Protocell Flow Reactors for Enzyme and Whole-Cell Mediated Biocatalysis.

The design and construction of continuous flow biochemical reactors comprising immobilized biocatalysts have generated great interest in the efficient synthesis of value-added chemicals. Living cells use compartmentalization and reaction-diffusion processes for spatiotemporal regulation of biocatalytic reactions, and implementing these strategies into continuous flow reactors can offer new opportunities in reactor design and application. Herein, the fabrication of protocell-based continuous flow reactors for enzyme and whole-cell mediated biocatalysis is demonstrated. Semipermeable membranized coacervate vesicles are employed as model protocells that spontaneously sequester enzymes or accumulate living bacteria to produce embodied microreactors capable of single- or multiple-step catalytic reactions. By packing millions of the enzyme/bacteria-containing coacervate vesicles in a glass column, a facile, cost-effective, and modular methodology capable of performing oxidoreductase, peroxidase and lipolytic reactions, enzyme-mediated L-DOPA synthesis, and whole-cell glycolysis under continuous flow conditions, is demonstrated. It is shown that the protocell-nested enzymes and bacterial cells exhibit enhanced activities and stability under deleterious operating conditions compared with their non-encapsulated counterparts. These results provide a step toward the engineering of continuous flow reactors based on cell-like microscale agents and offer opportunities in the development of green and sustainable industrial bioprocessing.

Functional Integration of Synthetic Cells into 3D Microfluidic Devices for Artificial Organ-on-Chip Technologies.

Microfluidics play a pivotal role in organ-on-chip technologies and in the study of synthetic cells, especially in the development and analysis of artificial cell models. However, approaches that use synthetic cells as integral functional components for microfluidic systems to shape the microenvironment of natural living cells cultured on-chip have not been explored. Here, we integrate colloidosome-based synthetic cells into 3D microfluidic devices, pioneering the concept of synthetic cell-based microenvironments for organs-on-chip. We devise methods to create dense and stable networks of silica colloidosomes, enveloped by supported lipid bilayers, within microfluidic channels. These networks promote receptor-ligand interactions with on-chip cultured cells. Furthermore, we introduce a technique for the controlled release of growth factors from the synthetic cells into the channels, using a calcium alginate-based hydrogel formation within the colloidosomes. To demonstrate the potential of the technology, we present a modular plug-and-play lymph-node-on-a-chip prototype that guides the expansion of primary human T cells by stimulating receptor ligands on the T cells and modulating their cytokine environment. This integration of synthetic cells into microfluidic systems offers a new direction for organ-on-chip technologies and suggests further avenues for exploration in potential therapeutic applications. This article is protected by copyright. All rights reserved.

Self-assembly of stabilized droplets from liquid-liquid phase separation for higher-order structures and functions.

Dynamic microscale droplets produced by liquid-liquid phase separation (LLPS) have emerged as appealing biomaterials due to their remarkable features. However, the instability of droplets limits the construction of population-level structures with collective behaviors. Here we first provide a brief background of droplets in the context of materials properties. Subsequently, we discuss current strategies for stabilizing droplets including physical separation and chemical modulation. We also discuss the recent development of LLPS droplets for various applications such as synthetic cells and biomedical materials. Finally, we give insights on how stabilized droplets can self-assemble into higher-order structures displaying coordinated functions to fully exploit their potentials in bottom-up synthetic biology and biomedical applications.

DNA as a universal chemical substrate for computing and data storage.

DNA computing and DNA data storage are emerging fields that are unlocking new possibilities in information technology and diagnostics. These approaches use DNA molecules as a computing substrate or a storage medium, offering nanoscale compactness and operation in unconventional media (including aqueous solutions, water-in-oil microemulsions and self-assembled membranized compartments) for applications beyond traditional silicon-based computing systems. To build a functional DNA computer that can process and store molecular information necessitates the continued development of strategies for computing and data storage, as well as bridging the gap between these fields. In this Review, we explore how DNA can be leveraged in the context of DNA computing with a focus on neural networks and compartmentalized DNA circuits. We also discuss emerging approaches to the storage of data in DNA and associated topics such as the writing, reading, retrieval and post-synthesis editing of DNA-encoded data. Finally, we provide insights into how DNA computing can be integrated with DNA data storage and explore the use of DNA for near-memory computing for future information technology and health analysis applications.

Superstructural ordering in self-sorting coacervate-based protocell networks.

Bottom-up assembly of higher-order cytomimetic systems capable of coordinated physical behaviours, collective chemical signalling and spatially integrated processing is a key challenge in the study of artificial multicellularity. Here we develop an interactive binary population of coacervate microdroplets that spontaneously self-sort into chain-like protocell networks with an alternating sequence of structurally and compositionally dissimilar microdomains with hemispherical contact points. The protocell superstructures exhibit macromolecular self-sorting, spatially localized enzyme/ribozyme biocatalysis and interdroplet molecular translocation. They are capable of topographical reconfiguration using chemical or light-mediated stimuli and can be used as a micro-extraction system for macroscale biomolecular sorting. Our methodology opens a pathway towards the self-assembly of multicomponent protocell networks based on selective processes of coacervate droplet-droplet adhesion and fusion, and provides a step towards the spontaneous orchestration of protocell models into artificial tissues and colonies with ordered architectures and collective functions.

Patterning of Protein-Sequestered Liquid-Crystal Droplets Using Acoustic Wave Trapping.

Development of spatially organized structures and understanding their role in controlling kinetics of multistep chemical reactions are essential for the successful design of efficient systems and devices. While studies that showcase different types of methodologies for the spatial organization of various colloidal systems are known, design and development of well-defined hierarchical assemblies of liquid-crystal (LC) droplets and subsequent demonstration of biological reactions using such assemblies still remain elusive. Here, we show reversible and reconfigurable one-dimensional (1D) assemblies of protein-bioconjugate-sequestered monodisperse LC droplets by combining microfluidics with noninvasive acoustic wave trapping technology. Tunable spatial geometries and lattice dimensions can be achieved in an aqueous medium comprising ≈19 or 62 µm LC droplets. Different assemblies of a mixed population of larger and smaller droplets sequestered with glucose oxidase (GOx) and horseradish peroxidase (HRP), respectively, exhibit spatially localized enzyme kinetics with higher initial rates of reaction compared with GOx/HRP cascades implemented in the absence of an acoustic field. This can be attributed to the direct substrate transfer/channeling between the two complementary enzymes in close proximity. Therefore, our study provides an initial step toward the fabrication of LC-based devices for biosensing applications.

Exergaming and education: a relational model for games selection and evaluation.

Exergaming, or technology-driven physical exercise, has gained popularity in recent years. Its applications include physical education, health promotion, and rehabilitation. Although studies have obtained promising results regarding the positive effects of exergaming, the outcomes of exergaming for different populations remain undetermined. Inconsistencies in the literature on this topic have multiple potential explanations, including the content and demand of the exergames and the capability of the exergamer. A model with a sound theoretical framework is required to facilitate matching between games and gamers. This article proposes a relational model based on a matrix of Bloom's taxonomy of learning domains and the performance components of exergames. Appropriate matching of the physical demands of an exergame and the ability of the exergamer would enhance the effective usage of exergaming for individuals with various needs. This theory-based exergame model is developed to promote the general development, physical status, and psychosocial well-being of students, older adults, and individuals with rehabilitation needs. This model may provide a resource for future research on the application, effectiveness, and design of exergaming.

A Photo-degradable Crosslinker for the Development of Light-responsive Protocell Membranes.

The achievement of light-responsive behaviours is an important target for protocell engineering to allow control of fundamental protocellular processes such as communication via diffusible chemical signals, shape changes or even motility at the flick of a switch. As a step towards this ambitious goal, here we describe the synthesis of a novel poly(ethylene glycol)-based crosslinker, reactive towards nucleophiles, that effectively degrades with UV light (405 nm). We demonstrate its utility for the fabrication of the first protocell membranes capable of light-induced disassembly, for the photo-generation of patterns of protocells, and for the modulation of protocell membrane permeability. Overall, our results not only open up new avenues towards the engineering of spatially organised, communicating networks of protocells, and of micro-compartmentalised systems for information storage and release, but also have important implications for other research fields such as drug delivery and soft materials chemistry.

Autonomic Integration in Nested Protocell Communities.

The self-driven organization of model protocells into higher-order nested cytomimetic systems with coordinated structural and functional relationships offers a step toward the autonomic implementation of artificial multicellularity. Here, we describe an endosymbiotic-like pathway in which proteinosomes are captured within membranized alginate/silk fibroin coacervate vesicles by guest-mediated reconfiguration of the host protocells. We demonstrate that interchange of coacervate vesicle and droplet morphologies through proteinosome-mediated urease/glucose oxidase activity produces discrete nested communities capable of integrated catalytic activity and selective disintegration. The self-driving capacity is modulated by an internalized fuel-driven process using starch hydrolases sequestered within the host coacervate phase, and structural stabilization of the integrated protocell populations can be achieved by on-site enzyme-mediated matrix reinforcement involving dipeptide supramolecular assembly or tyramine-alginate covalent cross-linking. Our work highlights a semi-autonomous mechanism for constructing symbiotic cell-like nested communities and provides opportunities for the development of reconfigurable cytomimetic materials with structural, functional, and organizational complexity.

DNA storage in thermoresponsive microcapsules for repeated random multiplexed data access.

DNA has emerged as an attractive medium for archival data storage due to its durability and high information density. Scalable parallel random access to information is a desirable property of any storage system. For DNA-based storage systems, however, this still needs to be robustly established. Here we report on a thermoconfined polymerase chain reaction, which enables multiplexed, repeated random access to compartmentalized DNA files. The strategy is based on localizing biotin-functionalized oligonucleotides inside thermoresponsive, semipermeable microcapsules. At low temperatures, microcapsules are permeable to enzymes, primers and amplified products, whereas at high temperatures, membrane collapse prevents molecular crosstalk during amplification. Our data show that the platform outperforms non-compartmentalized DNA storage compared with repeated random access and reduces amplification bias tenfold during multiplex polymerase chain reaction. Using fluorescent sorting, we also demonstrate sample pooling and data retrieval by microcapsule barcoding. Therefore, the thermoresponsive microcapsule technology offers a scalable, sequence-agnostic approach for repeated random access to archival DNA files.

Signal Transduction in Artificial Cells.

In recent years, significant progress has been made in the emerging field of constructing biomimetic soft compartments with life-like behaviors. Given that biological activities occur under a flux of energy and matter exchange, the implementation of rudimentary signaling pathways in artificial cells (protocells) is a prerequisite for the development of adaptive sense-response phenotypes in cytomimetic models. Herein, recent approaches to the integration of signal transduction modules in model protocells prepared by bottom-up construction are discussed. The approaches are classified into two categories involving invasive biochemical signals or non-invasive physical stimuli. In the former mechanism, transducers with intrinsic recognition capability respond with high specificity, while in the latter, artificial cells respond through intra-protocellular energy transduction. Although major challenges remain in the pursuit of a sophisticated artificial signaling network for the orchestration of higher-order cytomimetic models, significant advances have been made in establishing rudimentary protocell communication networks, providing novel organizational models for the development of life-like microsystems and new avenues in protoliving technologies.

Algal cell bionics as a step towards photosynthesis-independent hydrogen production.

The engineering and modulation of living micro-organisms is a key challenge in green bio-manufacturing for the development of sustainable and carbon-neutral energy technologies. Here, we develop a cellular bionic approach in which living algal cells are interfaced with an ultra-thin shell of a conductive polymer along with a calcium carbonate exoskeleton to produce a discrete cellular micro-niche capable of sustained photosynthetic and photosynthetic-independent hydrogen production. The surface-augmented algal cells induce oxygen depletion, conduct photo-induced extracellular electrons, and provide structural and chemical stability that collectively give rise to localized hypoxic conditions and concomitant hydrogenase activity under daylight in air. We show that assembly of the living cellular micro-niche opens a direct extracellular photoelectron pathway to hydrogenase resulting in photosynthesis-independent hydrogen evolution for 200 d. In addition, surface-conductive dead algal cells continue to produce hydrogen for up to 8 d due to their structural stability and retention of functional hydrogenases. Overall, the integration of artificial biological hydrogen production pathways and natural photosynthesis in surface-augmented algal cells provides a cellular bionic approach to enhanced green hydrogen production under environmentally benign conditions and could pave the way to new opportunities in sustainable energy production.

Total hip arthroplasty for displaced femoral neck fracture: Survey of orthopaedic surgeons in Ontario, Canada.

BACKGROUND: Total hip arthroplasty (THA) for displaced femoral neck fractures in older patients remains a controversial topic. This study describes patient and surgeon factors that are associated with surgeons' recommendation of THA for this patient population. Furthermore, this study explores surgeon perceptions on why most patients are treated with hemiarthroplasty over THA. METHODS: In October 2019, a cross-sectional survey was mailed to practicing orthopaedic surgeons in Ontario, Canada. The questionnaire included paper patient cases to capture surgical practice variation using a full factorial, vignette-based experimental design. Multilevel linear regression and multivariable linear regression were used to determine patient and surgeon factors that are associated with treatment recommendations. RESULTS: Of a target population of 494 practicing surgeons, 302 (61.1%) responded. Sixty percent of respondents worked in the community, and most respondents (89.4%) had fellowship training. Surgeon-level predictors of treatment with THA included higher volume of THA for fracture in the last 12 months, having an elective THA practice, and increasing years in practice. Pre-existing hip arthritis increased likelihood to recommend THA, while increasing patient age and comorbidity burden decreased likelihood to recommend THA. There are medical, institutional, financial, and historic reasons why most patients are treated with hemiarthroplasty over THA. INTERPRETATION: This survey identified several patient and surgeon-level factors that were associated with treatment recommendation for THA. Hemiarthroplasty remains the more common treatment for this patient population for multiple reasons. There is potential for differential access to care when the factors driving treatment decisions are unrelated to the patient.

Chemical Communication and Protocell-Matrix Dynamics in Segregated Colloidosome Micro-Colonies.

Despite an emerging catalogue of collective behaviours in communities of homogeneously distributed cell-like objects, microscale protocell colonies with spatially segregated populations have received minimal attention. Here, we use microfluidics to fabricate Janus-like calcium alginate hydrogel microspheres with spatially partitioned populations of enzyme-containing inorganic colloidosomes and investigate their potential as integrated platforms for domain-mediated chemical communication and programmable protocell-matrix dynamics. Diffusive chemical signalling within the segregated communities gives rise to increased initial enzyme kinetics compared with a homogeneous distribution of protocells. We employ competing enzyme-mediated hydrogel crosslinking and decrosslinking reactions in different domains of the partitioned colonies to undertake selective expulsion of a specific protocell population from the community. Our results offer new possibilities for the design and construction of spatially organized cytomimetic consortia capable of endogenous chemical processing and protocell-environment interactivity.

Osmotic-Induced Reconfiguration and Activation in Membranized Coacervate-Based Protocells.

The design and construction of synthetic protocells capable of stimuli response and homeostatic regulation is an important challenge for synthetic protobiology. Here, we develop a step toward the construction of model protocells capable of a hypotonic stress-induced volume response that facilitates an increase in membrane permeability and the triggering of endogenous enzyme reactions. We describe a facile self-transformation process for constructing single- or multichambered molecularly crowded protocells based on the osmotic reconfiguration of lipid-coated coacervate droplets into multicompartmentalized coacervate vesicles. Hypotonic swelling broadens membrane permeability and increases transmembrane transport such that protease-based hydrolysis and enzyme cascades can be triggered and enhanced within the protocells by osmotically induced expansion. Specifically, we demonstrate how the enhanced production of nitric oxide (NO) within the swollen coacervate vesicles can be used to induce in vitro blood vessel vasodilation in thoracic artery rings. Our approach provides opportunities for designing reconfigurable model protocells capable of homeostatic volume regulation, dynamic structural reorganization, and adaptive functionality in response to changes in environment osmolarity, and could find applications in biomedicine, cellular diagnostics, and bioengineering.

Acoustic Trapping: An Emerging Tool for Microfabrication Technology.

The high throughput deposition of microscale objects with precise spatial arrangement represents a key step in microfabrication technology. This can be done by creating physical boundaries to guide the deposition process or using printing technologies; in both approaches, these microscale objects cannot be further modified after they are formed. The utilization of dynamic acoustic fields offers a novel approach to facilitate real-time reconfigurable miniaturized systems in a contactless manner, which can potentially be used in physics, chemistry, biology, as well as materials science. Here, the physical interactions of microscale objects in an acoustic pressure field are discussed and how to fabricate different acoustic trapping devices and how to tune the spatial arrangement of the microscale objects are explained. Moreover, different approaches that can dynamically modulate microscale objects in acoustic fields are presented, and the potential applications of the microarrays in biomedical engineering, chemical/biochemical sensing, and materials science are highlighted alongside a discussion of future research challenges.

Total hip arthroplasty versus hemiarthroplasty for treatment of femoral neck fractures : a population-based analysis of practice variation in Ontario, Canada.

AIMS: This study aimed to describe practice variation in the use of total hip arthroplasty (THA) for older patients with femoral neck fracture and to determine the association between patient, surgeon, and institution factors and treatment with THA. METHODS: We performed a cross-sectional analysis of 49,597 patients aged 60 years and older from Ontario, Canada, who underwent hemiarthroplasty or THA for femoral neck fracture between 2002 and 2017. This population-based study used routinely collected healthcare databases linked through ICES (formerly known as the Institute for Clinical Evaluative Sciences). Multilevel logistic regression modelling was used to quantify the association between patient, surgeon, and institution-level variables and whether patients were treated with THA. Variance partition coefficient and median odds ratios were used to estimate the variation attributable to higher-level variables and the magnitude of effect of higher-level variables, respectively. RESULTS: Over the study period, 9.4% of patients (n = 4,638) were treated with THA. Patient factors associated with higher likelihood of treatment by THA included: younger age, male sex, and diagnosis with rheumatoid arthritis. Long-term care residence, use of home care services prior to hip fracture, diagnosis of dementia, higher comorbidity burden, and the most marginalized group were negatively associated with treatment by THA. Treating surgeon and institution accounted for 54.2% and 17.8% of the total variation in treatment with THA, respectively. Surgeon volume of THA procedures in the 365 days prior to surgery was the strongest higher-level predictor of treatment with THA. Specific treating surgeons and institutions still accounted for significant proportions of the variability in treatment with THA (40.3% and 19.5% of total observed variation, respectively) after controlling for available patient, surgeon, and institution-level variables. CONCLUSION: The strongest predictors for treatment of patients with femoral neck fracture with THA were patient age, treating surgeon, and treating institution. This practice variation highlights differential access to care for patients.Cite this article: Bone Joint J 2023;105-B(2):180-189.

Modular Metal-Quinone Networks with Tunable Architecture and Functionality.

Nanostructured materials with tunable structures and functionality are of interest in diverse areas. Herein, metal ions are coordinated with quinones through metal-acetylacetone coordination bonds to generate a class of structurally tunable, universally adhesive, hydrophilic, and pH-degradable materials. A library of metal-quinone networks (MQNs) is produced from five model quinone ligands paired with nine metal ions, leading to the assembly of particles, tubes, capsules, and films. Importantly, MQNs show bidirectional pH-responsive disassembly in acidic and alkaline solutions, where the quinone ligands mediate the disassembly kinetics, enabling temporal and spatial control over the release of multiple components using multilayered MQNs. Leveraging this tunable release and the inherent medicinal properties of quinones, MQN prodrugs with a high drug loading (>89 wt %) are engineered using doxorubicin for anti-cancer therapy and shikonin for the inhibition of the main protease in the SARS-CoV-2 virus.

Comparative Effectiveness of Total Hip Arthroplasty and Hemiarthroplasty for Femoral Neck Fracture: A Propensity-Score-Matched Cohort Study.

BACKGROUND: The optimal treatment of older patients with a displaced femoral neck fracture remains a controversial topic. This study aimed to compare clinical outcomes across a matched group of patients with a femoral neck fracture treated with either hemiarthroplasty or total hip arthroplasty (THA). METHODS: Routinely collected health-care databases were linked to create a population-based cohort of 49,597 patients ≥60 years old from Ontario, Canada, who underwent hemiarthroplasty or THA for a femoral neck fracture between 2002 and 2017. A propensity-score-matched cohort was created using relevant and available predictors of treatment assignment and outcomes of interest. Clinical outcomes consisting of hip dislocation, revision surgery, hospital readmission, and death were compared in the matched cohort using survival analysis. RESULTS: Over 99% of THA patients (4,612) were adequately matched 1:1 to hemiarthroplasty patients (total matched cohort = 9,224). Patients treated with THA were at higher risk for hip dislocation at 30 days and 1 and 2 years postoperatively (2-year risk, 1.8% for THA versus 0.8% for hemiarthroplasty; p < 0.001). There was no difference in the short-term (30-day) or long-term (up to 10-year) risk of revision surgery between treatment groups. There was no significant difference in the risk of 30-day hospital readmission between groups. The risk of death at 1 year and 2 years postoperatively was lower for patients treated with THA. CONCLUSIONS: For patients with a hip fracture, shared decision-making should involve discussion of the potential higher risk of short-term hip dislocation after THA compared with hemiarthroplasty. The risk of revision surgery was similar between treatment groups at up to 10 years of follow-up. LEVEL OF EVIDENCE: Therapeutic Level III . See Instructions for Authors for a complete description of levels of evidence.