Cancer is an increasingly recognized problem associated with immunosuppression. Recent reports, however, suggest that the immunosuppressive agent rapamycin has anti-cancer properties that could address this problem. Thus far, rapamycin's effects on immunity and cancer have been studied separately. Here we tested the effects of rapamycin, versus cyclosporine A (CsA), on established tumors in mice simultaneously bearing a heart allograft. In one tumor-transplant model, BALB/c mice received subcutaneous syngenic CT26 colon adenocarcinoma cells 7 days before C3H ear-heart transplantation. Rapamycin or CsA treatment was initiated with transplantation. In a second model system, a B16 melanoma was established in C57BL/6 mice that received a primary vascularized C3H heart allograft. In vitro angiogenic effects of rapamycin and CsA were tested in an aortic ring assay. Results show that CT26 tumors grew for 2 weeks before tumor complications occurred. However, rapamycin protected allografts, inhibited tumor growth, and permitted animal survival. In contrast, CsA-treated mice succumbed to advancing tumors, albeit with a functioning allograft. Rapamycin's antitumor effect also functioned in severe combined immunodeficient BALB/c mice. Similar effects of the drugs occurred with B16 melanomas and primary vascularized C3H allografts in C57BL/6 mice. Furthermore, in this model, rapamycin inhibited the tumor growth-enhancing effects of CsA. Moreover, in vitro experiments showed that CsA promotes angiogenesis by a transforming growth factor-beta-related mechanism, and that this effect is abrogated by rapamycin. This study demonstrates that rapamycin simultaneously protects allografts from rejection and attacks tumors in a complex transplant-tumor situation. Notably, CsA protects allografts from rejection, but cancer progression is promoted in transplant recipients.

Natural killer (NK) cells are important effector cells in the immune response to cancer. Clinical trials on adoptively transferred NK cells in patients with solid tumors, however, have thus far been unsuccessful. As NK cells need to pass stringent safety evaluation tests before clinical use, the cells are cryopreserved to bridge the necessary evaluation time. Standard degranulation and chromium release cytotoxicity assays confirm the ability of cryopreserved NK cells to kill target cells. Here, we report that tumor cells embedded in a 3-dimensional collagen gel, however, are killed by cryopreserved NK cells at a 5.6-fold lower rate compared to fresh NK cells. This difference is mainly caused by a 6-fold decrease in the fraction of motile NK cells after cryopreservation. These findings may explain the persistent failure of NK cell therapy in patients with solid tumors and highlight the crucial role of a 3-D environment for testing NK cell function.

We describe a quantitative, high-precision, high-throughput method for measuring the mechanical properties of cells in suspension with a microfluidic device, and for relating cell mechanical responses to protein expression levels. Using a high-speed (750 fps) charge-coupled device camera, we measure the driving pressure Δp, maximum cell deformation ε, and entry time t of cells in an array of microconstrictions. From these measurements, we estimate population averages of elastic modulus E and fluidity β (the power-law exponent of the cell deformation in response to a step change in pressure). We find that cell elasticity increases with increasing strain ε according to E ∼ ε, and with increasing pressure according to E ∼ Δp. Variable cell stress due to driving pressure fluctuations and variable cell strain due to cell size fluctuations therefore cause significant variability between measurements. To reduce measurement variability, we use a histogram matching method that selects and analyzes only those cells from different measurements that have experienced the same pressure and strain. With this method, we investigate the influence of measurement parameters on the resulting cell elastic modulus and fluidity. We find a small but significant softening of cells with increasing time after cell harvesting. Cells harvested from confluent cultures are softer compared to cells harvested from subconfluent cultures. Moreover, cell elastic modulus increases with decreasing concentration of the adhesion-reducing surfactant pluronic. Lastly, we simultaneously measure cell mechanics and fluorescence signals of cells that overexpress the GFP-tagged nuclear envelope protein lamin A. We find a dose-dependent increase in cell elastic modulus and decrease in cell fluidity with increasing lamin A levels. Together, our findings demonstrate that histogram matching of pressure, strain, and protein expression levels greatly reduces the variability between measurements and enables us to reproducibly detect small differences in cell mechanics.

Summary1. Optical recordings are ubiquitous in current scientific research. As recording and storing images and videos have become simpler and cheaper over the past years, new and more efficient tools are needed to analyse the growing amount of data. While current scientific image analysis tools focus on medical 3D data sets, a tool for efficiently handling large-scale 2D time-series recordings is still missing. 2. We developed ClickPoints, an expandable, open-source, Python-based software to fill this gap. ClickPoints combines and streamlines the three main steps of image analysis -visualization, annotation and evaluation -in one program. 3. ClickPoints enables the user to (i) efficiently review large data sets of images and videos, (ii) annotate interesting findings and (iii) facilitate manual and automatic evaluations. ClickPoints is highly versatile, ranging from clicking and selecting objects with simple markers, drawing masks and computing trajectories, up to (iv) customwritten Python add-ons for adapting or extending the available features. These add-ons (v) reduce development time by utilizing ClickPoints' display, interface and storage solutions. In addition, ClickPoints provides an (vi) extensive data base application programming interface to facilitate efficient storage and retrieval of results. 4. ClickPoints is designed to simplify repetitive and labour-intensive scientific tasks, especially when dealing with large image and video time-series recordings. ClickPoints offers cross-platform support for Windows and Linux and is released under the GPLv3 license. Download, documentation and numerous examples are available at http://clickpoints.readthedocs.io.

Scientific applications often require an exact reconstruction of object positions and distances from digital images. Therefore, the images need to be corrected for perspective distortions. We present CameraTransform, a python package that performs a perspective image correction whereby the height, tilt/roll angle and heading of the camera can be automatically obtained from the images if additional information such as GPS coordinates or object sizes are provided. We present examples of images of penguin colonies that are recorded with stationary cameras and from a helicopter. Camera MatrixAll information about the mapping of real-world points to image points are stored in a camera matrix. The camera matrix is expressed in projective coordinates, and can be split into two parts: the intrinsic matrix and the extrinsic matrix [3]. The intrinsic matrix depends on the camera sensor and lens, the extrinsic matrix depends on the camera's position and orientation. Projective coordinatesProjective coordinates, also known as homogeneous coordinates, are used to represent projective transformations as matrix multiplications [6]. They are a mathematical trick that extends the 1 arXiv:1712.07438v1 [cs.MS]