Three-dimensional (3D) hydrogels made from synthetic polymers have emerged as in vitro cell culture platforms capable of representing the extracellular geometry, modulus, and water content of tissues ...in a tunable fashion. Hydrogels made from these otherwise non-bioactive polymers can be decorated with short peptides derived from proteins naturally found in tissues to support cell viability and direct phenotype. We identified two key limitations that limit the ability of this class of materials to recapitulate real tissue. First, these environments typically display between 1 and 3 bioactive peptides, which vastly underrepresents the diversity of proteins found in the extracellular matrix (ECM) of real tissues. Second, peptides chosen are ubiquitous in ECM and not derived from proteins found in specific tissues, per se. To overcome this critical limitation in hydrogel design and functionality, we developed an approach to incorporate the complex and specific protein signature of bone marrow into a poly (ethylene glycol) (PEG) hydrogel. This bone marrow hydrogel mimics the elasticity of marrow and has 20 bone marrow-specific and cell-instructive peptides. We propose this tissue-centric approach as the next generation of 3D hydrogel design for applications in tissue engineering and beyond.
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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.
The polymer network of thiol-maleimide hydrogels assembles faster than individual components can be uniformly mixed due to their fast gelation kinetics. The lack of homogeneity can result in variable cell-based assay results, resulting in batch-to-batch variability and limiting their use in predictive screening assays. Although these hydrogels are incredibly useful in tissue engineering, this network heterogeneity is a known problem in the field. We screened a variety of possible techniques to slow down the reaction speed and improve the homogeneity of these hydrogels, without sacrificing the viability and distribution of encapsulated cells. As others have reported, an electronegative crosslinker was the most effective technique to slow the reaction, but the chemical modification required is technically challenging. Of interest to a broad community, we screened buffer type, strength, and crosslinker electronegativity to find an optimal reaction speed that allows for high cell viability and small molecule diffusion, without allowing cells to settle during gelation, allowing application of these materials to the drug screening industry and tissue engineering community.
Reducing the foreign body response (FBR) to implanted biomaterials will enhance their performance in tissue engineering. Poly(ethylene glycol) (PEG) hydrogels are increasingly popular for this ...application due to their low cost, ease of use, and the ability to tune their compliance via molecular weight and cross-linking densities. PEG hydrogels can elicit chronic inflammation in vivo, but recent evidence has suggested that extremely hydrophilic, zwitterionic materials and particles can evade the immune system. To combine the advantages of PEG-based hydrogels with the hydrophilicity of zwitterions, we synthesized hydrogels with comonomers PEG and the zwitterion phosphorylcholine (PC). Recent evidence suggests that stiff hydrogels elicit increased immune cell adhesion to hydrogels, which we attempted to reduce by increasing hydrogel hydrophilicity. Surprisingly, hydrogels with the highest amount of zwitterionic comonomer elicited the highest FBR. Lowering the hydrogel modulus (165 to 3 kPa), or PC content (20 to 0 wt %), mitigated this effect. A high density of macrophages was found at the surface of implants associated with a high FBR, and mass spectrometry analysis of the proteins adsorbed to these gels implicated extracellular matrix, immune response, and cell adhesion protein categories as drivers of macrophage recruitment. Overall, we show that modulus regulates macrophage adhesion to zwitterionic-PEG hydrogels, and demonstrate that chemical modifications to hydrogels should be studied in parallel with their physical properties to optimize implant design.
Mechanics of intact bone marrow Jansen, Lauren E.; Birch, Nathan P.; Schiffman, Jessica D. ...
Journal of the mechanical behavior of biomedical materials,
10/2015, Letnik:
50
Journal Article
Recenzirano
Odprti dostop
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 to 24.7kPa 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 medicine.
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•Bone marrow is a benign material with a large amount of intra- and inter-tissue heterogeneity.•The EEff of intact porcine marrow ranges from 0.3 to 24.7 kPa at physiological temperature.•Bulk rheology is the best method to capture the effects of temperature on tissue viscoelasticity.•Indentation is the ideal method for quantifying local tissue heterogeneity.•Cavitation mitigates destructive sample preparation, so tissue mechanics can be tested in situ.
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.
•Recombinant manganese peroxidase (rMnP), produced in the yeast Pichia pastoris, can degrade furfural and 5-hydroxymethyl-furfural (HMF), which are produced during biomass pretreatment and are ...inhibitory to yeast fermentations for biofuels production.•Treatment with rMnP or manganese(III) acetate reduced furfural and HMF concentrations in a dose-dependent manner.•Furfural disappearance was accompanied by malonate disappearance and accumulation of four distinct degradation products.•Growth assays using Saccharomyces cerevisiae, a commonly used yeast in biofuels fermentations, indicated that rMnP treatment reduced the toxicity of furfural and HMF.
Biomass pretreatment-derived degradation compounds, such as furfural and 5-hydroxymethyl-furfural (HMF), inhibit the growth of fermentation microorganisms that utilize biomass to produce fuels and chemicals. Here we report that recombinant manganese peroxidase (rMnP) produced from the yeast Pichia pastoris can degrade furfural and HMF making them less toxic to microorganisms. Treatment with rMnP or manganese(III) acetate reduced furfural and HMF concentrations in a dose-dependent manner. Furfural disappearance was accompanied by malonate disappearance and accumulation of four distinct degradation products. Furfural was more readily degraded by rMnP and manganese(III) acetate than HMF. Growth assays using Saccharomyces cerevisiae indicated that rMnP treatment reduced the toxicity of furfural and HMF. This work provides an avenue to use rMnP to increase the growth of fermentation microorganisms that are inhibited by toxic compounds derived from pretreatment of biomass.
During February 7─September 3, 2022, a total of 39 US states experienced outbreaks of highly pathogenic avian influenza A(H5N1) virus in birds from commercial poultry farms and backyard flocks. Among ...persons exposed to infected birds, highly pathogenic avian influenza A(H5) viral RNA was detected in 1 respiratory specimen from 1 person.
The B.1.1.529 (Omicron) variant of SARS-CoV-2 (the virus that causes COVID-19) was first detected in specimens collected on November 11, 2021, in Botswana and on November 14 in South Africa;* the ...first confirmed case of Omicron in the United States was identified in California on December 1, 2021 (1). On November 29, the Nebraska Department of Health and Human Services was notified of six probable cases
of COVID-19 in one household, including one case in a man aged 48 years (the index patient) who had recently returned from Nigeria. Given the patient's travel history, Omicron infection was suspected. Specimens from all six persons in the household tested positive for SARS-CoV-2 by reverse transcription-polymerase chain reaction (RT-PCR) testing on December 1, and the following day genomic sequencing by the Nebraska Public Health Laboratory identified an identical Omicron genotype from each specimen (Figure). Phylogenetic analysis was conducted to determine if this cluster represented an independent introduction of Omicron into the United States, and a detailed epidemiologic investigation was conducted. This activity was reviewed by CDC and was conducted consistent with applicable federal law and CDC policy.
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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 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.
The Engineering Education Research Network in Africa (EERN-Africa) was created to enable connections between practitioners and researchers with a shared interest in African engineering education ...contexts. Recognising the importance of developing an African voice in the engineering education research space, the EERN-Africa community has interacted in a dynamic and dialogic way with our own teaching and research practices across diverse African contexts, with an ethical commitment to democratic and inclusive community-building. The objective of this paper is to reflect on the current status of the Community of Practice (CoP), and the challenges and opportunities in sustaining and growing the CoP. A collaborative analysis of perspectives on this emerging identity is presented, using an Appreciative Inquiry (AI) methodology and drawing on collective written reflections and discussions. Six broad themes on the value that the CoP has for both individuals and the group were identified: networking, capacity development, emotional support, impact on professional identity, social and environmental impact, and breaking borders. This paper contributes an approach for collaborative capacity-building in EER through a virtual CoP, underpinned by the spirit of ubuntu.
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