Integrin-mediated traction force enhances paxillin molecular associations and adhesion dynamics that increase the invasiveness of tumor cells into a three-dimensional extracellular matrix.

Metastasis requires tumor cells to navigate through a stiff stroma and squeeze through confined microenvironments. Whether tumors exploit unique biophysical properties to metastasize remains unclear. Data show that invading mammary tumor cells, when cultured in a stiffened three-dimensional extracellular matrix that recapitulates the primary tumor stroma, adopt a basal-like phenotype. Metastatic tumor cells and basal-like tumor cells exert higher integrin-mediated traction forces at the bulk and molecular levels, consistent with a motor-clutch model in which motors and clutches are both increased. Basal-like nonmalignant mammary epithelial cells also display an altered integrin adhesion molecular organization at the nanoscale and recruit a suite of paxillin-associated proteins implicated in invasion and metastasis. Phosphorylation of paxillin by Src family kinases, which regulates adhesion turnover, is similarly enhanced in the metastatic and basal-like tumor cells, fostered by a stiff matrix, and critical for tumor cell invasion in our assays. Bioinformatics reveals an unappreciated relationship between Src kinases, paxillin, and survival of breast cancer patients. Thus adoption of the basal-like adhesion phenotype may favor the recruitment of molecules that facilitate tumor metastasis to integrin-based adhesions. Analysis of the physical properties of tumor cells and integrin adhesion composition in biopsies may be predictive of patient outcome

Progressing Mechanobiology from a Simplified to a More Complex System: Development of Novel Platforms and Investigation of Actin Cytoskeletal Remodeling

Compression is a common mechanical stimulus in our body that results from gravity and tissue growth. Inability for cells to maintain force homeostasis can lead to various diseases, and cells may experience an increase in compressive stress in diseases such as hypertension and cancer. Actin cytoskeleton provides structural and mechanical strength for cells to withstand external mechanical forces. It is also very sensitive to deformation and plays an important role in mechanosensing, due to the tensegrity nature of the cytoskeleton. Despite being an important component, actin remodeling and molecular signaling in response to compression have not been very well studied. From a molecular perspective, the actin network is regulated by actin accessory proteins into different structures and functions. At a cellular level, different actin networks are organized and regulated by various signaling pathways. At an intercellular level, cells are mechanically coupled that enable the transmission of force via actin networks between cells. In this dissertation, I investigated the sensing and transduction of compression stimulus in cells from molecular to intercellular scales, I developed new emerging platforms for the compression studies in in vitro reconstitution and single-cell system, and investigated actin remodeling and its related mechanisms under compression in 2D cell population. First, I developed an approach that combines purified actin cytoskeletal proteins or mammalian cell-free expression and ultra-thin double emulsion template for constructing a simplified model of a cell through in vitro reconstitution. This was used to study the mechanical properties of a specific and isolated cytoskeleton structure. We demonstrated the formation of bundled actin filaments inside the lipid vesicles. By using the mammalian cell-free expression system, we found that actin structures inside the system were precipitating with the droplet stabilizing surfactant polyvinyl alcohol, leading to a compensation of protein production and vesicle stability. Nevertheless, this approach provides a simplified yet insightful cell-like model for future cell mechanics investigation. I also developed a pneumatic-controlled, two-layered microfluidic platform for applying compression and aspiration to double emulsion droplets as a model cell. I further improved the microfluidic device to apply controlled compression to single cells that can be used to study the heterogeneity of mechanical properties and responses of cells. The device was designed through optimization by simulation and was characterized experimentally. Static and cyclic compressions were applied to single cells seeded inside the device. Finally, I investigated actin cytoskeletal remodeling, mechanosensing and mechanotransduction of epithelial cells under compression, in a 2D cell population context. I discovered the formation of actin protrusions at the apical surface of the cells under 1200 Pa compression. The actin protrusions were structurally similar to invadopodia but not functionally. Src signaling was found to be an important signaling pathway in these actin protrusions. This discovery opens up new direction of research and may explain why cells become more invasive under compression. This work could also shed light on the heterogeneity of cancer tissues and may inspire a new treatment paradigm. In this dissertation, I developed experimental platforms and studied the remodeling of actin networks under compression using in vitro reconstitution, single-cell and cell population systems, taking advantages of each experimental system. The platforms developed in this dissertation provide novel techniques for investigating actin cytoskeleton response of cell under compression, and the discovery of actin protrusions reported in this dissertation reveals a new cellular response under high compression.PHDMechanical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/144103/1/kennetho_1.pd

At the interface: biotic-abiotic interactions between substrates and a model epithelium

The need for determining the fundamental mechanisms that define the interaction of biological systems with underlying materials, both natural and synthetic, is important as humanity endeavors to improve the quality of life of individuals through technology. Recently, much work has focused on the role of material properties on the behavior of cells. Most of these studies have concentrated their efforts on fibroblastic cell lines and more recently different kinds of stems cells. While these cells represent an important subset of cells in complex organisms, they do not manifest cell-cell interactions, a feature of epithelial cells, the most abundant cell type. Epithelial cells represent the largest cell type in the body and introduce an intrinsic complexity when researching the interaction of biological systems with materials. Adherens junctions (AJ) play a significant role in many signaling pathways, and therefore there is need to investigate how physical interactions with underlying substrates affect cell-cell interactions, such as the adhesion properties between cells, as well as how cell-substrate interactions influence the morphology and growth of epithelial cells. In this work I seek to determine the effects and identify mechanisms that epithelial cells use to “read” their environment. To do this I examined changes in cell behavior (growth, morphological, adhesion) of a model epithelium on substrates that have similar composition but significant differences in surface organization. In such a manner, I probed the limitations at which the nanoscale differences in substrate topography affect cellular behavior

Agro-techniques for lodging stress management in maize-soybean intercropping system—A review

Lodging is one of the most chronic restraints of the maize-soybean intercropping system, which causes a serious threat to agriculture development and sustainability. In the maize-soybean intercropping system, shade is a major causative agent that is triggered by the higher stem length of a maize plant. Many morphological and anatomical characteristics are involved in the lodging phenomenon, along with the chemical configuration of the stem. Due to maize shading, soybean stem evolves the shade avoidance response and resulting in the stem elongation that leads to severe lodging stress. However, the major agro-techniques that are required to explore the lodging stress in the maize-soybean intercropping system for sustainable agriculture have not been precisely elucidated yet. Therefore, the present review is tempted to compare the conceptual insights with preceding published researches and proposed the important techniques which could be applied to overcome the devastating effects of lodging. We further explored that, lodging stress management is dependent on multiple approaches such as agronomical, chemical and genetics which could be helpful to reduce the lodging threats in the maize-soybean intercropping system. Nonetheless, many queries needed to explicate the complex phenomenon of lodging. Henceforth, the agronomists, physiologists, molecular actors and breeders require further exploration to fix this challenging problem.National Key Research and Development Program of China | Ref. 2018YFD1000905Sichuan Innovation Team Project of National Modern Agricultural Industry Technology System | Ref. SCCXTD−2020−2

Optogenetic Brain Interfaces

The brain is a large network of interconnected neurons where each cell functions as a nonlinear processing element. Unraveling the mysteries of information processing in the complex networks of the brain requires versatile neurostimulation and imaging techniques. Optogenetics is a new stimulation method which allows the activity of neurons to be modulated by light. For this purpose, the cell-types of interest are genetically targeted to produce light-sensitive proteins. Once these proteins are expressed, neural activity can be controlled by exposing the cells to light of appropriate wavelengths. Optogenetics provides a unique combination of features, including multimodal control over neural function and genetic targeting of specific cell-types. Together, these versatile features combine to a powerful experimental approach, suitable for the study of the circuitry of psychiatric and neurological disorders. The advent of optogenetics was followed by extensive research aimed to produce new lines of light-sensitive proteins and to develop new technologies: for example, to control the distribution of light inside the brain tissue or to combine optogenetics with other modalities including electrophysiology, electrocorticography, nonlinear microscopy, and functional magnetic resonance imaging. In this paper, the authors review some of the recent advances in the field of optogenetics and related technologies and provide their vision for the future of the field.United States. Defense Advanced Research Projects Agency (Space and Naval Warfare Systems Center, Pacific Grant/Contract No. N66001-12-C-4025)University of Wisconsin--Madison (Research growth initiative; grant 101X254)University of Wisconsin--Madison (Research growth initiative; grant 101X172)University of Wisconsin--Madison (Research growth initiative; grant 101X213)National Science Foundation (U.S.) (MRSEC DMR-0819762)National Science Foundation (U.S.) (NSF CAREER CBET-1253890)National Institutes of Health (U.S.) (NIH/NIBIB R00 Award (4R00EB008738)National Institutes of Health (U.S.) (NIH Director’s New Innovator award (1-DP2-OD002989))Okawa Foundation (Research Grant Award)National Institutes of Health (U.S.) (NIH Director’s New Innovator Award (1DP2OD007265))National Science Foundation (U.S.) (NSF CAREER Award (1056008)Alfred P. Sloan Foundation (Fellowship)Human Frontier Science Program (Strasbourg, France) (Grant No. 1351/12)Israeli Centers of Research Excellence (I-CORE grant, program 51/11)MINERVA Foundation (Germany

Development Of Carbon Based Neural Interface For Neural Stimulation/recording And Neurotransmitter Detection

Electrical stimulation and recording of neural cells have been widely used in basic neuroscience studies, neural prostheses, and clinical therapies. Stable neural interfaces that effectively communicate with the nervous system via electrodes are of great significance. Recently, flexible neural interfaces that combine carbon nanotubes (CNTs) and soft polymer substrates have generated tremendous interests. CNT based microelectrode arrays (MEAs) have shown enhanced electrochemical properties compared to commonly used electrode materials such as tungsten, platinum or titanium nitride. On the other hand, the soft polymer substrate can overcome the mechanical mismatch between the traditional rigid electrodes (or silicon shank) and the soft tissues for chronic use. However, most fabrication techniques suffer from low CNT yield, bad adhesion, and limited controllability. In addition, the electrodes were covered by randomly distributed CNTs in most cases. In this study, a novel fabrication method combining XeF2 etching and parylene deposition was presented to integrate the high quality vertical CNTs grown at high temperature with the heat sensitive parylene substrate in a highly controllable manner. Lower stimulation threshold voltage and higher signal to noise ratio have been demonstrated using vertical CNTs bundles compared to a Pt electrode and other randomly distributed CNT films. Adhesion has also been greatly improved. The work has also been extended to develop cuff shaped electrode for peripheral nerve stimulation. Fast scan cyclic voltammetry is an electrochemical detection technique suitable for in-vivo neurotransmitter detection because of the miniaturization, fast time response, good sensitivity and selectivity. Traditional single carbon fiber microelectrode has been limited to single detection for in-vivo application. Alternatively, pyrolyzed photoresist film (PPF) is a good candidate for this application as they are readily compatible with the microfabrication process for precise fabrication of microelectrode arrays. By the oxygen plasma treatment of photoresist prior to pyrolysis, we obtained carbon fiber arrays. Good sensitivity in dopamine detection by this carbon fiber arrays and improved adhesion have been demonstrated

Two Distinct Filopodia Populations at the Growth Cone Allow to Sense Nanotopographical Extracellular Matrix Cues to Guide Neurite Outgrowth

The process of neurite outgrowth is the initial step in producing the neuronal processes that wire the brain. Current models about neurite outgrowth have been derived from classic two-dimensional (2D) cell culture systems, which do not recapitulate the topographical cues that are present in the extracellular matrix (ECM) in vivo. Here, we explore how ECM nanotopography influences neurite outgrowth