. We here provide a subsample of the drone images acquired. To spare the time needed to visually count the litter objects in the thousands of drone images acquired, these were automatically screened using an object detection algorithm, specifically a Faster R-CNN, able to perform a binary classification in litter and non-litter and to categorize the objects in classes. The multi-class classification, however, is a challenging problem and, hence, it was conducted only on the 15 beaches that showed the highest performance after the binary classification. The performance of the algorithm was calculated by visually screening a subsample of images and it was used to correct the output of the Faster R-CNN. The described steps allowed to obtain an estimate of the litter density in 44 beaches and the litter composition in 15 beaches. By multiplying the relative abundance of each litter class and the median weight of objects belonging to each class, we obtained an estimate of the total mass of plastic beached on 15 beaches. Possible predictors of litter density and mass are the population and marine traffic densities at the site, the exposure of the beach to the prevailing wind and the wind speed, the fetch length and the presence of vegetation where litter could get trapped. Making such raw data (i.e. litter density and composition and their predictors) available can help building the base for a robust global estimate of anthropogenic litter in coastal environments and it is particularly important if data regards an understudied region like the Arabian Peninsula. Moreover, we share a subsample of the original drone images to allow usage from stakeholders.
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Beach litter assessments rely on time inefficient and high human cost protocols, mining the attainment of global beach litter estimates. Here we show the application of an emerging technique, the use of drones for acquisition of high-resolution beach images coupled with machine learning for their automatic processing, aimed at achieving the first national-scale beach litter survey completed by only one operator. The aerial survey had a time efficiency of 570 ± 40 m2 min-1 and the machine learning reached a mean (±SE) detection sensitivity of 59 ± 3% with high resolution images. The resulting mean (±SE) litter density on Saudi Arabian shores of the Red Sea is of 0.12 ± 0.02 litter items m-2, distributed independently of the population density in the area around the sampling station. Instead, accumulation of litter depended on the exposure of the beach to the prevailing wind and litter composition differed between islands and the main shore, where recreational activities are the major source of anthropogenic debris.
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Praias , Plásticos , Monitoramento Ambiental , Humanos , Oceano Índico , Aprendizado de Máquina , Arábia Saudita , Resíduos/análiseRESUMO
Traumatic brain injury (TBI) causes both physical disruption of the blood brain barrier (BBB) and altered immune responses that can lead to significant secondary brain injury and chronic inflammation within the central nervous system (CNS). Cell therapies, including mesenchymal stromal cells (MSC), have been shown to restore BBB integrity and augment endogenous splenic regulatory T cells (Treg), a subset of CD4+ T cells that function to regulate immune responses and prevent autoimmunity. We have recently shown that infusion of human cord blood-derived Treg decreased neuroinflammation after TBI in vivo and in vitro. However, while both cells have demonstrated anti-inflammatory and regenerative potential, they likely utilize differing, although potentially overlapping, mechanisms. Furthermore, studies investigating these two cell types together, as a combination therapy, are lacking. In this study, we compared the ability of Treg+MSC combination therapy, as well as MSC and Treg monotherapies, to improve BBB permeability in vivo and suppress inflammation in vitro. While Treg+MSC combination did not significantly augment potency in vivo, our in vitro data demonstrates that combination therapy may augment therapeutic potency and immunosuppressive potential compared to Treg or MSC monotherapy.