Imperial College London

Professor Molly Stevens

Faculty of EngineeringDepartment of Materials

Professor of Biomedical Materials and Regenerative Medicine
 
 
 
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Contact

 

+44 (0)20 7594 6804m.stevens

 
 
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Location

 

208Royal School of MinesSouth Kensington Campus

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Summary

 

Publications

Publication Type
Year
to

408 results found

Maynard SA, Pchelintseva E, Zwi-Dantsis L, Nagelkerke A, Gopal S, Korchev YE, Shevchuk A, Stevens Met al., 2021, IL-1β mediated nanoscale surface clustering of integrin α5β1 regulates the adhesion of mesenchymal stem cells, Scientific Reports, ISSN: 2045-2322

Journal article

Xianyu Y, Lin Y, Chen Q, Belessiotis-Richards A, Stevens M, Thomas Met al., 2021, Iodide-Mediated Rapid and Sensitive Surface Etching of Gold Nanostars for Biosensing, Angewandte Chemie International Edition, ISSN: 1433-7851

Journal article

Kim N, Kim E, Kim H, Thomas M, Najer A, Stevens Met al., 2021, Tumor-targeting cholesterol-decorated DNA nanoflowers for intracellular ratiometric aptasensing, Advanced Materials, Vol: 33, Pages: 1-10, ISSN: 0935-9648

Probing endogenous molecular profiles is of fundamental importance to understand cellular function and processes. Despite the promise of programmable nucleic‐acid‐based aptasensors across the breadth of biomolecular detection, target‐responsive aptasensors enabling intracellular detection are as of yet infrequently realized. Several challenges remain, including the difficulties in quantification/normalization of quencher‐based intensiometric signals, stability issues of the probe architecture, and complex sensor operations often necessitating extensive structural modeling. Here, the biomimetic crystallization‐empowered self‐assembly of a tumor‐targetable DNA–inorganic hybrid nanocomposite aptasensor is presented, which enables Förster resonance energy transfer (FRET)‐based quantitative interpretation of changes in the cellular target abundance. Leveraging the design programmability and high‐throughput fabrication of rolling circle amplification‐driven DNA nanoarchitecture, this designer platform offers a method to self‐assemble a robust nanosensor from a multifunctionality‐encoded template that includes a cell‐targeting aptamer, a ratiometric aptasensor, and a cholesterol‐decorating element. Taking prostate cancer cells and intracellular adenosine triphosphate molecules as a model system, a synergistic effect in the targeted delivery by cholesterol and aptamers, and the feasibility of quantitative intracellular aptasensing are demonstrated. It is envisioned that this approach provides a highly generalizable strategy across wide‐ranging target systems toward a biologically deliverable nanosensor that enables quantitative monitoring of the abundance of endogenous biomolecules.

Journal article

Guagliardo R, Herman L, Penders J, Zamborlin A, De Keersmaecker H, Van de Vyver T, Verstraeten S, Merckx P, Mingeot-Leclercq M-P, Echaide M, Pérez-Gil J, Stevens MM, De Smedt SC, Raemdonck Ket al., 2021, Surfactant Protein B Promotes Cytosolic SiRNA Delivery by Adopting a Virus-like Mechanism of Action., ACS Nano

RNA therapeutics are poised to revolutionize medicine. To unlock the full potential of RNA drugs, safe and efficient (nano)formulations to deliver them inside target cells are required. Endosomal sequestration of nanocarriers represents a major bottleneck in nucleic acid delivery. Gaining more detailed information on the intracellular behavior of RNA nanocarriers is crucial to rationally develop delivery systems with improved therapeutic efficiency. Surfactant protein B (SP-B) is a key component of pulmonary surfactant (PS), essential for mammalian breathing. In contrast to the general belief that PS should be regarded as a barrier for inhaled nanomedicines, we recently discovered the ability of SP-B to promote gene silencing by siRNA-loaded and lipid-coated nanogels. However, the mechanisms governing this process are poorly understood. The major objective of this work was to obtain mechanistic insights into the SP-B-mediated cellular delivery of siRNA. To this end, we combined siRNA knockdown experiments, confocal microscopy, and focused ion beam scanning electron microscopy imaging in an in vitro non-small-cell lung carcinoma model with lipid mixing assays on vesicles that mimic the composition of (intra)cellular membranes. Our work highlights a strong correlation between SP-B-mediated fusion with anionic endosomal membranes and cytosolic siRNA delivery, a mode of action resembling that of certain viruses and virus-derived cell-penetrating peptides. Building on these gained insights, we optimized the SP-B proteolipid composition, which dramatically improved delivery efficiency. Altogether, our work provides a mechanistic understanding of SP-B-induced perturbation of intracellular membranes, offering opportunities to fuel the rational design of SP-B-inspired RNA nanoformulations for inhalation therapy.

Journal article

Kit-Anan W, Mazo M, Wang BX, Leonardo V, Pence I, Gopal S, Gelmi A, Becce M, Chiappini C, Harding SE, Terracciano C, Stevens Met al., 2021, Multiplexing physical stimulation on single human induced pluripotent stem cell-derived cardiomyocytes for phenotype modulation, Biofabrication, Vol: 13, Pages: 1-16, ISSN: 1758-5082

Traditional in vitro bioengineering approaches whereby only individual biophysical cues are manipulated at any one time are highly inefficient, falling short when recapitulating the complexity of the cardiac environment. Multiple biophysical cues are present in the native myocardial niche and are essential during development, as well as in maintenance of adult cardiomyocyte (CM) phenotype in both health and disease. This study establishes a novel biofabrication workflow to study and manipulate hiPSC-CMs and to understand how these cells respond to a multiplexed biophysical environment, namely microscopic topography (3D shape resembling that of adult CM) and substrate stiffness, at a single cell level. Silicon masters were fabricated and developed to generate pillars of the desired 3D shapes, which would be used to mould the designed microwell arrays into a hydrogel. Polyacrylamide was modified with the incorporation of acrylic acid to provide a carboxylic group conjugation site for adhesion motifs, without comprising its capacity to modulate the stiffness. In this manner, individual parameters can be finely tuned independently within the hydrogel: the dimension of 3D shaped microwell and its stiffness. The design allows the platform to isolate single hiPSC-CMs to study solely biophysical cues in an absence of cell-cell physical interaction. Under physiologic-like physical conditions (3D shape resembling that of adult CM and 9.83 kPa substrate stiffness), isolated single hiPSC-CMs exhibit increased Cx-43 density, cell Peer reviewed version of the manuscript published in final form at Biofabrication (2020). membrane stiffness and calcium transient amplitude; co-expression of the subpopulation-related MYL2- MYL7 proteins; while displaying higher anisotropism in comparison to pathologic-like conditions (flat surface and 112 kPa substrate stiffness). This demonstrates that supplying a physiological or pathological microenvironment to an isolated single hiPSC-CM in absen

Journal article

Pinna A, Baghbaderani MT, Hernandez VV, Naruphontjirakul P, Li S, McFarlane T, Hachim D, Stevens MM, Porter AE, Jones JRet al., 2021, Nanoceria provides antioxidant and osteogenic properties to mesoporous silica nanoparticles for osteoporosis treatment, ACTA BIOMATERIALIA, Vol: 122, Pages: 365-376, ISSN: 1742-7061

Journal article

Horgan C, Bergholt MS, Thin MZ, Nagelkerke A, Kennedy R, Kalber TL, Stuckey D, Stevens Met al., 2021, Image-guided Raman spectroscopy probe-tracking for tumor margin delineation, Journal of Biomedical Optics, ISSN: 1083-3668

Journal article

Lin Y, Penna M, Spicer CD, Higgins SG, Gelmi A, Kim N, Wang S-T, Wojciechowski JP, Pashuck ET, Yarovsky I, Stevens MMet al., 2021, High-throughput peptide derivatization toward supramolecular diversification in microtiter plates, ACS Nano, Vol: 15, Pages: 4034-4044, ISSN: 1936-0851

The evolution of life on earth eventually leads to the emergence of species with increased complexity and diversity. Similarly, evolutionary chemical space exploration in the laboratory is a key step to pursue the structural and functional diversity of supramolecular systems. Here, we present a powerful tool that enables rapid peptide diversification and employ it to expand the chemical space for supramolecular functions. Central to this strategy is the exploitation of palladium-catalyzed Suzuki-Miyaura cross-coupling reactions to direct combinatorial synthesis of peptide arrays in microtiter plates under an open atmosphere. Taking advantage of this in situ library design, our results unambiguously deliver a fertile platform for creating a set of intriguing peptide functions including green fluorescent protein-like peptide emitters with chemically encoded emission colors, hierarchical self-assembly into nano-objects, and macroscopic hydrogels. This work also offers opportunities for quickly surveying the diversified peptide arrays and thereby identifying the structural factors that modulate peptide properties.

Journal article

Li C, Ouyang L, Armstrong J, Stevens Met al., 2021, Advances in the fabrication of biomaterials for gradient tissue engineering, Trends in Biotechnology, Vol: 39, Pages: 150-164, ISSN: 0167-7799

Natural tissues and organs exhibit an array of spatial gradients, from the polar-ized neural tube during embryonic development to the osteochondral interfacepresent at articulating joints. The strong structure–function relationships inthese heterogeneous tissues have sparked intensive research into the develop-ment of methods that can replicate physiological gradients in engineered tis-sues. In this Review, we consider different gradients present in natural tissuesand discuss their critical importance in functional tissue engineering. Using thisbasis, we consolidate the existing fabrication methods into four categories: addi-tive manufacturing, component redistribution, controlled phase changes, andpostmodification. We have illustrated this with recent examples, highlightedprominent trends in thefield, and outlined a set of criteria and perspectives forgradient fabrication.

Journal article

Puetzer JL, Ma T, Sallent I, Gelmi A, Stevens Met al., 2021, Driving hierarchical collagen fiber formation for functional tendon, ligament and meniscus replacement, Biomaterials, Vol: 269, Pages: 1-10, ISSN: 0142-9612

Hierarchical collagen fibers are the primary source of strength in musculoskeletal tendons, ligaments, and menisci. It has remained a challenge to develop these large fibers in engineered replacements or in vivo after injury. The objective of this study was to investigate the ability of restrained cell-seeded high density collagen gels to drive hierarchical fiber formation for multiple musculoskeletal tissues. We found boundary conditions applied to high density collagen gels were capable of driving tenocytes, ligament fibroblasts, and meniscal fibrochondrocytes to develop native-sized hierarchical collagen fibers 20–40 μm in diameter. The fibers organize similar to bovine juvenile collagen with native fibril banding patterns and hierarchical fiber bundles 50–350 μm in diameter by 6 weeks. Mirroring fiber organization, tensile properties of restrained samples improved significantly with time, reaching ~1 MPa. Additionally, tendon, ligament, and meniscal cells produced significantly different sized fibers, different degrees of crimp, and different GAG concentrations, which corresponded with respective juvenile tissue. To our knowledge, these are some of the largest, most organized fibers produced to date in vitro. Further, cells produced tissue specific hierarchical fibers, suggesting this system is a promising tool to better understand cellular regulation of fiber formation to better stimulate it in vivo after injury.

Journal article

Solanki A, Lali F, Autefage H, Agarwal S, Nommeots-Nomm A, Metcalfe A, Stevens M, Jones Jet al., 2021, Bioactive glasses and electrospun composites that release cobalt to stimulate the HIF pathway for wound healing applications, Biomaterials Research, Vol: 25, ISSN: 2055-7124

BackgroundBioactive glasses are traditionally associated with bonding to bone through a hydroxycarbonate apatite (HCA) surface layer but the release of active ions is more important for bone regeneration. They are now being used to deliver ions for soft tissue applications, particularly wound healing. Cobalt is known to simulate hypoxia and provoke angiogenesis. The aim here was to develop new bioactive glass compositions designed to be scaffold materials to locally deliver pro-angiogenic cobalt ions, at a controlled rate, without forming an HCA layer, for wound healing applications.MethodsNew melt-derived bioactive glass compositions were designed that had the same network connectivity (mean number of bridging covalent bonds between silica tetrahedra), and therefore similar biodegradation rate, as the original 45S5 Bioglass. The amount of magnesium and cobalt in the glass was varied, with the aim of reducing or removing calcium and phosphate from the compositions. Electrospun poly(ε-caprolactone)/bioactive glass composites were also produced. Glasses were tested for ion release in dissolution studies and their influence on Hypoxia-Inducible Factor 1-alpha (HIF-1α) and expression of Vascular Endothelial Growth Factor (VEGF) from fibroblast cells was investigated.ResultsDissolution tests showed the silica rich layer differed depending on the amount of MgO in the glass, which influenced the delivery of cobalt. The electrospun composites delivered a more sustained ion release relative to glass particles alone. Exposing fibroblasts to conditioned media from these composites did not cause a detrimental effect on metabolic activity but glasses containing cobalt did stabilise HIF-1α and provoked a significantly higher expression of VEGF (not seen in Co-free controls).ConclusionsThe composite fibres containing new bioactive glass compositions delivered cobalt ions at a sustained rate, which could be mediated by the magnesium content of the glass. The dis

Journal article

Horgan C, Bergholt M, Nagelkerke A, Thin MZ, Pence IJ, Kauscher U, Kalber TL, Stuckey D, Stevens Met al., 2021, Integrated photodynamic Raman theranostic system for cancer diagnosis, treatment, and post-treatment molecular monitoring, Theranostics, Vol: 11, Pages: 2006-2019, ISSN: 1838-7640

Theranostics, the combination of diagnosis and therapy, has long held promise as a means to achieving personalised precision cancer treatments. However, despite its potential, theranostics has yet to realise significant clinical translation, largely due the complexity and overriding toxicity concerns of existing theranostic nanoparticle strategies.Methods: Here, we present an alternative nanoparticle-free theranostic approach based on simultaneous Raman spectroscopy and photodynamic therapy (PDT) in an integrated clinical platform for cancer theranostics.Results: We detail the compatibility of Raman spectroscopy and PDT for cancer theranostics, whereby Raman spectroscopic diagnosis can be performed on PDT photosensitiser-positive cells and tissues without inadvertent photosensitiser activation/photobleaching or impaired diagnostic capacity. We further demonstrate that our theranostic platform enables in vivo tumour diagnosis, treatment, and post-treatment molecular monitoring in real-time.Conclusion: This system thus achieves effective theranostic performance, providing a promising new avenue towards the clinical realisation of theranostics.

Journal article

Sero JE, Stevens MM, 2021, Nanoneedle-Based Materials for Intracellular Studies., Pages: 191-219

Nanoneedles, defined as high aspect ratio structures with tip diameters of 5 to approximately 500 nm, are uniquely able to interface with the interior of living cells. Their nanoscale dimensions mean that they are able to penetrate the plasma membrane with minimal disruption of normal cellular functions, allowing researchers to probe the intracellular space and deliver or extract material from individual cells. In the last decade, a variety of strategies have been developed using nanoneedles, either singly or as arrays, to investigate the biology of cancer cells in vitro and in vivo. These include hollow nanoneedles for soluble probe delivery, nanocapillaries for single-cell biopsy, nano-AFM for direct physical measurements of cytosolic proteins, and a wide range of fluorescent and electrochemical nanosensors for analyte detection. Nanofabrication has improved to the point that nanobiosensors can detect individual vesicles inside the cytoplasm, delineate tumor margins based on intracellular enzyme activity, and measure changes in cell metabolism almost in real time. While most of these applications are currently in the proof-of-concept stage, nanoneedle technology is poised to offer cancer biologists a powerful new set of tools for probing cells with unprecedented spatial and temporal resolution.

Book chapter

Potter M, Najer A, Kloeckner A, Zhang S, Holme MN, Nele V, Che J, Penders J, Saunders C, Doutch JJ, Edwards A, Ces O, Stevens Met al., 2020, Controlled dendrimersome nanoreactor system for localised hypochlorite-induced killing of bacteria, ACS Nano, Vol: 14, Pages: 17333-17353, ISSN: 1936-0851

Antibiotic resistance is a serious global health problem necessitating new bactericidal approaches such as nanomedicines. Dendrimersomes (DSs) have recently become a valuable alternative nanocarrier to polymersomes and liposomes due to their molecular definition and synthetic versatility. Despite this, their biomedical application is still in its infancy. Inspired by the localized antimicrobial function of neutrophil phagosomes and the versatility of DSs, a simple three-component DS-based nanoreactor with broad-spectrum bactericidal activity is presented. This was achieved by encapsulation of glucose oxidase (GOX) and myeloperoxidase (MPO) within DSs (GOX-MPO-DSs), self-assembled from an amphiphilic Janus dendrimer, that possesses a semipermeable membrane. By external addition of glucose to GOX-MPO-DS, the production of hypochlorite (−OCl), a highly potent antimicrobial, by the enzymatic cascade was demonstrated. This cascade nanoreactor yielded a potent bactericidal effect against two important multidrug resistant pathogens, Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa), not observed for H2O2 producing nanoreactors, GOX-DS. The production of highly reactive species such as –OCl represents a harsh bactericidal approach that could also be cytotoxic to mammalian cells. This necessitates the development of strategies for activating –OCl production in a localized manner in response to a bacterial stimulus. One option of locally releasing sufficient amounts of substrate using a bacterial trigger (released toxins) was demonstrated with lipidic glucose-loaded giant unilamellar vesicles (GUVs), envisioning, e.g., implant surface modification with nanoreactors and GUVs for localized production of bactericidal agents in the presence of bacterial growth.

Journal article

Maynard S, Gelmi A, Skaalure S, Pence I, Lee-Reeves C, Sero J, Whittaker T, Stevens Met al., 2020, Nanoscale molecular quantification of stem cell-hydrogel interactions, ACS Nano, Vol: 14, Pages: 17321-17332, ISSN: 1936-0851

A common approach to tailoring synthetic hydrogels for regenerative medicine applications involves incorporating RGD cell adhesion peptides, yet assessing the cellular response to engineered microenvironments at the nanoscale remains challenging. To date, no study has demonstrated how RGD concentration in hydrogels affects the presentation of individual cell surface receptors. Here we studied the interaction between human mesenchymal stem cells (hMSCs) and RGD-functionalized poly(ethylene glycol) hydrogels, by correlating macro- and nanoscale single-cell interfacial quantification techniques. We quantified RGD unbinding forces on a synthetic hydrogel using single cell atomic force spectroscopy, revealing that short-term binding of hMSCs was sensitive to RGD concentration. We also performed direct stochastic optical reconstruction microscopy (dSTORM) to quantify the molecular interactions between integrin α5β1 and a biomaterial, unexpectedly revealing that increased integrin clustering at the hydrogel-cell interface correlated with fewer available RGD binding sites. Our complementary, quantitative approach uncovered mechanistic insights into specific stem cell-hydrogel interactions, where dSTORM provides nanoscale sensitivity to RGD-dependent differences in cell surface localization of integrin α5β1. Our findings reveal that it is possible to precisely determine how peptide-functionalized hydrogels interact with cells at the molecular scale, thus providing a basis to fine-tune the spatial presentation of bioactive ligands.

Journal article

Belessiotis-Richards A, Higgins S, Sansom MSP, Alexander-Katz A, Stevens Met al., 2020, Coarse Grained Simulations Suggest the Epsin N-Terminal Homology Domain Can Sense Membrane Curvature Without its Terminal Amphipathic Helix, ACS Nano, Vol: 14, Pages: 16919-6928, ISSN: 1936-0851

Nanoscale membrane curvature is a common feature in cell biology required for functions such as endocytosis, exocytosis and cell migration. These processes require the cytoskeleton to exert forces on the membrane to deform it. Cytosolic proteins contain specific motifs which bind to the membrane, connecting it to the internal cytoskeletal machinery. These motifs often bind charged phosphatidylinositol phosphate lipids present in the cell membrane which play significant roles in signaling. These lipids are important for membrane deforming processes, such as endocytosis, but much remains unknown about their role in the sensing of membrane nanocurvature by protein domains. Using coarse-grained molecular dynamics simulations, we investigated the interaction of a model curvature active protein domain, the epsin N-terminal homology domain (ENTH), with curved lipid membranes. The combination of anionic lipids (phosphatidylinositol 4,5-bisphosphate and phosphatidylserine) within the membrane, protein backbone flexibility, and structural changes within the domain were found to affect the domain's ability to sense, bind, and localize with nanoscale precision at curved membrane regions. The findings suggest that the ENTH domain can sense membrane curvature without the presence of its terminal amphipathic α helix <i>via</i> another structural region we have denoted as H3, re-emphasizing the critical relationship between nanoscale membrane curvature and protein function.

Journal article

J C, Najer A, Blakney A, McKay P, Bellahcene M, Winter C, Sintou A, Tang J, Keane TJ, Schneider M, Shattock R, Sattler S, Stevens Met al., 2020, Neutrophils enable local and non-invasive liposome delivery to inflamed skeletal muscle and ischemic heart, Advanced Materials, Vol: 32, Pages: 1-10, ISSN: 0935-9648

Uncontrolled inflammation is a major pathological factor underlying a range of diseases including autoimmune conditions, cardiovascular disease, and cancer. Improving localized delivery of immunosuppressive drugs to inflamed tissue in a non‐invasive manner offers significant promise to reduce severe side effects caused by systemic administration. Here, a neutrophil‐mediated delivery system able to transport drug‐loaded nanocarriers to inflamed tissue by exploiting the inherent ability of neutrophils to migrate to inflammatory tissue is reported. This hybrid system (neutrophils loaded with liposomes ex vivo) efficiently migrates in vitro following an inflammatory chemokine gradient. Furthermore, the triggered release of loaded liposomes and reuptake by target macrophages is studied. The migratory behavior of liposome‐loaded neutrophils is confirmed in vivo by demonstrating the delivery of drug‐loaded liposomes to an inflamed skeletal muscle in mice. A single low‐dose injection of the hybrid system locally reduces inflammatory cytokine levels. Biodistribution of liposome‐loaded neutrophils in a human‐disease‐relevant myocardial ischemia reperfusion injury mouse model after i.v. injection confirms the ability of injected neutrophils to carry loaded liposomes to inflammation sites. This strategy shows the potential of nanocarrier‐loaded neutrophils as a universal platform to deliver anti‐inflammatory drugs to promote tissue regeneration in inflammatory diseases.

Journal article

Armstrong JPK, Keane TJ, Roques AC, Stephen Patrick P, Mooney CM, Kuan W-L, Pisupati V, Oreffo ROC, Stuckey D, Watt FM, Forbes SJ, Barker RA, Stevens Met al., 2020, A blueprint for translational regenerative medicine, Science Translational Medicine, Vol: 12, ISSN: 1946-6234

The last few decades have produced a large number of proof-of-concept studies in regenerative medicine.However, the route to clinical adoption is fraught with technical and translational obstacles that frequentlyconsign promising academic solutions to the so-called “valley of death.” This review is intended to serve as ablueprint for translational regenerative medicine: we suggest principles to help guide cell and materialselection, present key in vivo imaging modalities and argue that the host immune response should beconsidered throughout therapeutic development. Finally, we suggest a pathway to navigate the oftencomplex regulatory and manufacturing landscape of translational regenerative medicine.

Journal article

Hogset H, Horgan C, Bergholt M, Armstrong J, Torraca V, Chen Q, Keane TJ, Bugeon L, Dallman M, Mostowy S, Stevens Met al., 2020, In vivo biomolecular imaging of zebrafish embryos using confocal Raman spectroscopy, Nature Communications, Vol: 11, Pages: 1-12, ISSN: 2041-1723

Zebrafish embryos provide a unique opportunity to visualize complex biological processes, yet conventional imaging modalities are unable to access intricate biomolecular information without compromising the integrity of the embryos. Here, we report the use of confocal Raman spectroscopic imaging for the visualization and multivariate analysis of biomolecular information extracted from unlabeled zebrafish embryos. We outline broad applications of this method in: (i) visualizing the biomolecular distribution of whole embryos in three dimensions, (ii) resolving anatomical features at subcellular spatial resolution, (iii) biomolecular profiling and discrimination of wild type and ΔRD1 mutant Mycobacterium marinum strains in a zebrafish embryo model of tuberculosis and (iv) in vivotemporal monitoring of the wound response in living zebrafish embryos.Overall, this study demonstrates the application of confocal Raman spectroscopic imaging for the comparative bimolecular analysis in fully intact and living zebrafish embryos.

Journal article

Finbloom JA, Sousa F, Stevens MM, Desai TAet al., 2020, Engineering the drug carrier biointerface to overcome biological barriers to drug delivery, Advanced Drug Delivery Reviews, Vol: 167, Pages: 89-108, ISSN: 0169-409X

Micro and nanoscale drug carriers must navigate through a plethora of dynamic biological systems prior to reaching their tissue or disease targets. The biological obstacles to drug delivery come in many forms and include tissue barriers, mucus and bacterial biofilm hydrogels, the immune system, and cellular uptake and intracellular trafficking. The biointerface of drug carriers influences how these carriers navigate and overcome biological barriers for successful drug delivery. In this review, we examine how key material design parameters lead to dynamic biointerfaces and improved drug delivery across biological barriers. We provide a brief overview of approaches used to engineer key physicochemical properties of drug carriers, such as morphology, surface chemistry, and topography, as well as the development of dynamic responsive materials for barrier navigation. We then discuss essential biological barriers and how biointerface engineering can enable drug carriers to better navigate and overcome these barriers to drug delivery.

Journal article

Maynard S, Winter C, Cunnane E, Stevens Met al., 2020, Advancing cell instructive biomaterials through increased understanding of cell receptor spacing and material surface functionalization, Regenerative Engineering and Translational Medicine, ISSN: 2364-4133

Regenerative medicine is aimed at restoring normal tissue function and can benefit from the application of tissue engineering and nano-therapeutics. In order for regenerative therapies to be effective, the spatiotemporal integration of tissue-engineered scaffolds by the native tissue, and the binding/release of therapeutic payloads by nano-materials, must be tightly controlled at the nanoscale in order to direct cell fate. However, due to a lack of insight regarding cell–material interactions at the nanoscale and subsequent downstream signaling, the clinical translation of regenerative therapies is limited due to poor material integration, rapid clearance, and complications such as graft-versus-host disease. This review paper is intended to outline our current understanding of cell–material interactions with the aim of highlighting potential areas for knowledge advancement or application in the field of regenerative medicine. This is achieved by reviewing the nanoscale organization of key cell surface receptors, the current techniques used to control the presentation of cell-interactive molecules on material surfaces, and the most advanced techniques for characterizing the interactions that occur between cell surface receptors and materials intended for use in regenerative medicine.Lay SummaryThe combination of biology, chemistry, materials science, and imaging technology affords exciting opportunities to better diagnose and treat a wide range of diseases. Recent advances in imaging technologies have enabled better understanding of the specific interactions that occur between human cells and their immediate surroundings in both health and disease. This biological understanding can be used to design smart therapies and tissue replacements that better mimic native tissue. Here, we discuss the advances in molecular biology and technologies that can be employed to functionalize materials and characterize their interaction with biological entities to facilita

Journal article

Higgins S, Lo Fiego A, Patrick I, Creamer A, Stevens Met al., 2020, Organic bioelectronics: using highly conjugated polymers to interface with biomolecules, cells and tissues in the human body, Advanced Materials Technologies, Vol: 5, Pages: 1-35, ISSN: 2365-709X

Conjugated polymers exhibit interesting material and optoelectronic properties that makethem well-suited to the development of biointerfaces. Their biologically relevant mechanicalcharacteristics, ability to be chemically modified, and mixed electronic and ionic chargetransport are captured within the diverse field of organic bioelectronics. Conjugated polymershave been used in wide range of device architectures, and cell and tissue scaffolds. Thesedevices enable biosensing of many biomolecules, such as metabolites, nucleic acids and more.Devices can be used to both stimulate and sense the behavior of cells and tissues. Similarly,tissue interfaces permit interaction with complex organs, aiding both fundamental biologicalunderstanding and providing new opportunities for stimulating regenerative behaviors andbioelectronic based therapeutics. Applications of these materials are broad, and muchcontinues to be uncovered about their fundamental properties. This report covers the currentunderstanding of the fundamentals of conjugated polymer biointerfaces and their interactionswith biomolecules, cells and tissues in the human body. An overview of current materials anddevices is presented, along with highlighted major in vivo and in vitro applications. Finally,open research questions and opportunities are discussed.

Journal article

Nele V, Wojciechowski J, Armstrong J, Stevens Met al., 2020, Tailoring gelation mechanisms for advanced hydrogel applications, Advanced Functional Materials, Vol: 30, Pages: 1-22, ISSN: 1616-301X

Hydrogels are one of the most commonly explored classes of biomaterials. Their chemical and structural versatility has enabled their use across a wide range of applications, including tissue engineering, drug delivery, and cell culture. Hydrogels form upon a sol–gel transition, which can be elicited by different triggers designed to enable precise control over hydrogelation kinetics and hydrogel structure. The chosen hydrogelation trigger and chemistry can have a profound effect on the success of the targeted application. In this Progress Report, a critical overview of recent advances in hydrogel design is presented, with a focus on the available strategies used to trigger the formation of hydrogel networks (e.g., temperature, light, ultrasound). These triggers are presented within a new classification system, and their suitability for six key hydrogel‐based applications is assessed. This Progress Report is intended to guide trigger selection for new hydrogel applications and inspire the rational design of new hydrogelation trigger mechanisms.

Journal article

Ritzau-Reid K, Spicer C, Gelmi A, Grigsby CL, Ponder Jr J, Bemmer V, Creamer A, Vilar R, Serio A, Stevens Met al., 2020, An electroactive oligo-EDOT platform for neural tissue engineering, Advanced Functional Materials, Vol: 30, Pages: 1-11, ISSN: 1616-301X

The unique electrochemical properties of the conductive polymer poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS) make it an attractive material for use in neural tissue engineering applications. However, inadequate mechanical properties, and difficulties in processing and lack of biodegradability have hindered progress in this field. Here, we have improved the functionality of PEDOT:PSS for neural tissue engineering by incorporating 3,4-ethylenedioxythiophene (EDOT) oligomers, synthesised using a novel end-capping strategy, into block co-polymers. By exploiting end-functionalised oligoEDOT constructs as macroinitiators for the polymerization of poly(caprolactone) (PCL), we produce a block co-polymer that is electroactive, processable, and bio-compatible. By combining these properties, we were able to produce electroactive fibrous mats for neuronal culture via solution electrospinning and melt electrospinning writing (MEW). Importantly, we also show that neurite length and branching of neural stem cells can be enhanced on our materials under electrical stimulation, demonstrating the promise of these scaffolds for neural tissue engineering.

Journal article

Zwi Dantsis L, Winter CW, Kauscher U, Ferrini A, Wang B, Whittaker T, Hood SR, Terracciano C, Stevens Met al., 2020, Highly purified extracellular vesicles from human cardiomyocytes demonstrate preferential uptake by human endothelial cells, Nanoscale, Vol: 12, Pages: 19844-19854, ISSN: 2040-3364

Extracellular vesicles (EVs) represent a promising cell-free alternative for treatment of cardiovascular diseases. Nevertheless, the lack of standardised and reproducible isolation methods capable of recovering pure, intact EVs presents a significant obstacle. Additionally, there is significant interest in investigating the interactions of EVs with different cardiac cell types. Here we established a robust technique for the production and isolation of EVs harvested from an enriched (>97% purity) population of human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) with size exclusion chromatography. Utilizing an advanced fluorescence labelling strategy, we then investigated the interplay of the CM-EVs with the three major cellular components of the myocardium (fibroblasts, cardiomyocytes and endothelial cells) and identified that cardiac endothelial cells show preferential uptake of these EVs. Overall, our findings provide a great opportunity to overcome the translational hurdles associated with the isolation of intact, non-aggregated human iPSC-CM EVs at high purity. Furthermore, understanding in detail the interaction of the secreted EVs with their surrounding cells in the heart may open promising new avenues in the field of EV engineering for targeted delivery in cardiac regeneration.

Journal article

Majid QA, Fricker ATR, Gregory DA, Davidenko N, Hernandez Cruz O, Jabbour RJ, Owen TJ, Basnett P, Lukasiewicz B, Stevens M, Best S, Cameron R, Sinha S, Harding SE, Roy Iet al., 2020, Natural biomaterials for cardiac tissue engineering: a highly biocompatible solution., Frontiers in Cardiovascular Medicine, Vol: 7, Pages: 1-32, ISSN: 2297-055X

Cardiovascular diseases (CVD) constitute a major fraction of the current major global diseases and lead to about 30% of the deaths, i.e., 17.9 million deaths per year. CVD include coronary artery disease (CAD), myocardial infarction (MI), arrhythmias, heart failure, heart valve diseases, congenital heart disease, and cardiomyopathy. Cardiac Tissue Engineering (CTE) aims to address these conditions, the overall goal being the efficient regeneration of diseased cardiac tissue using an ideal combination of biomaterials and cells. Various cells have thus far been utilized in pre-clinical studies for CTE. These include adult stem cell populations (mesenchymal stem cells) and pluripotent stem cells (including autologous human induced pluripotent stem cells or allogenic human embryonic stem cells) with the latter undergoing differentiation to form functional cardiac cells. The ideal biomaterial for cardiac tissue engineering needs to have suitable material properties with the ability to support efficient attachment, growth, and differentiation of the cardiac cells, leading to the formation of functional cardiac tissue. In this review, we have focused on the use of biomaterials of natural origin for CTE. Natural biomaterials are generally known to be highly biocompatible and in addition are sustainable in nature. We have focused on those that have been widely explored in CTE and describe the original work and the current state of art. These include fibrinogen (in the context of Engineered Heart Tissue, EHT), collagen, alginate, silk, and Polyhydroxyalkanoates (PHAs). Amongst these, fibrinogen, collagen, alginate, and silk are isolated from natural sources whereas PHAs are produced via bacterial fermentation. Overall, these biomaterials have proven to be highly promising, displaying robust biocompatibility and, when combined with cells, an ability to enhance post-MI cardiac function in pre-clinical models. As such, CTE has great potential for future clinical solutions and he

Journal article

Ouyang L, Armstrong J, Lin Y, Wojciechowski J, Lee-Reeves C, Hachim D, Zhou K, Burdick JA, Stevens Met al., 2020, Expanding and optimizing 3D bioprinting capabilities using complementary network bioinks, Science Advances, Vol: 6, Pages: 1-13, ISSN: 2375-2548

A major challenge in 3D bioprinting is the limited number of bioinks that fulfill the physiochemical requirements of printing, while also providing a desirable environment for encapsulated cells.Here, we address this limitation by temporarily stabilizing bioinks with a complementary thermo-reversible gelatin network. This strategy enablesthe effective printing of biomaterials that would typically not meet printing requirements, with instrument parameters and structural output largely independent of the base biomaterial. This approach is demonstrated across a library of photo-crosslinkable bioinks derived from natural and synthetic polymers, including gelatin, hyaluronic acid, chondroitin sulfate, dextran, alginate, chitosan, heparin,and poly(ethylene glycol). A range of complex and heterogeneous structures are printed, including soft hydrogel constructs supporting the 3D culture of astrocytes. This highly generalizable methodology expands the palette of available bioinks, allowing the biofabrication of constructs optimized to meet the biological requirements of cell culture and tissue engineering.

Journal article

Massi L, Najer A, Chapman R, Spicer C, Nele V, Che J, Booth MA, Doutch JJ, Stevens Met al., 2020, Tuneable peptide cross-linked nanogels for enzyme-triggered protein delivery, Journal of Materials Chemistry B, Vol: 8, Pages: 8894-8907, ISSN: 2050-750X

Many diseases are associated with the dysregulated activity of enzymes, such as matrixmetalloproteinases (MMPs). This dysregulation can be leveraged in drug delivery to achieve disease- orsite-specific cargo release. Self-assembled polymeric nanoparticles are versatile drug carrier materialsdue to the accessible diversity of polymer chemistry. However, efficient loading of sensitive cargo, suchas proteins, and introducing functional enzyme-responsive behaviour remain challenging. Herein,peptide-crosslinked, temperature-sensitive nanogels for protein delivery were designed to respond toMMP-7, which is overexpressed in many pathologies including cancer and inflammatory diseases. Theincorporation of N-cyclopropylacrylamide (NCPAM) into N-isopropylacrylamide (NIPAM)-basedcopolymers enabled us to tune the polymer lower critical solution temperature from 33 to 44 1C,allowing the encapsulation of protein cargo and nanogel-crosslinking at slightly elevated temperatures.This approach resulted in nanogels that were held together by MMP-sensitive peptides for enzymespecificprotein delivery. We employed a combination of cryogenic transmission electron microscopy(cryo-TEM), dynamic light scattering (DLS), small angle neutron scattering (SANS), and fluorescencecorrelation spectroscopy (FCS) to precisely decipher the morphology, self-assembly mechanism,enzyme-responsiveness, and model protein loading/release properties of our nanogel platform. Simplevariation of the peptide linker sequence and combining multiple different crosslinkers will enable us toadjust our platform to target specific diseases in the future.

Journal article

Whittaker T, Nagelkerke A, Nele V, Kauscher U, Stevens Met al., 2020, Experimental artefacts can lead to misattribution of bioactivity from soluble mesenchymal stem cell paracrine factors to extracellular vesicles, Journal of Extracellular Vesicles, Vol: 9, ISSN: 2001-3078

It has been demonstrated that some commonly used Extracellular Vesicle (EV) isolation techniques can lead to substantial contamination with non-EV factors. Whilst it has been established that this impacts the identification of biomarkers, the impact on apparent EV bioactivity has not been explored. Extracellular vesicles have been implicated as critical mediators of therapeutic human mesenchymal stem cell (hMSC) paracrine signalling. Isolated hMSC-EVs have been used to treat multiple in vitro and in vivo models of tissue damage. However, the relative contributions of EVs and non-EV factors have not been directly compared. The dependence of hMSC paracrine signalling on EVs was first established by ultrafiltration of hMSC-conditioned medium to deplete EVs, which led to a loss of signalling activity. Here, we show that this method also causes depletion of non-EV factors, and that when this is prevented proangiogenic signalling activity is fully restored in vitro. Subsequently, we used size-exclusion chromatography (SEC) to separate EVs and soluble proteins to directly and quantitatively compare their relative contributions to signalling. Non-EV factors were found to be necessary and sufficient for the stimulation of angiogenesis and wound healing in vitro. EVs in isolation were found to be capable of potentiating signalling only when isolated by a low-purity method, or when used at comparatively high concentrations. These results indicate a potential for contaminating soluble factors to artefactually increase the apparent bioactivity of EV isolates and could have implications for future studies on the biological roles of EVs.

Journal article

Li S, Tallia F, Mohammed AA, Stevens MM, Jones JRet al., 2020, Scaffold channel size influences stem cell differentiation pathway in 3-D printed silica hybrid scaffolds for cartilage regeneration, Biomaterials Science, Vol: 8, Pages: 4458-4466, ISSN: 2047-4830

We report that 3-D printed scaffold channel size can direct bone marrow derived stem cell differentiation. Treatment of articular cartilage trauma injuries, such as microfracture surgery, have limited success because durability is limited as fibrocartilage forms. A scaffold-assisted approach, combining microfracture with biomaterials has potential if the scaffold can promote articular cartilage production and share load with cartilage. Here, we investigated human bone marrow derived stromal cell (hBMSC) differentiation in vitro in 3-D printed silica/poly(tetrahydrofuran)/poly(ε-caprolactone) hybrid scaffolds with specific channel sizes. Channel widths of ∼230 μm (210 ± 22 μm mean strut size, 42.4 ± 3.9% porosity) provoked hBMSC differentiation down a chondrogenic path, with collagen Type II matrix prevalent, indicative of hyaline cartilage. When pores were larger (∼500 μm, 229 ± 29 μm mean strut size, 63.8 ± 1.6% porosity) collagen Type I was dominant, indicating fibrocartilage. There was less matrix and voids in smaller channels (∼100 μm, 218 ± 28 μm mean strut size, 31.2 ± 2.9% porosity). Our findings suggest that a 200–250 μm pore channel width, in combination with the surface chemistry and stiffness of the scaffold, is optimal for cell–cell interactions to promote chondrogenic differentiation and enable the chondrocytes to maintain their phenotype.

Journal article

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