A synthetic, three-dimensional bone marrow hydrogel [preprint]

Three-dimensional (3D) synthetic hydrogels have recently emerged as desirable in vitro cell culture platforms capable of representing the extracellular geometry, elasticity, and water content of tissue in a tunable fashion. However, they are critically limited in their biological functionality. Hydrogels are typically decorated with a scant 1-3 peptide moieties to direct cell behavior, which vastly underrepresents the proteins found in the extracellular matrix (ECM) of real tissues. Further, peptides chosen are ubiquitous in ECM, and are not derived from specific proteins. We developed an approach to incorporate the protein complexity of specific tissues into the design of biomaterials, and created a hydrogel with the elasticity of marrow, and 20 marrow-specific cell-instructive peptides. Compared to generic PEG hydrogels, our marrow-inspired hydrogel improves stem cell differentiation and proliferation. We propose this tissue-centric approach as the next generation of 3D hydrogel design for applications in tissue engineering

The Predictive Link between Matrix and Metastasis

Cancer spread (metastasis) is responsible for 90% of cancer-related fatalities. Informing patient treatment to prevent metastasis, or kill all cancer cells in a patient\u27s body before it becomes metastatic is extremely powerful. However, aggressive treatment for all non-metastatic patients is detrimental, both for quality of life concerns, and the risk of kidney or liver-related toxicity. Knowing when and where a patient has metastatic risk could revolutionize patient treatment and care. In this review, we attempt to summarize the key work of engineers and quantitative biologists in developing strategies and model systems to predict metastasis, with a particular focus on cell interactions with the extracellular matrix (ECM), as a tool to predict metastatic risk and tropism

A Biomaterial Screening Approach Reveals Microenvironmental Mechanisms of Drug Resistance

Traditional drug screening methods lack features of the tumor microenvironment that contribute to resistance. Most studies examine cell response in a single biomaterial platform in depth, leaving a gap in understanding how extracellular signals such as stiffness, dimensionality, and cell–cell contacts act independently or are integrated within a cell to affect either drug sensitivity or resistance. This is critically important, as adaptive resistance is mediated, at least in part, by the extracellular matrix (ECM) of the tumor microenvironment. We developed an approach to screen drug responses in cells cultured on 2D and in 3D biomaterial environments to explore how key features of ECM mediate drug response. This approach uncovered that cells on 2D hydrogels and spheroids encapsulated in 3D hydrogels were less responsive to receptor tyrosine kinase (RTK)-targeting drugs sorafenib and lapatinib, but not cytotoxic drugs, compared to single cells in hydrogels and cells on plastic. We found that transcriptomic differences between these in vitro models and tumor xenografts did not reveal mechanisms of ECM-mediated resistance to sorafenib. However, a systems biology analysis of phospho-kinome data uncovered that variation in MEK phosphorylation was associated with RTK-targeted drug resistance. Using sorafenib as a model drug, we found that co-administration with a MEK inhibitor decreased ECM-mediated resistance in vitro and reduced in vivo tumor burden compared to sorafenib alone. In sum, we provide a novel strategy for identifying and overcoming ECM-mediated resistance mechanisms by performing drug screening, phospho-kinome analysis, and systems biology across multiple biomaterial environments

A cell–ECM screening method to predict breast cancer metastasis

Breast cancer preferentially spreads to the bone, brain, liver, and lung. The clinical patterns of this tissue-specific spread (tropism) cannot be explained by blood flow alone, yet our understanding of what mediates tropism to these physically and chemically diverse tissues is limited. While the micro- environment has been recognized as a critical factor in governing metastatic colonization, the role of the extracellular matrix (ECM) in mediating tropism has not been thoroughly explored. We created a simple biomaterial platform with systematic control over the ECM protein density and composition to determine if integrin binding governs how metastatic cells differentiate between secondary tissue sites. Instead of examining individual behaviors, we compiled large patterns of phenotypes associated with adhesion to and migration on these controlled ECMs. In combining this novel analysis with a simple biomaterial platform, we created an in vitro fingerprint that is predictive of in vivo metastasis. This rapid biomaterial screen also provided information on how b1, a2, and a6 integrins might mediate metastasis in patients, providing insights beyond a purely genetic analysis. We propose that this approach of screening many cell–ECM interactions, across many different heterogeneous cell lines, is predictive of in vivo behavior, and is much simpler, faster, and more economical than complex 3D environments or mouse models. We also propose that when specifically applied toward the question of tissue tropism in breast cancer, it can be used to provide insight into certain integrin subunits as therapeutic targets

A global model for predicting the arrival of imported dengue infections

With approximately half of the world's population at risk of contracting dengue, this mosquito-borne disease is of global concern. International travellers significantly contribute to dengue's rapid and large-scale spread by importing the disease from endemic into non-endemic countries. To prevent future outbreaks and dengue from establishing in non-endemic countries, knowledge about the arrival time and location of infected travellers is crucial. We propose a network model that predicts the monthly number of dengue-infected air passengers arriving at any given airport. We consider international air travel volumes to construct weighted networks, representing passenger flows between airports. We further calculate the probability of passengers, who travel through the international air transport network, being infected with dengue. The probability of being infected depends on the destination, duration and timing of travel. Our findings shed light onto dengue importation routes and reveal country-specific reporting rates that have been until now largely unknown. This paper provides important new knowledge about the spreading dynamics of dengue that is highly beneficial for public health authorities to strategically allocate the often limited resources to more efficiently prevent the spread of dengue.Comment: 32 pages, 20 figure

Control of thiol-maleimide reaction kinetics in PEG hydrogel networks

Michael-type addition reactions are widely used to polymerize biocompatible hydrogels. The thiol-maleimide modality achieves the highest macromer coupling efficiency of the reported Michael-type pairs, but the resulting hydrogel networks are heterogeneous, because polymerization is faster than the individual components can be manually mixed. The reactivity of the thiol dictates the overall reaction speed, which can be slowed in organic solvents and acidic buffers. Since these modifications also reduce the biocompatibility of resulting hydrogels, we investigated a series of biocompatible buff­ers and crosslinkers to decelerate gelation while maintaining high cell viability. We found that lowering the polymer weight percentage (wt%), buffer concentration, and pH slowed gelation kinetics, but crosslinking with an electronegative peptide was optimal for both kinetics and cell viability. Including a high glucose medium supplement in the polymer solvent buffer improved the viability of the cells being encapsulated without impacting gelation time. Slowing the speed of polymerization resulted in more uniform hydrogels, both in terms of visual inspection and the diffusion of small molecules through the network. However, reactions that were too slow resulted in non-uniform particle dispersion due to settling, thus there is a trade-off in hydrogel network uniformity versus cell distribution in the hydrogels when using these networks in cell applications

Anomalously Diffusing and Persistently Migrating Cells in 2D and 3D Culture Environments

Appropriately chosen descriptive models of cell migration in biomaterials will allow researchers to characterize and ultimately predict the movement of cells in engineered systems for a variety of applications in tissue engineering. The persistent random walk (PRW) model accurately describes cell migration on two-dimensional (2D) substrates. However, this model inherently cannot describe subdiffusive cell movement, i.e. migration paths in which the root mean square displacement increases more slowly than the square root of the time interval. Subdiffusivity is a common characteristic of cells moving in confined environments, such as three-dimensional (3D) porous scaffolds, hydrogel networks, and in vivo tissues. We demonstrate that a generalized anomalous diffusion (AD) model, which uses a simple power law to relate the mean square displacement (MSD) to time, more accurately captures individual cell migration paths across a range of engineered 2D and 3D environments than does the more commonly used PRW model. We used the AD model parameters to distinguish cell movement profiles on substrates with different chemokinetic factors, geometries (2D vs 3D), substrate adhesivities, and compliances. Although the two models performed with equal precision for superdiffusive cells, we suggest a simple AD model, in lieu of PRW, to describe cell trajectories in populations with a significant subdiffusive fraction, such as cells in confined, 3D environments

Mechanics of Intact Bone Marrow

The current knowledge of bone marrow mechanics is limited to its viscous properties, neglecting the elastic contribution of the extracellular matrix. To get a more complete view of the mechanics of marrow, we characterized intact yellow porcine bone marrow using three different, but complementary techniques: rheology, indentation, and cavitation. Our analysis shows that bone marrow is elastic, and has a large amount of intra- and inter-sample heterogeneity, with an effective Young’s modulus ranging from 0.25-24.7 kPa at physiological temperature. Each testing method was consistent across matched tissue samples, and each provided unique benefits depending on user needs. We recommend bulk rheology to capture the effects of temperature on tissue elasticity and moduli, indentation for quantifying local tissue heterogeneity, and cavitation rheology for mitigating destructive sample preparation. We anticipate the knowledge of bone marrow elastic properties for building in vitro models will elucidate mechanisms involved in disease progression and regenerative medicin

Complementary, Semi-automated Methods for Creating Multi-dimensional, PEG-based Biomaterials

Tunable biomaterials that mimic selected features of the extracellular matrix (ECM), such as its stiffness, protein composition, and dimensionality, are increasingly popular for studying how cells sense and respond to ECM cues. In the field, there exists a significant trade-off for how complex and how well these biomaterials represent the in vivo microenvironment, versus how easy they are to make and how adaptable they are to automated fabrication techniques. To address this need to integrate more complex biomaterials design with high-throughput screening approaches, we present several methods to fabricate synthetic biomaterials in 96-well plates and demonstrate that they can be adapted to semiautomated liquid handling robotics. These platforms include 1) glass bottom plates with covalently attached ECM proteins, and 2) hydrogels with tunable stiffness and protein composition with either cells seeded on the surface, or 3) laden within the three-dimensional hydrogel matrix. This study includes proof-of-concept results demonstrating control over breast cancer cell line phenotypes via these ECM cues in a semi-automated fashion. We foresee the use of these methods as a mechanism to bridge the gap between high-throughput cell-matrix screening and engineered ECM-mimicking biomaterials

Learning through linguistic citizenship: finding the “I” of the essay

In recent years, the South African higher education system has seen growing calls for broadened epistemic access, decolonised curricula and transformed institutions. Scholars across South Africa have taken up the challenge and are working on new theoretical approaches to teaching and learning in higher education. In this paper, we reflect on students’ experiences of a multilingual, multimodal module called Reimagining Multilingualisms, which was jointly offered by the Universities of the Western Cape and Stellenbosch in April and May of 2018. In this paper, we provide an overview of the module and the different types of activities it involved. We reflect on these experiences using the theoretical lenses of decolonial scholar Mignolo (2009) on the ‘locus of enunciation’, and Stroud (2018) on ‘Linguistic Citizenship’. We present extracts from focus group interviews with students from both campuses to illustrate the involvement of ‘the body’ in ‘knowing’ and the ways in which the module enabled different ‘voices’ to emerge. We focus particularly on the role played by students’ perceived ‘vulnerability’ in the transformative benefits of the module and discuss this by way of conclusion. In sum, we suggest how the centring of multilingualism and diversity – not only as core pedagogic principles, but also as a methodology for transformation – may be used to enhance access and recapture voice in the building of a more integrated and just society