PlGFMMP9-engineered iPS cells supported on a PEGfibrinogen hydrogel scaffold possess an enhanced capacity to repair damaged myocardium

Cell-based regenerative therapies are significantly improved by engineering allografts to express factors that increase vascularization and engraftment, such as placental growth factor (PlGF) and matrix metalloproteinase 9 (MMP9). Moreover, the seeding of therapeutic cells onto a suitable scaffold is of utmost importance for tissue regeneration. On these premises, we sought to assess the reparative potential of induced pluripotent stem (iPS) cells bioengineered to secrete PlGF or MMP9 and delivered to infarcted myocardium upon a poly(ethylene glycol)-fibrinogen scaffold. When assessing optimal stiffness of the PEG-fibrinogen (PF) scaffold, we found that the appearance of contracting cells after cardiogenic induction was accelerated on the support designed with an intermediate stiffness. Revascularization and hemodynamic parameters of infarcted mouse heart were significantly improved by injection into the infarct of this optimized PF scaffold seeded with both MiPS (iPS cells engineered to secrete MMP9) and PiPS (iPS cells engineered to secrete PlGF) cells as compared with nonengineered cells or PF alone. Importantly, allograft-derived cells and host myocardium were functionally integrated. Therefore, survival and integration of allografts in the ischemic heart can be significantly improved with the use of therapeutic cells bioengineered to secrete MMP9 and PlGF and encapsulated within an injectable PF hydrogel having an optimized stiffness

Quantification of the Temporal Evolution of Collagen Orientation in Mechanically Conditioned Engineered Cardiovascular Tissues

Load-bearing soft tissues predominantly consist of collagen and exhibit anisotropic, non-linear visco-elastic behavior, coupled to the organization of the collagen fibers. Mimicking native mechanical behavior forms a major goal in cardiovascular tissue engineering. Engineered tissues often lack properly organized collagen and consequently do not meet in vivo mechanical demands. To improve collagen architecture and mechanical properties, mechanical stimulation of the tissue during in vitro tissue growth is crucial. This study describes the evolution of collagen fiber orientation with culture time in engineered tissue constructs in response to mechanical loading. To achieve this, a novel technique for the quantification of collagen fiber orientation is used, based on 3D vital imaging using multiphoton microscopy combined with image analysis. The engineered tissue constructs consisted of cell-seeded biodegradable rectangular scaffolds, which were either constrained or intermittently strained in longitudinal direction. Collagen fiber orientation analyses revealed that mechanical loading induced collagen alignment. The alignment shifted from oblique at the surface of the construct towards parallel to the straining direction in deeper tissue layers. Most importantly, intermittent straining improved and accelerated the alignment of the collagen fibers, as compared to constraining the constructs. Both the method and the results are relevant to create and monitor load-bearing tissues with an organized anisotropic collagen network

Writing Russia's future: paradigms, drivers, and scenarios

The development of prediction and forecasting in the social sciences over the past century and more is closely linked with developments in Russia. The Soviet collapse undermined confidence in predictive capabilities, and scenario planning emerged as the dominant future-oriented methodology in area studies, including the study of Russia. Scenarists anticipate multiple futures rather than predicting one. The approach is too rarely critiqued. Building on an account of Russia-related forecasting in the twentieth century, analysis of two decades of scenarios reveals uniform accounts which downplay the insights of experts and of social science theory alike

Comparing political futures: the rise and use of scenarios in future-oriented area studies

The predictive ability of scholars of politics has long been a subject of theoretical debate and methodological development. In theoretical debate, prediction represents a central issue regarding the extent to which the study of politics is scientific. In methodological development, much effort and resource have been devoted to a diverse range of predictive approaches, with varying degrees of success. Expectations that scholars forecast accurately come as much from the policy and media worlds as from the academy. Since the end of the Cold War, scenario development has become prevalent in future-oriented research by area studies scholars. This approach is long due critical re-assessment. For all its strengths as a policy tool, scenario development tends towards a bounded methodology, driving the process of anticipating futures along predetermined paths into a standardised range of options, and paying insufficient attention to theoretical and contextual understandings available within the relevant scholarly disciplines

Screening out irrelevant cell-based models of disease

The common and persistent failures to translate promising preclinical drug candidates into clinical success highlight the limited effectiveness of disease models currently used in drug discovery. An apparent reluctance to explore and adopt alternative cell-and tissue-based model systems, coupled with a detachment from clinical practice during assay validation, contributes to ineffective translational research. To help address these issues and stimulate debate, here we propose a set of principles to facilitate the definition and development of disease-relevant assays, and we discuss new opportunities for exploiting the latest advances in cell-based assay technologies in drug discovery, including induced pluripotent stem cells, three-dimensional (3D) co-culture and organ-on-a-chip systems, complemented by advances in single-cell imaging and gene editing technologies. Funding to support precompetitive, multidisciplinary collaborations to develop novel preclinical models and cell-based screening technologies could have a key role in improving their clinical relevance, and ultimately increase clinical success rates

Visualizing cell-laden fibrin-based hydrogels using cryogenic scanning electron microscopy and confocal microscopy

The present investigation explores the microscopic aspects of cell-laden hydrogels at high resolutions, using three-dimensional cell cultures in semi-synthetic constructs that are of very high water content (>98% water). The study aims to provide an imaging strategy for these constructs, while minimizing artefacts. Constructs of poly(ethylene glycol)-fibrinogen and fibrin hydrogels containing embedded mesenchymal cells (human dermal fibroblasts) were first imaged by confocal microscopy. Next, high-resolution scanning electron microscopy (HR-SEM) was used to provide images of the cells within the hydrogels, at submicron resolutions. Because it was not possible to obtain artefact-free images of the hydrogels using room-temperature HR-SEM, a cryogenic HR-SEM imaging methodology was employed to visualize the sample while preserving the natural hydrated state of the hydrogel. The ultrastructural details of the constructs were observed at subcellular resolutions, revealing numerous cellular components, the biomaterial in its native configuration, and the uninterrupted cell membrane as it relates with the biomaterial in the hydrated state of the construct. Constructs containing microscopic albumin microbubbles were also imaged using these methodologies to reveal fine details of the interaction between the cells, the microbubbles, and the hydrogel. Taken together with the confocal microscopy, this imaging strategy provides a more complete picture of the hydrated state of the hydrogel network with cells inside. As such, this methodology addresses some of the challenges of obtaining this information in amorphous hydrogel systems containing a very high water content (>98%) with embedded cells. Such insight may lead to better hydrogel-based strategies for tissue engineering and regeneration

Fibrinogen-Based Hydrogel Modulus and Ligand Density Effects on Cell Morphogenesis in Two-Dimensional and Three-Dimensional Cell Cultures

There is a need to further explore the convergence of mechanobiology and dimensionality with systematic investigations of cellular response to matrix mechanics in 2D and 3D cultures. Here, a semisynthetic hydrogel capable of supporting both 2D and 3D cell culture is applied to investigate cell response to matrix modulus and ligand density. The culture materials are fabricated from adducts of polyethylene glycol (PEG) or PluronicF127 and fibrinogen fragments, formed into hydrogels by free-radical polymerization, and characterized by shear rheology. Control over the modulus of the materials is accomplished by changing the concentration of synthetic PEG-diacrylate crosslinker (0.5% w/v), and by altering the molecular length of the PEG (10 and 20 kDa). Control over ligand density is accomplished by changing fibrinogen concentrations from 3 to 12 mg mL-1 . In 2D culture, cell motility parameters, including cell speed and persistence time are significantly increased with increasing modulus. In both 2D and 3D culture, cells express vinculin and there is evidence of focal adhesion formation in the high stiffness materials. The modulus- and ligand-dependent morphogenesis response from the cells in 2D culture is contradictory to the same measured response in 3D culture. In 2D culture, anchorage-dependent cells become more elongated and significantly increase their size with increasing ligand density and matrix modulus. In 3D culture, the same anchorage-dependent cells become less spindled and significantly reduce their size in response to increasing ligand density and matrix modulus. These differences arise from dimensionality constraints, most notably the encapsulation of cells in a non-porous hydrogel matrix. These insights underscore the importance of mechanical properties in regulating cell morphogenesis in a 3D culture milieu. The versatility of the hydrogel culture environment further highlights the significance of a modular approach when developing materials that aim to optimize the cell culture environment

PlGFMMP9-engineered iPS cells supported on a PEGfibrinogen hydrogel scaffold possess an enhanced capacity to repair damaged myocardium

Cell-based regenerative therapies are significantly improved by engineering allografts to express factors that increase vascularization and engraftment, such as placental growth factor (PlGF) and matrix metalloproteinase 9 (MMP9). Moreover, the seeding of therapeutic cells onto a suitable scaffold is of utmost importance for tissue regeneration. On these premises, we sought to assess the reparative potential of induced pluripotent stem (iPS) cells bioengineered to secrete PlGF or MMP9 and delivered to infarcted myocardium upon a poly(ethylene glycol)fibrinogen scaffold. When assessing optimal stiffness of the PEGfibrinogen (PF) scaffold, we found that the appearance of contracting cells after cardiogenic induction was accelerated on the support designed with an intermediate stiffness. Revascularization and hemodynamic parameters of infarcted mouse heart were significantly improved by injection into the infarct of this optimized PF scaffold seeded with both MiPS (iPS cells engineered to secrete MMP9) and PiPS (iPS cells engineered to secrete PlGF) cells as compared with nonengineered cells or PF alone. Importantly, allograft-derived cells and host myocardium were functionally integrated. Therefore, survival and integration of allografts in the ischemic heart can be significantly improved with the use of therapeutic cells bioengineered to secrete MMP9 and PlGF and encapsulated within an injectable PF hydrogel having an optimized stiffness. © 2014 Macmillan Publishers Limited