Characterizing the melanoma brain metastasis microenvironment using CyTOF IMC and the adenosine pathway in melanoma

Introduction: Le mélanome est le type de cancer de la peau le plus fréquent et les métastases du système nerveux central en sont une complication fréquente et grave. Les cellules de mélanome interagissent avec une grande variété de types de cellules dans le microenvironnement tumoral (MET), ce qui peut entraîner des effets pro- ou antitumoral. Plusieurs voies immunosuppressives ont été récemment découvertes comme des cibles médicamenteuses prometteuses, notamment la voie de l'adénosine. L'adénosine extracellulaire s'accumule dans le MET suite à l'hydrolyse de l'ATP par les ectonucléotidases CD39 et CD73. Les principaux régulateurs de la voie de l'adénosine sont CD39, CD73, et les récepteurs A2a et A2b. Matériel et Méthodes: Pour caractériser spatialement le MET des métastases cérébrales du mélanome (MCM), nous avons quantifié l'expression de 35 marqueurs protéiques à l'aide du time of flight (CyTOF) Imaging Mass Cytometry (IMC) dans 21 MCM, et segmenté et classé plus de 130 000 cellules. Ensuite, pour évaluer les effets du ciblage du récepteur A2b et du CD73 dans la voie de l'adénosine sur le développement du mélanome, nous avons utilisé les tests de prolifération IncuCyte et MTS pour évaluer la prolifération des cellules de mélanome. Résultats: Dans notre ensemble de données, les caractéristiques immunitaires du MET étaient hétérogènes dans tous les échantillons et le type de cellule le plus courant après les cellules cancéreuses du mélanome était les macrophages dérivés de la moelle osseuse (MDMO). Les échantillons à propagation leptoméningée avaient significativement moins de neutrophiles, de MDMO de type M1, d'autres cellules T et plus de cellules cancéreuses dans leur microenvironnement. Nous avons observé que la stimulation du récepteur A2b a un effet antiprolifératif sur les cellules cancéreuses du mélanome. Conclusion: Cette recherche met en évidence le rôle du MET dans la progression du mélanome et l'importance du MET comme base pour le développement de nouvelles thérapies pour les patients atteints de cancer.Background: Melanoma is the most frequent type of skin cancer and metastasis to the central nervous system is a common and serious complication of it. Melanoma cells interact with a wide variety of cell types in the tumor microenvironment (TME) which can lead to tumor-promoting or tumor suppressive effects. Several immunosuppressive pathways have emerged as promising drug targets, including the adenosine pathway. The extracellular adenosine accumulates in the TME as the result of ATP hydrolysis by the ectonucleotidases CD39 and CD73. Key regulators of the adenosine pathway are CD39, CD73, A2a and A2b receptor. Methods: To spatially characterize the TME of melanoma brain metastases (MBM), we quantified the expression of 35 protein markers using time of flight (CyTOF) Imaging Mass Cytometry (IMC) in 21 MBMs, and segmented and classified over 130 000 cells. Then, to evaluate the effects of targeting the A2b receptor and CD73 in the adenosine pathway on the development of melanoma, we used the IncuCyte and MTS proliferation assays to assess the proliferation of melanoma cells. Results: In our dataset, the immune landscape of the TME was heterogeneous across all samples and the most common cell type after melanoma cancer cells were bone marrow derived macrophages (BMDM). Samples with leptomeningeal spread had significantly less neutrophils, M1-like BMDM, T other cells and more cancer cells in their microenvironment. We observed that stimulation of the A2b receptor has an antiproliferative effect on melanoma cancer cells. Conclusion: This research highlights the role of the TME in the progression of melanoma and the importance of the TME as grounds for development of new therapies for cancer patients

Bacterial communities of the specialty crop phyllosphere: response to biological soil amendment use, rainfall, and insect visitation

Microorganisms in the plant rhizosphere, the zone under the influence of roots, and phyllosphere, the aboveground plant habitat, exert a strong influence on plant growth, health, and protection. Tomatoes and cucumbers are important players in produce safety, and the microbial life on their surfaces may contribute to their fitness as hosts for foodborne pathogens such as Salmonella enterica and Listeria monocytogenes. External factors such as agricultural inputs and environmental conditions likely also play a major role. However, the relative contributions of the various factors at play concerning the plant surface microbiome remain obscure, although this knowledge could be applied to crop protection from plant and human pathogens. Recent advances in genomic technology have made investigations into the diversity and structure of microbial communities possible in many systems and at multiple scales. Using Illumina sequencing to profile particular regions of the 16S rRNA gene, this study investigates the influences of climate and crop management practices on the field-grown tomato and cucumber microbiome. The first research chapter (Chapter 3) involved application of 4 different soil amendments to a tomato field and profiling of harvest-time phyllosphere and rhizosphere microbial communities. Factors such as water activity, soil texture, and field location influenced microbial community structure more than soil amendment use, indicating that field conditions may exert more influence on the tomato microbiome than certain agricultural inputs. In Chapter 4, the impact of rain on tomato and cucumber-associated microbial community structures was evaluated. Shifts in bacterial community composition and structure were recorded immediately following rain events, an effect which was partially reversed after 4 days and was strongest on cucumber fruit surfaces. Chapter 5 focused on the contribution of insect visitors to the tomato microbiota, finding that insects introduced diverse bacterial taxa to the blossom and green tomato fruit microbiome. This study advances our understanding of the factors that influence the microbiomes of tomato and cucumber. Farms are complex environments, and untangling the interactions between farming practices, the environment, and microbial diversity will help us develop a comprehensive understanding of how microbial life, including foodborne pathogens, may be influenced by agricultural conditions

The acute effects of subtalar neutral on pre-activation levels of the peroneus longus, time to stabilization, and stiffness in unstable and stable ankles

The purpose of this study was to examine if a more affordable over the counter orthotic can be used by the public to assist people with unstable ankles with ankle joint stiffness, pre-activation levels of the peroneus longus (PL), and time to stabilization (TTS). Forty individuals (20 unstable ankles and 20 stable ankles) repeated three randomized tasks with and without orthotics in a pretest-posttest design. The tasks included: 1) A single leg drop landing task in a) frontal and b) sagittal planes and 2) ankle joint stiffness utilizing an ankle cradle with inversion perturbation. Results revealed no interaction effects on ankle joint stiffness, pre-activation levels of the PL, and TTS in both the sagittal and frontal planes. We observed the unstable ankle group took longer to stabilize in both the sagittal and frontal planes. We also noted that ankle joint stiffness decreased in both groups in the orthotic condition

Management of COVID-19 in the community and the role of primary care: how the pandemic has shone light on a fragmented health system

[Extract] The Australian health care system is well regarded on the global stage in terms of the balance between investment in health care and outcomes delivered, particularly in terms of universal access, quality and safety.1 However, there is considerable fragmentation and poor coordination of care and communication between hospitals and primary care, which limits further improvement.2, 3 Geographical barriers, workforce shortages and issues relating to acceptability of services limit health care access for residents of rural, regional and remote communities, Aboriginal peoples and Torres Strait Islanders, and together with an inadequate focus on prevention, limit progress towards health equity. Australian responses to the coronavirus disease 2019 (COVID-19) pandemic through both public health responses and the acute health sector have been viewed as among the best in the world. Nevertheless, challenges in the structure, organisatIon and financing of the Australian health care system have been brought into stark relief by the evolution of responses to the pandemic

Conflicting Ideologies of Mexican Immigrant English Across Levels of Schooling

This article explores how language ideologies—beliefs about immigrant students’ language use—carry conflicting images of Spanish speakers in one New Latino Diaspora town. We describe how teachers and students encounter, negotiate, and appropriate divergent ideologies about immigrant students’ language use during routine schooling practices, and we show how these ideologies convey different messages about belonging to the community and to the nation. Although the concept of language ideology often assumes stable macrolevel beliefs, our data indicate that ideologies can vary dramatically in one town. Elementary educators and students had a positive, “bilinguals-in-the-making” ideology about Spanish-speaking students, while secondary educators used more familiar deficit accounts. Despite their differences, we argue that both settings tended toward subtractive schooling, and we offer suggestions for how educators could more effectively build upon emergent bilinguals’ language skills and practices

Evaluation of a patient self-stratification methodology to identify those in need of shielding during COVID-19

The logistical challenges of rapidly and accurately identifying those patients who needed to shield during the COVID-19 pandemic were unprecedented. We report our experiences of meeting this challenge for &gt;9,000 patients with rheumatic and musculoskeletal disease at our centre, incorporating an element of guided patient self-stratification. Our results indicate that patients are able to stratify their own risk accurately using the BSR COVID-19 risk stratification guidance.</p

Data citation in the wild

Consistent attribution of research data upon reuse is necessary to reward the original data-producing investigators, reconstruct provenance, and inform data sharing policies, tool requirements, and funding decisions. Unfortunately, norms for data attribution are varied and often weak. As part of the DataONE 2010 summer internship program, three interns studied the policies, practice, and implications of current data attribution behavior in the environmental sciences. We found that few policies recommend robust data citation practices: in our preliminary evaluation, only one-third of repositories (n=26), 6% of journals (n=307), and 1 of 53 funders suggested a best practice for data citation. We manually reviewed 500 papers published between 2000 and 2010 across six journals; of the 198 papers that reused datasets, only 14% reported a unique dataset identifier in their dataset attribution, and a partially-overlapping 12% mentioned the author name and repository name. Few citations to datasets themselves were made in the article references section. In multivariate analysis, citation patterns were more correlated with repository (with citations to Genbank being most complete) than journal or datatype. Attribution patterns were found to be steady over time. Consistent with these findings, dataset reuse was difficult to track through standard retrieval resources. Searching by repository name retrieved many instances of data submission rather than data reuse, combing the citation history of data creation articles was time consuming, and searching citation databases for the few early-adopter dataset DOIs and HDLs in reference lists failed due to apparent limitations in database query capabilities and structured extraction of DOIs. We hope these descriptions of the current data attribution environment will highlight outstanding issues and motivate change in policy, tools, and practice. This research was done as open science (http://openwetware.org/wiki/DataONE:Notebook/Summer_2010): ask us about it

Detection and Identification of Salmonella enterica, Escherichia coli, and Shigella spp. via PCR-ESI-MS: Isolate Testing and Analysis of Food Samples

An assay to identify the common food-borne pathogens Salmonella, Escherichia coli, Shigella, and Listeria monocytogenes was developed in collaboration with Ibis Biosciences (a division of Abbott Molecular) for the Plex-ID biosensor system, a platform that uses electrospray ionization mass spectroscopy (ESI-MS) to detect the base composition of short PCR amplicons. The new food-borne pathogen (FBP) plate has been experimentally designed using four gene segments for a total of eight amplicon targets. Initial work built a DNA base count database that contains more than 140 Salmonella enterica, 139 E. coli, 11 Shigella, and 36 Listeria patterns and 18 other Enterobacteriaceae organisms. This assay was tested to determine the scope of the assay\u27s ability to detect and differentiate the enteric pathogens and to improve the reference database associated with the assay. More than 800 bacterial isolates of S. enterica, E. coli, and Shigella species were analyzed. Overall, 100% of S. enterica, 99% of E. coli, and 73% of Shigella spp. were detected using this assay. The assay was also able to identify 30% of the S. enterica serovars to the serovar level. To further characterize the assay, spiked food matrices and food samples collected during regulatory field work were also studied. While analysis of preenrichment media was inconsistent, identification of S. enterica from selective enrichment media resulted in serovar-level identifications for 8 of 10 regulatory samples. The results of this study suggest that this high-throughput method may be useful in clinical and regulatory laboratories testing for these pathogens