Imperial College London

Professor Molly Stevens FREng

Faculty of EngineeringDepartment of Materials

Prof of Biomedical Materials&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

348 results found

Wang S, Lin Y, Todorova N, Xu Y, Mazo M, Rana S, Leonardo V, Amdursky N, Spicer CD, Alexander BD, Edwards AA, Matthews SJ, Yarovsky I, Stevens MMet al., 2017, Facet-dependent interactions of islet amyloid polypeptide with gold nanoparti-cles: implications for fibril formation and peptide-induced lipid membrane dis-ruption, Chemistry of Materials, Vol: 29, ISSN: 1520-5002

A comprehensive understanding of the mechanisms of interaction between proteins or peptides and nanomaterials is crucial for the development of nanomaterial-based diagnos-tics and therapeutics. In this work, we systematically explored the interactions between citrate-capped gold nanoparticles (AuNPs) and islet amyloid polypeptide (IAPP), a 37-amino acid peptide hormone co-secreted with insulin from the pancreatic islet. We uti-lized diffusion-ordered spectroscopy, isothermal titration calorimetry, localized surface plasmon resonance spectroscopy, gel electrophoresis, atomic force microscopy, transmis-sion electron microscopy (TEM), and molecular dynamics (MD) simulations to systemati-cally elucidate the underlying mechanism of the IAPP−AuNP interactions. Because of the presence of a metal-binding sequence motif in the hydrophilic peptide domain, IAPP strongly interacts with the Au surface in both the monomeric and fibrillar states. Circular dichroism showed that AuNPs triggered the IAPP conformational transition from random coil to ordered structures (α-helix and β-sheet), and TEM imaging suggested the accelera-tion of IAPP fibrillation in the presence of AuNPs. MD simulations revealed that the IAPP−AuNP interactions were initiated by the N-terminal domain (IAPP residues 1−19), which subsequently induced a facet-dependent conformational change in IAPP. On a Au(111) surface, IAPP was unfolded and adsorbed directly onto the Au surface, while for the Au(100) surface, it interacted predominantly with the citrate adlayer and retained some helical conformation. The observed affinity of AuNPs for IAPP was further applied to reduce the level of peptide-induced lipid membrane disruption.

Journal article

Ehademhosseini A, Chan WWC, Chhowalla M, Glotzer SC, Gogotsi Y, Hafner JH, Hammond PT, Hersam MC, Javey A, Kagan CR, Kotov NA, Lee S-T, Li Y, Mohwald H, Mulvaney PA, Nel AE, Parak WJ, Penner RM, Rogach AL, Schaak RE, Stevens MM, Wee ATS, Brinker J, Chen X, Chi L, Crommie M, Dekker C, Farokhzad O, Gerber C, Ginger DS, Irvine DJ, Kiessling LL, Kostarelos K, Landes C, Lee T, Leggett GJ, Liang X-J, Liz-Marzan L, Millstone J, Odom TW, Ozcan A, Prato M, Rao CNR, Sailor MJ, Weiss E, Weiss PSet al., 2017, Nanoscience and Nanotechnology Cross Borders, Publisher: AMER CHEMICAL SOC

Other

Lin Y, Pashuck ET, Thomas MR, Amdursky N, Wang S-T, Chow LW, Stevens MMet al., 2017, Plasmonic chirality imprinting on nucleobase-displaying supramolecular nanohelices by metal-nucleobase recognition, ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, Vol: 56, Pages: 2361-2365, ISSN: 1433-7851

Supramolecular self-assembly is an important process that enables the conception of complex structures mimicking biological motifs. Herein, we constructed helical fibrils through chiral self-assembly of nucleobase–peptide conjugates (NPCs), where achiral nucleobases are helically displayed on the surface of fibrils, comparable to polymerized nucleic acids. Selective binding between DNA and the NPC fibrils was observed with fluorescence polarization. Taking advantage of metal–nucleobase recognition, we highlight the possibility of deposition/assembly of plasmonic nanoparticles onto the fibrillar constructs. In this approach, the supramolecular chirality of NPCs can be adaptively imparted to metallic nanoparticles, covering them to generate structures with plasmonic chirality that exhibit significantly improved colloidal stability. The self-assembly of rationally designed NPCs into nanohelices is a promising way to engineer complex, optically diverse nucleobase-derived nanomaterials.

Journal article

Spicer C, Booth M, Mawad D, Armgarth A, Nielsen C, Stevens MMet al., 2017, Synthesis of hetero-bifunctional, end-capped oligo-EDOT derivatives, Chem, Vol: 2, Pages: 125-138, ISSN: 2451-9294

Conjugated oligomers of 3,4-ethylenedioxythiophene (EDOT) are attractive materials for tissue engineering applications, and as model systems for studying the properties of the widely used polymer PEDOT. We report here the facile synthesis of a series of keto-acid end-capped oligo-EDOT derivatives (n = 2-7) through a combination of a glyoxylation end capping strategy and iterative direct arylation chain extension. Importantly, these structures not only represent the longest oligo-EDOTs reported, but are also bench stable in contrast to previous reports on such oligomers. The constructs reported here can undergo subsequent derivatization for integration into higher order architectures, such as those required for tissue engineering applications. The synthesis of hetero-bifunctional constructs, as well as those containing mixed monomer units is also reported, allowing further complexity to be installed in a controlled manner. Finally, we describe the optical and electrochemical properties of these oligomers and demonstrate the importance of the keto-acid in determining their characteristics.

Journal article

Parmar PA, St-Pierre JP, Chow LW, Spicer CD, Stoichevska V, Peng YY, Werkmeister JA, Ramshaw JAM, Stevens MMet al., 2017, Enhanced articular cartilage by human mesenchymal stem cells in enzymatically mediated transiently RGDS–functionalized collagen–mimetic hydrogels, Acta Biomaterialia, Vol: 51, ISSN: 1878-7568

Recapitulation of the articular cartilage microenvironment for regenerative medicine applications faces significant challenges due to the complex and dynamic biochemical and biomechanical nature of native tissue. Towards the goal of biomaterial designs that enable the temporal presentation of bioactive sequences, recombinant bacterial collagens such as Streptococcal collagen-like 2 (Scl2) proteins can be employed to incorporate multiple specific bioactive and biodegradable peptide motifs into a single construct. Here, we first modified the backbone of Scl2 with glycosaminoglycan-binding peptides and cross-linked the modified Scl2 into hydrogels via matrix metalloproteinase 7 (MMP7)-cleavable or non-cleavable scrambled peptides. The cross-linkers were further functionalized with a tethered RGDS peptide creating a system whereby the release from an MMP7-cleavable hydrogel could be compared to a system where release is not possible. The release of the RGDS peptide from the degradable hydrogels led to significantly enhanced expression of collagen type II (3.9-fold increase), aggrecan (7.6-fold increase), and SOX9 (5.2-fold increase) by human mesenchymal stem cells (hMSCs) undergoing chondrogenesis, as well as greater extracellular matrix accumulation compared to non-degradable hydrogels (collagen type II; 3.2-fold increase, aggrecan; 4-fold increase, SOX9; 2.8-fold increase). Hydrogels containing a low concentration of the RGDS peptide displayed significantly decreased collagen type I and X gene expression profiles, suggesting a major advantage over either hydrogels functionalized with a higher RGDS peptide concentration, or non-degradable hydrogels, in promoting an articular cartilage phenotype. These highly versatile Scl2 hydrogels can be further manipulated to improve specific elements of the chondrogenic response by hMSCs, through the introduction of additional bioactive and/or biodegradable motifs. As such, these hydrogels have the possibility to be used for other

Journal article

Armstrong JPK, Holme MN, Stevens MM, 2017, Re-Engineering Extracellular Vesicles as Smart Nanoscale Therapeutics, ACS Nano, Vol: 11, Pages: 69-83, ISSN: 1936-0851

In the past decade, extracellular vesicles(EVs) have emerged as a key cell-free strategy for thetreatment of a range of pathologies, including cancer,myocardial infarction, and inflammatory diseases. Indeed,the field is rapidly transitioning from promising in vitroreports toward in vivo animal models and early clinicalstudies. These investigations exploit the high physicochemicalstability and biocompatibility of EVs as well as theirinnate capacity to communicate with cells via signaltransduction and membrane fusion. This review focuseson methods in which EVs can be chemically or biologicallymodified to broaden, alter, or enhance their therapeuticcapability. We examine two broad strategies, which havebeen used to introduce a wide range of nanoparticles, reporter systems, targeting peptides, pharmaceutics, and functionalRNA molecules. First, we explore how EVs can be modified by manipulating their parent cells, either through genetic ormetabolic engineering or by introducing exogenous material that is subsequently incorporated into secreted EVs. Second,we consider how EVs can be directly functionalized using strategies such as hydrophobic insertion, covalent surfacechemistry, and membrane permeabilization. We discuss the historical context of each specific technology, presentprominent examples, and evaluate the complexities, potential pitfalls, and opportunities presented by different reengineeringstrategies.

Journal article

Chan WWC, Chhowalla M, Glotzer S, Gogotsi Y, Hafner JH, Hammond PT, Hersam MC, Javey A, Kagan CR, Khademhosseini A, Kotov NA, Lee ST, Li Y, Möhwald H, Mulvaney PA, Nel AE, Nordlander PJ, Parak WJ, Penner RM, Rogach AL, Schaak RE, Stevens MM, Wee ATS, Willson CG, Fernandez LE, Weiss PSet al., 2016, Nanoscience and Nanotechnology Impacting Diverse Fields of Science, Engineering, and Medicine

Other

Fiocco L, Li S, Stevens MM, Bernardo E, Jones JRet al., 2016, Biocompatibility and bioactivity of porous polymer-derived Ca-Mg silicate ceramics., Acta Biomaterialia, Vol: 50, Pages: 56-67, ISSN: 1878-7568

Magnesium is a trace element in the human body, known to have important effects on cell differentiation and the mineralisation of calcified tissues. This study aimed to synthesise highly porous Ca-Mg silicate foamed scaffolds from preceramic polymers, with analysis of their biological response. Akermanite (Ak) and wollastonite-diopside (WD) ceramic foams were obtained from the pyrolysis of a liquid silicone mixed with reactive fillers. The porous structure was obtained by controlled water release from selected fillers (magnesium hydroxide and borax) at 350°C. The homogeneous distribution of open pores, with interconnects of modal diameters of 160-180μm was obtained and maintained after firing at 1100°C. Foams, with porosity exceeding 80%, exhibited compressive strength values of 1-2MPa. In vitro studies were conducted by immersion in SBF for 21days, showing suitable dissolution rates, pH and ionic concentrations. Cytotoxicity analysis performed in accordance with ISO10993-5 and ISO10993-12 standards confirmed excellent biocompatibility of both Ak and WD foams. In addition, MC3T3-E1 cells cultured on the Mg-containing scaffolds demonstrated enhanced osteogenic differentiation and the expression of osteogenic markers including Collagen Type I, Osteopontin and Osteocalcin, in comparison to Mg-free counterparts. The results suggest that the addition of magnesium can further enhance the bioactivity and the potential for bone regeneration applications of Ca-silicate materials. STATEMENTS OF SIGNIFICANCE: Here, we show that the incorporation of Mg in Ca-silicates plays a significant role in the enhancement of the osteogenic differentiation and matrix formation of MC3T3-E1 cells, cultured on polymer-derived highly porous scaffolds. Reduced degradation rates and improved mechanical properties are also observed, compared to Mg-free counterparts, suggesting the great potential of Ca-Mg silicates as bone tissue engineering materials. Excellent biocompatibility of the

Journal article

Mawad D, Mansfield C, Lauto A, Perbellini F, Nelson G, Tonkin J, Bello S, Carrod D, Micolich A, Mahat M, Simonotto J, Payne D, Lyon A, Gooding J, Harding S, Terracciano C, Stevens MMet al., 2016, A conducting polymer with enhanced electronic stability applied in cardiac models, Science Advances, Vol: 2, ISSN: 2375-2548

Electrically active constructs can have a beneficial effect on electroresponsive tissues, such as the brain, heart, and nervous system. Conducting polymers (CPs) are being considered as components of these constructs because of their intrinsic electroactive and flexible nature. However, their clinical application has been largely hampered by their short operational time due to a decrease in their electronic properties. We show that, by immobilizing the dopant in the conductive scaffold, we can prevent its electric deterioration. We grew polyaniline (PANI) doped with phytic acid on the surface of a chitosan film. The strong chelation between phytic acid and chitosan led to a conductive patch with retained electroactivity, low surface resistivity (35.85 ± 9.40 kilohms per square), and oxidized form after 2 weeks of incubation in physiological medium. Ex vivo experiments revealed that the conductive nature of the patch has an immediate effect on the electrophysiology of the heart. Preliminary in vivo experiments showed that the conductive patch does not induce proarrhythmogenic activities in the heart. Our findings set the foundation for the design of electronically stable CP-based scaffolds. This provides a robust conductive system that could be used at the interface with electroresponsive tissue to better understand the interaction and effect of these materials on the electrophysiology of these tissues.

Journal article

Pashuck ET, Duchet BJR, Hansel CS, Maynard SA, Chow LW, Stevens MMet al., 2016, Controlled sub-nanometer epitope spacing in a three-dimensional self-assembled peptide hydrogel, ACS Nano, Vol: 10, Pages: 11096-11104, ISSN: 1936-0851

Cells in the body use a variety of mechanisms to ensure the specificity and efficacy of signal transduction. One way that this is achieved is through tight spatial control over the position of different proteins, signaling sequences, and biomolecules within and around cells. For instance, the extracellular matrix protein fibronectin presents RGDS and PHSRN sequences that synergistically bind the α5β1 integrin when separated by 3.2 nm but are unable to bind when this distance is >5.5 nm.1 Building biomaterials to controllably space different epitopes with subnanometer accuracy in a three-dimensional (3D) hydrogel is challenging. Here, we synthesized peptides that self-assemble into nanofiber hydrogels utilizing the β-sheet motif, which has a known regular spacing along the peptide backbone. By modifying specific locations along the peptide, we are able to controllably space different epitopes with subnanometer accuracy at distances from 0.7 nm to over 6 nm, which is within the size range of many protein clusters. Endothelial cells encapsulated within hydrogels displaying RGDS and PHSRN in the native 3.2 nm spacing showed a significant upregulation in the expression of the alpha 5 integrin subunit compared to those in hydrogels with a 6.2 nm spacing, demonstrating the physiological relevance of the spacing. Furthermore, after 24 h the cells in hydrogels with the 3.2 nm spacing appeared to be more spread with increased staining for the α5β1 integrin. This self-assembling peptide system can controllably space multiple epitopes with subnanometer accuracy, demonstrating an exciting platform to study the effects of ligand density and location on cells within a synthetic 3D environment.

Journal article

Pacheco-Moreno CM, Schreck M, Scaccabarozzi AD, Bourgun P, Wantz G, Stevens MM, Dautel OJ, Stingelin Net al., 2016, The importance of materials design to make ions flow: toward novel materials platforms for bioelectronics applications, Advanced Materials, Vol: 29, ISSN: 0935-9648

Chemical design criteria for materials for bioelectronics applications using a series of copolymer derivatives based on poly(3-hexylthiophene) are established. Directed chemical design via side-chain functionalization with polar groups allows manipulation of ion transport and ion-to-electron transduction. Insights gained will permit increased use of the plethora of materials employed in the organic electronics area for application in the bioelectronics field.

Journal article

Wood CS, Stevens MM, 2016, MATERIALS SCIENCE Improving the image of nanoparticles, Nature, Vol: 539, Pages: 505-506, ISSN: 0028-0836

A biocompatible probe that combines fluorescent nanodiamonds and gold nanoparticles allows cells to be imaged using both optical and electron microscopy techniques, opening up fresh opportunities for biological research.

Journal article

Bergholt M, St-Pierre J, Offeddu G, Parmar P, Albro M, Puetzer J, Oyen M, Stevens MMet al., 2016, Raman spectroscopy reveals new insights into the zonal organization of native and tissue-engineered articular cartilage, ACS Central Science, Vol: 2, Pages: 885-895, ISSN: 2374-7951

Tissue architecture is intimately linked with its functions, and loss of tissue organization is often associated with pathologies. The intricate depth-dependent extracellular matrix (ECM) arrangement in articular cartilage is critical to its biomechanical functions. In this study, we developed a Raman spectroscopic imaging approach to gain new insight into the depth-dependent arrangement of native and tissue-engineered articular cartilage using bovine tissues and cells. Our results revealed previously unreported tissue complexity into at least six zones above the tidemark based on a principal component analysis and k-means clustering analysis of the distribution and orientation of the main ECM components. Correlation of nanoindentation and Raman spectroscopic data suggested that the biomechanics across the tissue depth are influenced by ECM microstructure rather than composition. Further, Raman spectroscopy together with multivariate analysis revealed changes in the collagen, glycosaminoglycan and water distributions in tissue-engineered constructs over time. These changes were assessed using simple metrics that promise to instruct efforts towards the regeneration of a broad range of tissues with native zonal complexity and functional performance.

Journal article

Stevens MM, Campagnolo P, Gormley AJ, Chow L, Guex A, Parmar P, Puetzer JL, Steele JAM, Breant A, Madeddu Pet al., 2016, Pericyte seeded dual peptide scaffold with improved endothelialization for vascular graft tissue engineering, Advanced Healthcare Materials, Vol: 5, Pages: 3046-3055, ISSN: 2192-2640

The development of synthetic vascular grafts for coronary artery bypass is challenged by insufficient endothelialization, which exposes to the risk of thrombosis, and lack of native cellular constituents, which favours pathological remodelling. Here, an bifunctional electrospun poly(ε-caprolactone) (PCL) scaffold with potential for synthetic vascular graft applications is presented. This scaffold incorporates two tethered peptides: the osteopontin-derived peptide (Adh) on the ‘luminal’ side and a heparin-binding peptide (Hep) on the ‘abluminal’ side. Additionally, the ‘abluminal’ side of the scaffold is seeded with saphenous vein-derived pericytes (SVPs) as a source of pro-angiogenic growth factors. The Adh peptide significantly increase endothelial cell adhesion, while the Hep peptide promote accumulation of vascular endothelial growth factor (VEGF) secreted by SVPs. SVPs increase endothelial migration both in a transwell assay and a modified scratch assay performed on the PCL scaffold. Seeding of SVPs on the ‘abluminal’/Hep side of the scaffold further increase endothelial cell density, indicating a combinatory effect of the peptides and pericytes. Lastly, SVP-seeded scaffolds are preserved by freezing in a xeno-free medium, maintaining good cell viability and function. In conclusion, this engineered scaffold combines patient-derived pericytes and spatially organized functionalities, which synergistically increase endothelial cell density and growth factor retention.

Journal article

Kim E, Howes PD, Crowder SW, Stevens MMet al., 2016, Multi-Amplified Sensing of MicroRNA by a Small DNA Fragment-Driven Enzymatic Cascade Reaction, ACS Sensors, Vol: 2, Pages: 111-118, ISSN: 2379-3694

Combining technological developments such asnanomaterials, DNA nanotechnology, and functional enzymeshas great potential to facilitate next generation high performancemolecular diagnostic systems. In this work, we describe amicroRNA (miRNA) detection assay that combines targetrecycling and isothermal amplification in an elegantly designedenzyme-mediated cascade reaction. Target recycling is drivenby the action of duplex-specific nuclease (DSN), resulting inhighly amplified translation of input miRNA to short outputDNA fragments. These fragments act as highly specificinitiators of rolling circle amplification (RCA), an isothermalreaction that outputs a large volume of polymeric DNAzymesper initiator, and finally a fluorogenic output signal. Based oncareful electrophoretic analysis we observed that the DSN produces ca. 10 nt DNA fragments from DNA/miRNA duplexes,regardless of the length of DNA strands. Target recycling yielded ca. 5 orders of magnitude amplification through the DSNassistedrecycling system on magnetic particles, and the RCA yielded a further 2 orders of magnitude. The final assay exhibited alimit of detection of 1.8 fM of miRNA spiked into 20% human serum, and showed excellent selectivity for miR-21 versus singlebase-mismatched sequences and other cancer-related miRNAs. The developed assay was further employed to determine accurateamounts of miR-21 in total RNA samples extracted from human cancer cell lines and normal cells, confirming the applicability ofthe assay for direct and absolute quantification of mature specific miRNA in real biological samples.

Journal article

Flamant Q, Caravaca C, Meille S, Gremillard L, Chevalier J, Biotteau-Deheuvels K, Kuntz M, Chandrawati R, Herrmann IK, Spicer CD, Stevens MM, Anglada Met al., 2016, Selective etching of injection molded zirconia-toughened alumina: towards osseointegrated and antibacterial ceramic implants., Acta Biomaterialia, Vol: 46, Pages: 308-322, ISSN: 1878-7568

Due to their outstanding mechanical properties and excellent biocompatibility, zirconia-toughened alumina (ZTA) ceramics have become the gold standard in orthopedics for the fabrication of ceramic bearing components over the last decade. However, ZTA is bioinert, which hampers its implantation in direct contact with bone. Furthermore, periprosthetic joint infections are now the leading cause of failure for joint arthroplasty prostheses. To address both issues, an improved surface design is required: a controlled micro- and nano-roughness can promote osseointegration and limit bacterial adhesion whereas surface porosity allows loading and delivery of antibacterial compounds. In this work, we developed an integrated strategy aiming to provide both osseointegrative and antibacterial properties to ZTA surfaces. The micro-topography was controlled by injection molding. Meanwhile a novel process involving the selective dissolution of zirconia (selective etching) was used to produce nano-roughness and interconnected nanoporosity. Potential utilization of the porosity for loading and delivery of antibiotic molecules was demonstrated, and the impact of selective etching on mechanical properties and hydrothermal stability was shown to be limited. The combination of injection molding and selective etching thus appears promising for fabricating a new generation of ZTA components implantable in direct contact with bone.

Journal article

Maçon ALB, Li S, Chung J, Nommeots-Nomm A, Solanki AK, Stevens MM, Jones JRet al., 2016, Ductile silica/methacrylate hybrids for bone regeneration, Journal of Materials Chemistry B, Vol: 4, Pages: 6032-6042, ISSN: 2050-7518

Bioglass® was the first synthetic material capable of bonding with bone without fibrous encapsulation, and fulfils some of the criteria of an ideal synthetic bone graft. However, it is brittle and toughness is required. Here, we investigated hybrids consisting of co-networks of high cross-linking density polymethacrylate and silica (class II hybrid) as a potential new generation of scaffold materials. Poly(3-(methoxysilyl)propyl methacrylate) (pTMSPMA) and tetraethyl orthosilicate (TEOS) were used as sol–gel precursors and hybrids were synthesised with different inorganic to organic ratios (Ih). The hybrids were nanoporous, with a modal pore diameter of 1 nm. At Ih = 50%, the release of silica was controlled by varying the molecular weight of pTMSPMA while retaining a specific surface area above 100 m2 g−1. Strain to failure increased to 14.2%, for Ih = 50% using a polymer of 30 kDa, compared to 4.5% for pure glass. The modulus of toughness (UT) increased from 0.73 (pure glass) to 2.64 GPa. Although, the hybrid synthesised in this report did not contain calcium, pTMSPMA/SiO2 hybrid was found to nucleate bone-like mineral on its surface after 1 week of immersion in simulated body fluid (SBF), whereas pure silica sol–gel glass did not. This increase in apatite forming ability was due to the ion–dipole complexation of calcium with the ester moieties of the polymer that were exposed after release of soluble silica from TEOS. No adverse cytotoxicity for MC3T3-E1 osteoblast-like cells was detected and improved cell attachment was observed, compared to a pure silica gel. pTMSPMA/SiO2 hybrids have potential for the regeneration of hard tissue as they overcome the major drawbacks of pure inorganic substrates while retaining cell attachment.

Journal article

Chung J, Li S, Stevens MM, Georgiou T, Jones JRet al., 2016, Tailoring mechanical properties of sol-gel hybrids for bone regeneration through polymer structure, Chemistry of Materials, Vol: 28, Pages: 6127-6135, ISSN: 1520-5002

Bioglass® was the first synthetic biomaterial that formed a chemical bond to bone. Although bioactive glassscaffolds can mimic bone’s porous structure, they are brittle. Sol-gel derived hybrids could overcome this problem becausetheir nanoscale co-networks of silica and organic polymer have the potential to provide unique physical propertiesand controlled homogenous biodegradation. Copolymers of methyl methacrylate (MMA) and 3-(trimethoxysilyl)propylmethacrylate (TMSPMA) has been used as an organic source for hybrids to take advantage of its self-hardening property.However, the effect of well-defined poly(MMA-co-TMSPMA) architecture in the hybrid system has not been investigated.Here, linear, randomly branched and star shaped methacrylate based copolymers were synthesized via reversible addition-fragmentationchain transfer (RAFT) polymerization method. These copolymers were then used to fabricate hybrids.The 3-D polymer structure had a significant effect on mechanical properties, providing higher strain to failure whilemaintaining a compressive strength similar to sol-gel glass. Star copolymer-SiO2 hybrids had a modulus of toughness 9.6fold greater, and Young’s modulus 4.5 fold lower than a sol-gel derived bioactive glass. During in vitro cell culture,MC3T3-E1 osteoblast precursor cells adhered on the surface regardless of the polymer structure. Introducing star polymersto inorganic-organic hybrids opens up possibilities for the fine-tuning physical properties of bone scaffold materials

Journal article

Reznikov N, Steele JAM, Fratzl P, Stevens MMet al., 2016, A materials science vision of extracellular matrix mineralization, Nature Reviews Materials, Vol: 1, ISSN: 2058-8437

From an engineering perspective, skeletal tissues are remarkable structures because they are lightweight, stiff and tough, yet produced at ambient conditions. The biomechanical success of skeletal tissues is largely attributable to the process of biomineralization — a tightly regulated, cell-driven formation of billions of inorganic nanocrystals formed from ions found abundantly in body fluids. In this Review, we discuss nature's strategies to produce and sustain appropriate biomechanical properties in mineralizing (by the promotion of mineralization) and non-mineralizing (by the inhibition of mineralization) tissues. We review how perturbations of biomineralization are controlled over a continuum that spans from the desirable (or defective in disease) mineralization of the skeleton to pathological cardiovascular mineralization, and to mineralization of bioengineered constructs. A materials science vision of mineralization is presented with an emphasis on the micro- and nanostructure of mineralized tissues recently revealed by state-of-the-art analytical methods, and on how biomineralization-inspired designs are influencing the field of synthetic materials.

Journal article

Mawad D, Artzy-Schnirman A, Tonkin J, Ramos J, Inal S, Mahat M, Darwish N, Zwi-Dantsis L, Malliaras G, Gooding J, Lauto A, Stevens MMet al., 2016, Electroconductive Hydrogel Based on Functional Poly(EthylenedioxyThiophene), Chemistry of Materials, Vol: 28, Pages: 6080-6088, ISSN: 1520-5002

Poly(ethylene dioxythiophene) with functional pendant groups bearing double bonds is synthesized and employed for the fabrication of electroactive hydrogels with advantageous characteristics; covalently crosslinked porous 3D scaffolds with notable swelling ratio, appropriate mechanical properties, electroactive in physiological conditions, and suitable for prolifera-tion and differentiation of C2C12 cells. This is a new approach for the fabrication of conductive engineered constructs.

Journal article

Nitiputri K, Ramasse QM, Autefage H, McGilvery CM, Boonrungsiman S, Evans ND, Stevens MM, Porter AEet al., 2016, Nanoanalytical Electron Microscopy Reveals a Sequential Mineralization Process Involving Carbonate-Containing Amorphous Precursors, ACS Nano, Vol: 10, Pages: 6826-6835, ISSN: 1936-086X

A direct observation and an in-depth characterization of the steps by which bone mineral nucleates and grows in the extracellular matrix during the earliest stages of maturation, using relevant biomineralization models as they grow into mature bone mineral, is an important research goal. To better understand the process of bone mineralization in the extracellular matrix, we used nanoanalytical electron microscopy techniques to examine an in vitro model of bone formation. This study demonstrates the presence of three dominant CaP structures in the mineralizing osteoblast cultures: <80 nm dense granules with a low calcium to phosphate ratio (Ca/P) and crystalline domains; calcium phosphate needles emanating from a focus: “needle-like globules” (100–300 nm in diameter) and mature mineral, both with statistically higher Ca/P compared to that of the dense granules. Many of the submicron granules and globules were interspersed around fibrillar structures containing nitrogen, which are most likely the signature of the organic phase. With high spatial resolution electron energy loss spectroscopy (EELS) mapping, spatially resolved maps were acquired showing the distribution of carbonate within each mineral structure. The carbonate was located in the middle of the granules, which suggested the nucleation of the younger mineral starts with a carbonate-containing precursor and that this precursor may act as seed for growth into larger, submicron-sized, needle-like globules of hydroxyapatite with a different stoichiometry. Application of analytical electron microscopy has important implications in deciphering both how normal bone forms and in understanding pathological mineralization.

Journal article

Ren J, Blackwood KA, Doustgani A, Poh PP, Steck R, Stevens MM, Woodruff MAet al., 2016, Melt-electrospun polycaprolactone strontium-substituted bioactive glass scaffolds for bone regeneration., Publisher: Wiley

Other

Macon ALB, Jacquemin M, Page SJ, Li S, Bertazzo S, Stevens MM, Hanna JV, Jones JRet al., 2016, Lithium-silicate sol-gel bioactive glass and the effect of lithium precursor on structure-property relationships, Journal of Sol-Gel Science and Technology, Vol: 81, Pages: 84-94, ISSN: 1573-4846

This work reports the synthesis of lithium-silicate glass, containing 10 mol% of Li 22 O by the sol–gel process, intended for the regeneration of cartilage. Lithium citrate and lithium nitrate were selected as lithium precursors. The effects of the lithium precursor on the sol–gel process, and the resulting glass structure, morphology, dissolution behaviour, chondrocyte viability and proliferation, were investigated. When lithium citrate was used, mesoporous glass containing lithium as a network modifier was obtained, whereas the use of lithium nitrate produced relatively dense glass-ceramic with the presence of lithium metasilicate, as shown by X-ray diffraction, 2929 Si and 77 Li MAS NMR and nitrogen sorption data. Nitrate has a better affinity for lithium than citrate, leading to heterogeneous crystallisation from the mesopores, where lithium salts precipitated during drying. Citrate decomposed at a lower temperature, where the crystallisation of lithium-silicate crystal is not thermodynamically favourable. Upon decomposition of the citrate, a solid-state salt metathesis reaction between citrate and silanol occurred, followed by the diffusion of lithium within the structure of the glass. Both glass and glass-ceramic released silica and lithium ions in culture media, but release rate was lower for the glass-ceramic. Both samples did not affect chondrocyte viability and proliferation.

Journal article

Liu NJ, Chapman R, Lin Y, Bentham A, Tyreman M, Philips N, Khan SA, Stevens MMet al., 2016, Phospholipase A2 as a point of care alternative to serum amylase and pancreatic lipase, Nanoscale, Vol: 8, Pages: 11834-11839, ISSN: 2040-3372

Acute pancreatitis is a relatively common and potentially fatal condition, but the presenting symptoms are non-specific and diagnosis relies largely on the measurement of amylase activity by the hospital clinical laboratory. In this work we develop a point of care test for pancreatitis measuring concentration of secretory phospholipase A2 group IB (sPLA2-IB). Novel antibodies for sPLA2-IB were raised and used to design an ELISA and a lateral flow device (LFD) for the point of care measurement of sPLA2-IB concentration, which was compared to pancreatic amylase activity, lipase activity, and sPLA2-IB activity in 153 serum samples. 98 of these samples were obtained from the pathology unit of a major hospital and classified retrospectively according to presence or absence of pancreatitis, and the remaining 55 were obtained from commercial sources to serve as high lipase (n = 20), CA19-9 positive (n = 15), and healthy (n = 20) controls. sPLA2-IB concentration correlated well with the serum activity of both amylase and lipase, and performed at least as well as either markers in the differentiation of pancreatitis from controls.

Journal article

Shevchuk A, Tokar S, Gopal S, Sanchez Alonso JL, Tarasov A, Vélez-Ortega AC, Chiappini C, Rorsman P, Stevens MM, Gorelik J, Frolenkov GI, Klenerman D, Korchev Yet al., 2016, Angular approach Scanning Ion Conductance Microscopy, Biophysical Journal, Vol: 110, Pages: 2252-2265, ISSN: 1542-0086

Scanning ion conductance microscopy (SICM) is a super-resolution live imagingtechnique that uses a glass nanopipette as an imaging probe to produce 3D images of cell surface.SICM can be used to analyze cell morphology at nanoscale, follow membrane dynamics, preciselyposition an imaging nanopipette close to a structure of interest, and use it to obtain ion channelrecordings or locally apply stimuli or drugs. Practical implementations of these SICM advantages,however, are often complicated due to the limitations of currently available SICM systems that“inherited” their design from other scanning probe microscopes in which the scan assembly isplaced right above the specimen. Such arrangement makes the setting of optimal illuminationnecessary for phase contrast or the use of high magnification upright optics difficult. Here wedescribe the designs that allow mounting SICM scanhead on a standard patch-clampmicromanipulator and imaging the sample at an adjustable approach angle. This angle could be asshallow as the approach angle of a patch-clamp pipette between a water immersion objective andthe specimen. Using this angular approach SICM, we obtained topographical images of cells grownon non-transparent nanoneedle arrays, of islets of Langerhans, and of hippocampal neurons under 2upright optical microscope. We also imaged previously inaccessible areas of cells such as the sidesurfaces of the hair cell stereocilia and the intercalated disks of isolated cardiac mocytes, andperformed targeted patch-clamp recordings from the latter. Thus, our new angular approach SICMallows imaging of living cells on non-transparent substrates and a seamless integration with mostpatch-clamp setups on either inverted or upright microscopes, which would facilitate research incell biophysics and physiology.

Journal article

Stevens MM, Parmar P, St-Pierre J, Chow L, Puetzer J, Stoichevska V, Peng Y, Werkmeister J, Ramshaw Jet al., 2016, Harnessing the versatility of bacterial collagen to improve the chondrogenic potential of porous collagen scaffolds, Advanced Healthcare Materials, Vol: 5, Pages: 1656-1666, ISSN: 2192-2640

Collagen type I foams have been used extensively in the clinic as scaffolds to promote articular cartilage repair by providing a bioactive environment for cells with chondrogenic potential. However, collagen type I as a base material does not allow for precise control over the bioactive regions interfaced with cells. On the other hand, recombinant bacterial collagens can be used as ‘blank slate’ collagen molecules to offer a versatile platform for controllable incorporation of selected bioactive sequences and fabricated into three–dimensional scaffolds. Here, we demonstrate the potential of Streptococcal collagen–like 2 (Scl2) protein foams modified with peptide sequences designed to specifically and non–covalently bind hyaluronic acid and chondroitin sulfate to significantly improve chondrogenesis of human mesenchymal stem cells (hMSCs), when compared to collagen type I foams. Specific compositions of the functionalized Scl2 proteins led to improved chondrogenesis compared to both non–functionalized Scl2 and collagen type I foams as indicated by differences in gene expression, extracellular matrix accumulation, and compression moduli. Furthermore, hMSCs cultured in functionalized Scl2 foams exhibited decreased collagen types I and X gene and protein expression, suggesting a major advantage over collagen type I foams in promoting an articular chondrocyte phenotype. These highly modular foams can be further modified to improve specific aspects of the chondrogenic response through careful selection of additional biological moieties. As such, these scaffolds also have the potential to be tailored for other regenerative medicine applications.

Journal article

Stevens MM, Gliddon H, Howes P, Kaforou M, Levin Met al., 2016, A nucleic acid strand displacement system for the multiplexed detection of tuberculosis-specific mRNA using quantum dots, Nanoscale, Vol: 8, Pages: 10087-10095, ISSN: 2040-3372

The development of rapid, robust and high performance point-of-care diagnostics relies on the advancement and combination of various areas of research. We have developed an assay for the detection of multiple mRNA molecules that combines DNA nanotechnology with fluorescent nanomaterials. The core switching mechanism is toehold-mediated strand displacement. We have used fluorescent quantum dots (QDs) as signal transducers in this assay, as they bring many benefits including bright fluorescence and multiplexing abilities. The resulting assay is capable of multiplexed detection of long RNA targets against a high concentration background of non-target RNAs, with high sensitivity and specificity and limits of detection in the nanomolar range using only a standard laboratory plate reader. We demonstrate the utility of our QD-based system for the detection of two genes selected from a microarray-derived tuberculosis-specific gene expression signature. Levels of up- and downregulated gene transcripts comprising this signature can be combined to give a disease risk score, making the signature more amenable for use as a diagnostic marker. Our QD-based approach to detect these transcripts could pave the way for novel diagnostic assays for tuberculosis.

Journal article

Paresh P, Skaalure S, Chow L, St-Pierre JP, Stoichevska V, Peng YY, Werkmeister JAM, Stevens MMet al., 2016, Temporally degradable collagen–mimetic hydrogels tuned to chondrogenesis of human mesenchymal stem cells, Biomaterials, Vol: 99, Pages: 56-71, ISSN: 1878-5905

Tissue engineering strategies for repairing and regenerating articular cartilage face critical challenges to recapitulate the dynamic and complex biochemical microenvironment of native tissues. One approach to mimic the biochemical complexity of articular cartilage is through the use of recombinant bacterial collagens as they provide a well–defined biological ‘blank template’ that can be modified to incorporate bioactive and biodegradable peptide sequences within a precisely defined three–dimensional system. We customized the backbone of a Streptococcal collagen–like 2 (Scl2) protein with heparin–binding, integrin–binding, and hyaluronic acid–binding peptide sequences previously shown to modulate chondrogenesis and then cross–linked the recombinant Scl2 protein with a combination of matrix metalloproteinase 7 (MMP7)– and aggrecanase (ADAMTS4)–cleavable peptides at varying ratios to form biodegradable hydrogels with degradation characteristics matching the temporal expression pattern of these enzymes in human mesenchymal stem cells (hMSCs) during chondrogenesis. hMSCs encapsulated within the hydrogels cross–linked with both degradable peptides exhibited enhanced chondrogenic characteristics as demonstrated by gene expression and extracellular matrix deposition compared to the hydrogels cross–linked with a single peptide. Additionally, these combined peptide hydrogels displayed increased MMP7 and ADAMTS4 activities and yet increased compression moduli after 6 weeks, suggesting a positive correlation between the degradation of the hydrogels and the accumulation of matrix by hMSCs undergoing chondrogenesis. Our results suggest that including dual degradation motifs designed to respond to enzymatic activity of hMSCs going through chondrogenic differentiation led to improvements in chondrogenesis. Our hydrogel system demonstrates a bimodal enzymatically degradable biological platform that can mi

Journal article

He M, Callanan A, Lagaras K, Steele JA, Stevens MMet al., 2016, Optimization of SDS exposure on preservation of ECM characteristics in whole organ decellularization of rat kidneys., Journal of Biomedical Materials Research Part B: Applied Biomaterials, Vol: 105, Pages: 1352-1360, ISSN: 1552-4973

Renal transplantation is well established as the optimal form of renal replacement therapy but is restricted by the limited pool of organs available for transplantation. The whole organ decellularisation approach is leading the way for a regenerative medicine solution towards bioengineered organ replacements. However, systematic preoptimization of both decellularization and recellularization parameters is essential prior to any potential clinical application and should be the next stage in the evolution of whole organ decellularization as a potential strategy for bioengineered organ replacements. Here we have systematically assessed two fundamental parameters (concentration and duration of perfusion) with regards to the effects of differing exposure to the most commonly used single decellularizing agent (sodium dodecyl sulphate/SDS) in the perfusion decellularization process for whole rat kidney ECM bioscaffolds, with findings showing improved preservation of both structural and functional components of the whole kidney ECM bioscaffold. Whole kidney bioscaffolds based on our enhanced protocol were successfully recellularized with rat primary renal cells and mesenchymal stromal cells. These findings should be widely applicable to decellularized whole organ bioscaffolds and their optimization in the development of regenerated organ replacements for transplantation. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2016.

Journal article

Fiocco L, Li S, Bernardo E, Stevens MM, Jones JRet al., 2016, Highly porous polymer-derived wollastonite–hydroxycarbonate apatite ceramics for bone regeneration, Biomedical Materials, Vol: 11, ISSN: 1748-605X

Journal article

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