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The Arp 2/3 complex, a nucleator of branched actin, is shown to provide multiple distinct mechanical advantages to migrating glioblastoma cells. These include both decreased substrate stress and increased parallel focal adhesion interactions, preventing failure of cell–-substrate interactions.

Heart disease is one of the largest burdens to human health worldwide and has very limited therapeutic options. Engineered three-dimensional (3D) vascularized cardiac tissues have shown promise in rescuing cardiac function in diseased hearts and may serve as a whole organ replacement in the future. One of the major obstacles in reconstructing these thick myocardial tissues to a clinically applicable scale is the integration of functional vascular networks capable of providing oxygen and nutrients throughout whole engineered constructs. Without perfusion of oxygen and nutrient flow throughout the entire engineered tissue not only is tissue viability compromised, but also overall tissue functionality is lost. There are many supporting technologies and approaches that have been developed to create vascular networks such as 3D bioprinting, co-culturing hydrogels, and incorporation of soluble angiogenic factors. In this state-of-the-art review, we discuss some of the most current engineered vascular cardiac tissues reported in the literature and future directions in the field. IMPACT STATEMENT: The field of cardiac tissue engineering is rapidly evolving and is now closer than ever to having engineered tissue models capable of predicting preclinical responses to therapeutics, modeling diseases, and being used as a means of rescuing cardiac function following injuries to the native myocardium. However, a major obstacle of engineering thick cardiac tissue remains to be the integration of functional vasculature. In this review, we highlight seminal and recently published works that have influenced and pushed the field of cardiac tissue engineering toward achieving vascularized functional tissues.

Summary Research in low Earth orbit (LEO) has become more accessible. The 2020 Biomanufacturing in Space Symposium reviewed space-based regenerative medicine research and discussed leveraging LEO to advance biomanufacturing for regenerative medicine applications. The symposium identified areas where financial investments could stimulate advancements overcoming technical barriers. Opportunities in disease modeling, stem-cell-derived products, and biofabrication were highlighted. The symposium will initiate a roadmap to a sustainable market for regenerative medicine biomanufacturing in space. This perspective summarizes the 2020 Biomanufacturing in Space Symposium, highlights key biomanufacturing opportunities in LEO, and lays the framework for a roadmap to regenerative medicine biomanufacturing in space., Sharma and colleagues recap the 2020 Biomanufacturing in Space Symposium, which reviewed space-based regenerative medicine research and discussed opportunities to leverage low Earth orbit (LEO) to advance biomanufacturing for regenerative medicine. This perspective will highlight key biomanufacturing opportunities in LEO, note current technical gaps, and discuss next steps in developing a roadmap to biomanufacturing in space for regenerative medicine applications.

Poly(dimethylsiloxane) (PDMS) is a commonly used polymer in organ-on-a-chip devices and microphysiological systems. However, due to its hydrophobicity and permeability, it absorbs drug compounds, preventing accurate drug screening applications. Here, we developed an effective and facile method to prevent the absorption of drugs by utilizing a PDMS-PEG block copolymer additive and drug pretreatment. First, we incorporated a PDMS-PEG block copolymer into PDMS to address its inherent hydrophobicity. Next, we addressed the permeability of PDMS by eliminating the concentration gradient via pretreatment of the PDMS with the drug prior to experimentally testing drug absorption. The combined use of a PDMS-PEG block copolymer with drug pretreatment resulted in a mean reduction of drug absorption by 91.6% in the optimal condition. Finally, we demonstrated that the proposed method can be applied to prevent drug absorption in a PDMS-based cardiac microphysiological system, enabling more accurate drug studies.

Skeletal muscle atrophy is a well-known consequence of spaceflight. Because of the potential significant impact of muscle atrophy and muscle dysfunction on astronauts and their mission, a thorough understanding of the mechanisms of this atrophy and the development of effective countermeasures is critical. Spaceflight-induced muscle atrophy is similar to atrophy seen in many terrestrial conditions, and therefore our understanding of this form of atrophy may also contribute to the treatment of atrophy in humans on Earth. The unique environmental features humans encounter in space include the weightlessness of microgravity, space radiation, and the distinctive aspects of living in a spacecraft. The disuse and unloading of muscles in microgravity are likely the most significant factors that mediate spaceflight-induced muscle atrophy and have been extensively studied and reviewed. However, there are numerous other direct and indirect effects on skeletal muscle that may be contributing factors to the muscle atrophy and dysfunction seen as a result of spaceflight. This review offers a novel perspective on the issue of muscle atrophy in space by providing a comprehensive overview of the unique aspects of the spaceflight environment and the various ways in which they can lead to muscle atrophy. We systematically review the potential contributions of these different mechanisms of spaceflight-induced atrophy and include findings from both actual spaceflight and ground-based models of spaceflight in humans, animals, and in vitro studies.

Migration of meiosis-I (MI) spindle from the cell center to a sub-cortical location is a critical step for mouse oocytes to undergo asymmetric meiotic cell division. In this study, we investigate the mechanism by which formin-2 (FMN2) orchestrates the initial movement of MI spindle. By defining protein domains responsible for targeting FMN2, we show that spindle-periphery localized FMN2 is required for spindle migration. The spindle-peripheral FMN2 nucleates short actin bundles from vesicles derived likely from the endoplasmic reticulum (ER) and concentrated in a layer outside the spindle. This layer is in turn surrounded by mitochondria. A model based on polymerizing actin filaments pushing against mitochondria, thus generating a counter force on the spindle, demonstrated an inherent ability of this system to break symmetry and evolve directional spindle motion. The model is further supported through experiments involving spatially biasing actin nucleation via optogenetics and disruption of mitochondrial distribution and dynamics., Mammalian oocytes divide asymmetrically during meiotic maturation. Here, the authors show that spindle movement away from oocyte center depends on actin filaments nucleated from the spindle periphery pushing against surrounding mitochondria, which polarizes spontaneously to produce directional spindle motion.

In humankind’s endeavor to explore beyond our planet and travel further into space, we are now at the threshold of an era in which it is possible to move to and from low Earth orbit (LEO) with increasing ease and reduced cost. Through the International Space Station (ISS) U.S. National Laboratory, investigators from industry, academia, and government can easily access the unique LEO environment on the ISS to conduct research and development (R&D) activities in ways not possible on Earth. A key advantage of the LEO environment for life sciences research is the ability to conduct experiments in sustained microgravity conditions. The ability to conduct long-term research in microgravity enables opportunities for novel, fundamental studies in tissue engineering and regenerative medicine, including research on stem cell proliferation and differentiation, biofabrication, and disease modeling using microphysiological systems (MPS) that build on prior research using simulated microgravity conditions (Grimm, D., et al. 2018). Over the last decade, space-based research has demonstrated that microgravity informs our knowledge of fundamental biology and accelerates advancements in health care and medical technologies (International Space Station 2019). The benefits provided by conducting biomedical research in LEO may lead to breakthroughs not achievable on Earth. We are now at a transition point, in which nations are changing their approach to space-based R&D. The focus is shifting from government-funded fundamental science toward the expansion of privately funded R&D with terrestrial application and economic value that will drive a robust marketplace for innovation and manufacturing in LEO. Making this long-term transition requires public-private participation and near-term funding to support critical R&D to leverage the benefits of the LEO environment and de-risk space-based research. Studies conducted on the ISS over the past several years have indicated that one area with potential significant economic value and benefit to life on Earth is space-based biomanufacturing, or the use of biological and nonbiological materials to produce commercially relevant biomolecules and biomaterials for use in preclinical, clinical, and therapeutic applications. We must take advantage of the remaining lifetime of the ISS as a valuable LEO platform to demonstrate this economic value and Earth benefit. By facilitating access to the space station, the ISS National Lab is uniquely positioned to enable the R&D necessary to bridge the gap between the initial discovery phase of space-based biomedical research and the development of a sustainable, investment-worthy biomanufacturing market in LEO supported by future commercial platforms. Through a joint effort, the Center for the Advancement of Science in Space (CASIS), which manages the ISS National Lab, and the University of Pittsburgh’s McGowan Institute for Regenerative Medicine brought together thought leaders from around the U.S. for a Biomanufacturing in Space Symposium that consisted of a series of working sessions to review data from past space-based tissue engineering and regenerative medicine research, discuss relevant current space-based R&D in this area, and consider potential future markets to address the questions: What are the most promising opportunities to leverage the ISS to advance space-based biomanufacturing moving forward? What are the current gaps or barriers that, if overcome, could clear pathways toward private investment in LEO as a valued site for research, development, and production activity? And, most importantly: For which opportunities do the most compelling value propositions exist? The goal of the Biomanufacturing in Space Symposium was to help identify the specific areas in which government and industry investment would be most likely to stimulate advancements that overcome barriers. This would lead to a more investment-ready landscape for private interests to enter the market and fuel exponential growth. The symposium was meant to serve as the first step in developing a roadmap to a sustainable market for biomanufacturing in space. The symposium identified and prioritized multiple key R&D opportunities to advance space-based biomanufacturing. These opportunities fall in the areas of disease modeling, stem cells and stem-cell-derived products, and biofabrication. Additionally, symposium participants highlighted the critical need for additional data to help validate and de-risk these opportunities and concluded that approaches such as automation, artificial intelligence (AI), and machine learning will be needed to produce and capture the required data. Symposium participants also came to a consensus that public-private partnerships and funding will be needed to advance the opportunities toward a biomanufacturing marketplace in LEO. This paper will summarize the current state of the science and technology on the ISS and in the fields of tissue engineering and regenerative medicine; provide an overview of biomanufacturing R&D in space to date; review the goals of the Biomanufacturing in Space Symposium; highlight the key commercial opportunities and gaps identified during the symposium; provide information on potential market sizes; and briefly discuss the next steps in developing a roadmap to biomanufacturing in space.

Aneuploidy, which refers to unbalanced chromosome numbers, represents a class of genetic variation that is associated with cancer, birth defects and eukaryotic micro-organisms1–4. Whereas it is known that each aneuploid chromosome stoichiometry can give rise to a distinct pattern of gene expression and phenotypic profile4,5, it remains a fundamental question as to whether there are common cellular defects that are associated with aneuploidy. Here we show the existence in budding yeast of a common aneuploidy gene-expression signature that is suggestive of hypo-osmotic stress, using a strategy that enables the observation of common transcriptome changes of aneuploidy by averaging out karyotype-specific dosage effects in aneuploid yeast-cell populations with random and diverse chromosome stoichiometry. Consistently, aneuploid yeast exhibited increased plasma-membrane stress that led to impaired endocytosis, and this defect was also observed in aneuploid human cells. Thermodynamic modelling showed that hypo-osmotic-like stress is a general outcome of the proteome imbalance that is caused by aneuploidy, and also predicted a relationship between ploidy and cell size that was observed in yeast and aneuploid cancer cells. A genome-wide screen uncovered a general dependency of aneuploid cells on a pathway of ubiquitin-mediated endocytic recycling of nutrient transporters. Loss of this pathway, coupled with the endocytic defect inherent to aneuploidy, leads to a marked alteration of intracellular nutrient homeostasis. A common aneuploidy gene-expression signature is identified in yeast that is suggestive of hypo-osmotic stress, and which leads to cells that exhibit increased plasma-membrane stress and impaired endocytosis.

The ability of cells to sense and respond to mechanical cues from the surrounding environment has been implicated as a key regulator of cell differentiation, migration, and proliferation. The extracellular matrix (ECM) is an oft-overlooked component of the interface between cells and their surroundings. Cells assemble soluble ECM proteins into insoluble fibrils with unique mechanical properties that can alter the mechanical cues a cell receives. In this study, we construct a model that predicts the dynamics of cellular traction force generation and subsequent assembly of fibrils of the ECM protein fibronectin (FN). FN fibrils are the primary component in primordial ECM and, as such, FN assembly is a critical component in the cellular mechanical response. The model consists of a network of Hookean springs, each representing an extensible domain within an assembling FN fibril. As actomyosin forces stretch the spring network, simulations predict the resulting traction force and FN fibril formation. The model accurately predicts FN fibril morphometry and demonstrates a mechanism by which FN fibril assembly regulates traction force dynamics in response to mechanical stimuli and varying surrounding substrate stiffness.

High-grade gliomas are especially difficult tumors to treat due to their invasive behavior. This has led to extensive research focusing on arresting glioma cell migration. Cell migration involves the sensing of a migratory cue, followed by polarization in the direction of the cue, and reorganization of the actin cytoskeleton to allow for a protrusive leading edge and a contractile trailing edge. Transmission of these forces to produce motility also requires adhesive interactions of the cell with the extracellular microenvironment. In glioma cells, transmembrane receptors such as CD44 and integrins bind the cell to the surrounding extracellular matrix that provides a substrate on which the cell can exert the requisite forces for cell motility. These various essential parts of the migratory machinery are potential targets to halt glioma cell invasion. In this review, we discuss the mechanisms of glioma cell migration and how they may be targeted in anti-invasion therapies.

Cells respond to mechanical cues from the substrate to which they are attached. These mechanical cues drive cell migration, proliferation, differentiation, and survival. Previous studies have highlighted three specific mechanisms through which substrate stiffness directly alters cell function: increasing stiffness drives (1) larger contractile forces; (2) increased cell spreading and size; and (3) altered nuclear deformation. While studies have shown that substrate mechanics are an important cue, the role of the extracellular matrix (ECM) has largely been ignored. The ECM is a crucial component of the mechanosensing system for two reasons: (1) many ECM fibrils are assembled by application of cell-generated forces, and (2) ECM proteins have unique mechanical properties that will undoubtedly alter the local stiffness sensed by a cell. We specifically focused on the role of the ECM protein fibronectin (FN), which plays a critical role in de novo tissue production. In this study, we first measured the effects of substrate stiffness on human embryonic fibroblasts by plating cells onto microfabricated pillar arrays (MPAs) of varying stiffness. Cells responded to increasing substrate stiffness by generating larger forces, spreading to larger sizes, and altering nuclear geometry. These cells also assembled FN fibrils across all stiffnesses, with optimal assembly occurring at approximately 6 kPa. We then inhibited FN assembly, which resulted in dramatic reductions in contractile force generation, cell spreading, and nuclear geometry across all stiffnesses. These findings suggest that FN fibrils play a critical role in facilitating cellular responses to substrate stiffness.

The extracellular matrix (ECM) is an assembly of proteins that surround cells, and serves as the cell substrate in vivo. A primary component of newly synthesized ECM is the fibronectin (FN). Despite many years of research, the mechanism of FN assembly is still not completely understood. While it is recognized that FN assembly requires application of traction force to expose a buried FN-FN binding site, such a site has never been elucidated. We hypothesize that assembly of fibronectin (FN) fibrils is a complex event: each of the 15 Type III domains in FN is made up of a sandwich of 7 beta strands; when relaxed, the Type III domains are folded such that the beta strands are twisted, blocking the non-specific binding of other proteins. Application of force straightens these beta-strands, allowing for binding of other FNs via a beta-strand addition mechanism. This suggests that all 15 domains are capable of binding FN molecules in a growing fibril. To investigate this hypothesis, we present a mechanistic computational model of cell/FN/substrate biomechanical interactions, which accounts for the unique, nonlinear mechanical properties of each domain and the stochastic binding between molecular clutches and the moving actin bundle. Monte Carlo simulations predict that increasing substrate stiffness leads to longer and thicker fibrils. Additionally, the model demonstrates complex time-dependent dynamics governing the size of the growing fibril, the domains’ stiffness, and traction forces; that is, at low force, a small subset of domains open, allowing for minimal fibril assembly, while large forces unfold a considerable fraction of domains and create large, thick fibrils. Simulation outputs are compared to experimental data in which traction force, FN assembly, and domain opening are quantified using microfabricated pillar arrays and cysteine-labeling of recombinant FNs with introduced buried cysteines.