Extracellular vesicle-carried Jagged-1 inhibits HUVEC sprouting in a 3D microenvironment.

NOTCH signalling is an evolutionarily conserved juxtacrine signalling pathway that is essential in development. Jagged1 (JAG1) and Delta-like ligand 4 (DLL4) are transmembrane NOTCH ligands that regulate angiogenesis by controlling endothelial cell (EC) differentiation, vascular development and maturation. In addition, DLL4 could bypass its canonical cell-cell contact-dependent signalling to influence NOTCH signalling and angiogenesis at a distance when it is packaged into extracellular vesicles (EVs). However, it is not clear whether JAG1 could also be packaged into EVs to influence NOTCH signalling and angiogenesis. In this work, we demonstrate that JAG1 is also packaged into EVs. We present evidence that JAG1-EVs inhibit NOTCH signalling and regulate EC behaviour and function. JAG1-EVs inhibited VEGF-induced HUVEC proliferation and migration in 2D culture condition and suppressed sprouting in a 3D microfluidic microenvironment. JAG1-EV treatment of HUVECs leads to a reduction of Notch1 intracellular domain (N1-ICD), and the proteasome and the intracellular domain of JAG1 (JAG1-ICD) are both required for this reduction to occur. These findings reveal a novel mechanism of JAG1 function in NOTCH signalling and ECs through EVs.

Long-chain glucosylceramides crosstalk with LYN mediates endometrial cell migration.

Endometriosis is a disease characterized by regurgitated lesions which are invasive and migratory, embedding at ectopic, extra-uterine locations. Extracellular glucosylceramides (GlcCers), bioactive sphingolipids potentiating signals for cell migration, are found in elevated levels in endometriosis; however underlying mechanisms that result in cellular migration are poorly defined. Here, we demonstrated that internalized GlcCer induced migratory activity in immortalized human endometrial stromal cells (HESCs), with highest potency observed in long-chain GlcCer. Long-chain ceramide (Cer) similarly induced cellular migration and mass spectrometry results revealed that the migratory behavior was contributed through glycosylation of ceramides. Cells treated with GlcCer synthase inhibitor, or RNAi-mediated knockdown of glucosylceramide synthase (GCS), the enzyme catalyzing GlcCer production attenuated cell motility. Mechanistic studies showed that GlcCer acts through stromal cell-derived factor-1 alpha and its receptor, CXC chemokine receptor 4 (SDF-1α-CXCR4) signaling axis and is dependent on phosphorylation of LYN kinase at Tyr396, and dephosphorylation of Tyr507. Migration was prominently attenuated in cells exposed to CXCR4 antagonist, AMD3100, yet can be rescued with diprotin A, which prevents the degradation of SDF-1α. Furthermore, blocking of LYN kinase activity in the presence of SDF-1α and GlcCer reduced HESC migration, suggesting that LYN acts downstream of GlcCer-SDF-1α-CXCR4 axis as part of its intracellular signal transduction. Our results reveal a novel role of long-chain GlcCer and the dialog between GlcCer, LYNpTyr396 and SDF-1α-CXCR4 in inducing HESC migration. This finding may improve our understanding how endometriotic lesions invade to their ectopic sites, and the possibility of using GlcCer to modulate the SDF-1α-CXCR4-LYNpTyr396 axis in endometriosis.

Quantification of magnetically induced changes in ECM local apparent stiffness.

The stiffness of the extracellular matrix (ECM) is known to influence cell behavior. The ability to manipulate the stiffness of ECM has important implications in understanding how cells interact mechanically with their microenvironment. This article describes an approach to manipulating the stiffness ECM, whereby magnetic beads are embedded in the ECM through bioconjugation between the streptavidin-coated beads and the collagen fibers and then manipulated by an external magnetic field. It also reports both analytical results (obtained by formal modeling and numerical simulation) and statistically meaningful experimental results (obtained by atomic force microscopy) that demonstrate the effectiveness of this approach. These results clearly suggest the possibility of creating desired stiffness gradients in ECM in vitro to influence cell behavior.

An Electromagnetic System for Inducing a Localized Force Gradient in an ECM and Its Influence on HMVEC Sprouting.

Mechanical properties of the extracellular matrix (ECM) have been observed to influence the behavior of cells. Investigations on such an influence commonly rely on using soluble cues to alter the global intrinsic ECM properties in order to study the subsequent response of cells. This article presents an electromagnetic system for inducing a localized force gradient in an ECM, and reports the experimentally observed effect of such a force gradient on in vitro angiogenic sprouting of human microvascular endothelial cells (HMVECs). This force gradient is realized through the induction of magnetic forces on the superparamagnetic microparticle-embedded ECM ( sECM). Both analytical and statistically meaningful experimental results demonstrate the effectiveness of this approach in influencing the behavior of a targeted HMVEC sprout without affecting that of other sprouts nearby. These results suggest the possibility of selectively controlling the in vitro behavior of cells by the induction of a localized force gradient in the ECM.

A Magneto-Microfluidic System for Investigating the Influence of an Externally Induced Force Gradient in a Collagen Type I ECM on HMVEC Sprouting.

Advances in mechanobiology have suggested that physiological and pathological angiogenesis may be differentiated based on the ways in which the cells interact with the extracellular matrix (ECM) that exhibits partially different mechanical properties. This warrants investigating the regulation of ECM stiffness on cell behavior using angiogenesis assays. In this article, we report the application of the technique of active manipulation of ECM stiffness to study in vitro angiogenic sprouting of human microvascular endothelial cells (HMVECs) in a microfluidic device. Magnetic beads were embedded in the ECM through bioconjugation (between the streptavidin-coated beads and collagen fibers) in order to create a pretension in the ECM when under the influence of an external magnetic field. The advantage of using this magneto-microfluidic system is that the resulting change in the local deformability of the collagen fibers is only apparent to a cell at the pericellular level near the site of an embedded bead, while the global intrinsic material properties of the ECM remain unchanged. The results demonstrate that this system represents an effective tool for inducing noninvasively an external force on cells through the ECM, and suggest the possibility of creating desired stiffness gradients in the ECM for manipulating cell behavior in vitro.

Image classification of unlabeled malaria parasites in red blood cells.

This paper presents a method to detect unlabeled malaria parasites in red blood cells. The current "gold standard" for malaria diagnosis is microscopic examination of thick blood smear, a time consuming process requiring extensive training. Our goal is to develop an automate process to identify malaria infected red blood cells. Major issues in automated analysis of microscopy images of unstained blood smears include overlapping cells and oddly shaped cells. Our approach creates robust templates to detect infected and uninfected red cells. Histogram of Oriented Gradients (HOGs) features are extracted from templates and used to train a classifier offline. Next, the ViolaJones object detection framework is applied to detect infected and uninfected red cells and the image background. Results show our approach out-performs classification approaches with PCA features by 50% and cell detection algorithms applying Hough transforms by 24%. Majority of related work are designed to automatically detect stained parasites in blood smears where the cells are fixed. Although it is more challenging to design algorithms for unstained parasites, our methods will allow analysis of parasite progression in live cells under different drug treatments.

Characterization of uniaxial stiffness of extracellular matrix embedded with magnetic beads via bio-conjugation and under the influence of an external magnetic field.

In this paper, we study the deformation, and experimentally quantify the change in stiffness, of an extracellular matrix (ECM) embedded with magnetic beads that are bio-conjugated with the collagen fibers and under the influence of an external magnetic field. We develop an analytical model of the viscoelastic behavior of this modified ECM, and design and implement a stretch test to quantify (based on statistically meaningful experiment data) the resulting changes in its stiffness induced by the external magnetic field. The analytical results are in close agreement with that obtained from the experiments. We discuss the implication of these results that point to the possibility of creating desired stiffness gradients in an ECM in vitro to influence cell behavior.

Exosomes in Cancer Microenvironment and Beyond: have we Overlooked these Extracellular Messengers?

Cancer is a complex organ whose behavior is not only influenced by genetic and epigenetic changes in cancer cells but also by stromal cells, local extracellular matrix and specific tissue architecture. Intercellular communications within the cancer microenvironment are critical to coordinate the assembly of multiple cell types for an amalgamated form and function of a cancer. Exosomes are small membrane vesicles with an endosome origin that are released by cells into the extracellular environment. They carry a cargo of proteins, lipids, and nucleic acids and transfer their cargo to recipient cells and altering the recipient cells' biochemical composition, signaling pathways, and gene regulation. Exosomes can thus serve as extracellular messengers mediating cell-cell communication. Both cancer cells and stromal cells release exosomes not only into the cancer microenvironment but also into the circulation. In this review, we summarize the research done so far on cancer-derived exosomes and assess their roles as extracellular messengers facilitating cancer progression and metastasis.

A hybrid continuum-discrete modelling approach to predict and control angiogenesis: analysis of combinatorial growth factor and matrix effects on vessel-sprouting morphology.

Angiogenesis is crucial during many physiological processes and is influenced by various biochemical and biomechanical factors. Models have proved useful in understanding the mechanisms of angiogenesis and also the characteristics of the capillaries formed as part of the process. We have developed a three-dimensional hybrid, agent-field model where individual cells are modelled as sprout-forming agents in a matrix field. Cell independence, cell-cell communication and stochastic cell response are integral parts of the model. The model simulations incorporate probabilities of an individual cell to transition into one of four stages--quiescence, proliferation, migration and apoptosis. We demonstrate that several features, such as continuous sprouts, cell clustering and branching, that are observed in microfluidic experiments conducted under controlled conditions using few angiogenic factors can be reproduced by this model. We also identify the transition probabilities that result in specific sprout characteristics such as long continuous sprouts and specific branching patterns. Thus, this model can be used to cluster sprout morphology as a function of various influencing factors.

Dimensionality reduction of cellular actuator arrays using the concept of synergy for driving a robotic hand.

This paper presents a method of reducing dimensionality of cellular actuator arrays. The cellular actuator arrays are used to drive a multi-axis system, i.e. robotic hand. The actuator arrays use Segmented Binary Control (SBC) which simplifies the control of nonlinear artificial muscle actuators. Although the SBC simplifies the control, the number of cells required to create motions is increased dramatically, thereby increasing the dimension of the actuator arrays. Therefore, reducing the number of controls, or dimension is needed. The coordinated motion of the robotic hand enables the coupling of actuators. In biological systems, synergies, a strategy of grouping output variables to simplify the control of a large number of muscles and joints is used to explain the coordinated motion created by the muscles. Similarly we can apply this concept to group the cells of the actuator array to be turned on or off together. Each group of cells, called segments can be designed to perform a certain set of desired postures. Data from fifteen different tasks is used in the design. The gathered joint angle data is transformed into actuator displacement data and used to generate a segmentation design of the actuator. For segmentation design, feature extraction method, similar to non-negative matrix factorization with binary filter is proposed. The segmentation design reduces 96 separately controlled segments to 6, while maintaining the ability to accomplish all desired postures. A prototype robotic hand with five fingers, designed and fabricated using the FDM process, is driven with this actuator system.

Cable-Free Body Area Network using Conductive Fabric Sheets for Advanced Human-Robot Interaction.

Wearable sensing networks offer a multitude of benefits for the user. One field that can benefit from such technologies is in-home robotics. Much work has been dedicated to the general area of interaction between robots and humans, and more specifically to gesture recognition. We propose a wearable monitoring network composed of conductive fabrics that can more easily facilitate robot interaction. It creates additional interaction by visual indication, and electronically by way of a local PC. This system creates a more natural human-robot interface and makes feasible many new forms of gesture recognition. Finally, we can address safety concerns; the garment gives us a method of locating the human and minimizing the possibility for robots to strike the user. In this paper, we lay out the architecture for such a system, and perform some of the initial characterization.

Novel design for a wearable, rapidly deployable, wireless noninvasive triage sensor.

This paper presents a unique design for a low-power, continuous non-invasive sensor capable of remotely monitoring the five major vital signs of a patient. In particular, the sensor is designed for rapid attachment to the fingerbase of a patient by utilizing a clip-type mechanism and is comprised of a photoplethysmograph (PPG), a MEMS accelerometer, a temperature sensor, and a wireless node. Although hastily placed by a medic, the finger sensor will automatically find the location of a digital artery and acquire a clear, pulse signal: a micro-sensor array accommodates the location of the sensor attachment. Additionally, the PPG signal, although corrupted with the patient's motion in chaotic environment, will be recovered by using the MEMS accelerometer and an Active Noise Cancellation algorithm.