P148 CD68 expression in inflammatory cell infiltration of nonspecific invasive breast cancer

BackgroundTumor-associated macrophages play a main role in tumor progression and dissemination. Taking into account the high heterogeneicity of tumor the different clinical impact of macrophages, infiltrating different sites of tumor, could be expected. The aim was to detect the level of CD68+ cells (macrophages) in the different site of stroma in breast tumor in comparison to clinical course.Materials and methodsOne thirty-six women with nonspecific invasive breast cancer T1-4N0-3M0, who were treated in General Oncology Department of Tomsk Cancer Research Institute (Tomsk, Russia), were included in the present study. Patients did not receive preoperative treatment. The material was fixed in 10–12% neutral formalin. Preparation of the histological material was carried out according to standard procedures. Morphological examination of the surgical specimens was performed by the standard method using a light microscope “Carl Zeiss Axio Lab.A1” (Germany) and slidescanner “MiraxMidiZeiss” (Germany). Metastatic lesion was detected in regional lymph nodes. Immunohistochemical study was performed according to standard procedures. Cytoplasmic expression of these markers was determined in the inflammatory cell infiltrate of different tumor segments: (1) in areas with soft fibrous stroma; (2) in areas with coarse fibrous stroma; (3) in the areas of the so-called “maximum stromal-and-parenchymal relationship” where the individual tumor cells, short strands and groups of tumor cells arranged in soft fibrous stroma; (4) among parenchymal elements; (5) in gaps of ductal tumor structures. Double-stained immunofluorescence was performed according to standard procedures using Leica TCS SP2 laser-scanning spectral confocal microscope (Germany). The following primary antibodies were used: mouse monoclonal anti-human CD68 (BD Biosciences) and rabbit polyclonal anti-stabilin-1 or RS1 (marker of M2 macrophages).ResultsThe highest expression of CD68 in the inflammatory cell infiltrate was detected more frequently in areas with soft fibrous stroma (54%) or the so-called “maximum stromal-and-parenchymal relationship” (79%) in patients with breast cancer. The lowest expression of CD68 was observed in areas with coarse fiber stroma (23%). The CD68-positive cells of the inflammatory infiltrate were located between parenchymal elements of tumor (88%). Inverse correlation (R=−0.67; p=0.02) observed between tumor size and the expression of CD68 in the cells of the inflammatory infiltrate in gaps of tubular tumor structures. The CD68 expression in cells of the inflammatory tumor infiltrate was correlated with the presence of metastatic regional lymph nodes. It was found that in the case of the lymph node metastases the average score of CD68 expression in cells of ductal gaps tumor structures was lower (1.4±0.5) in comparison with the negative lymph nodes case (3.1±1.0; F=10.9; p=0.007). Same time no correlation between the CD68 expression in the inflammatory cell tumor infiltrate and the rate of tumor malignancy was found. Using confocal microscopy domination of CD68+/RS1+ cells were found.ConclusionSo, low CD68 expression level in ductal gaps tumor structures is associated with the presence of metastatic regional lymph nodes

Macrophage and tumor cell responses to repetitive pulsed X-ray radiation

To study a response of tumor cells and macrophages to the repetitive pulsed low-dose X-ray radiation. Methods. Tumor growth and lung metastasis of mice with an injected Lewis lung carcinoma were analysed, using C57Bl6. Monocytes were isolated from a human blood, using CD14+ magnetic beads. IL6, IL1-betta, and TNF-alpha were determined by ELISA. For macrophage phenotyping, a confocal microscopy was applied. "Sinus-150" was used for the generation of pulsed X-ray radiation (the absorbed dose was below 0.1 Gy, the pulse repetition frequency was 10 pulse/sec). The irradiation of mice by 0.1 Gy pulsed X-rays significantly inhibited the growth of primary tumor and reduced the number of metastatic colonies in the lung. Furthermore, the changes in macrophage phenotype and cytokine secretion were observed after repetitive pulsed X-ray radiation. Conclusion. Macrophages and tumor cells had a different response to a low-dose pulsed X-ray radiation. An activation of the immune system through changes of a macrophage phenotype can result in a significant antitumor effect of the low-dose repetitive pulsed X-ray radiation

In Vitro Evaluation of a Specific Radiochemical Compound Based on 99mTc-labeled DARPinG3 for Radionuclide Imaging of Tumors Overexpressing Her-2/neu

It is still necessary to search for new informative diagnostic methods to detect malignant tumors with overexpression of Her-2/neu, which are characterized by the aggressive course of the disease, rapid rate of tumor growth and low rates of relapse-free and overall survival. In recent years, the radioisotope techniques for detection of specific tumor targets have been developing actively. Purpose: to develop a chemically stable radiochemical compound for the targeted imaging of cells overexpressing Her-2/neu. Material and methods: The study was performed using 2 cell lines .The human breast adenocarcinoma HER2-overexpressing cell line BT-474 was chosen to detect specific binding. As a control, HER2-negative human breast adenocarcinoma MCF-7 was used. The human breast adenocarcinoma BT-474 and MCF-7 cell lines were seeded in chamber-slides at the density of 35,000 cells/ml in trypsin-EDTA (PanEco) medium and grown overnight at 37°C. After that both cell lines were washed with Phosphate buffered saline (PBS) and distributed into test tubes to 1 ml (5 millions cells in each). After adding 100 [mu]l (70 MBq) studied complex of 99mTc-DPAH-DARPinG3 was incubated for 40 min at +4°C. Washing was performed three times with buffer PBS and 5% Bovine Serum Albumin (BSA). The characteristics of the binding specificity of the test set with the HER-2/neu receptor were determined by direct radiometric and planar scintigraphy. Nonparametric Mann-Whitney test was used to assess the differences in the quantitative characteristics between groups. Results: The output of the labeled complex was more than 91%, with a radiochemical purity of more than 94%. When carrying out a visual scintigraphic assessment much greater intensity accumulation of radiotracer was observed in the studied cell culture surface receptor overexpressing Her-2/neu. The results of direct radiometric also showed higher accumulation of the radiopharmaceutical in the adenocarcinoma cell line BT-474 human breast cancer overexpressing Her-2/neu compared to the control group. Conclusion: The preclinical studies demonstrated a high in vitro stability of the study compound, as well as its accumulation in the cell group overexpressing Her-2/neu

Clinically relevant morphological structures in breast cancer represent transcriptionally distinct tumor cell populations with varied degrees of epithelial-mesenchymal transition and CD44+CD24- stemness

Intratumor morphological heterogeneity in breast cancer is represented by different morphological structures (tubular, alveolar, solid, trabecular, and discrete) and contributes to poor prognosis; however, the mechanisms involved remain unclear. In this study, we performed 3D imaging, laser microdissection-assisted array comparative genomic hybridization and gene expression microarray analysis of different morphological structures and examined their association with the standard immunohistochemistry scorings and CD44+CD24- cancer stem cells. We found that the intratumor morphological heterogeneity is not associated with chromosomal aberrations. By contrast, morphological structures were characterized by specific gene expression profiles and signaling pathways and significantly differed in progesterone receptor and Ki-67 expression. Most importantly, we observed significant differences between structures in the number of expressed genes of the epithelial and mesenchymal phenotypes and the association with cancer invasion pathways. Tubular (tube-shaped) and alveolar (spheroid-shaped) structures were transcriptionally similar and demonstrated co-expression of epithelial and mesenchymal markers. Solid (large shapeless) structures retained epithelial features but demonstrated an increase in mesenchymal traits and collective cell migration hallmarks. Mesenchymal genes and cancer invasion pathways, as well as Ki-67 expression, were enriched in trabecular (one/two rows of tumor cells) and discrete groups (single cells and/or arrangements of 2-5 cells). Surprisingly, the number of CD44+CD24- cells was found to be the lowest in discrete groups and the highest in alveolar and solid structures. Overall, our findings indicate the association of intratumor morphological heterogeneity in breast cancer with the epithelial-mesenchymal transition and CD44+CD24- stemness and the appeal of this heterogeneity as a model for the study of cancer invasion

The makewaves tsunami tests and their relevance to tsunami engineering and risk management

MAKEWAVES is an international multi-partner collaborative project bringing together nine academic institutions and two commercial consultancies. The objective of the collaboration is to develop experimental data and associated numerical modelling on tsunami inundation and interaction with boulders, buildings, natural and engineered barriers, towards the development of new internationally accepted guidance for structural codes and standards. Using a pneumatic tsunami simulator (TS) developed jointly by HR Wallingford and UCL the team conducted experiments between November 2022 and April 2023 within a highly instrumented 100m long flume. The TS is capable of simulating realistic trough and crest-led tsunami waves at 1:50, including traces from the The TS is capable of generating very long trough and crest-led waves, and can reproduce at 1:50 scale waves from real life events such as the Mercator trace from the 2004 Indian Ocean event and the and 2011 Tohoku tsunamis. The TS capability has been further extended to include bore-waves. The characteristics of the waves are controlled by adjusting the flow rate and total volume of water drawn in and discharged by the TS. The experimental campaign is was subdivided into discrete research areas, each aimed at furthering knowledge on how different tsunami wave characteristics affect their interaction with manmade and natural structures environments. These include tests aimed at understanding: (1) how roughness representative of coastal forests and mangroves affects tsunami inundation characteristics, (2) how tsunami interact with boulders (3) the effectiveness of offshore breakwaters as tsunami barriers (4) how structural loads and foundation scour are affected by building permeability. This paper presents an overview of the tests conducted and some of the important early observations made that are relevant to future engineering standards and to tsunami disaster management

ЭКСПРЕССИЯ CXCR4 В РАЗЛИЧНЫХ ПОПУЛЯЦИЯХ ЦИРКУЛИРУЮЩИХ И ОДИНОЧНЫХ ОПУХОЛЕВЫХ КЛЕТОК РАКА МОЛОЧНОЙ ЖЕЛЕЗЫ

The aim of this study was to assess CXCR4 expression in different subsets of CTCs and single (detached) breast cancer cells.Materials and methods. Thirty five patients with invasive breast carcinoma of no specialtype (IC NST) (T1-4N0-2M0), between 29 and 69 years of age were included in this study. Different subsets of CTCs with CXCR4 expression were evaluated by flow cytometry. A  confocal microscopy was used to assess CXCR4 expression in different subsets of single (detached) cancer cells in breast tissue.Results. The CXCR4 was expressed in CTCs without stem-like and EMT phenotype, in CTCs  with EMT but not stem markers and in stem-like CTCs without EMT features. In all blood  samples, the CXCR4 expression in CTCs with stem-like and EMT phenotype was absent. In  breast tumor the CXCR4 was expressed in the non stemlike single (detached) breast cancer  cells with EMT features, in the single (detached) breast cancer cells with stem and EMT  features. In all tumor samples the stem-like or non stem-like single (detached) breast  cancer cells without EMT features were absent.Conclusions. Different subsets of the CTCs exhibited CXCR4. The CXCR4 expression did not  depend on the presence or absence of stem or/and EMT features in tumor cells. We showed that some subsets of single (detached) breast cancer cells in the primary tumor  were characterized by the ability to express CXCR4 and may be a source of the respective CTC subsets.Целью исследования явилось определение экспрессии CXCR4 в различных популяциях циркулирующих (ЦОК) и одиночных (дискретных) опухолевых клеток рака молочной железы.Материал и методы. В исследование были включены 35 пациенток с инвазивной карциномой неспецифического типа молочной железы (T1–4N0–2M0) в возрасте от 29 до 69 лет. Экспрессию CXCR4 в  различных популяциях ЦОК оценивали методом проточной цитометрии. Для оценки экспрессии CXCR4 в  аналогичных популяциях одиночных (дискретных) опухолевых клеток в первичной опухоли использовали метод конфокальной микроскопии.Результаты. Нами было установлено, что CXCR4 экспрессировался ЦОК без признаков стволовости и  эпителиально-мезенхимального перехода (ЭМП), ЦОК с признаками ЭМП, но без маркеров стволовости, а  также ЦОК с признаками стволовости, но без признаков ЭМП. У всех пациенток в крови ЦОК с признаками  стволовости и ЭМП не экспрессировался CXCR4. В первичной опухоли молочной железы CXCR4  обнаруживался как на одиночных (дискретных) опухолевых клетках без признаков стволовости с  признаками ЭМП, так и на клетках с маркерами стволовости и ЭМП. У всех пациенток в образцах первичной  опухоли отсутствовали стволовые и нестволовые клетки без признаков ЭМП.Заключение. Таким образом, CXCR4 экспрессируются на различных популяциях ЦОК. Экспрессия CXCR4  не зависит от наличия или отсутствия признаков стволовости и/или ЭМП в опухолевых клетках. Также мы  показали, что некоторые популяции одиночных (дискретных) опухолевых клеток в первичной опухоли  характеризуются способностью презентировать на своей мембране CXCR4 и могут являться источником соответствующих популяций ЦОК

A Comparative Study on the Nonlinear Interaction Between a Focusing Wave and Cylinder Using State-of-the-art Solvers: Part A

This paper presents ISOPE’s 2020 comparative study on the interaction between focused waves and a fixed cylinder. The paper discusses the qualitative and quantitative comparisons between 20 different numerical solvers from various universities across the world for a fixed cylinder. The moving cylinder cases are reported in a companion paper as part B (Agarwal, Saincher, et al., 2021). The numerical solvers presented in this paper are the recent state of the art in the field, mostly developed in-house by various academic institutes. The majority of the participants used hybrid modeling (i.e., a combination of potential flow and Navier–Stokes solvers). The qualitative comparisons based on the wave probe and pressure probe time histories and spectral components between laminar, turbulent, and potential flow solvers are presented in this paper. Furthermore, the quantitative error analyses based on the overall relative error in peak and phase shifts in the wave probe and pressure probe of all the 20 different solvers are reported. The quantitative errors with respect to different spectral component energy levels (i.e., in primary, sub-, and superharmonic regions) capturing capability are reported. Thus, the paper discusses the maximum, minimum, and median relative errors present in recent solvers as regards application to industrial problems rather than attempting to find the best solver. Furthermore, recommendations are drawn based on the analysis